AU2019247099A1 - Methods and kits for detecting an infectious agent - Google Patents

Abstract

Abstract The present invention provides methods and kits for determining the presence, absence, or level of an infectious agent in a sample. Specifically, the present invention provides methods and kits for detecting or quantifying certain target polynucleotides of the infectious agent. In certain embodiments, the present invention provides for such detection without the need for amplification (e.g., replication) of the target molecule and/or without the need for labor intensive purification procedures. In certain embodiments, the present invention provides positive control and housekeeping gene for normalization and quantatively detection of the copy numbers of infectious agent in a sample. In these or other embodiments, the invention allows for such detection with the desired sensitivity and/or specificity, even where the polynucleotide is present in the sample at low copy number.

Description

FIELD OF THE INVENTION [0001] This invention relates to detecting the presence, absence, or level of an infectious agent in a sample.

BACKGROUND OF THE INVENTION [0002] Although many tests have been developed to diagnose infection, many of these tests are only specific and/or sensitive when there is a high load of the infectious agent present, i.e. in the case of late or advanced stages of infection. In addition, some of these tests require highly technical personnel or specialized knowledge to carry out the tests.

[0003] For example, to date, there is no robust test available for detecting high risk strains of HPV in pap smears, and which is both clinically sensitive and specific, as well as convenient, easy and inexpensive, without the need for highly trained personnel. (Kurtycz et al., Comparison of Methods Trial for High-Risk HPV Diagnostic Cytopathology 38: 104-108 (2009); Ginocchio et al., Comparison of the Third Wave Invader Human Papilloma (HPV) Assay and the Digene HPV Hybrid Capture 2 Assay for Detection of High-Risk HPV DNA J. Clin. Microbiol. 46: 1641-1646 (2008)). The Pap or Papanicolau test is currently the test of choice for the initial screening of pre-malignant or malignant cervical cancers caused by HPV. The Pap test is a cytology test which requires highly trained personnel to discriminate between normal cells and cells undergoing malignant lesions under a light microscope. The Diagene-HC2 DNA test, which is the only FDA-market approved HPV test, relies on the capture of DNA/RNA hybrids on a solid phase. The captured DNA/RNA hybrids are detected using an enzyme-linked antibody upon amplification. Although the test can detect up to thirteen HPV types, it has poor sensitivity (5000 copies/mL) and specificity. Furthermore, the test requires purification of DNA from the sample and it cross-hybridizes with low-risk and high-risk HPV types when the viral load in the sample is high.

[0004] Other tests for HPV include GenProbe's Aptima HPV test, Third Wave's Invader HPV DNA test, Ventana's Inform 1 HPV in situ hybridization test, and other PCR-based tests, including Roche's Amplicor or Linear Array HPV DNA tests. These tests require purification of the nucleic acid from the samples and/or amplification of the target nucleic

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PCT/US2010/040649 acids prior to detection, which can result in poor sensitivity or specificity. In addition, these tests can be expensive and time-consuming. Furthermore, since most of these tests require amplification of the target polynucleotides, a highly clean and dedicated environment is required to ensure that there is no cany over or cross contamination between samples.

2019247099 14 Oct 2019 [0005] Accordingly, there is a need for an efficient, rapid, reliable, and inexpensive method and/or assay for detecting an infectious agent, such as HPV, and which lias the desired level of sensitivity and specificity for screening and/or evaluating samples for the presence or level of the infectious agent.

SUMMARY OF THE INVENTION [0006] The present invention provides methods and kits for determining the presence, absence, or level of an infectious agent in a sample. Specifically, the present invention provides methods and kits for detecting or quantifying certain target polynucleotides of the infectious agent. In certain embodiments, the present invention provides for such detection without the need for amplifica tion (e.g., replication) of the target molecule and/or without the need for labor and/or time intensive procedures. In these or other embodiments, the invention allows for such detection with the desired sensitivity and/or specificity, even where the polynucleotide is present in the sample at low copy number.

[0007] In one aspect, the invention provides a method for detecting the presence, absence, or level of an infectious agent in a sample, such as a biological sample. The method comprises capturing a target polynucleotide, if present, from the sample, where the target polynucleotide is indi cati ve of the presence of the infectious agent. The target polynucleotide is then detected directly or indirectly by hybridization with one or a series of signalamplifying polynucleotide probes (e.g., branched DNA.), The method allows detection of the target polynucleotide, with the desired specificity and/or sensitivity, and at low copy number, to thereby allow for detection of even early stage infection.

[0008] The target polynucleotide may be captured from the sample by hybridization to a Capture Extender probe having at least a first sequence and a second sequence, the first sequence hybridizing to the target polynucleotide and the second sequence hybridizing to an immobilized capture probe. The capture probe may be immobilized via any solid support (e.g., chip, bead, or well).

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2019247099 14 Oct 2019 [0009] The captured target polynucleotide is then detected by hybridizing one or a series of signal-amplifying polynucleotide probe(s) directly or indirectly to the target. The series of signal-amplifying probes may comprise branched DNA. For example, the series of signalamplifying probes may comprise a pre-Amplifier probe, an Amplifier probe, and a Label probe. Specifically, the pre-Amplifier probe hybridizes to the target through a Label Extender probe. The Label Extender probe generally has a first sequence and a second sequence, the first sequence hybridizing to the target polynucleotide and the second sequence hybridizing to the pre-Amplifier probe. Consecutive hybridizations of Amplifier probes and Label probes may then be conducted to amplify the signal. The signal-amplifying probes may allow for amplification of the signal 40 times or more, and in some embodiments, 200 times or more (e.g., relative to the number of hybridized Label Extender probes).

[0010 j In various embodiments, the target polynucleotide or biomarker is a polynucleotide involved in the replication machinery of the infectious agent (e.g., cis- or trans-acting factor), a polynucleotide encoding a protein involved in replication or transcription, or a polynucleotide encoding a cell surface protein, integral membrane protein, etc. For example, in the case of HPV the target polynucleotides may be independently selected from E6 and E7 sequences. Exemplary Capture Extender and Label Extender probes for various infectious agent targets, including HPV, HBV, HIV, influenza H1N1, HCV, SARS, Mycobacterium, Syphilis, Aspergillus, Candida, and Cryptococcus are described herein.

[0011] In another aspect, the present invention provides a kit for detecting target polynucleotides. The kit provides one or more Capture Extender probes and one or more Label Extender probes for the detection of particular target polynucleotide(s), and exemplary pairs or “sets” of Capture Extender probes and Label Extender probes are disclosed herein. In some embodiments, the kit may further comprise one or more capture probes, a solid support, and/or a signal amplifying probe set (e.g., the sei comprising a pre-Amplifier probe, an Amplifier probe, and a Label probe). The kit may comprise (in addition to the Capture Extender and Label Extender probes for detecting the target polynucleotide), Capture Extender and Label Extender probes for detecting one or more positive and/or negative control sequences. The positive control probes may be used to normalize between samples and/or to calculate the copy number of polynucleotides present. Exemplary positive control probes may comprise sequences from housekeeping genes, such as but not limited to glyceraldehyde 3-phosphate dehydrogenase (GAPDH/GAPD).

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PCT/US2010/040649 [0012] Other aspects and embodiments of the invention will be apparent to the skilled artisan in view of the following detailed description.

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BRIEF DESCRIPTION OF THE DRAWI NGS [0013] FIG. 1 (A, B, and C) illustrates the components of the test system for detecting target polynucleotides in connection with the invention.

[0014] FIG. 2 shows a standard curve for detection of HPV type 16 ssDNA.

f0015| FIG. 3 shows the relative light units (RLU) emitted from Alkaline Phosphataselabeled probe, for detecting HPV biomarkers in Pap smears of HPV patients during various disease states. AUCUS is atypical squamous cells of undetermined significance. CEN is cervical intraepithelial neoplasia. BK is background.

[0016] FIG. 4 shows detection of HIV target polynucleotides that are present in amounts as low as 25 copies.

[0017] FIG. 5 shows a standard curve for the detection for HBV polynucleotides.

[0018] FIG. 6 shows a standard curve for the detection of HCV (lop panel), and quantitation of HC V particles from sera of six patients (bottom panel).

[0019] FIG. 7 shows a standard curve for the detection of SARS.

[0020] FIG. 8A and B shows standard curves for the detection of ssDNA complementary to 18S RNA from fungal cells (Candida albicans).

[0021] FIG. 9 shows quantifying different species and strains of Aspergillus and Candida.

[0022] FIG. 10 shows quantifying Candida from three clinical blood samples.

DETAILED DESCRIPTION OF THE INVENTION [0023] T he present invention provides methods and kits for determining the presence, absence, or level of an infectious agent in a sample. Specifically, the present invention provides methods and kits for detecting or quantifying certain target polynucleotides of the infectious agent. In certain embodiments, the present invention provides for such detection

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PCT/US2010/040649 without the need for amplification (e.g., replication) of the target molecule and/or without the need for labor and/or time intensive procedures, such as nucleic acid purification steps. In these or other embodiments, the invention allows for rapid detection of the target, with the desired sensitivity and/or specificity, even where the polynucleotide is present in the sample at low copy number.

2019247099 14 Oct 2019 [0024] In one aspect, the invention provides a method for detecting the presence, absence, or level of an infectious agent, such as a biological sample. The method comprises capturing one or more target polynucleotide/s), if present, from the sample, wherein the target polynucleotide is indicative of the presence of the infectious agent, In one embodiment, a single polynucleotide of interest is detected (single plex detection). In another embodiment, two polynucleotides of interest are detected (two-plex detection), or more than two polynucleotides of interest are detected (multiplex detection). The target polynucleotide is then detected directly or indirectly by hybridization with one or a series of signal-amplifying polynucleotide probes. The method may further comprise detecting the level of one or more control polynucleotides, for example, to normalize signals across samples.

[0025] For example, the target polynucleotide may be captured by hybridization to a Capture Extender probe having at least a first sequence and a second sequence, the first sequence hybridizing to the target polynucleotide and the second sequence hybridizing to an immobilized capture probe. The capture probe may be immobilized via any solid support, including but not limited to a chip (e.g., an array), well, bead, or other solid support or matrix.

[0026] The captured target polynucleotide is then detected by hybridizing one or a series of signal-amplifying polynucleotide probes, either directly to the target polynucleotide, or indirectly through a Label Extender probe. A Label Extender probe generally has a first sequence and a second sequence, the first sequence hybridizing to the target polynucleotide and the second sequence hybridizing to a signal-amplifying polynucleotide probe. The signal-amplifying probe may comprise branched DNA, e.g., may include a pre-Amplifier probe, an Amplifier probe, and a Label probe.

[0027] The sample may be a liquid sample, or a biological sample, such as, for example, a body fluid. In various embodiments, the sample is blood, plasma, serum, urine, vaginal secretion, pap smear, semen, nasal swabbing, lung lavage, pleural effusion, sputum, and throat swab. In other embodiments liquid samples can be obtained from swabs of objects enrh a« tnilM, seats, pond water, tree sap, and insect or plant extracts. Other types of samples s

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PCT/US2010/040649 for which detection of infectious agents is desired may be employed in connection with the invention. In certain embodiments, a nucleic acid purification step may be performed prior to detection, such as the use of any commercially available filter-based method or kit for purification of nucleic acids. In other embodiments, no such purification step is required.

2019247099 14 Oct 2019 [0028] The method of detection may be carried out using the principles set forth in U.S. Patent Number 7,709,198, which is hereby incorporated by reference. For example, detection generally takes place with the use of a series of signal amplifying polynucleotide probes (e.g., branched DNA), which is hybridized directly to the target polynucleotide, or indirectly through a series of hybridization reactions. The signal-amplifying probes ob viate the need for thermal cycling or amplification of the target sequences prior to detection. Thus, in such embodiments, the method intensifies the signal of hybridization by multiple layers of probe hybridization, instead of any actual nucleotide sequence amplification of the target polynucleotide.

[0029] In various embodiments, the target polynucleotide (which may be linear or circular) is captured on a solid support by hybridizing to one or more sets of probes. The set of probes generally comprises a Capture Extender and a Label Extender, and optionally a Blocking Label Extender (or blocking probe).

[0030] The Capture Extender indirectly captures the target polynucleotide by hybridizing, simultaneously, to the target polynucleotide and a Capture Probe attached to a solid support, e.g., bead, chip, etc. The Capture Extender generally has a first sequence that hybridizes to the target polynucleotide and a second sequence that hybridizes to the Capture Probe. The Capture Extender optionally comprises a linking sequence between the first sequence and the second sequence. The first sequence of the Capture Extender, which hy bridizes to the target polynucleotide, is generally from about 15 to about 30 nucleotides in length, or about 20 to about 27 nucleotides in length. The sequence and length is generally selected to hybridize under the hybridization conditions selected or described herein. The second sequence of the Capture Extender, which may hybridize to the capture probe, may be any capturable sequence, but is generally selected to hybridize under the selected hybridization conditions. The capture sequence is generally about 10 to about 30 nucleotides in length, and exemplary capture sequences are disclosed herein. The linking sequence is selected to provide for the physical independence of the first and second sequences, and is selected to avoid significant

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2019247099 14 Oct 2019 structural constraints. The linking sequence may be short, for example, from about 2 to about nucleotides. The linking sequence may be poly(T) or poly(Aj in certain embodiments.

[0031] Various Capture Extender sequences (and sets of Label Extender probes) are disclosed herein for the detection of numerous infectious agents. See Tables 3, 6, 9, 12, 15, 18, 21,24, 27, 30, 33, 36, 39, 42, 45, 48, 51, 54, 57, 60, 63, 66, 69, 72, 75, 78, 81, 84, 87, 90, and 93. It is understood that various modifications of these sequences may be made within the spirit of the invention, such as the addition or deletion of from 1 to 4 nucleotides to any or each of the target-hybridizing sequence, capture sequence, and/or linking sequence, or the addition of from 1 to 5 degenerate nucleotides.

[0032] A Blocking Label Extender may optionally be employed to block certain sequences. Such optional blocking probes (and sets of Blocking probes) for use with the invention are described in Tables 5, 8, 11, 14, 17, 20, 23, 26, 29, 32, 35, 38, 41, 44, 47, 50, 53, 56, 59, 62, 65, 68, 71, 74, 77, 80, 83, 86, 89, 92, and 95. It is understood that various modifications of these sequences may be made within the spirit of the invention, such as the addition or deletion of from 1 to 4 nucleotides, or the addition of from 1 to 5 degenerate nucleotides.

[0033] The Label Extender hybridizes to the target polynucleotide and a signalamplifying polynucleotide simultaneously. For example, the Label Extender generally comprises a first sequence that hybridizes to the target sequence, and a second sequence that hybridizes to a signal amplifying probe. The Label Extender may optionally have a linking sequence. The first sequence which hybridizes to the target sequence may generally be from about 15 to about 30 nucleotides in length, or about 20 to about 27 nucleotides in length. The sequence and length of the first sequence is generally selected to hybridize under the hybridization conditions selected or described herein. The second sequence of the Label Extender, which may hybridize to the signal amplifying probe, is generally selected to hybridize under the selected hybridization conditions. The second sequence is generally about 10 to about 30 nucleotides in length, and exemplary sequences are disclosed herein. The linking sequence is selected to provide for the physical independence of the first and second sequences, and is selected to avoid significant structural constraints. The linking sequence may be short, for example, from about 2 to about 10 nucleotides. The linking sequence may be poly(T) or poly(A) in certain embodiments.

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2019247099 14 Oct 2019 [0034] Various Label Extender sequences (including Label Extender probe sets) for the detection of certain infectious agents are disclosed herein. See Tables 1, 2, 4, 7, 10, 13, 16, 19, 22, 25, 28, 31, 34, 37, 40, 43, 46, 49, 52, 55, 58, 61, 64, 67, 70, 73, 76, 79, 82, 85, 88, 91, and 94. It is understood that various modifications of these sequences may be made within the spirit of the invention, such as the addition or deletion of from 1 to 4 nucleotides to any or each of the target-hybridizing sequence, capture sequence, anchor linking sequence, or the addition of from 1 to 5 degenerate nucleotides.

[0035] The hybridization conditions for the assay include selected temperature, time, and reagents so as to allow for rapid, sensitive, and/or specific detection. For example, the hybridization temperatures may be independently selected in the range of about 45°C to about 65°C, or about 50°C to about 60°C, such as about 55°C. The hybridization buffer is generally 3x SSC, or comparable hybridization buffer.

[0036] 'rhe length of time for hybridization of the Capture Extender and the Label Extender probes may be selected based on the type of virus suspected, the stage of infection, the copy number of virus present in the sample, etc. In some embodiments, the Capture Extender and Label Extender probes are allowed to hybridize to the target for at least about 5 minutes, such as within the range of about 10 minutes to about 120 minutes, such as about 20 to about 90 minutes, or about 30 or 60 minutes. In certain embodiments, the Capture Extender and Label Extender probes are allowed to hybridize to the target for about 60 minutes to about 4 hours, 8 hours, 12 hours or overnight (e.g., from 18 to 24 hours). With samples suspected of representing early stages of an infection (or where the copy number of target polynucleotides is expected to be low), the Capture Extender and the Label Extender probes are allow to hybridize to the target for a longer period of time, than for samples suspected of representing advanced stages of infection (or where a high copy number of the target polynucleotides is expected). For example, the hybridization time may be from about 1 hour to about 4 hours, from about 3 hours to 8 hours, from about 7 hours to about 12 hours, or from about 12 hours to about 24 hours or longer for samples obtained from the early stages of infection or where the copy number of infectious agents are low. Where the copy number of infectious agents are high or during the late or advance stages of infection, the length of time needed for the probes to hybridize to the target polynucleotide may be from about 5 minutes to about one hour.

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2019247099 14 Oct 2019 [0037] A robust detection of the hybridization reactions is obtained by either directly detecting the multiple hybridizations of Label Extenders to the target polynucleotide, or by further hybridization of one or a series of signal-amplifying probes. For example, detection may occur by hybridizing a pre-Amplifier probe to the label extender probe, and hybridizing Amplifier probes to the pre-Amplifier probe, which complex may be detected with Label probes. Such reactions may take place in consecutive hybridizations, e.g., at conditions in the range of 40 to 60° C, in 3x SSC (or comparable hybridization buffer), and for about 30 to about 60 minutes each. Exact conditions may be selected based upon the sequences and melting temperature of each probe. Exemplary sequences/structures for the pre-Amplifier probe, Amplifier probe, and label probes are known. For illustration, an exemplary sequence of the preamplifier is 5’

AGGCATAGGACCCGTGTCTttttttttttAGGCATAGGACCCGTGTCTtttttATGCTTTGACT CAG AAAACGGTAACTTC 3’ (SEQ ID NO:1). The underlined sequences are complementary to sequences in the GAPD Label Extenders (used as a positive control as described herein).

[0038] The configuration and hybridization of test components, such as Label Extenders, may be as described in U.S. Patent No 7,709,198, which is hereby incorporated by reference. For example, Label Extenders may be described with reference to the “cruciform” configuration or the “double Z” configuration. See FIG. 1C. Generally, one end of the Label Extender hybridizes to the preamplifier probe, and the other end hybridizes to the target polynucleotide. The pre-amplifier probe may hybridize to a single Label Extender probe as described herein (e.g., cruciform configuration). In some embodiments using the double Zconfiguration, the preamplifier probe hybridizes to two Label Extender probes.

[0039] An exemplary Label Extender probe set in the cruciform configuration is set-forth in Table 1 and Table 94, for glyceraldehyde 3-phosphate dehydrogenase (GAPD), which may be used as a control for normalizing across samples, and for quantifying polynucleotide copy number.

[0040] The sequences complementary to the preamplifier sequence of SEQ ID NO: 1 is underlined for each of the Label Extenders shown in Table 1.

Table 1. GADP Label Extenders with Cruciform Configuration

I SEQ ID NO;

I

PROBE

IDENTIFIER

LABEL EXTENDER NUCLEOTIDE SEQ

OF GLYCERALDEHYDE 3-PHOSPHATE

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		DEHYDROGENASE
SEQ ID NO:2	GAPD 127	ccagtggactccacgacgtacTTTTTgaagttaccgtttt	CPI tail
SEQ ID NO:3	GAPD 128	ctgagtcaaagcat’1T'TTTttctccatggtggtgaagacg	CP2 head
SEQ ID NO:4	GAPD 129	tcttgaggctgttgtcatacttctTTTTTgaagttaccgtttt	CPI tail
SEQ ID NO:5	GAPD130	ctgagtcaaagcatTTTTTgcaggaggcattgctgatga	CP2 head
i SEQ IDNO:6	GAPD131	cagtagaggcagggatgatgttcTTTTTgaagttaccgtttt	CPI tail
SEQ ID NO:7	GAPD 132	ctgagtcaaagcafΤΙ TTTcacagccttggcagcgc	CP2 head

[0041] Table 2 shows an exemplary Label Extender probe set for GAPD in the double Z configuration. The sequences complementary to the preamplifier sequence is underlined for each of the Label Extenders shown in Table 2.

Table 2. GAPD Label Extenders with Dnsible Z configuration

SEQ ID NO: ί i i	PROBE IDENTIFIER	LABEL EXTENDER NUCLEOTIDE SEQ OF GLYCERALDEHYDE 3-PHOSPHATE DEHYDROGENASE
i SEQ ID NO:8	GAPD217	ccagtggactccacgacgtacTTTTTgaagttaccgtttt	CPI tail
ί SEQ IDNO:9	GAPD218	ttctccatggtggtgaagacgTTTTTctgagtcaaagcat	CP2 tail
SEQ ID NO: 10	GAPD219	tcttgaggctgltgtcatacttctTTTTTgaagttac c gtttt	CPI tail
| SEQ ID NO: 11	GAPD220	gcaggaggcattgctgatgaTTTTTctgagtcaaagcal	CP2 tail
| SEQ ID NO: 12	GAPD221	cagtagaggcagggatgatgttcTTTTTgaagttaccgtttt	CPI tail
| SEQ ID NO: 13	GAPD222	cacagccttggcagcgcTTTTTctgagtcaaagcat	CP2 tail

[0042] The Amplifier probe or Label probe, in particular embodiments, is conjugated with Alkaline Phosphatase, allowing its detection with glow luminescence. Alternatively, an Amplifier probe or label probe may have a chromogenic or fluorescent generating moiety (e.g., enzyme) to provide an intensified signal in the presence of the appropriate substrate. An exemplary format is shown in Figure 1, illustrating an Alkaline Phosphatase Label probe and corresponding luminescent substrate.

[0043] The number of Amplifier probes that hybridize to the growing complex, per the number of hybridized pre-Amplifier probes, may be within the range of 5:1 to 20:1. The number of Label probes that hybridize to the growing complex, per the number of hybridized Amplifier probes, may be withm the range of 2:1 to about 5:1.

[0044] The Capture Extender and the Label Extender are designed to comprise at least one sequence that recognizes (hybridizes) to the target polynucleotide. The level of

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PCT/US2010/040649 complementarity is appropriately selected based upon the desired hybridization conditions and selectivity, and in some embodiments, the complementary sequence is fully complementary to its intended target. In certain embodiments, the Capture Extender and/or

Labe! Extender includes from 1 to about 5 degenerate nucleotides in the target-hybridizing sequence, so as to provide for the desired selectivity or cross-reactivity.

2019247099 14 Oct 2019 [0045] The Capture Extender and Label Extender may be designed to recognize and bind to such target sequences as inverted terminal repeats (e.g., of a virus genome); bacterial 5S, 16S, or 23 S ribosomal RNA sequences, terminal inverted repeat sequences (TIR), miniature inverted repeat transposable elements (MITE), CpG sequences in prokaryotic cells such as bacteria; or 18S or 28S ribosomal RNA sequences in eukaryotic cells, such as fungi or parasites.

[0046] In various embodiments, the sensitivity of the detection is less than about 1000 copies, less than about 500 copies, less than about 250 copies, or less than about 100 copies (e.g., about 25 copies) of the target sequence. The full test may take less than about 7 hours, and in some embodiments, may take less than about 6 hours, or less than about 5 hours.

[0047] Detection moeities for the target polynucleotide and controls may be the same or different, and may be independently selected from a luminescence-generating moiety, such as but not limited to alkaline phosphatase having a luminescent substrate, and a chromogenic generating moiety, such as but not limited to a horseradish peroxidase (HRP). In some embodiments, detection of the one or more target polynucleotides may be earned out simultaneously or sequentially with the detection of the positive control or normalization control (e.g., GAPDH), that is, in the same or parallel reaction. In some embodiments, detection signals for the target polynucleotide and the normalization control may be read sequentially from a patient sample in the same reaction. In such embodiments, after detection of a luminescent or chromogenic reaction, the sample is washed, and a different luminescent or chromogenic reagent is added.

[0048] In various embodiments, the target polynucleotide is indicative of the presence of a virus of the family Paramyxoviridae (e.g., parainfluenza, mumps, measles); Orthomyxoviridae (e.g., influenza); Hepadnaviridae (e.g., hepatitis); Adenoviridae (e.g., acute respiratory disease); Poxviridae (e.g., small pox); Herpesviridae (e.g., HSV, Varicella Zoster, Karposi sarcoma); Papillomaviridae (e.g., HPV); Polyomaviridae (e.g., cystitis or miM nr acute respiratory diseases); Parvoviridae; Rhabdoviridae (e.g., rabies); Filoviridae 11

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2019247099 14 Oct 2019 (e.g., hemorrhagic fever caused by Ebola virus and Marburg virus); Bunyaviridae (e.g., encephalitis. Hantavirus respiratory syndrome, Rift Valley fever); Arenaviridae (e.g., aseptic meningitis, encephalitis, meningoencephalitis, Lassa fever); Coronaviridae (e.g., severe acute respiratory syndrome or SARS); Flaviviradae (e.g., Dengue hemorrhagic fever); Togaviridae; Picomaviridae; Caliciviridae (e.g., winter vomiting disease); Astroviridae (e.g., gastroenteritis); Retroviridae (e.g., HIV, HTLV) and Reoviridae (e.g., Colorado Tick fever).

[0049] In certain embodiments, the target polynucleotide is an HPV target polynucleotide, and may be a high risk type HPV, such as type 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, or 68; or may be a low risk type HPV, such as type 6, 11, 40, 42, 43, or 44. High risk sexually transmitted HPV may lead to development of cervical intraepithelial neoplasia (ON), vulvar intraepithelial neoplasia (VIN), penile intraepithelial neoplasia (PIN), and/or anal intraepithelial neoplasia (AIN), whereas low risk sexually transmitted HPV may lead to genital warts and are not life-threatening since they do not lead to cervical cancer. Of the high risk type HPV, types 16 and 18 are the most dangerous since they cause about 70% of cervical cancer. For example, in a NIH study, it was found that 10% of women infected with type 16 or 18 HPV developed advance precancerous cervical disease (CIN3) within 3 years while 20% did so in 10 years, [0050] The invention in certain embodiments allows for the determination of a specific type of HPV present in a sample, and/or distinguishes between early course of the infection, and late or latent stage of HPV infection.

[0051] In various embodiments, the target polynucleotide sequence is a portion of an HPV late viral gene (e.g., LI and/or L2), or an early viral gene (e.g., El and/or E2), which encode proteins responsible for replicating and maintaining the viral DNA as a circular episome. In one exemplary embodiment, the target polynucleotide is a portion of the gene encoding the E6 and/or E7 proteins of the HPV. In particular, the target polynucleotides are derived from the mRNA regions encoding the E6 and/or E7 proteins that mediate degradation of the tumor suppressors, p53 and retinoblastoma (RB), by interfering with the regulation of the cell cycle.

[0052] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence of HPV type 16. In such embodiments, at least one Capture Extender (e.g., at least two, three, four nr fivA comprise a target-hybridizing sequence selected from Table 3 (underlined). At 12

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2019247099 14 Oct 2019 least one Capture Extender may comprise, consist essentially of, or consist of a sequence independently selected from SEQ ID NOS: 14 to 21 (Tables 3). In these or other embodiments, at least one Label Extender (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 4 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 22 to 37 (Table 4). The method may further employ a Blocking Label Extender (BL), e.g., at least one, two, or three BLs each comprising, consisting essentially of, or consisting of a sequence selected from SEQ ID NOS: 38 to 48 (Table 5),

Table 3: Capture Extender of HPV type 16

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF HPV TYPE-16
14.	ctcctgtgggtcctgaaacattTTTTTctcttggaaagaaagt
15.	cacgtcgcagtaactgttgcttTTTTTctcttggaaagaaagt
16.	tgttgttccatacaaactataacaataatTTTTTctcttggaaagaaagt
17.	aatctaacatatattcatgcaatgtaggTTTTTctcttggaaagaaagt
18.	atattgtaatgggctctgtccgTTTTTctcttggaaagaaagt
19.	cccattaacaggtcttccaaagtaTTTTTctcttggaaagaaagt
20.	ggtagattatggtttctgagaacagatTT Τ' ΤΊ ctcttggaaagaaagt
21.	cccgtaccctcttccccattTTTTTctcttggaaggaaagt

Table 4: Label Extender of HPV type 16

SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HPV TYPE-16
22.	gtctgcttttatactaaccggtttcTTTTTgaagttaccgtttt
23.	gcagttctcttttggtgcataaaatTTTTTctgagtcaaagcat
24.	aactgtggtaactttctgggtcgTTTTTgaagttaccgtttt
25.	agttgtttgcagctctgtgcatTTTTTctgagtcaaagcat
26.	ttcccatctctatatactatgcataaatTTTTTgaagttaccgtttt
27.	aacatttatcacatacagcatatggaTTTTTctgagtcaaagcat
28.	cggtttgttgtattgctgttctaaTTTTTgaagttaccgtttt
29.	taatacacctaattaacaaatcacacaaTTTTTctgagtcaaagcat
30.	ggacacagtggcttttgacagtTTTTTgaagttaccgtttl
31.	ccagatgtctttgcttttcttcaTTTTTctgagtcaaagcat
32.	gatetgcaacaagacatacatcgaTTTTTgaagttaccgtttt
33.	tgggtttctctacgtgttcttgalTTTTTctgagtcaaagcat
34.	gagatcagltgtctctggttgcaTTTTTgaagltaccgtttt
35.	RCtgtcatttaattgckyttaacagtaTTTTTctgagtcaaagcat
36.	tcacacttgcaacaaaaggttacaTTTTTgaagttaccgtttt
37.	cgcacaaccgaagcgtagagTTTTTctgagtcaaagcat

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Table 5: Blocking Label Extender of HPV type 16

[ SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HPV TYPE-16
| 38.	gcagtacacacattctaatattatatcatgtat
| 39.	cccgaaaagcaaaglcatatacct
40.	gtctatactcactaatlttagaataaaacttta
i 41.	ttalattalggaatctttgctttttgt
1 42.	C ύ g t ύ C clC ύ lie c ύ C
I 43.	tgtatctccatgcatgattacagc
44.	catctatttcatcctectcctctga
1 45.	gttctgcltgtccagctggac
1 46.	cga.atgtctacgtgt.gtgc tttgta
47.	ggggcacacaattcctagtgtg
[ 48.	ggtacctgcaggatcagccat

[0053] In certain embodiments, the target polynucleotide is an E6 or E7 sequence from HPV type 18. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 6 (underlined). Thus, at least one Capture Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 49 to 56 (Tables 6), At least one Label Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 7 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 57 to 72 (Table 7). The test may further employ Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 73 to 75 (Table 8).

Table 6: Capture Extender of HPV type 18

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF HPV TYPE-18
49.	cgtttttcattaaggtgtctaagtttttTTTTTctcttggaaagaaagt
| 50.	tgctcggttecagcacgTTTTTctcttggaaagaaagt
51.	tgttgccttaggtccatgcataTTTTTctcttggaaagaaagt
i 52.	gctttctactactagcttaattctggcTTTTTctcttggaaagaaagt
! 53.	ciggatcagccattgrtgctt'ΓΤΙΊ Tctcttggaaagaaagt
i 54.	tactacctgctaatcttctatgagcttTTTTTctcttggaaagaaagt
55.	tggccagggaagatccactTTTTTctcttggaaagaaagt
56.	ttgcagctcaatacaaaacggTTTTTctcttggaaagaaagt

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Table 7: Label Extender of'HPV type 18

ί SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HPV TYPE-18
i 57.	cccagctatgttgtggaatcgtTTTTTgaagttaccgtttt
i 58.	aatggcactggcctctatagtgTTTTTctgagtcaaagcat
| 59.	tcgttggagtctttcctgtcgTTTTTgaagttaccgtttt
| 60.	cttaatattatacttgtgtttctc tgc gTTTTTctgagtcaaagcai
1 61.	taaatgcaatacaatgtcttgcaaTTTTTgaagttaccgtttt
ί 62.	ggaatttcattttggggctcTTTTTctgagtcaaagcat
i 63.	gctcgtgacatagaaggtcaaccTTTTTgaagttaccgtttt
| 64.	tttcttcctctgagtcgcttaattTTTTTctgagtcaaagcat
1 65.	atgattaactccatctatttcatcgtTTTTTgaagttaccgtttt
j 66.	cgtcgggctggtaaatgttgTTTTTctgagtcaaagcat
! 67.	gctcgaaggtcgtclgctgaTTTTTgaagttaccglttt
68.	tettcagaaacagctgctggaatTTTTTctgagtcaaagcat
| 69.	cggacacacaaaggacagggTTTTTgaagttaccgtttt
70.	actgctgggatgcacaccaTTTTTctgagtcaaagcat
71.	cgtcagtttcctcalctgaaaacttTTTTTgaagttaccgtttt
72.	cgtcaagtcatttaagct.tggtaccTTTTTctgagtcaaagcat

Table 8: Blocking Label Extender of HPV type 18

SEQ ID NO	BLOCKING LABEL EXTENDER 'NUCLEOTIDE SEQUENCE OF HPV TYPE- 18
| 73.	tgtgtgacgttgtggttcagct
i 74.	ttcacacttacaacacatacacaacat
\Lf	gaattaggagtcagc agtcattattc

[0054] In certain embodiments, the target polynucleotide is an E6 or E7 sequence from HPV type 31. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 9 (underlined). Thus, at least one Capture Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 76 to 82 (Tables 9). At least one Label Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 10 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 83 to 94 (Table 10). The test may further employ Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 95 to 99 (Table 11).

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Table 9: Capture Extender of HPV type 31

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 31
76.	gtctttctgcaggatttttgaacTTTTTctottggaaagaaagt
77.	gcttagttcatgcaatttccgagTTTTTctcttggaaagaaagt
78.	££t£tgttt£tgtta^tgag£tt^TTTTTctcttggaaagaaagt
79.	atotaaattSSfittacttttgMtaa^TTTTTctcttggaaagaaagt
80.	catatacctttgtttgtcaatttttctTTTTTctcttggaaagaaagt
81.	acgcatgtttacacttgggtttTTITrctcttggaaagaaagt
82.	tctgagctgtcgggtaattgcTTTTTctcttg^aagaaagt

Table 10: Label Extender of HPV type 31

SEQ ID NO	LABEL EXTENDER OF HPV TYPE 31
83.	gtagggtatttccaatgccgaTTTTTgaagtt ac c g it it
84.	cagtagacacaattcaatdtagttcatcTTTTT c tgagt c aaagc at
85.	gtggtgtgtcgtccctatatactattTTTTTgaagttaccgtttt
86.	cttaaac^tttgm^ractecgtTTTTTctgagtcaaagcat
87.	acacgttatacacctaattaacaaatcaTTTTTgaagttaccgtttt
88.	tcttctggacacaacggtctttgTTTTTctgagtcaaagcat
89.	tctttttetccaaatgtctttgttttTTTTTgaagttaccgtttt
90.	tt cctcctatg ttgt ggaatc gt tTTTTT ctgagtcaaagcat
91.	cttgcaacgtaggtgtttclccTTTTTgaagttaccgtttt
92.	ctcaggttgcaaatctaacacatagtTTlTTctgagtcaaagcat
93.	ggactgtctatgacatcctcc.tcaTTTTTga agtta ccgtttt
9T	ccggttctgcttgtccagctlTrTTctgagtcaaagcat

Table 11: Blocking Label Extender of HPV type 31

SEQ ID NO	BLOCKING EXTENDER OF HPV TYPE 31
95.	gttaaatctgiaaatgcaaaatctaata
96.	aatgttgttccatacacactatatctalacc
97.	ctatgcaacgtcctgtccacc
98.	cagtacgaggtcttctccaacatg
99.	tcataacagtggaggtcagttgc

[0055} In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from HPV type 35. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 12 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 100 to 105 (Tables 12), In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 13 (underlined). At least one Label F.xtft-nrler nrnbe may comprise, consist essentially of, or consist of a sequence selected from 16

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SEQ ID NOS: 106 to 117 (Table 13). The test may further employ a Blocking Label (BL),

e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS:

118 to 127 (Table 14).

2019247099 14 Oct 2019

Table 12: Capture Extender of HPV type 35

SEQ ID NO !	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF HPV TYPE-35
I loo.	caaacaaatttcatggatgctttcTTTTTctcttggaaagaaagt
101.	tactccatatggctggccttcTTTTTctcttggaaagaaagt
102.	tgtttttctaacgtttctccatacacTTTTTctcttggaaagaaagt
1 103.	caccgtccaccgalgttatgTTTTTctcttggaaagaaagt
104.	tccagctggaccgtcaatagtatTTTTTctcttggaaagaaagt
| 105.	ccatlaataaatcttccaaittacgtaTTTTT ct c i tggaaagaaagt

Table 13: Label Extender of HPV type 35

SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HPV TYPE-35
106.	gcagtttgtaaggtcgttcagctTTTTTgaagttaccgtttt
i 107.	ttctacctcgtt^cacaaatcatTTTTTctgagtcaaagcat
108.	taatt.cttgttLgcagtataca.caat.tTT T ΤΊ’gaagttaccgtttt
! 109.	aaagtcatatacctcactccgctgTTTTTctgagtcaaagcat
| no.	aataaatgacataactgtttgttgcatTTTTTgaagttaccgtttt
| ill.	ggtttttgacatgtaatacacctaattlTTTTctgagtcaaagcat
112.	tctaaaacatagtcttgcaatgtagttatlTITTgaagttaccgtttt
113.	agttgcctcgggttccaaaTTTTTctgagtcaaagcat
i 114.	acacaattgctcataacagtataggtcTTTTTgaagttaccgtttt
i 115.	ctt£CfCOtcctetg.agctgtcTTTTTctgagtcaaagcat
116.	ggaggtgtctggttttgcttgTTTTTgaagttaccgtttt
| 117.	ttacaacaggacgttacaatattataattTTTTTctgagteaaagcat

Table 14: Blocking Label Extender of HPV type 35

SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HPV ΊΎΡΕ35
ί 118.	tctatatactatacacaaatcatagcatgc
| 119.	gaataaaattttaaacatttcatgca
| 120.	actatatctataccatctatattcacttattttt
1 121.	ttgcttttcaactggacacagc
i 122.	gaatcgttttttttcttctaaatgtct
| 123.	aacaggacatacaccgacctgtc
124.	ggtttctctacgtgttggtttcc
I 125.	ttctccatgcatgattacacctc
| 126.	cagacgtagtgtcgcctcacat
	tgtcaatgtgtgtgctctgtacaca

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2019247099 14 Oct 2019 [0056] In certain embodiments, the target polynucleotide is an E6 or E7 sequence from HPV type 39. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 15 (underlined). Thus, at least one Capture Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 128 to 134 (Tables 15). At least one Label Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 7 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 135 to 145 (Table 16). The test may further employ Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 146 to 154 (Table 17).

Table 15: Capture Extender of HPV type 39

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 39
128.	tacctcggtttgctgtagtggTTTTTctcttggaaagaaagt
129.	ggttccccgtccctolatactalTrTT c tctiggaaagaaagt
1 130.	tttatgaaatcttcgtttgctatttTTTTTctcttggaaagaaagt
ί Ϊ3Ϊ.	ggtccacgcatetctgatgttataTTTTTctcttggaaagaaagt
132.	tttccigcaaggtgggctttTTTTTctctlggaaagaaagl
I 133.	itct^ctak.'C£i^c tTTTTTcl.c 33.3^1
134.	^gSstoctagJgagtccateaac^TTTTTctcttggaaagaaagt

Table 16: Label Extender of HPV type 39

SEQ ID NO 1	LABEL EXTENDER OF HPV TYPE 39
| 135.	cccgtattttagcataaaaAtttataT'rTTTctgagicaaagcat
| 136.	ccgagtccgagtaatatcgtagctTTTTTgaagttaccgtttt
1 137.	ttagttatatt.ttctaatgtagttgcatacaTTTTTctgagtcaaagcat
| 138.	cacagcggtttcagacaacacaTTTTTgaagttaccgtttt
i 139.	aggtetcttaatttttctgctggaTTTTTctgagtcaaagcat
[ 140. [ 14 L I “142?	ttcattgtaaggacataaatctaatacaaTTTTTgaagttaccgtttt catacaaggtcaaccggetgtatTTTTTctgagtcaaagcat gtcgggttcatotatttcatgctTTTTTgaagttaccgtttt
143.	ttgatgttggtgatmctgcatgT IT TTctgagtc aaagcat
144.	ggttc^ccgtctggcggKgTTTTTgaagttaccgtttt
1 145.	gaacactgtattgtgtgacgctgtTTTTTctgagtcaaagcat

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Table 17: Blocking Labe? Extender of HPV type 39

SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 39
I 146.	catataaatcactaaatgcaaattcata
i 147.	tgC^CCtl£ltt.H3t3.3.3tt3t3 L3.3Cttigt3
I 148.	actgtccigtatagcttcctgctat
i 149.	tggtccagcaccgtcgac
| 150.	tgcggtcctcccgttttg
i 151.	cttgggtttcicttcgtgttagtc
i 152.	ctgaclcicctaatigcicgiga
i 153.	gcagtgtgttgttacacttacaacac
154.	ctgctgtagttgtcgcagagtatc

[0057] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from HPV type 45. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 18 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 155 to 160 (Tables 18). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 19 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 161 to 172 (Table 19). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 173 to 180 (Table 20).

Table 18: Capture Extender of HPV type 45

| SEQ ID NO	CAPTURE EXTENDER OF HPV7 TYPE 45
j 155.	tatacctctgtgcgttccaatgtTTTTTctcttggaaagaaagt
| 156.	tataaataaatctttaaaagcaaaitgaTTrTTctcttggaaagaaagt
157.	cagtgttgctcggggtccaTTTTTctcttggaaagaaagt
1 158.	tgactaacgccatctgcttcatTTTTTctettggaaagaaagt
159.	agctcaattctgccgtocacTTTTTctcttggaaagaaagt
1 160.	catctgccgagctctctactgtaTTTTTctcttggaaagaaagt

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Table 19: Label Extender of HPV type 45

SEQ ID NO	LABEL EXTENDER OF HPV TYPE 45
161.	tttctgctgg^caatggttlTTTTgaagttaccgtttt
162.	c gttt gtccttaaggi gtctacgtt TTTTT ctgagtc aaagcat
163.	igtattacactgccctcggtactgTTTTTgaagttaccgtttt
164.	ggcgtg.cctggteacaacaTT'T’rTctgagtcaaagcat
165.	cctacgtctgcgaagtctttcttTTTTTgaagttaccgtttt
166.	tgcatacttattgctatactigtgtttoTTTTTctgagtcaaagcat
167.	ttccaaatgcaatacaatttottgTTTTTgaagttacc.gtttt
168.	caacaggatctaattcattctgaggTTTTTctgagtcaaagcai
169.	ttaattgctcgtaacacaacaggtTTTTTgaagttaccgtttt
h ΪΜ	cgttttcctcctctgactcgcITrTTctgagtcaaagcat
171.	tcgggctggtagttgigcalTTTTgaagttaccgtttt
172.	cgctgtggttcggctcgTTTTTctgagtcaaagcat

Table 20: Blocking Label Extender of HPV type 45

SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 45
173.	gcagcatatgctatacagictctatacac
174.	ggaataaaagtctatacattiatggcat
175.	gagtttgaataatatcttaattctctaattct
176.	ttttttccagtgtctctccatataca
177.	cttattaacaaattatacaactctgtattagtta
178.	tctggcaccgcaggcac
179.	tc c agct atgc tgtgga at c tt
180.	ttacaacatacacacaaaattttgtga

[0058] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from HPV type 51. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 21 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 181 to 187 (Tables 21). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 22 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 188 to 201 (Table 22). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 202 to 206 (Table 23).

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Table 21: Capture Extender of HPV type 51

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 51
181.	ggtctbccctcttgtcltcgaaTTTTTctcttggaaagaaagt
182.	tgcatagaaacgttc aaagc ttc'TTTTT c t c ttggaaagaaag t
183.	tttgaataaaacagtaaacattgtttgTTTTTctcttggaaagaaagt
184.	te^taaatcatataagctttttttagtaaTTTTTctcttggaaagaaagt
185.	cgcattgccccgtccaa 1 till ctcttggaaagaaagt
186.	cggtacgttgccsgca^^TTTTTctcttggaaagaaagt
187.	g^^gttoaattctgtaacacgtaTTTTTctcttggaaagaaagt

Table 22: Label Extender of HPV type 51

SEQ ID NO	LABEL EXTENDER OF HPV TYPE 51
188.	ttacaatacacacacactacctgtatattgTrTITgaagttaccgtttt
189.	ttatatacatctgctctacataattcctttTTTTTctgagtcaaagcat
190.	atacaatcttaatttcagtaaatgctecaTTTTTgaagttaccgtttt
191.	catactgcatatggatlattatecctatTTTTTctgagtcaaagcat
192.	acctgctataacgtctatactctctaattTTTTTgaagitaccgtttt ttgcctctaatgtagtacciitacacagTTTTTctgagtcaaagcat
194.	gtggtctttgacatctatgacaccttaTTTTTgaagttaccgtttt
195.	tt tgcttttcttcaggc cc aaTTTTTc tgagt c aaag cat
196.	cg^ttgggtttcgttacg^TTTTTgaagttaccgtttt
197.	cattaccacgcatggctttattaTTTTTctgagtcaaagcat
198.	tgcaatactacatettttaattgtggtaTTTTTgaagttaccgtttt
199.	agtcaatttcagtctgtggtgttaaaTTTTTctgagtcaaagcat
200.	tcaaattgctcgtagcattgcalTrTTgaagttaccgtttt
201.	tacttcatcctcctcctctgagctg'nTTTctgagtcaaagcat

Table 23: Blocking Label Extender of HPV type 51

SEQ ID NO	BLOCKING EXTENDER OF HPV TYPE 51
202.	acataatlcatgcagcgttcgt
203.	ctttttttttcgtccaccaatt
204.	cgtcccgctatttcatggaac
205.	ctggtagctggteacgcatattatc
206.	gcctgtccagcccgtcttt

[0059] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from HPV type 52. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 24 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 207 to 212 (Tables 24). In such

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Table 24: Capture Extender of HPV type 52

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 52
207.	gtgcagggtccggggtcTTTTTctcttggaaagaaagt
208.	cactotgaacagcgccctgtTTTTTctcttggaaagaaagt
209.	cacaggtcggggtctccaaTTTTTctcttggaaagaaagt
210.	cagtgctatgaatgcatagccgTTTTTctcttggaaagaaagt
211.	tgtgcccaacagcatttgcTTTTTctcttggaaagaaagt
r 2Ϊ2.	ttcagggfcctocattgcagTTTTTctcttggaaagaaagt

Table 25: Label Extender of HPV type 52

SEQ ID NO	LABEL EXTENDER OF HPV TYPE 52
213.	cttecagcacctcaocMtoTTTTTgaagttaccgtttt
214.	gccttatttcatgcaccgattlTTTTctgagtcaaagcat
215.	tttgcactgcacacactgcaTTTTTgaagttaccgtttt
216.	tatacctctcttcgttgtagctctttTTTTTctgagtcaaagcat
217.	tttctttttcttcaggacataatggTTTTTgaagttaccgtttt
218.	tcgcttgtttgcattaacatgtcITTTT c tgagt c aaagc at
219.	cgcatgacgttacacttgggtITITTgaagtt accgtttt
220.	aatcttttatagttgctttgtctccaTTTTTctgagtcaaagcat
221.	gccggtccacaccatctgtatTTTTTgaagttaccgtttt
222.	gcttgttctgcttgtcc^tgTTTTTc^agtcaaagcat
223.	atgtcacaatgtagtaattgcttgtg'rTI'TT'gaagttaccgtttt
2M	tagtgtgctatcacaactgigacaatTTTTTctgagtcaaagcat

Table 26: Blocking Label Extender of HPV type 52

SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 52
225.	c t attc gtaaat c t gtaaatagaaacttg
226.	cgccatatggattattgtctctatata
227.	LEICcIvlI
228.	tgataatgcctatattcacttatcttagata
229.	tctaatgttttcccatacagtgaatat
230.	cacttaatggtttttttaccctctct
231.	cgtttgacaaattatacatctaatagtiattt
232.	ccaacgacccataatattatgaaa
0Ί7	ttgttlcaggttgcagatctaatatal

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SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 52
234.	aattgctcatagcagtgtaggteag
235.	cctcctcatctgagctgtcacct
236.	tgtagagtacgaaggtccgtcg
237.	ccggggcacacaacttgtaa
238.	ggttgtttatagccgtgcacag

[0060] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from HPV type 56. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 27 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 239 to 245 (Tables 27). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 28 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 246 to 257 (Table 28). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 258 to 262 (Table 29).

Table 27: Capture Extender of HPV type 56

SEQ ID NO	CAPTURE. EXTENDER OF HPV TYPE 56
239.	gttagttcttttttgcaatatacacatgTTTTTctcttggaaagaaagt
240.	atgcaaaattatatacctcagcacgtTTTTTctcttggaaagaaagt
241.	gcacactgcataaggaaaatcaTTTTTctcttggaaagaaagt
242.	csg^^c^ttgcttttectgTTTTTctcttg^aagaaagt
Λ 't	ctattagatgaaatcgtctttttctgtTTTTTctcttggaaagaaagt
244.	gtetccagcaccccaaacatTTTTTctcttggaaagaaagt
245.	aggtcaatttctgtttgaggtgttaTTTTTctcttggaaagaaagt

Table 28: Label Extender of HPV type 56

SEQ ID NO	LABEL EXTENDER OF HPV TYPE 56
246.	ca^ractgaataglca^aiacci^mTTTTTgaagttaccgUit
247.	gtttittagttatactttctagtgtagctcTTTTTctgagtcaaagcat
248.	gtagcaccttattaataaatoacataactTTTTTgaagttaccgtttt
249.	cggagitaacggactttgacatCL1 IT ITTctgagtcaaagcat
250.	tgtagattct.etaggt.totctagargtitTTTTT'gaagttaccgtttt
251.	ttggtactttaccatgcatgatialacTTTTTctgagtcaaagcat

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SEQ ID NO	LABEL EXTENDER OF HPV TYPE 56
252.	cctcatcctcatcctctgagctTTTTTgaagttaccgtttt
253.	gctcctgcaaatggtctacttcatTTTTTctgagtxiaaagcat
254.	cttgtctagcttgctgtggccTTTTTgaagttaccgtttt
255.	gtgtattaggtoacacgtatgttgittagTTTTTctgagtcaaagcat
256.	cacaaacttacactcacaacaaggtacTTTTTgaagttaccgtttt
257.	ggtactgtgaatgtccaactgcacTTTTTctgagtcaaagcat

Table 29; Blocking Label Extender of HPV type 56

SEQ ID NO	BLOCKING LABEL EXTENDER. OF HPV TYPE 56
258.	tccctatacactaagtttaattcagtgc
259.	tctaactttactataaaacaataaacatactct
260.	gacccggtccaaccatgtg
261.	gttctaatataacgtcttgcagcg
262.	gtccaattgctcattgcactgt

[0061] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from HPV type 58. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 30 (underlined). At least, one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 263 to 269 (Tables 30). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or live) may comprise a target-hybridizing sequence shown in Table 31 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 270 to 281 (Table 31). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 282 to 286 (Table 32).

Table 30: Capture Extender of HPV type 58

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 58
263.	ctcctctgcgtcctggaacaTTTTTctcttggaaagaaagt
264.	tcaattcgatttcatgcacagaTTTTTctcttggaaagaaagt
265.	gtctmtgcattcaacgcattTTTTT c tcttggaaagaaagt
266.	cactattcttaaatctgcaaatacaaagTTTTTctcttggaaagaaagt
267.	agcaatcgtaagcacactttacatacTTTTTctcttggaaagaaagt
268.	attaatafflcatttaaacacltttttagtgtTTTTTctottggaaagaaagt
269.	ccgtccaagcctatttcatcctTTTTTctcttggaaagaaagt

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Table 31: Label Extender of HPV type 58

ί SEQ ID NO	LABEL EXTENDER OF HPV TYPE 58
ί 270.	atcatgcaatgtccgtggtttTTTTTgaagttaccgtttt
I 271.	tgtctccaacgcctgacacaaTTTTT c igagtcaaagcat
| 272.	ataattataatgtctatactcacttattttagatTTTTTgaagttaccgtttt
| 273.	.ttjgttcteatgtgtctccatatagcaaTTrTTctgagtcaaagcat
I 274..	caai^gtetttgac^ataatacateta.TTTTTgaagttaccgtttt
ί 275.	LgcciitLltLilcί tglggaca 111 I Iclgagteaaagcal
| 276.	cctgtccaacgacccgaaatatTTTTTgaagttaccgtttt
i 277.	tccaacacactgcacagcgcTTTTTctgagtcaaagcat
1 278.	tgtgtttgtctacgteggggtcTTTTTgaagttaccgtitt
279.	atj^cgttgttacaggttacactTTTTTctgagtcaaagcat
280.	toatagcagaataggtcagttggtTITITgaagttaccgtttt
1 281.	cgtctgagctgtcacataattgcTTTTT ctgagtcaaagcat

Table 32: Blocking Label Extender of HPV type 58

SEQ ID NO i	BLOCKING LABEL EXTENDER OF HPV TYPE 58
| 282.	tc atatacct c agate getgeaaa
| 283.	tgcaaatggatttccatctctata
I 284.	tatgaaaccttttgtttaaatccaca
| 285.	gcgttgggttgtttectctc
1 286.	tc aggatgtaaatc taaaat a ta it etc tta

[0062] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from HPV type 59. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 33 (underlined). At least, one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 287 to 292 (Tables 33). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Tabic 34 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 293 to 304 (Table 34). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 305 to 317 (Table 35).

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Table 33: Capture Extender of HPV type 59

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 59
287.	ggatcctcaaagcgtgccalTTTT c t cttgga aagaaag t
288.	tgatgcgaatatcatgcagaggTTTTTctcttggaaagaaagt
289.	l?jateacag.cgtetcag.cagctcTTTTTctcttggaaagaaagt
290.	tKttg.gacatagaggttttaggcaTTTTTctctt.ggaaagaaagt
291.	taggtgtcttgctegggtccTTTTTctcttggaaagaaagt
292.	aaagtgttgcttttggtecatgTTrTTctcttggaaagaaagt

Table 34: Label Extender of HPV type 59

SEQ ID NO	LABEL EXTENDER OF HPV TYPE 59
___________________________________ 293.	cccctttgcaaaacacacaatTTTTTgaagttaccgtttt
294.	tcaaatacctctctttcttgcagttITrTT c tgagt c aaagc at
295.	cctctaatgtttctccatacacggTTTTTgaagttaccgtttt
296.	atgteacggtgtcttggtttcagTTTTTctgagtcaaagcat
297.	gcgcttgtcg^ctgtctTTTTTgaagttaccgtttt
298.	cattgttttacaccagtgtttcactacTTTTTctgagtcaaagcat
299.	ggttccaaatetaaaacaatetcacTTTTTgaagttaccgtttt.
300.	aaggtcaacttcctcataaitttgtTTITr c tgagtc aaagcat
301.	tcaggtaattgctegtagcacacTTTTTgaagttaccgtttt
302.	ttttcattcicggagtc^agTTTTTctgagtcaaagcat
303.	gcgaggrttctactactagctgaagTTTTT gaagtta c cgtitt
304.

Table 35: Blocking Label Extender of HPV type 59

SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 59
305.	ggcagtttgtatggtcgttgtgta
306.	aatattcaatgttgtgctcaaaica
307.	tacaciataaataagtcattaaaagcaaat
308.	gctgcatacggtgtacagtctcta
309.	cataaaatgaaatgcatttcagacac
310.	aatctctataatatcttaattctcttactcttg
311.	cttttttcagttatatgctttaatttatc
312.	tatatattccagctatattatggaatctt
313.	gacacccacgacactgtoctg
314.	gatgattaactccatctggttcatct
315.	cagctcgtctagctagtagcaaag
316.	acaatgttgtgacgctgtggtt
317.	ttgattattacacttacaacacacacac

[0063] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from HPV type 68. In such embodiments, at least one Capture Extender probe (e.g., at least

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2019247099 14 Oct 2019 two, three, four, or five) comprises a target hybridizing sequence selected from Table 36 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 318 to 324 (Tables 36). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 37 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 325 to 338 (Table 37). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 339 to 347 (Table 38).

Table 36; Capture Extender of HPV type 68

SEQ ID NO	CAPTURE EXTENDER. OF HPV TYPE 68
318.	^cntgcaatgtagtgtccaatgtTTTTTctcttggaaagaaagt
319.	gtteggtgcctggtttttc^TTTTTctctt^aaagaaagt
i 320.	f&cttactggtccagi^gtgcTT'fTTctcttggaaagaaagt
321.	cggtgggctttggtccatgTTTTTcicttggaaagaaagt
i 322.	atagctctaacacaatttcctgcaTTTTTctcttggaaagaaagt
| 323.	cccgcgacgcttctactactlTTTT c t c ttgga aagaaag i
324.	caaaatttagteagtccataaacagcTTTTTctcttggaaagaaagt

Table 37: Label Extender of HPV type 68

1 SEQ ID NO i	LABEL EXTENDER OF HPV TYPE 68
I 325.	cttctgcaatagacacagtetattgtaacTTTTTgaagttaccgtttt
i 326.	catatacctctgtccgttgtagttgcTTTTTctgagtcaaagcat
i 327.	attiaatacatgattgscatgcagTTTTTgaagttaccgtitt
| 328.	cgWttcccgtattttagcataaa’T’T’rT’T’ctgagtcaaagcat
1 329.	gcat^ccgattccgagmtatTTTTTgaagttaccgtttt
1 330.	ctttgtatlagttatggtltetaatgtagttTTTTTctgagtcaaagcat
331.	aclcatgcacctLatcaataaattatataa'TTTTTgaagttaccgltU
i 332.	tggacacaatggtttcaggcalTTTTctgagfcaaagcat
[ 333.	cggctgtatttcattgtatggacTTTTTgaagttaccgtttt
i 334.	tgctcgtgacatacaaggtcaacTTTTTctgagtcaaagcat
[ 335.	ctatttc.atcgtctgaatctoctaatTTTTTgaagttaccgtttt
| 336.	aactgcatggtcgggttcatTTTTT ctgagtcaa agcat
| 337.	gctgttgttcgtoccgtctgTTTTTgaagttaccgtttt
338.	caacacagacactgaattctgtgacTTTTTctgagtcaaagcat

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Table 38: Blocking Label Extender of HPV type 68

SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 68
339.	ctacacataggtcac taaaggcaaatt
340.	caaatggtaccccgtctctataca
341.	gctattttatgtaatcttcgttttgt
342.	cgacactgtcctgtaaagtttcct
r 343.	gtatgcgtctgcggtcctct
344.	c atagttaetta aacttgtgttt ettga c
345.	gctagtagtagatgttggtggtgatt
346.	agttgc agtgc cttgttac aetta
347.	tgttgtagtgtccgcaggttgt

[0064] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from low risk-HPV type 6. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 39 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 348 to 354 (Tables 39). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 40 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 355 to 368 (Table 40). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 369 to 375 (Table 41).

Table 39: Capture Extender of HPV type 6

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 6
348.	tggatagccgcctcgaaaTTTTTctcttggaaagaaagt
349.	cacgcgcaggctgcataTTTTTgtcttggaaagaaagt
350.	tttttccatgaaattctaggcagTTTTTctcttggaaagaaagt
351.	taggcagcgacccttccaTTTTTctottggaaagaaagt
352.	caatatccttta^gtaacatgtcttcTTTTTctcttggaaagaaagt
__ J53. __ __ ...... ...	cagaagctgttgcacttctctgaTTTTT etc ttggaaagaaagt ggtcttcggtgegcagatgTTTTTctettggaaagaaagt

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Table 40: Label Extender of HPV type 6

[ SEQ ID NO	LABEL EXTENDER OF HPV TYPE 6
| 355.	altaatttgcaacgtalgcatagaiaTTTTTgaagttaccgttU
i 356.	gtgcattcttgcaaaacacacaTTTTT etgagte aaagcat
357.	tatgaataaatetetgctgtggtcaTTTTT gaagtt ac c g tttt
i 358.	caggacctttaggtgtttaiatgcaTTTTTctgagtcaaagcat
i 359.	tottc^tgttgttgc^tetccaTTTTTgaagttaccgtttt
j 360.	cwgtotea^tgtettgttegtectTTTTTctgagte-aaagcat
361.	cgcgttggttagtetetgttttacctTTTTTgaagttaccgtttt
| 362.	cgtacaatttagctttatgaaccgTTTTTctgagtcaaagcat
| 363.	ggtotggaggttgcaggtctaata'lTrTTgaagttaccgtttt
i 364.	tgctcatagcaatgtaaccctacagTTTTTctgagtcaaagcat
365.	acctcatcttctgagctgtctactaatTTTTTgaagtt ac c gtttt
| 366.	cttgtccgtccacttcgtccTTTTTctgagtoaaagcat
! 367.	cagtcgaacgttgctgtcacatTTTTTgaagttaccgtttt
368.	tgtctgtttctgtacactgcacaacTTTTTctgagtcaaagcat

Table 41: Blocking Label Extender of HPV type 6

SEQ ID NO i	BLOCKING LABEL EXTENDER. OF HPV TYPE 6
i 369.	gcataat c aaagtgtctatattggttta
i 370.	tgacacaggtagcaccgaattag
371.	tttctacttcacacagcggtttg
i 372,	catgcatgttgtccagcagtg
| 373.	gaaatgttgttitaaaggttgtgaat
ί 374.	ccacagcaacaggtcactatttg
i 375.	ggacacactatgtttagtgttcccaa

[0065] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from low risk-HPV type 11. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 42 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 376 to 382 (Tables 42), In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 43 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 383 to 398 (Table 43). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 399 to 403 (Table 44).

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Table 42: Capture Extender of HPV type 11

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 11
376.	caaagaaagattaaacgtcttecacTTTTTctcttggaaagaaagt
377.	cgcactgaatttgcagagtgtgTmTctottggaaagaaagt
378.	tggtttcttcttctactgtaggtgcTTTTTctcttggaaagaaagt
379.	cgaatt aac acttttaaaatat c tteat'TTTTT ct cttgga aagaaagt
380.	agcgigjxtttcccaata^TTTTT ct cttggaaag aaagt
381.	accctacagg^aggaggctTTTTTctcttggaaagaaagt
382.	gtgcgtottgtttgtocacctTTTTTctctt^^iagaaagt

Table 43: Label Extender of HPV type 11

SEQ ID NO	LABEL EXTENDER OF HPV TYPE 11
383.	tggaggcatctttactttecataaTTTTTgaagttaccgtttt |
384.	aactggtctatagatgttgcagacgTTTTTctgagtcaaagcat |
385.	atatgcataiatctctgcggtggTTTTTgaagttaccgtttt |
386.	ac^aacctttaggttcttataggcTTTTTctgagtcaaagcat i
387.	^gcaacaggcacacgctgTTTTT gaagttaccgtttt |
388.	ggtta^tgi^tt^agttgtlTrTT ctgagtcaaagcat |
389.
390.	tgctttagttttlctatttcacacaaTTTTTctgaotcaaagcat i
391.	cttccactggttatttagttttatgaTTTTTgaagttaccgtttt |
392.	ccagcagtgtaagcaacgaccTTTTT ctgagtcaaagcat |
393.	gtcttctaattgctcatagcaatgtaTTTTT gaagttaccgtttt |
394.	tgtccacctcatcttctgagctTTTTTctgagtcaaagcat |
395.	gtatttggtaatgttgtgtiaaaggttTTTTTgaagtiaccgittt |
396.	cacatccacagcaacaggtcaTTTTTctgagtcaaagcat |
397.	caaccagtcggacgttgctgtTTTTTgaagttaccgtttt |
398.	atgtotccgtctgtgcactccaTT'TTT ctgagtcaaagcat |

Table 44: Blocking Label Extender of HPV type 11

SEQ ID NO	BLOCKING LABEL EXTEND	ER OF HPV TYPE 11
399.	tcagtgcattcctgcaaaaca
400.	caaagggaaagttgtctcgcc
401.	atatgcagcataattaaagtgtctatatt
402.	taacaagtcttccatgcatgttgt
403.	gcaggtctagtactatatcctttaggg

[0066] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from low risk-HPV type 40. In such embodiments, at least one Capture Extender probe (e.g..

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2019247099 14 Oct 2019 at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 45 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 404 to 410 (Tables 45). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 46 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 411 to 424 (Table 46). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 425 to 432 (Table 47).

Table 45; Capture Extender of HPV type 40

SEQ ID NO	CAPTURE EXTENDER. OF HPV TYPE 40
I 404.	tcatacagggtcctggcctgTTTTTctcttggaaagaaagt
405.	g^ccgtot^caaaacarac TT TTTctcttggaaagaaagt
] 406.	.cgtggacatgt’ggcgtgtT'rT'FTctcttggaaagaaagt
407.	aaagaattgcgtcttctttacaatatTTTTTctettggaaagaaagt
i 408.	agtttagacatacaggttcagggtgTTTTTctcttggaaagaaagt
: 409.	acaccgagttactactttaaatgattgTrnTctcttggaaagaaagt
410.	tgatggaacaatgcactgctaagTTTTTctcttggaaagaaagt

Table 46: Label Extender of HPV type 40

i SEQ ID NO i	LABEL EXTENDER OF HPV TYPE 40
1 411.	tgtaatattgcactggtcacacagfmTT gaagtt acc gtt it
i 412.	aatcaatttgcaacgtaggcaaTTTTT ctgagtcaaagcat
i 413.	ggccagtacctragctgtttttaTTTTT gaagtt ac c gtttt
| 414.	£gcaacatataaU.UctaaaggcaaaTITI'Tctgagtcaaagcat
| 415.	ccgtgcaggtccaggcacTTTTTgaagttaccgtttt
1 416.	alctaaagtltctgtattggtttacttUTTTTTctgagtcaaagcat
417.	gttaaictftgtctcttcltccacgTTTTTgaagttaccgtttt.
i 418.	cagcatctaatccttacttgtaaaalgTTTTTctgagtcaaagcat
[ 4'19.	ccctgtccacRaatcttttaatttTTTTT gaagttacc gtttt
420.	catttcttccagcaatgtagacagtaTTTTTctgagtcaaagcat
[ 42 Ϊ.	gagctgtetaattgctegttgcTTTTTgaagttaccgtttt
i 422.	ct gtt catggt catottctga gtctTTTTT etgagt caaagcat
| 423.	fctactg^taagctgtetogttggtcTTTTTgaagttaccgtttt
424.	cgtgggttgclcacgctcTTTTTctgagtcaaagcat

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Table 47: Blocking Label Extender of HPV type 40

ί SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 40
| 425.	ggaaagtcgtcgcgcca
I 426.	gttggtgcataggclgcgi
| 427.	gacaaaggcttgtggcacttg
428.	ggitggttttttccacggga
I 429.	gcgttggcctttctccatg
| 430.	caggtttaacacaatgtctccga
| 431.	caaatttacttgcaggtcctgttg
432.	cgcaccaaacactgacaaaatac

[0067] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from low risk-HPV type 42. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 48 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 433 to 439 (Tables 48). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 49 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 440 to 451 (Table 49). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 452 to 462 (Table 50).

Table 48: Capture Extender of HPV type 42

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 42
433.	acattctgccttaactgggaccTTTTTctcttggaaagaaagt
j 434.	££tgtteagtg£tttttg£2££.TTTTTctcttggaaagaaagt
| 435.	acgcgagcacctctg^TTTTTctcttggaaagaaagt
436.	caatataaattgaaatcttgtacctgtatcITTTTctottggaaagaaagt
! 437.	cattgtectctgcaatgcgtaTTTTTctcttggaaagaaagt
438.	cgctgtatgtcctgtttggctTTTITctcttggaaagaaagt
j 439.	cttatgtccgcctctgtacactgTTTTTctcttggaaagaaagt

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Table 49: Label Extender of HPV type 42

ί SEQ ID NO	LABEL EXTENDER OF HPV TYPE 42
i 440.	ctgtgatgaggcagatgtacctgTTTTTgaagttaccgtttt
i 441.	tacacaattggtataatgtgcgtggTTTTTctgagtcaaagcat
| 442.	gcaatgtcagc.ccaaattce.tTTTTT gaagttac cgtttt
443.	aaHig;-i vSgj.irui.U q'7'1'J'T' c tgagt c aaagcat
I 444.	caasgrgct^tetttcgtag^tTTTTTgaagttaccgtttt
i 445.	agtocagtttctttctccactgtatac'r'TTTTctgagtcaaagcat
| 446.	gataacggcttttgacacaaggTTTTTgaagttaccgtttt
i 447.	aatatgatggtttttttcgctetgtlTrTTctgagteaaagcat
I 448.	atgtcaaacaaaacaatgtcctttaTTTTTgaagttaccgttit
449.	atgggtgtctcacacgttggtTTTTT c tgagtcaaagcat
450.	caaaagcatctgttgcaggtttTTTTTgaagttaccgtttt
1 451.	ggacacacaatatccagtgtgccTTTTTctgagtcaaagcat

Table 50: Blocking Label Extender of HPV type 42

SEQ ID NO 1	BLOCKING LABEL EXTEN DER OF HPV TYPE 42
| 452.	caccactaccaaatctttaaaatggt
| 453.	gcatatggaaagtccttcctcca
I 454.	attctaaacaaaatgcacatgca
1 455.	cgcagtgcacaaattttagaattaa
j 456.	cacatetaatttgttgttcttctaaaagt
| 457.	caccgacccgtccactgaca
i 458.	gggtaggcgtctctccacg
! 459.	caattgttcatagcaatacaggtca
I 460.	tggtcatcttcatctgagclgtc
| 461.	ctgtgtacacacacacagiatlctgiaa
462.	ca.ca.acgagtftaacagacttgtaa.ca

[0068] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from low risk-HPV type 43. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 51 (underlined). At least one Capture Extender probe may comprise, consist essentially of or consist of a sequence selected from SEQ ID NOS: 463 to 468 (Tables 51). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 52 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 469 to 482 (Table 52). The test, may further employ a Blocking Label (BL.),

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e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS:

483 to 488 (Table 53).

Table 51: Capture Extender of HPV type 43

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 43
463.	agcatgcagcaaacggatatcTTTTTctcttggaaagaaagt
464.	ccateaaactgtagacaggccaTTTTTctcttggaaagaaagt
465.	teaaagtgccUtattgautatttttTTTTTctcltggaaagaaagt
466.	acagtatctgcatatgctgcgtagITrTTctcttggaaagaaagt
467.	atgcatgatttocagcaatgtagTnTTctcttggaaagaaagt
468.	ttaatgtgcccaacagcaggTTTTTctcttggaaagaaagt

Table 52: Label Extender of HPV type 43

SEQ ID NO	LABEL. EXTENDER OF HPV TYPE 43
469.	catcacacaactcaaatatagtccgTTTTTgaagttaccgtttt
470.	gcagagtegg^^gttatgttacactTTTTTctgagtcaaagcat
471.	acttccgtggteagt^ccactteTTTTTgaagttaccgtttt
472.	tttaaatctctaaatgcaaacgataatTTTTTctgagtcaaagcat
473.	aacacigtttgcttagtttcttcttctlTTTTgaagttaccgtttt
474.	cttacagcateta^ca^^caTTTTTctgagtcaaagcat
475.	tggtgataatggcttgtggca IT IT fgaagttaccgtttt
476.	gcacaatatgctgtactttttccacTTTTTctgagtcaaagcat
477.	atgtettttaaagaattgtgccmiTTTTTgaagttaccgtttt
478.	gcagtatcctttccacacgctTTTTTctgagtcaaagcat
479.	tggttgcatagttagcacatagtctTTTTTgaagttaccgtttt
480.	cgtocaggJtea^ttetaggtteTTTTTctgagtcaaagcat
481.	attacacacagacaggatgtgcac IT IT Tgaagttaccgtttt
482.	agagcactgcacaacaagtcgaTTTTTctgagtcaaagcat

Table 53: Blocking Label Extender of HPV type 43

SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 43
483.	ctgaicgtcggcttttttcc
484.	tctgagtctgagctgtctaattgct
485.	gggttgctcacgctcatcc
486.	acttgctggtcctgttgcgt
487.	tctgttacaactctgtaaacttgtagattc
488.	tcttctagcttcttgatgtcactgtc

[0069] In certain embodiments, the target polynucleotide is an E6 and/or E7 sequence from low risk-HPV type 44. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table

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PCT/US2010/040649 (underlined), At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 489 to 494 (Tables 54). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Table 55 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 495 to 506 (Table 55). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 507 to 510 (Table 56).

Table 54: Capture Extender of HPV type 44

SEQ ID NO	CAPTURE EXTENDER OF HPV TYPE 44
489.	cctctgcagtacttaacgtttttotgTTTTTctcltggaaagaaagt
490.	gcacgtccagaattgacttatttgtTTTTTctcttggaaagaaagt
491.	tttaatgaatcgcgccttgtcTTTTTctcttggaaagaaagt
492.	cgacccttocaggtatcttgtaaTTTTTctcttggaaagaaagt
493.	cctt^ggtag^agtttccatg^TTTTTctcttggaaagaaagt
494.	Sggttocagctgtaaaaca^TTTTTctcttggaaagaaagt

Table 55: Label Extender of HPV type 44

SEQ ID NO	LABEL EXTENDER OF HPV TYPE 44
495.	gcagacgtggaggcatt^cTTTTTgaagttaccgtttt
496.	ccttgcacaactggtctatactttgtTTTTTctgagtcaaagcat
497.	attgtgcataggaatgftgcactlTTTTgaagttaccgtttt
498.	caaaacacgcataaaatttgcagTTTTT c tgagteaaage at
499.	acaaatggcacaggctgcaTTTTTgaagttaccgtttt
500.	tgattgaccttaccttgtagttctaaTTTTTctgagtcaaagcat
501.	ccgcgtagttaaaatgcctaaatTTTTTgaagtt ac c gtttt
502.	.ttcitcitccactgttactgcatatcTTTTTctgagtcaaagcat
503.	ggcacaaatagcagcgt^alTTTTgaagttaccgtttt
504.	cacgtggcacaatggtttgtTTTTTctgagtcaaagcat
505.	caatgtaggcctacagggtcagTTTTTgaagttaccgtttt
506.	totgagctgtctaattgctcattgTTTTTctgagtcaaagcat

Table 56: Blocking Label Extender of HPV type 44

SEQ ID NO	BLOCKING LABEL EXTENDER OF HPV TYPE 44
507.	ctacatataactgtttatatgcgaatgaataaa
508.	aatggaaagtttcc tcggtaca
509.	caatatgiggcgcaccttttc
510.	tgatgtccaaca.fttggaa.gcag

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2019247099 14 Oct 2019 [0070] In other embodiments, the target polynucleotide is hepadnavirus polynucleotide, such as HBV. In such embodiments, the target polynucleotide is one or more polynucleotide sequences encoding the viral core protein, HBcAg; the proteolytic processing protein, HBeAg, and/or the viral surface antigen, HBsAg. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence as selected from Table 57 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 511 to 519 (Tables 57). At least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 58 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 520 to 535 (Table 58). The test may optionally employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 536 to 553 (Table 59).

Table 57: Capture Extender of HBV

SEQ ID NO	CAPTURE· EXTENDER NUCLEOTIDE SEQUENCE OF HBV
511.	ggggagWccgcgtaaagagatttttctcttggaaagaaagt
512.	tgcRacgtgcagaggtgaatttttctcttgga.aagaaagt
513.	RagtccaagagtcctcttatgYaagactttttctcttggaaagaaagt
514.	cagaggtgaaaaagttgcatgRtttttctcttggaaagaaagt
515.	aMggRtcaatgtccatgcctttttctcttggaaagaaagt
516.	tcagaaggcaaaaaMgagagtaacttttttctcttggaaagaaagt
517.	atDgcttgcctKagtgcHgtatgtttttctettggaaagaaagt
518.	cggaagtgttgataagataggggtttttctcttggaaagaaagt
519.	ctKcgtctgcgaggcgagtttttctcttggaaagaaagt

Table 58: Label Extender of HBV

SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HBV
520.	ggcagatgagaaggcacagactttUgaagttaccgttlt
521.	gcgaagtgcacacggWcctttttctgagtcaaagcat
522.	tcggtcgttgacattRcWgRtttttgaagttaccgtttt
523.	cagtctttgaagtaKgcctcaaggtttttctgagtcaaagcat
524.	gcctacagcctccYaRtacaaagactttttgaagttaccgtttt
525.	tgMtggtgMRcaSaccaatttattttttctgagtcaaagcat
526.	ggacatgWacaWgagatgatYaggtttttgaagttaccgtttt
527.	ca^cttggaggcttgaacagtRtttttctgagtcaaagcat
528.	agactctaaggcY tcY cgatactttttgaagttaccgtttt
529.	RtgaggtgaRcaatgYtcMggtttttctgagtcaaagcat
530.	cctcKtcgtctaacaacagtagtYtctttttgaagttaccgtttt

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SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HBV
531.	cagcttggaggcttgaacagtRtttttctgagtcaaagcat
532.	cgacgcggcgattgagaYtttttgaagttaccgtttt
| 533.	attcccgagattgagatctYctgtttttctgagtcaaagcat
i 534.	gWgtccaaggRatactaacattgagtttttgaagttaccgtttt
\ 535.	cccMgtaaaRtttcccaccttattttttctgagtcaaagcat

Table 59: Blocking Label Extender of HBV

1 SEQ ID NO ! !	BLOCKING LABEL EXTENDER NUCI	.EOTIDE SEQUENCE OF HBV
| 536.	gcgttcacggtggtctcca
i 537.	cttgggcaRgWHB Y S DYgg
j 538.	aRctcctcccaStcHKtaaaYaMa
| 539.	ctttaaYctaRtctcctccccc
! 540.	cYaaagccaYccaaggca
[ 54Ϊ.	ccacagWagctccaaa ttctttat
| 542.	gDagatcIIcKDaYDgaaggaaagaag
543.	agagcWgaggcKgtRtcDa
544.	YatYaRB tcMcc ccaRcaVagR
545.	ccacccaggtRgMYaRaKt
[ 546.	tg Y WggRtcttccaaattaH YW c
i 547.	cataR Y tKactactaRD tccc iRga
I 548.	WYtttaRRcccatRttaRYRttRa
1 549.	aW a tRtgaaaccacaatagttgY ctRa
1 550.	gtYtctctYccaaaagtRagRcaRg
| 551.	cRaaagaSaccaaataYtcWaKDacH
| 552.	gWggagtgcgaatccacactc
| 553.	cattWggtggtctRtaDgcDg

[0071] The invention also provides a method for detecting a viral target polynucleotide from a RNA virus such as HIV or paramyxovirus (flu virus). The target polynucleotides for detecting a HIV infection can be HIV-A., -B or -F. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence of Table 60 (underlined). At least one Capture Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 554 to 558 (Table 60). At least one Label Extender (e.g., at least two, three, four, or five) may comprise a targethybridizing sequence selected from Table 61 (underlined). At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 559 to 567 (Table 61). The test may optionally including a Blocking Label (BI ), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 568 to 571 (Table 62).

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Table 60: Capture Extender of HIV-A

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF HIV-A
554.	ttcatttcctccaatccccttatgtgTTTTTctcttggaaagaaagt
555.	attgctgtgatatctttcatgttcttettgaTTTTTctcttggaaagaaagt
556.	actgctaccagaattacttteccttclaaatgTTTTTctcttggaaagaa^t
557.	cgctggtaaaattgctgccattgtctTTTTTctcttggaaagaaagt
558.	ctecttgactttgggg.attgtagggaTTTTTctcttggaaagaaa.gt

Table 61: Label Extender of HIV-A

SEQ ID NO	LABEL, EXTENDER NUCLEOTIDE SEQUENCE OF HIV-A
559.	ctgattccagagctgactaalttatctacttgTTTTTctgagtcaaagcatgaagttac
560.	gccttatetatcccatctaaaaatagtatcttcTTTTTctgagtcaaagcatgaagttac
561.	agattaaaatcactagccattgctetccaTTTTTctgagtcaaagcatgaagttac
562.	ggctactatttcctttgctactataggtggcTTTTTctgagtcaaagcatgaagttac
563.	gactacagtctacctgtccatgcatggcTTTTTctgagtcaaagcatgaagttac
__ 564. “'“565.	ctgcttctatatagccactggctacatggTTTTTctgagtcaaagcatgaagttac______________ cctgccctgtttctgctgggataacttTTTTTctgagtcaaagcatgaagttac
566.	gtgtgtactacttttactggccatcctccTTTTTctgagtcaaagcatgaagttac
567.	ccaccaacaggctgctttaaatgcagTTT'T Ίctgagtcaaagcatgaagttac

Table 62: Blocking Label Extender of HIV-A

SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HIV-A
568.	ttccccttttagttgacatttatcacagct
569.	tgtgcaatctaattgccacatecctg
570.	tgctaattttagcaaaaagtatgctgcct
571.	atcccaaattcctgttggatgtttgc

[0072] In another embodiment, the target polynucleotide is an HIV subtype B (HIV-B) polynucleotide. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 63 (underlined). At least one Capture Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 572 to 577 (Table 63). At least one Label Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 64 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 578 to 590 (Table 64). The test may optionally employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 591 to 595 (Table 65).

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Table 63: Capture Extender of HIV-B

[ SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF HIV-B
| 572.	tgcttctatatacccactggctacatgaactTTTTTctcttggaaagaaagt
i 573.	caacaggcggccttgaccgcTTTTTctcltggaaagaaagt
574.	tcctgcttgacccccgcccacTTTTTctcltggaaagaaagt
i 575.	gcactatagtccccaatcccccctcTTTTTctcttggaaagaaagt
576.	ctttccagaggagctttgctggtccTTTTTctcttggaaagaaagt
577.	actcttatcttgtattactactgccccttcacTTTTTctcttggaaagaaagt

Table 64: Label Extender of HIV-B

SEQ ID NO i	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HIV-B
| 578.	caatctagttgccatattcctggactacagtTTTTTctgagtcaaagcatgaagttac
§ 579.	gctaccaggataactcttccttctaaatgtgtaTTTTTctgagtcaaagcatgaagttac
| 580.	gccctgtctctgctgggatcacttcTTTTTctgagtcaaagcatgaagttac
j 58Ϊ.	tgtatgtattgtctttactggccatcttccTTTTTctgagtcaaagcatgaagttac
L 582.	ttctttattcatagattctactactccttgactTTTTTctgagtcaaagcatgaagttac
| 583.	gatctcttacctgfcctataattttctttaaTTTTTctgagtcaaagcatgaagttac
i 584.	tttgtactgctgtcttaagatgttcagcctTTTTTctgagtcaaagcatgaagttac
| 585.	ttcttttaaaattgtggatgaatactgccaTTTTTctgagtcaaagcatgaagttac
I 586.	ctgttgctattatgtetattattctctcccctTTTTTctgagtcaaagcatgaagttac
§ 587.	aatttgtttttgtaattctttggtttgtatgtTTTTTctgagtcaaagcatgaagttac
i 588.	tgtaatagacccgaaaattttgaatttttgtTTTTTctgagtcaaagcaigaagttac
589.	gccatctgttttccataatccctaatgatTTTTTctgagtcaaagcatgaagttac
| 590.	cctgtctacttgccatacaatcatcacctTTTTTctgagtcaaagcatgaagttac

Table 65: Blocking Label Extender of HIV-B

i SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HIV-B
591.	igctaattttaagagaaagtatgctgtttcct
' 592.	actactggtgaagttgcigccattgtc
| 593.	ttggggattgtagggaatgccaaat
594.	tttccaaagtggatctctgctgtccc
| 595.	ctttacttttcttcttggtactacttttatgtc

[0073] In a further embodiment, the target polynucleotide may be an HIV subtype F (HIV-F) polynucleotide. In such embodiments, at least one Capture Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 67 (underlined). At least one Capture Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 596 to 600 (Table 67). At least one Label Extender ( e.g., at least one, two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 68 (underlined). At least one Label Extender may comprise,

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68). The test may optionally employ a Blocking Label (BL), e.g., containing the sequence of

SEQ ID NO: 609 (Table 69).

Table 66: Capture Extender of HIV-F

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF HIV-F
596.	ttctttagctctttattcattgattctactactccTTTTTctcttggaaagaaagt
597.	ttcccctgcactgtaccccccaatcTTI T Tctcttggaaagaaagt
598.	gctgtccctgtaataaacccggaaattttTTTTTctcttggaaagaaagt
599.	tgccccttcacctttccagagtagctTTTTTctcttggaaagaaagt
600.	cgtcacctgccatctgttttccataatcTTTTTctcttggaaagaaagt

Table 67: Label Extender of HIV-F

SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HIV-F
601.	ttcagcttgatctcttacctgtcctatgatoTTTTTctgagtcaaagcatgaagttac
602.	atactgccatttgtactgctgtcttaagatgTTTTTctgagtcaaagcatgaagttac
603.	cccccttttcttttaaaattgtggatgaTTTTTctgagtcaaagcatgaagttac
604.	ttgtatgtctgttgctattatgtctattattctTTTTTctgagtcaaagcatgaagttac
605.	gaatttttgtaacttgtttttgtaattctttagtTTTTTctgagtcaaagcatgaagtta
606.	ttgctggtcctttccaaactgggtetctTTTTTctgagtcaaagcatgaagttac
607.	ctacttttatttcactattgtcttgtatgactac TTTTTctga.gtcaaa .gcatgaagtta
608.	tcatcctgtctacctgccacacaatTTTTTctgagtcaaagcatgaagttac

Table 68: Blocking Label Extender of HIV-F

SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HIV-F
609.	cctaatgatctttgcttttcttcttggca

[0074] The target polynucleotide may be a polynucleotide of a respiratory viruses, such as a influenza H1N1. For example, the target polynucleotide may be a sequence encoding a portion of hemaglutinin (HA), neuraminidase (NA), M protein, F protein, N protein, G protein, etc. In an exemplary embodiment, the target polynucleotide is an HA sequence. In such embodiments, at least one Capture Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 69 (underlined). At least one Capture Extender may comprise, consist essentially of, or consist, of a sequence selected from SEQ ID NOS: 610 to 629 (Tables 69). At least one Label Extender (e.g., at least two, three, four, or five) may comprise a target hybridizing sequence selected from Table 70 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 630 to 689 (Table 70). The test may optionally

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Table 69: Capture Extender of Ehs Virus, MINI

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF Flu Virus, H1N1
610.	ggttctattggaaaatgaaagaactttTTTTTctcttggaaagaaagt
611.	caaaatgagcaggggtcaggTTTTTctcttggaaagaaagt
612.	tgccggtttcattgaagggTTTTTctcttggaaagaaagt
613.	ccagcctcccatttcagaatatacTTTTTctcttggaaagaaagt
614.	acacccaagggtgctataaacaTTTTTctcttggaaagaaagt
615.	caagccggaaatagcaataagaTTTTTctcttggaaagaaagt
616.	aaaaggaaa.ttcatacccaaagcTTTTTctcttggaaagaaagt
617.	aaaggtgtaacggcagcatgtTTTTTctcttggaaagaaagt
618.	ctcagtgtcatcatttgaaaggtttTTTTTctcttggaaagaaagt
619.	gggtaaatgtaacattgctggctTTTTTctcttggaaagaaagt
620.	ggttctattggaaaatgaaagaactttTTTTTctcttggaaagaaagt
621.	caaaatgagcaggggtcaggTTTTTctcttggaaagaaagt
622.	tgccggtttcattgaagggTTTTTctcttggaaagaaagt
623.	ccagcctcccatttcagaatatacTTTTTctcttggaaagaaagt
624.	acacccaagggtgctataaacaTTTTTctcttggaaagaaagt
625.	caagccggaaatagcaataagaTTTTTctcttggaaagaaagt
626.	aaaaggaaattcatacccaaagcTTTTTctcttggaaagaaagt
627.	aaaggtgtaacggcagcatgtTTTTTctcttggaaagaaagt
628.	ctcagtgtcatcatttgaaaggtttT TTTTctcttggaaagaaagt
629.	ccagtaaactgattaactggttcttcaTTTTTctcttggaaagaaagt

Table 70: Label Extender of Flu Virus, H1N1

SEQ ID NO	LA BEL EXTENDER NUCLEOTIDE SEQUENCE OF Flu Virus, Hl N1
630.	tggacttacaatgccgaactgttTTTTTgaagttaccgtttt
631.	tgatgatggtttcctggacattTTTTTctgagtcaaagcat
632.	ggtaaagagttcaaccacctggaTTTTTgaagttaccgtttt
633.	aagatgaatacacagttcacagcagtaTTTTTctgagtcaaagcat
634.	cacacagaatgccattgacgagTTTTTgaagttaccgtttt
635.	atatgcagccgacctgaagagTTTTTctgagtcaaagcat
636.	tegatggtacggttalcaccatTTTTTgaagttaccgtttt
637.	gggtggacagggatggtagaTTTTTctgagtcaaagcat
638.	aggccta tttggggcca fTTTTT gaagttaccgtttt
639.	ggaatatcccgtctattcaatctagTTTTTctgagtcaaagcat
640.	tgagactggccacaggattgaTTTTTgaagttaccgtttt
641.	tccaaaatatgtaaaaagcacaaaatTTTTTctgagtcaaagcat
642.	cacgattgcaatacaacttgteaaTTTTTgaagttaccgtttt
643.	ggtattatcatttcagatacaccagtcTTTTTctgagtcaaagcat
644.	gtggtaccgagatatgcattcgTTTTTgaagttaccgtttt

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SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Flu Virus, H1N1
645.	cattcgaagcaactggaaatctaTTTTTctgagtcaaagcat
646.	agtagagccgggagacaaaataaTTTTTgaagttaccgtttt
647.	gggagaatgaactattactggacactTTTTTctgagtcaaagcat
648.	aatgcagatacatatgtttttgtggT TTT T gaagttaccgtttt
649.	tgctgaccaacaaagtctctatcagT TTTTctgagtcaaagcat
650.	ttctacaaaaatttaatatggctagttaaTTTTTgaagttaccgtttt
651.	cctcatgctggagcaaaaagcTTTTTctgagtcaaagcat
652.	ggcccaatcatgactcgaacTTTTTgaagttaccgtttt
653.	gagatattccccaagacaagttcatTTTTTctgagtcaaagcat
654.	tgaggagctaagagagcaattgagTTTTTgaagttaccgtttt
655.	tacccaggagatttcatcgattaTTTTTctgagtcaaagcat
656.	cctagttmgacaatggaacgtgtTTTTTgaagttaccgtttt
657.	catggtcctacattgtegaaacaTTTTTcigagtcaaagcat
658.	tgaatcactctccacagcaagctTTTTTgaagttaccgtttt
659.	ggatcctgggaaatccagagtgTTTTTctgagtcaaagcat
660.	tggacttacaatgccgaactgttTTTTTgaagttaccgtttt
661.	igatgatggtttcctggacattTTTTTctgagtcaaagcat
662.	ggtaaagagttcaaccacctggaTTTTTgaagttaccgtttt
663.	aagatgaatacacagttcacagcagtaTTTTTctgagtcaaagcat
664.	cacacagaatgccattgacgagT T TTTgaagttaccgtttt
665.	atatgcagccgacctgaagagTTTTTctgagtcaaagcat
666.	tggatggtacggttatcaccatTTTTTgaagttaccgtttt
667.	figgtggacagggatggtagaTTTTTctgagtcaaagcat
668.	aggcctatttggggccatTTTTTgaagttaccgtttt
669.	ggaatatcccgtctattcaatctagTTTTTctgagtcaaagcat
670.	tgagactggccacageattgaTTTTTgaagttaccgtttt
671.	tccaaaatatgtaaaaagcacaaaatTTTTTctgagtcaaagcat
.............67T.............	cacgattgcaatacaacttgtcaaTTTTTgaagttaccgtttt
673.	ggtattatcatttcagatacaccagtcTTTTTctgagtcaaagcat
674.	gtggtaccgagatatgcattcgTTTTT’gaagttaccgtttt
675.	cattcgaagcaactggaaatctaTTTTTctgagtcaaagcat
676.	agtagagccgggagacaaaataa'ΤΤΊTT’gaagttaccgtttt
.............677.............	gggagaatgaactattactggacactTT'TTTctgagtcaaagcat
678.	aatgcagatacatatgtttttgtggTTTTTgaagttaccgtttt
679.	tgctgaccaacaaagtctctatcagTTTIT ctgagtcaaagcat
680.	ttctacaaaaatttaatatggctagttaaTTTTTgaagttaccgtttt
681.	cctcatgctggagcaaaaagcTTTTTctgagtcaaagcat
682.	ggcccaatcatgactcgaacTTTTTgaagttaccgtttt
683.	gagatattccccaagacaagttc^TTTTTctgagtcaaagcat
684.	tgaggagctaagagagcaattgagTTTTTgaagttaccgtttt
685.	tacccaggagatttcatcgattaTTTTTctgagtcaaagcat
686.	cctagttcagacaatggaacgtgtTTTTTgaa.gttaccgtttt
687.	catggtcctacattgtggaaacal'TTTT ctgagtcaaagcat
688.	tgaatcactctccacagcaagctTTTTT gaagttaccgtttt
689.	ggatcctgggaaatccagagtgTTTTTctgagtcaaagcat

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Table 71: Blocking Label Extender of Flu Virus, HlN1

SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Flu Virus, H1NI
690.	aaaaagaatagagaatttaaataaaaaagt
691.	attactaacaaagt aaattctgttattgaa
692.	atccgatcacaaitggaaaatg
693.
694.	ccc-&a&gt.gagggatcaa.gaa
695.	ggtcatcaagatacagcaagaagtt
696.	gggcattcaccatccatctactag
697.	gggaaagaagtcclcgtgctaig
698.	tcagcaaatcctacattaatgataaa
699.	aaaaagaatagagaatttaaataaaaaagt
700.	attactaacaaagtaaattctgttattgaa
701.	atccgatcacaattggaaaatg
702.	caatggaaagaaatgciggatct
703.	cccaaagtgagggatcaagaa
704.	ggtcatcaagatacagcaagaagtt
705.	gggcattcaccatccatctactag
706.	gggaaagaagtcctcgtgctatg
707.	tcagcaaatcctacattaatgataaa

[0075] The present invention further provides a method for detecting viral target polynucleotides from a flavirus, for example, HCV. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 72 (underlined). At least one Capture Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 708 to 714 (Tables 72). At least one Label Extender (LE) (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 73 (underlined). At least one Label Extender may comprise, consist essentially of, or may consist of a sequence selected from SEQ ID NOS: 715 to 727 (Table 73). The test may optionally employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 728 to 732 (Table 74).

Table 72: Capture Extender of HCV

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF HCV
__ __708. _ 7097.......	taacgccatggctagacgctttctgcgtgaTTTTTctcttggaaagaaagt___________________ gcgggttgatccaagaaaggacccggtcTTTTTctcttggaaagaaagt
710.	ggcacgeccaaatctccaggcattgaTTTTTctcttggaaagaaagt
711.	agacctcccggggcactcgcaaTTTTTctcttggaaagaaagt
712.	cctaccctcgggctggcgagccTTTTTctcttggaaagaaagt

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SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF HCV
713.	caccccaagccctcattgccatagagTTTTTctcttggaaagaaagt
_	cgagcggaatgtaccccatgagatcggTTTTTctcttggaaagaaagt

Table 73: Label Extender of HCV

SEQ ID NO	LA BEL EXTENDER NUCLEOTIDE SEQUENCE OF HCV
715.	ggfcctggaggctgcacgacactcatacTTTTTctgagtcaaagcatgaagttac
716.	ccgcagaccactatggctctcccgTTTTTctgagtcaaagcatgaagttac
717.	gtcctggcaattccggtgtactcaccgTTTTTctgagtcaaagcatgaagttac
718.	caacactactcggctagcagtctcgcgggTTTTTctgagtcaaagcatgaagttac
719.	caccctatcaggcagtaccacaaggccttTTTTTctgagtcaaagcatgaagttac
720.	ttcgtgctcatggtgcacggtctacgTTTTTctgagtcaaagcatgaagttac
721.	ggtgttacgtttggtttttctttgaggtttagTTTTTctgagtcaaagcatgaagttac
722.	cccctgcgcggcaacaggtaaactccacTTTTTctgagtcaaagcatgaagttac
723.	gtcgcgcgcacacccaacctggTTTTTctgagtcaaagcatgaagttac
724.	ttggggadaggttgt^ccttccacggiTTTTTctgagtcaaagcatgaagttac
725.	acggggtgacaggagccatcctgccTTTTTctgagtcaaagcatgaagttac
726.	ccaaaltgcgcgacctacgccggggTTTTTctgagtcaaagcatgaagUac
727.	aagccgcacgtgagggtatcgatgaccttacTTTTTctgagtcaaagcatgaagttac

Table 74: Blocking Label Extender of HCV

SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF HCV
728.	acttgacgtcctgtgggcggcgg
729.	cgacgatctgaccaccgcccggga
730.	ggttgcgaccgctcggaagtcttccta
731.	ccaggggtacccgggctgagccca
732.	tggggccccaactaggccgagagcc

[0076] The present invention further provides a method for detecting viral target polynucleotides from a coronavirus, for example, severe acute respiratory syndrome (SARS) virus. In such embodiments, at least one Capture Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 75 (underlined). At least one Capture Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 733 to 738 (Table 75). At least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 76 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 739 to 750 (Table 76). The test may optionally employ a Blocking Label (BL), e.g., containing the sequence of SEQ ID NO: 751 (Table 77).

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Table 75: Capture Extender of SARS Virus

[ SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF SARS Virus
| 733.	ccagtaaactgattaactggttcttcaTTTTTctcttggaaagaaagt
i 734.	KtglatgttagcagcalttacaatcaTTTTTctcltggaaagaaagt
735.	ccttttgcalggcaccattgTTTTTctcttggaaagaaagt
i 736.	gcaagattatgtccagaaagcaaaTTTTTctcttggaaagaaagt
737.	gctgccttaagaagctggatgtTTTTTctcttggaaagaaagt
738.	ggtttagcaccaaatatgcctgTTTTTctcttggaaagaaagt

Table 76: Label Extender of SARS Virus

SEQ ID NO I	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF SARS Virus
| 739.	caatctctgattgctcagtagtatcatTTTTTgaagttaccgtttt
§ 740.	ggtgtaggttctggttctggctTTTTTctgagtcaaagcat
i 741.	ggcaacattgtcagtaagttttaaataaTTTTTgaagttaccgtttt
j 742.	tecttaacgatgtcaacacatttaatTTTTTctgagtcaaagcat
L 743.	gctacaccaccaccatgtttcagTTTTTgaagttaccgtttt
ί 744.	gttgccttgttga.gtgcacctTTTTTctgagtcaaagcat
745.	ccatttagcttaatgtaatcatcactcfTTTTTgaagttaccgtttt
746.	caagaccctcctactgtaagagggTTTTTctgagtcaaagcat
747.	ccaacaacatgcagacacttcttaTTTTTgaagttaccgtttt
748.	cctcacctgcatttaggttaggtTITTTctgagfcaaagcat
| 749.	tgtcctgtgaattgaaattttcatatTTTTTgaagttaccgtttt
I 750.	ctgacaacaatggtgcaagtaagaTTTTTctgagtcaaagcai

Table 77: Blocking Label Extender of SARS Virus

SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF SARS Virus
751.	ccataggattagcactttgtgcc
[1)077] In	one aspect of the invention, the method provides for detection of bacterial

target polynucleotides. Non-limiting examples of bacteria are: B. pertussis; Leptospira Pomona; S. paratyphi A and B; C. diphtheriae, C. tetani, C. botulinum, C. perfringens, C. feseri and other gas gangrene bacteria; B. anthracis; P. pestis; P. multocida; Neisseria meningitides; N. gonorrheae; Hemophilus influenzae; Actinomyces (e.g., Norcardia); Acmetobacter; Bacillaceae (e.g., Bacillus anthrasis); Bacteroides (e.g., Bacteroides fragilis); Blastomycosis; Bordetella (Bordetella pertusi); Borrelia (e.g., Borrelia burgdorferi); Brucella: Campylobacter; Chlamydia; Coccidioides; Corynebacterium (e.g., Corynebacterium diptheriae); E. coli (e.g., Enterotoxigenic E. coli and Enterohemorrhagic E. coli); Enterobactcr (e.g. Enterobacter aerogenes); Entcrobacteriaccae (Klebsiella, Salmonella (e.g., Salmonella typhi, Salmonella enteritidis, Serratia, Yersinia, Shigella);

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Erysipelothrix; Haemophilus (e.g., Haemophilus influenza type B); Helicobacter: Legionella (e.g., Legionella pneumophila)·, Leptospira; Listeria (e.g., Listeria monocytogenes)·, Mycoplasma; Mycobacterium (e.g., Mycobacterium leprae, Mycobacterium tuberculosis and Mycobacterium bovis); Vibrio (e.g., Vibrio cholerae): Pasteurellacea (Pasteurella haemolytica); Proteus; Pseudomonas (e.g., Pseudomonas aeruginosa); Rickettsiaceae; Spirochetes (e.g., Treponema spp., Leptospira spp., Borrelia spp.); Shigella spp.; Meningiococcus; Pneumococcus and Streptococcus (e.g., Streptococcus pneumoniae and Groups A, B, and C Streptococci); Ureaplasmas; Treponema pollidum; and Staphylococcus (Staphylococcus aureus and Staphylococcus epidermidis).

[1)078] The target polynucleotides from the bacteria can be a polynucleotide, DNA or a RNA (such as a ribosomal RNA) that is involved in the replication and transcription of the bacteria, housekeeping genes, repetitive sequences, terminal inverted repeat sequence (TIR), miniature inverted repeat transposable element (MITE), CpGs, and/or polynucleotide encoding other bacterial proteins that are expressed on the cell surface or as integral membrane proteins.

[0079] In one embodiment, the present invention provides a method for detecting mycobacterium. The target polynucleotide for detecting mycobacterium infection can be a 16S RNA sequence. At least one Capture Extender (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 78 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 752 to 756 (Table 78). At least one Label Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 79 (underlined). At least one Label Extender comprises, consists essentially of, or consists of a sequence selected from SEQ ID NOS: 757 to 770 (Table 79). The test may optionally employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 771 to 773 (Table 80).

Table 78: Capture Extender of Mycobacterium tuberculosis

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF Mycobacterium tuberculosis
752.	agcgctaaagcgctttcca'ri'Tri^rctcttggaaiigaaagt
753.	ccgcgggctcatcccacTTTTTctcttggaaagaaagt
754.	tggccggacaccctetcaTTTTTctcttggaaagaaagt_____________________________________
755.	agltaagcegtgagatticacgTIΎΊTctcttggaaagaaagt___________________________________
756.	tcgcccgcacgctcacTTTTTctcttggaaagaaagt

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Table 79: Label Extender of Mycobacterium tuberculosis

SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Mycobacterium tuberculosis
757.	cgccaggtaggccgtoTTTTTgaagtteccgtttt________________________________________
758.	ggccggctacccgtcgtT TT IT ctgagtcaaagcat
759.	ggccgtatcteagtccggtgn Ί TTgaagttaccgtttt________________________________________________________________________________________________________
760.	gctgectcccgtaggagtctg FI ΤΓ 1 ctgagtcaaagcat______________________________________
761.	ccattglRcaatattccccactl T1' TTgaagttaccgtttt
762.	gctgcatcaggcttgcgc' IT 1T Tctgagtcaaagcat
763.	ig.gcccacgcgg.cgtcl I T TTgaagttaccgtttt
764.	tacaacccgaaggccgtca T Ί1IT Tctgagtcaaagcat
765.	tgc ttcttctccacc taccgtc' 1ΊΤ1 Tgaagttaccgtttt
766.	tggcacgtagttggccgg IT 1ΊΤ ctgagtcaaagcat
767.	ctacgtattaccgcggctgc 11111 gaag ttaccgtttt
768.	ccg gacaacgctcgcacc 1 TIT 1 ctgagtcaaagcat
769.	cgagctctttacgcccagMT^
770.	aacaacg.cgacaaacca.cctal ΓΊ 1 I ctgagtcaaagcat

Table 80: Blocking Label Extender oi Mycobacterium tuberculosis

; SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Mycobacterium tuberculosis
I 771.	ccccaccaacaagctgatagg
! 772.	ttcgtcgatggtgaaagaggtt
I 773.	aatccgagagaacccggacc

[0080] In another embodiment, the target polynucleotide is indicative of a spirochete bacterium, for example, Treponema pallidum, which causes syphilis, a sexually transmitted disease. In one embodiment, the target polynucleotide for detecting spirochete infection is a 16S RNA sequence. At least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 81 (underlined). At least one Capture Extender comprises, consists essentially of, or consists of a sequence selected from SEQ ID NOS: 774 to 779 (Table 81). At least one Label Extender (LE) (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 82. At least one Label Extender may comprise, consist essentially of or consist of a sequence selected from SEQ ID NOS: 780 to 791 (Table 82). The test may optionally employ a Blocking Label (BL), e.g., containing the sequence of SEQ ID NO: 792 (Table 83).

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Table 81: Capture Extender of Syphilis

SEQ ID NO	C APTURE EXTENDER NUCLEOTIDE SEQUENCE OF Syphilis
I 774.	ccccttacgtgttaccgcgTTTTTctcttggaaagaaagt
| 775.	ccaggcttaccagtccgccTTTTTctcttggaaagaaagt
I 776.	cctccgtgattctagaccagcaTTTTTctcttggaaagaaagt
i 777.	ttc gccaccggtgttcttcTTTTT ctcttggaaagaaagt
i 778.	cgcaccteagcgtcaatcatTTTTTctctiggaaagaaagt
i 779.	taatgcgttcgcgtcggTTTTTctcttggaaagaaagt
Table 82: Label Extender of Syphilis

SEQ ID NO 1	LABEL EXTENDER NUCLEOTIDE SEQUENCE Of Syphilis
[ 780.	ggggcttattcgcacgacf T1Ί T T'gaagttaccgtttt
781.	gctgctggcacgtaattagccTTTTTctgagtcaaagcat
782.	aataattccgaacaacgctcgTTTTTgaagttaccgtttt
1 783.	tgcatgccctttacgcccTTTTTctgagtcaaagcat
| 784.	ttgagctcggggatttcacaTTTTTgaagttaccgtttt
785.	gtacccagtgcagttcccaagTTTTTctgagtcaaagcat
I 786.	cacttggaattccggtttccTTTT'Tgaagttaccgtttt
| 787.	caaatatc taca gattccacccctaTTTTTctgagtcaa agcat
1 788.	tgttcgctccccacaccttTTTTT gaagttaccgtttt
789.	tgtggactaccagggtatctaatccTTTTTctgagtcaaagcat
| 790.	caacacctagtgtacatcgtttactgTTTTTgaagttaccgtttt
| 791.	cgccgagactcatgcccTTTTTctgagtcaaagcat
	Table 83: Blocking Label Extender Syphilis

SEQ ID NO	BLOCKING LABEL EXTENDER. NUCLEOTIDE SEQUENCE OF Syphilis
ί 792	cggccagaaacccgcc

[0081] In another aspect of the present invention, the method also provides for detection of fungal target polynucleotide. Non-limiting examples of fungal infections include: aspergillosis (Aspergillus fumigatus, Aspergillus flavus, Aspergillus terreus, Aspergillus nidulans, and Aspergillus niger); blastomycosis (Blastomyces dermatitidis);, coccidioidomycosis (Coccidioides species); cryptococcosis (Cryptococcus neoformans, C. gattii); histoplasmosis (Histoplasma capsulatum): mucormycosis or zygoniycosis (Mucorales molds); paracoccidioidomycosis (Paracoccidioides brasiliensis)·, pneumocystis pneumonia sporotrichosis vche/ickd); etc.

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2019247099 14 Oct 2019 [0082] In one embodiment, die present invention provides a method for detecting fungal target polynucleotides from an Aspergillus, for example, Aspergillus fumigatus. A. fumigatus is a common, often lethal and difficult to diagnose cause of pathogenic disease in immunocompromised subjects such as in organ transplant recipients, bone marrow transplant and subjects diagnosed with cancer and AIDS. In one embodiment, the target polynucleotide for detecting aspergillus infection is a 18S RNA or 28S RNA sequence. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 84. At least one Capture Extender probe comprises, consists essentially of, or consists of a sequence selected from SEQ ID NOS: 793 to 798 (Table 84). At least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 85. At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 799 to 810 (Table 85). The test may optionally employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 811 to 813 (Table 86).

Table 84: Capture Extender of Aspergillus fumigatus

SEQ ID NO i	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF Aspergillus fumigatus
j 793.	actcgcggtgaggcggaTTTTTctcttggaaagaaagt
794.	aaggtccagccggaccagtTTTTTctcttggaaagaaagt
i 7QS	catgaggttccccagaaggaTTTTTctcttggaaagaaagt
j 796.	cagctgaatactgacgccccTTTTTctcttggaaagaaagt
| 797.	gaaaacatccttggcgaateTTTTTctcttggaaagaaagt
| 798.	cgagcgggtcatcatagaaacTTTTTctcttggaaagaaagt

Table 85: Label Extender Aspergillus fumigatus

| SEQ ID NO i	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Aspergillus fumigatus
799.	^gtteaactacgagctttttaactgTTTTTgaagttaccgtttt
i 800.	ccggccagccagacccaT'TTTTctgagtcaaagcat
801.	gcctgctttgaacactctaattttTTTTTgaagttaccgtttt
i 802.	catgctaa t gtattcgagca a a gTTTTTctgagtca aagcat
803.	cggt^.tagaaMcaacaaaatagaTTTTTgaagttaccgtttt
804.	cgactatccctattaatcattacggTTTTTctgagtcaaagcat
i 805.	aaatccaagaatttcacctctgaTTTTTgaagttaccgtttt
ΐ 806.	ctttcgcagtagttagtcttcagcTTTTTctgagtoaaagcat
807.	cc taac tttc gttcc c igattaatTTTTT gaagttaccgtttt
808.	cggtatctgatcgtcttcgatccTTTTTctgagtcaaagcat
809.	ggcatagtttatggttaagactacgaTTTTTgaagttaccgtttt
1 «tn	accgcccgatccctagtcTTTTTctgagtcaaagcat

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Table 86: Blocking Label Extender of Aspergillus fumigates

SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Aspergillus fumigatus
811.	ccccacagccagtgaaggc
812.	ttcacagtaaaagtcc tggttcc
813.	accgcacgtcctattctattatlc

[0083] In another embodiment, the fungal target polynucleotides are from Candida, for example, Candida albicans, an opportunistic yeast which causes oral and genital infections in human, particularly in immuno-compromised subjects (e.g. organ and bone marrow transplantation recipients, cancer and AIDS patients). In one embodiment, the target polynucleotide for detecting Candida infection is a 18S or 28S RNA sequence. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 87 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 814 to 819 (Table 87). At least one Label Extender probe (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 88 (underlined), At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 820 to 831 (Table 88). The test may optionally employ a Blocking Label (BL), e.g., at least one or two BLs, such as the BLs of SEQ ID NOS: 832 to 833 (Table 89).

Table 87: Capture Extender of Candida albicans

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF Candida albicans
814.	aaaaagatggaccggccagTTTTTctcttggaaagaaagt
815.	acccagaaggaaaggctcgTTTTTctcttggaaagaaagt
816.	tggttcgccataaatggctTTTTTctcttggaaagaaagt
817.	actgatacccccgaccgtcTTTTTctcttggaaagaaagt
818.	aaacgtccttggtaaatgctttcTTTTTctcttggaaagaaa.gt
819.	gtgccgattgcgtcaataaaaTTTTTctcttggaaagaaagt

Table 88: Label Extender of Candida albicans

SEQ ID NO	LA BEL EXTENDER NUCLEOTIDE SEQUENCE OF Candida albicans
820.	gagctttttaactgcaacaactttaataTTTTTgaagttaccgtttt
821.	ccaagcccaaggttcaactacTTTTTctgagtcaaagcat
822.	gagcaaaggcctgctttgaaTTTTTgaagttaccgtttt
823.	cctattctattattccatgctaatatattcTTTTTctgagtcaaagcat
824.	aaccaacaaaatagaaccataacgtTTTTTgaagttaccgtttt
80S	cctattaatcattacgatggtcctagaTTTTTctgagtcaaagcat

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SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Candida albicans
826.	agaatttcacctctgacaactgaatTTTTTgaagttaccgtttt
827.	gcagtagttagtcttc.agtaaatccaTTTTTctgagtcaaagcat
828.	cctaactttcgttettgattaatgaTTTTTgaagttaccgtttt
829.	cggtatctgatcatcttcgatccTTTTTctgagtcaaagcat
830.	ggca,tagtttatggttaagactacgaTTTTTgaagttaccgt.ttt ν^***“****Μν“*****“*****<Λ^*** CP CP CP
831.	gaacaacaacc gatccctagtcTTTTT ctgagtcaaagcat

Table 89: Blocking Label Extender of Candida albicans

SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Candida albicans
832.	gctggglccagtacgcatc
833.	cactctaattttttcaaagtaaaagtcc
[0084] In In particular,	another embodiment, the fungal target polynucleotides are from Cryptococcus. the target polynucleotides are from Cryptococcus neoformans, a medically

important species ofthe Crytococcus best known for causing a severe form of meningitis and meningo-encephalitis in subjects diagnosed with HIV-AIDS. The fungus is also known to infect and cause moderate to severe disease in patients undergoing cancer chemotherapy and metabolic immunosuppression etc. In one embodiment the target polynucleotide for detecting Cryptococcus infection is an 18S RNA sequence, In such embodiments, at least one Capture Extender (e.g., at least two, three, four, or five) comprises a target-hybridizing sequence selected from Table 90 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 834 to 839 (Table 90). At least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence selected from Table 91 (underlined). At least one Label Extender may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 840 to 851 (Table 91). The test may optionally employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 852 to 854 (Table 92).

Table 90: Capture Extender of Crytococcus neofomms

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUEf	4CE OF Crytococcus neoformans
834.	gcctcgccagacctgaagtfFTTTTctcttggaaagaaagt
835.	gcactccgtgaggaggaccTTTTTctcttggaaagaaagt
836.	ggttcccctgcacacccagTTTTTctcttggaaagaaagt
837.	gcgattgcctgctttgaacacTTTTTctcttggaaagaaagt
838.	cggcatcgtttactgttaagactaTTTTTctcttggaaagaaagt

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839.cgtgggccgatccctagtTTTTTctettggaaagaaagt

Table 91: Label Extender of Crytococcus neoformans

SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF Crytococcus neoformans
840.	gaggtaaggtccagcaagacagtTTTTTgaagttaccgtttt
841.	taaagagca .tacagga ccaccagTTTTTctgagtcaaagca t
842.	tattattccatgctaatgtattcggTTTTTgaagttaccgtttt
843.	aaatagaaccgcacgtcctaltcTTTTTclgagtcaaagcat
844.	cggcgatcctagaaaccaacaTTTTTgaagttaccgtttt
845,	ccgaccgtccctattaatcattaTTTTTctgagtcaaagcat
846.	ccgtcaatctaagaatttcacctctaTTTTTgaagttaccgtttt
847.	gctttcgcagttgttggtcttTTTTTctgagtcaaagcal
848.	ccctaaccttcgttcttgatca'T TTT Ί 'gaagttaccgtttt
849.	caac ggtatctante gtttttgateTT TTT etgagte aaagcat
850.	gccgacccaglcagagattgaTTTTTgaagttaccgtttt
851.	caaagactttgatttctcgtaaggtTTTTTctgagtcaaagcat

Table 92: Blocking Label Extender of Crytococcus neoformans

SEQ ID NO	BLOCKING LABEL EXTENDER. NUCLEOTIDE SEQUENCE OF Crytococcus neoformans
852.	totaattttttcaaggtaaaattcct
853.	gcaacggaataccaatgccc
854.	atgaaaacgtccttggcaaat

[0085] The present invention provides methods for detection of target polynucleotides from uni- and/or multicellular parasites or protozoa. Non-limiting examples of parasites or protozoa include: leishmaniasis (Leishmania tropica mexicana, Leishmania tropica, Leishmania major, Leishmania aethiopica, Leishmania braziliensis, Leishmania donovani, Leishmania infantum, Leishmania chagasi); trypanosomiasis (Trypanosoma brucei gambiense, Trypanosoma brucei rhodesiense); toxoplasmosis (Toxoplasma gondii); schistosomiasis (Schistosoma haematobium, Schistosoma japonic urn. Schistosoma mansoni, Schistosoma mekongi, Schistosoma intercalatuni); malaria (Plasmodium virax, Plasmodium falciparium, Plasmodium malariae and Plasmodium ovale); Amebiasis (Entamoeba histolytica); Babesiosis (Babesiosis microti); Cryptosporidiosis (Cryptosporidium parvum); Dientamoebiasis (Dientamoeba fragilis); Giardiasis (Giardia lamblid); Helminthiasis and Trichomonas (Trichomonas vaginalis).

[0086]

The method of the present invention described above may include detection of a housekeeping gene polynucleotide, where the level of the housekeeping gene polynucleotide is used for normalization (e.g., across samples) and/or calculation of the copy number of the

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PCT/US2010/040649 target polynucleotide. In certain embodiments, the housekeeping gene polynucleotide is glyceraldehyde 3-phosphate dehydrogenase (GAPDH) polynucleotide. In such embodiments, at least one Capture Extender probe (e.g., at least two, three, four, or five) comprises a target hybridizing sequence selected from Table 93 (underlined). At least one Capture Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 855 to 858 (Tables 93). In such embodiments, at least one Label Extender (e.g., at least two, three, four, or five) may comprise a target-hybridizing sequence shown in Tables, 1, 2 or 94 (underlined), At least one Label Extender probe may comprise, consist essentially of, or consist of a sequence selected from SEQ ID NOS: 2 to 7 (Table 1), 8 to 13 (Table 2). or 859 to 870 (Table 94). The test may further employ a Blocking Label (BL), e.g., at least one, two, or three BLs each containing a sequence selected from SEQ ID NOS: 871 to 875 (Table 95).

Table 93: Capture Extender of GAPDH

SEQ ID NO	CAPTURE EXTENDER NUCLEOTIDE SEQUENCE OF GAPDH
855.	ggtcaatgaaggggtcattgatTTTTTctcttggaaagaaagt
856.	ccttgacggtgccatggaatTTTTTctcttggaaagaaagt
857.	cgccccacttgattttggaTTTTTctcttggaaagaaagt
858.	gagatgatgacccttttggctcTTTTTctcttggaaagaaagt

Table 94: Label Extender of GAPDH

SEQ ID NO	LABEL EXTENDER NUCLEOTIDE SEQUENCE OF GAPDH
859.	ggagcagagagcgaagcggTTTTTgaagttaccgtttt
860.	cggctgactgtcgaacaggaTTTTTctgagtcaaagcat
861.	gagcgatgtggctcggcTTTTTgaagltaccgtttt
862.	tcaccttccccatggtgtctTTTTTctgagtcaaagcat
863.	ccaggcgcccaatacgacTTTTTgaagttaccgtttt
864.	agttaaaagcagccctggtgaTTTTTctgagtcaaagcat
865.	tggaacatgtaaaccatgtagttgaTTTTTgaagttaccgtttt
866.	ttgccatgggtggaatoatatTTTTTctgagtcaaagcat
867.	tgacaagcttcccgttctcagTT ΤΊ Tgaagttaccgtttt
868.	tegLsatgg.gatttccattgaTTTTTctgagtcaaagcat
869.	gacgtactcagcgccagcatTTTTTgaagttaccgtttt
870.	aagacgccagtggactccac' FIT ΤΊctgagtcaaagcat

Table 95: Blocking Label Extender of GAPDH
SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF GAPDH
871.	caaatccgttgactccgacct

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SEQ ID NO	BLOCKING LABEL EXTENDER NUCLEOTIDE SEQUENCE OF GAPDH
872.	ggcaacaatatccactttaccag
! 873.	gggatctcgctcctggaaga
874.	cagccttctccatggtggtg
| 875.	ccccctgcaaatgagccc

[0087] According to another aspect, the present invention provides a kit for detecting target polynucleotides, for example, associated with a pathogenic organism. The kit comprises at least one Capture Extender probe and at least one Label Extender probe specific for a target polynucleotide from an infectious agent as described above. The kit may comprise a set of Capture Extender and/or Label extender probes illustrated or represented by the Tables provided herein. The kit may further comprise at least one Blocking Label probe specific for the target polynucleotide, or a set of Blocking Label probes illustrated or represented by the Tables provided herein. The kit may further comprise at least one or a series of signal-amplifying probes as described.

[0088] In some embodiments, the kit further comprises components for conducting a positive control. For example, the positive control may be a cell lysate derived from HeLa cells, a cell-line derived from cervical cancer cells, or any commercially available cell line having a particular characteristic of interest.

[0089] In some embodiments, the kit further comprises a probe pair or set as a normalization control, to normalize across samples and to calculate the target polynucleotide copy number. The normalization control probes may comprise at least one Capture Extender probe and at least one Label Extender probe specific for one or more housekeeping gene polynucleotides, such as but not limited to, GAPDH (GAPD). Exemplary pairs or sets of Capture Extender probes and Label Extender probes for GAPDH are disclosed in Tables 1, 2, 93, and 94 above. The kit may further comprise Blocking Label Extenders disclosed in Table 95 above.

[0090] In certain embodiments, the kit is configured such that the level of target polynucleotide and level of normlaization control are read simultaneously or sequentially from a patient sample in the same reaction. Such may be accomplished by employing different labels, such as alkaline phosphatases having different luminescent substrates, or combinations of different signal generating labels (e.g., alkaline phosphatase and horseradish peroxidase).

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EXAMPLES

Example 1: Comparison of currently available HPV-tests [0091]

HPV-tests.

TABLE 96 shows a comparison of the invention with currently available

TABLE 96:

I Company and Product	HPV test based on	Advantages/Disadvantages
1 Present invention ! I I I 1 !	Use of primary probes hybridizing to the target sequence without amplification. Signal is intensified by additional probes which hybridize to the primary probes, creating a multilayer probe,	No purification of RNA or DNA required from PAP smears. No need for RT-PCT. Amplifies signals, not target.
j Access Genetics i	Standard PCR technology	Sensitive. Contamination issues
| Hologic (3rc wave) Invader® HPV i	3 specifically designed oligonucleotides and a fluorescent signal detection technology	Poor specificity
1 Qiagen/Diagene Hybrid | Capture 2® (HC2) assay I I i !	Enzyme-linked antibody detection of RNA/DNA hybrid technology	Requires purification of RNA or DNA. RNA/DNA hybrid must be captured on solid phase. Poor specificity and sensitivity Expensive
| Ventana’s Inform® HPV 1	In-situ hybridization technology	Requires intervention by eye Not sensitive

Example 2: Protocol [0092] The schematic in FIG, 1 illustrates the method of the invention. Briefly, the method involves capturing target polynucleotides from a sample by contacting with a Capture

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Extender and Label Extender under hybridizing conditions (e.g,, about 55°C for 120 minutes in 3x SSC, 10% dextransulfate, 0.2% casein, 10 qg/mL polyA and 100 pg/mL denatured salmon sperm DNA). Signal amplification is then carried out by sequentially hybridizing a pre-Amplifier, Amplifier Probe and Label Probe at 55°C, 55°C and 50°C respectively for 30 minutes each in 3x SSC, 10% dextran-sulfate, 0.2% casein, 10 pg/mL polyA and 100 pg/mL denatured salmon sperm DNA and with wash buffer: 20mmol/L Tris-HCL, 400 mmol/L lithium chloride, 1 mL/L Tween 20. The Label Probe may be directly conjugated with alkaline phosphatase, or biotinylated, labeled with fluorescein, or other light emitting dyes.

Example 3: Correlation of Relative Light Units (RLU) with HPV type 16 sequence concentration [0093] Samples containing various concentrations of HPV type 16 ss DNA sequences were incubated with Capture Probe-coated plates together with HPV type 16-specific Capture Extender (CE), Label Extender (LE) and Blocking Label (BL) for 1, 2, 3, 4 h and overnight. Probe sets are identified in Tables 3 to 14. TABLE 97 and Figure 2 below show the correlation of RLU with the concentration of viral DNA.

TABLE 97:

	RLU1	RLU2	AVE	AVE-BK	std
| 50amol	957.753	957.403	957.578	957.0605	0.25
i 5amol	862.611	662.097	762.354	761.8365	141.78
| O.Samol	220.362	234.933	227.6475	227.13	10.30
i 0.05amol	42.555	45.472	44.0135	43.496	2.06
i 0.005amol	5.301	5.373	5.337	4.8195	0.05
Background	0.46	0.575	0.5175	0	0.08

[0094] The results show that the method of the invention using the probe set described for detecting HPV has a sensitivity of about 3000 copies, which, given the high specificity of the probe set, is sufficient for accurate HPV detection.

Example 4: Detection of HPV in patient samples at various stages of infection [0095] FIG. 3 shows the detection of HPV target polynucleotides using the method of the invention. HPV target polynucleotides in patient Pap smear samples at various stages of HPV infection, i.e. at the CIN (cervical intraepithelial neoplasia) stage 1, 2, or 3; or at the ASC-US (atypical squamous cells of undetermined significance) stage in high risk and low risk natients were detected using the probe sets identified in Tables 3 to 14.

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The results in TABLE 98 and FIG. 3 show an increase in levels of HPV target polynucleotides in Pap smears of patients with C1N 3 infection.

TABLE 98:

I	ASCUS	ASCH	CIN1	CIN2	CIN3(1)	BK
1 High risk	0.8965	6.3085	15.0365	29.804	168.932	0.8675
Low risk	1.1925	0.9135	0.655	2.724	2.8915	0.8225

[0097] ASC, which are abnormal cells of uncertain significance, and CIN1 usually recover spontaneously. The CIN1 have a few cells showing intraepithelial neoplasia which can be precancerous. CIN3 lesions represent severe neoplasia involving the full thickness of the mucosa and this class includes carcinoma in situ, a pre-invasive cancer. CIN3 lesions are less likely to recover spontaneously and commonly progress to invasive cancer. Background levels (BK) were very close to that of the lowest abnormal cell of uncertain significance grade.

Example 5: Detection Sensitivity fur HIV [0098] Figure 4 shows the detection sensitivity for HIV target polynucleotides (HIV-A, HIV-B, and HIV-F). HIV target polynucleotides w^rere detected using the probe set identified in Tables 60 to 68 in serially diluted plasma samples, spiked with 25, 50, 100 and 200 copies of the HIV target polynucleotide.

[0099] The results show that this embodiment can detect as low as 25 copies. In addition, Figure 4 shows that the assay is linear for samples that are serially diluted.

Example 6: Detection Sensitivity for HBV [00100] Figure 5 shows the sensitivity of detection for HBV target polynucleotides. HBV target polynucleotides were detected using the probe set from Table 57, Table 58 and Table 59.

[00101] The results show that the method of detection in this embodiment, measured by log relative light units (RLU), is linearly correlated with the log number of copies of HBV.

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Example 7: Detection Sensitivity for HCV [00102] Figures 6A and B show the sensitivity of detection for HCV target polynucleotides. HCV target polynucleotides were detected using the probe set from Table 72, Table 73 and Table 74.

[00103] Figure 6A shows that, in this embodiment, log relative light units (RLU) is linearly correlated with the log number of copies of HCV.

[00104] Figure 6B show's the quantitation of HCV in 6 patient samples.

Example 8: Detection Sensitivity for Coronavirus (SARS) [00105] Figure 7 shows the detection sensitivity for coronavirus (SARS) target polynucleotides. SARS target polynucleotides were detected using the probe sets from Table 75, Table 76 and Table 77.

Example: 9 Lack of Cross-Reactivity for H1NI Assay [00106] An assay for hemaglutmin-neuraminidase (H1N1) of Influenza virus was conducted. The HIM target polynucleotides were detected using the following probe sets from Table 69, Table 70 and Table 71.

[00107] TABLE 99 shows that the H1N1 assay does not cross-react with various strains of Influenza A, Influenza B or other respiratory virus.

TABLE 99:

Viruses Tested in Samples	Result
Influenza A:
Brisbane/!0/2007 H3N2	negative
Brisbane/59/2007 Η INI	negative
Hong Kong/29/2006	negative
Netherlands/134/04 H1	negative
New Caledonia/20/99 H1N1	negative
Solomon Islands/03/2006 H1N1	negative
Taiwan/42/2006	negative
Victoria/3/75 H3	negative
Wisconsin,''67/05/H3N2	negative
Singapore/63/04	negative
Influenza B:
Florida/04/2006	negative

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Viruses Tested in Samples	Result
Florida/07/2004	negative
Malaysia/2506/2004	negative
Panama/45/90	negative
Yamanashi/166/98	negative
Other Respiratory Viruses’.
Parainfluenza 1	negative
Parainfluenza 2	negative
Parainfluenza 3	negative
Parainfluenza 4a	negative
Parainfluenza 4b	negative
Coronavirus 229E	negative
Coronavirus NL63	negative
Corona virus OC43	negative
SARS (200300592)	negative

Example: 10 ssDNA complementary to 18S RNA or Fungal cells as standard materials [00108] Figure 8 shows the correlation between the numbers of fungal cells and 18S RNA copies using the probe sets described in Tables 84 to 89. Both fungal cells and ssDNA complementary to 18S RNA were tested for their usefulness as standard materials. The results show a correlation between the numbers of fungal cells and 18S RNA copies with the level of luminescence (R> 0.95) detected.

Example: 11 Quantifying different species and strains of Aspergillus and Candida [00109] Figure 9 show s the quantitation of different species and strains of Aspergillus and Candida. Various species of Aspergillus and Candida were detected using the 18S RNA probe set for Aspergillus (Table 84, Table 85, Table 86) and for Candida (Table 87, Table 88, and Table 89 respectively and the method described above.

[00110] The results show that the method of detection measured by log relative light units (RLU) can detect as few as 10 cells/well and there is no cross-reactivity between the 16S RNA probes from Aspergillus and Candida. The results show that the Aspergillus 16S RNA probe-set. consistently detects various species of Aspergillus and not Candida, and the Candida 16S RNA probe-set detects only the Candida species and not Aspergillus. On the other hand, the common 18S RNA probe-set can detect either Candida or Aspergillus of various strains allowing for a single test that detects many potential fungal pathogens.

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Example: 12 Quantification Candida in Clinical Blood Samples [00111] Figure 10 shows the quantification of Candida. Clinical blood samples from patients infected with Candida were assayed for the Candida target polynucleotides using the 18S RNA probe set Candida and a “common probe” from Table 84. Table 85, Table 86, Table 87, Table 88, and Table 89.

[00112] The results show detection measured by log relative light units (RLU). A 50 ul blood sample for an uninfected patient (negative) or infected patient blood sample (positive) was tested. The clinical culture within 24-48 hr showed that two of them were negative and one positive. The results (Fig 10) of QuantiFungi^rM assay, which took only 5 hours, were consisten t with the results of clinical microbes culture.

[0100] All publications, patents and patent applications herein are incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

[0101] The foregoing detailed description has been given for clearness of understanding only and no unnecessary limitations should be understood therefrom as modifications will be obvious to those skilled in the art. It is not an admission that any of the information provided herein is prior art or relevant to the presently claimed inventions, or that any publication specifically or implicitly referenced is prior art.

[0102] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0103] While the invention has been described in connection with specific embodiments thereof, it will be understood that, it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains and as may be applied to the essential features hereinbefore set forth and as follows in the scope of the appended claims.

Claims (107)

1. We claim:

   1. A method for determining the presence, absence, or level of an infectious agent in a sample comprising the steps of:

   (a) capturing target polynucleotides, if present, from a sample, wherein the target polynucleotide is indicative of the presence of said infectious agent; and (b) detecting the target polynucleotides directly or indirectly by hybridization with a signal-amplifying polynucleotide probe, wherein the method allows detection of the target polynucleotide at low copy number to allow detection of early stage infection.
2. 2. The method of claim 1, wherein the signal-amplifying polynucleotide(s) comprise branched DNA.,
3. 3. The method of claim 1, further comprising a second target polynucleotide as a normalization control.
4. 4. The method of claim 3, wherein the normalization control polynucleotide is a GAPDH (GADH) polynucleotide.
5. 5. The method of claim 3 or 4, further comprising, determining a relative level or a copy number of the target pol ynucleotide across a plurality of samples,
6. 6. The method of any one of claims 1 to 5, further comprising, detecting the target polynucleotide and normalization control in a positive control sample, such as a HeLa cell lysate.
7. 7. The method of any one of claims 1 to 6, wherein two or more target polynucleotides are detected.
8. 8. The method of claim 7, wherein the two target polynucleotides are detected with an alkaline phosphatase-labeled probe and a horseradish peroxidaselabeled probe.
9. 9. The method of any one of claims 1 to 8, wherein the target polynucleotides are captured by hybridization to a Capture Extender probe having at least a first sequence and a second sequence, the first sequence hybridizing to the target polynucleotide and the second sequence hybridizing to an immobilized capture probe.

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10. 10. The method of claim 9, wherein a the Pre-Amplifier probe is hybridized to a Label Extender probe having a first sequence and a second sequence, the first sequence hybridizing to the target polynucleotide and the second sequence hybridizing to the Pre-Amplifier probe.
11. 11. The method of claim 10, wherein a plurality of Label Extender probes, with the same or different first sequence, are bound to the target polynucleotide.
12. 12. The method of claim 10, wherein the Label Extender probe is in a cruciform configuration or a double Z configuration.
13. 13. The method of claim 10, wherein the Pre-Amplifier comprises SEQ ID NO: 1.
14. 14. The method of claim 10, wherein Amp lifer probes are hybridized to the PreAmplifer probes, and wherein the ratio of hybridized Amplifier probes to hybridized Pre-Amplifier probes is at least 5:1.
15. 15. The method of claim 14, wherein Label probes are hybridized to the Amplifier probes, and wherein the ratio of hybridized Label probes to hybridized Amplifier probes is at least 10:1.
16. 16. The method of claim 15, wherein the Label probe comprises a detectable entity.
17. 17. The method of claim 16, wherein the detectable entity is a luminescence-, chromogenic- or fluorescence- generating moiety.
18. 18. The method of claim 17, wherein the detectable entity is alkaline phospatase, Horseradish Peroxidase, streptavidin, biotin, or fluorescein.
19. 19. The method of claim 10, wherein the signal-amplifying probe(s) amplify the signal by at least about 25x with respect to the number of hybridized Label Extender probes.
20. 20. The method of claim 19, wherein the signal-amplifying probe(s) amplify the signal by at least about 40x.
21. 21. The method of claim 20, wherein the signal-amplifying probe(s) amplify the signal by at least about 50x.
22. 22. The method of claim 20, wherein the signal-amplifying probe(s) amplify the signal by at least about 1 OOx,

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23. 23. The method of claim 20, wherein the signal-amplifying probe(s) amplify the signal by at least about 200x.
24. 24. The method of claim 20, wherein the signal-amplifying probe(s) amplify the signal by at least about 400x.
25. 25. The method of claim 20, wherein the signal-amplifying probe(s) amplify the signal by at least about 800x,
26. 26. The method of any one of claims 1 to 25, wherein the method detects less than 1000 copies of the target polynucleotide.
27. 27. The method of claim 26, wherein the method detects less than 800 copies of the target polynucleotide.
28. 28. The method of any one of claims 1 to 27, wherein the infectious agent is a vims, a bacterium or a fungus.
29. 29. The method of claim 28, wherein the infectious agent is a DNA virus or an RNA virus.
30. 30. The method of claim 28, wherein the infectious agent is a hepatitis virus or a papilloma virus.
31. 31. The method of claim 28, wherein the infectious agent is a human immunodeficiency virus (HIV), an orthomyxovirus (flu virus), or a corona virus (SARS).
32. 32. The method of claim 28, wherein the papilloma virus is a high risk HPV selected from HPV type 16, 18, 31. 33, 35, 39, 45, 51, 52, 56, 58, 59 and 68.
33. 33. The method of claim 28, wherein the papilloma virus is a low risk HPV selected from HPV type 6, .11,40, 42, 43, or 44.
34. 34. The method of claim 28, wherein the infectious agent is a hepatitis A virus, a hepatitis B virus (HBV), or a hepatitis C vims (HCV).
35. 35. The method of claim 32, wherein the detection of high risk HPV is carried out using at least one Capture Extender probe selected from SEQ ID NO.14 to 21, 49 to 56, 76 to 82, 100 to 105, .128 to 134, 155 to 160, 181 to 187, 207 to 212, 239 to 245, 263 to 269, 287 to 292 or 318 to 324, and at least one Label Extender probe selected from SEQ ID NO. 22 to 37, 57 to 72, 83 to 94, 106 to

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    117, 135 ίο 145, 161 to 172, 188 to 201, 213 to 224, 246 to 257, 270 to 281, 293 to 304, or 325 to 338.
36. 36. The method of claim 32, wherein the high risk HPV is HPV type-16, and the at least one Capture Extender probe is selected from SEQ ID NO. 14 to 21, and at least one Label Extender probe is selected from SEQ ID NO. 22 to 37 and optionally at least one blocking label extender is selected from SEQ ID NO. 38 to 48.
37. 37. The method of claim 32, wherein the high risk HPV is HPV type-18, and the at least one Capture Extender probe is selected from SEQ ID NO. 49 to 56, and at least one Label Extender probe is selected from SEQ ID NO. 57 to 72, and optionally at least one blocking label extender is selected from SEQ ID NO. 73 to 75.
38. 38. The method of claim 32, wherein the high risk HPV is HPV type-31 and the at least one Capture Extender is selected from SEQ ID NO. 76 to 82, and at least one Label Extender is selected from SEQ ID NO. 83 to 94, and optionally at least one blocking label extender is selected from SEQ ID NO. 95 to 99.
39. 39. The method of claim 32. wherein the high risk HPV is HPV type-35 and the at least one Capture Extender probe is selected from SEQ ID NO. 100 to 105, and at least one Label Extender probe is selected from SEQ ID NO. 106 to 117, and optionally at least one blocking label extender is selected from SEQ ID NO. 118 to 127.
40. 40. The method of claim 32, wherein the high risk HPV is HPV type-39 and the at least one Capture Extender probe is selected from SEQ ID NO. 128 to 134, and at least one Label Extender probe is selected from SEQ ID NO. 135 to 145, and optionally at least one blocking label extender is selected from SEQ ID NO. 146 to 154.
41. 41. The method of claim 32, wherein the high risk HPV is HPV type-45 and the at least one Capture Extender probe is selected from SEQ ID NO. 155 to 160, and at least one Label Extender probe is selected from SEQ ID NO. 161 to 172, and optionally at least one blocking label extender is selected from SEQ ID NO. 173 to 180.

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42. 42. The method of claim 32, wherein the high risk HPV is HPV type-51 and the at least one Capture Extender is selected from SEQ ID NO. 181 to 187, and at least one Label Extender is selected from SEQ ID NO. 188 to 201, and optionally at least one blocking label extender is selected from SEQ ID NO. 202 to 206.
43. 43. The method of claim 32, wherein the high risk HPV is HPV type-52 and the at least one Capture Extender is selected from SEQ ID NO. 207 to 212, and at least one Label Extender is selected from SEQ ID NO. 213 to 224, and optionally at least one blocking label extender is selected from SEQ ID NO. 225 to 238.
44. 44. The method of claim 32, wherein the high risk HPV is HPV type-56 and the at least one Capture Extender is selected from SEQ ID NO. 239 to 245, and at least one Label Extender is selected from SEQ ID NO. 246 to 257, and optionally at least one blocking label extender is selected from SEQ ID NO. 258 to 262.
45. 45. The method of claim 32, wherein the high risk HPV is HPV type-58 and the at least one Capture Extender is selected from SEQ ID NO. 263 to 269, and at least one Label Extender is selected from SEQ ID NO. 270 to 281, and optionally at least one blocking label extender is selected from SEQ ID NO. 282 to 286.
46. 46. The method of claim 32. wherein the high risk HPV is HPV type-59 and the at least one Capture Extender is selected from SEQ ID NO. 287 to 292, and at least one Label Extender is selected from SEQ ID NO. 293 to 304, and optionally at least one blocking label extender is selected from SEQ ID NO. 305 to 317.
47. 47. The method of claim 32, wherein the high risk HPV is HPV type-68 and the at least one Capture Extender is selected from SEQ ID NO. 318 to 324, and at least one Label Extender is selected from SEQ ID NO. 325 to 338, and optionally at least one blocking label extender is selected from SEQ ID NO. 339 to 347.
48. 48. The method of claim 33, wherein the detection of low risk HPV is carried out using at least one Capture Extender probes selected from SEQ ID NO. 348 to

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    354, 376 to 382, 404 to 410, 433 to 439, 463 to 468, or 489 to 494, and at least one Label Extender probe selected from SEQ ID NO. 355 to 368, 383 to 398, 411 to 424, 440 to 451, 469 to 482, or 495 to 506.
49. 49. The method of claim 33, wherein the low' risk HPV is HPV type-6, and the at least one Capture Extender probe is selected from SEQ ID NO. 348 to 354, and at least one Label Extender probe is selected from SEQ ID NO. 355 to 368 and optionally at least one blocking label extender is selected from SEQ ID NO. 369 to 375.
50. 50. The method of claim 33, wherein the low risk HPV is HPV type-11, and the at least one Capture Extender probe is selected from SEQ ID NO. 376 to 382, and at least one Label Extender probe is selected from SEQ ID NO. 383 to 398, and optionally at least one blocking label extender is selected from SEQ ID NO. 399 to 403.
51. 51. The method of claim 33, wherein the low risk HPV is HPV type-40 and the at least one Capture Extender is selected from SEQ ID NO. 404 to 410, and at least one Label Extender is selected from SEQ ID NO. 411 to 424, and optionally at least one blocking label extender is selected from SEQ ID NO. 425 to 432.
52. 52. The method of claim 33, wherein the low risk HPV is HPV type-42 and the at least one Capture Extender probe is selected from SEQ ID NO. 433 to 439, and at least one Label Extender probe is selected from SEQ ID NO. 440 to 451, and optionally at least one blocking label extender is selected from SEQ ID NO. 452 to 462.
53. 53. The method of claim 33, wherein the low risk HPV is HPV type-43 and the at least one Capture Extender probe is selected from SEQ ID NO. 463 to 468, and at least one Label Extender probe is selected from SEQ ID NO. 469 to 482, and optionally at least one blocking label extender is selected from SEQ ID NO. 483 to -488.
54. 54. The method of claim 33, wherein the low risk H PV is HPV type-44 and the at least one Capture Extender probe is selected from SEQ ID NO. 489 to 494, and at least one Label Extender probe is selected from SEQ ID NO. 495 to

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    506, and optionally at least one blocking label extender is selected from SEQ ID NO. 507 to 510.
55. 55. The method of claim 33, wherein the detection of HBV is carried out using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 511 to 519, and at least one Label Extender selected from SEQ ID NO. 520 to 535, and optionally at least one blocking label extender is selected from SEQ ID NO. 536 to 553.
56. 56. The method of claim 33, wherein the detection of HCV is carried out using a probe set comprising at least one Capture Extenders selected from SEQ ID NO. 708 to 714, and at least one Label Extender selected from SEQ ID NO. 715 to 727, and optionally at least one blocking label extender is selected from SEQ ID NO. 728 to 732.
57. 57. The method of claim 28, wherein the HIV is HIV-A, HIV-B and HI V-F.
58. 58. The method of claim 57, wherein the detection of HIV-A is carried out using a probe set comprising at least one Capture Extenders selected from SEQ ID NO. 554 to 558, and at least one Label Extender selected from SEQ ID NO. 559 to 567, and optionally at least one blocking label extender is selected from SEQ ID NO. 568 to 571.
59. 59. The method of claim 57, wherein the detection of HIV-B is carried out using a probe set comprising at one Capture Extender selected from SEQ ID NO. 572 to 577, and at least one Label Extender selected from SEQ ID NO. 578 to 590, and optionally at least one blocking label extender is selected from SEQ ID NO. 591 to 595.
60. 60. The method of claim 57, wherein the detection of HIV-F is carried out using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 596 to 600, and at least one Label Extender selected from SEQ ID NO. 601 to 608, and optionally at least one blocking label extender is selected from SEQ ID NO. 609.
61. 61. The method of claim 31, wherein the detection of the orthomyxovirus (flu virus) is carried out using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 610 to 629, and at least one Label Extender

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    2019247099 14 Oct 2019 selected from SEQ ID NO. 630 to 689, and optionally at least one blocking label extender is selected from SEQ) ID NO. 690 to 707.
62. 62. The method of claim 31, wherein the detection of SARS virus is carried out using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 733 to 738, and at least one Label Extender selected from SEQ ID NO. 739 to 750, and optionally at least, one blocking label extender is selected from SEQ ID NO. 751.
63. 63. The method of claim 28, wherein the infectious agent is a Mycobacterium or a spirochete.
64. 64. The method of claim 63, wherein the detection of Mycobacterium is carried out using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 752 to 756, and at least one Label Extender selected from SEQ ID NO. 757 to 770, and optionally at least one blocking label extender is selected from SEQ ID NO. 771 to 773,
65. 65. The method of claim 63, wherein the spirochete is syphilis and wherein the detection of syphilis is carried out using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 774 to 779, and at least one Label Extender selected from SEQ ID NO. 780 to 791, and optionally at least one blocking label extender is selected from SEQ ID NO. 792.
66. 66. The method of claim 28, wherein the infectious agent is a Aspergillus, a Candida, or a Cryptococcus.
67. 67. The method of claim 66, wherein the detection of Apergillus is carried out using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 793 to 798, and at least one Label Extender selected from SEQ ID NO. 799 to 810, and optionally at least one blocking label extender is selected from SEQ ID NO. 811 to 813.
68. 68. The method of claim 66, wherein the detection of Candida is carried out. using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 814 to 819, and at least one Label Extender selected from SEQ ID NO. 820 to 831, , and optionally at least one blocking label extender is selected from SEQ ID NO. 832 to 833.

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69. 69. The method of claim 66, wherein the detection of Cryptococcus is carried out using a probe set comprising at least one Capture Extender selected from SEQ ID NO. 834 to 839, and at least one Label Extender selected, from SEQ ID NO. 840 to 851, , and optionally at least one blocking label extender is selected from SEQ ID NO. 852 to 854.
70. 70. A kit for determining the presence, absence, or level of an infectious agent in a sample comprising:

    (a) a Capture Extender probe having a target-hybridizing sequence selected from Table 1, 4, 7, 10, 14, 17, 20, 23, 26, 29, 32, 35, 38, 41, and 44; and (b) and Label Extender probe having a target-hybridizing sequence selected from Table 2, 5, 8, 11, 15,18,21, 24, 27, 30, 33, 36, 39, 42, and 45, wherein the Capture Extender and Label Extender are each designed to hybridize to the same target polynucleotide.
71. 71. The kit of claim 70, further comprising a Capture Extender probe and Label Extender probe having a sequence that hybridizes to a control sequence.
72. 72. The method of claim 71, wherein the control sequence is a GAPDH (GADH) sequence.
73. 73. The method of claim 72, wherein the detection of a GAPDH sequence is carried out using a probe set comprising at least one Capture Extender selected from Table 93, and at least one Label Extender selected from Tables 1, 2 or 94.
74. 74. The method of claim 73, wherein the probe set further comprises the Blocking Label Extender of Table 95.
75. 75. The kit of claim 70, further comprising a set of signal-amplifying polynucleotide probes.
76. 76. The kit of claim 75 wherein the signal-amplifying polynucleotide probes comprise a Pre-Amplifier probe, an Amplifier probe, and a Label probe.
77. 77. The kit of any one of claims 70 to 76, having a set of Capture Extender and Label Extender probes disclosed herein.

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78. 78. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 14 to 21, and at least one Label Extender probe is selected from SEQ ID NO. 22 to 37 and optionally at least one blocking label extender is selected from SEQ ID NO. 38 to 48 for the detection of a high risk type 16 human papilloma virus (HPV).
79. 79. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 49 to 56, and at least one Label Extender probe is selected from SEQ ID NO. 57 to 72, and optionally at least one blocking label extender is selected from SEQ ID NO. 73 to 75 for the detection of a high risk type 18 human papilloma virus (HPV).
80. 80. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 76 to 82, and at least one Label Extender is selected from SEQ ID NO. 83 to 94, and optionally at least one blocking label extender is selected from SEQ ID NO. 95 to 99 for the detection of a high risk type 31 human papilloma virus (HPV).
81. 81. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 100 to 105, and at least one Label Extender probe is selected from SEQ ID NO. 106 to 117, and optionally at least one blocking label extender is selected from SEQ ID NO. 118 to 127 for the detection of a high risk type 35 human papilloma virus (HPV).
82. 82. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 128 to 134, and at least one Label Extender probe is selected from SEQ ID NO. 135 to 145, and optionally at least one blocking label extender is selected from SEQ ID NO. 146 to 154 for the detection of a high risk type 39 human papilloma virus (HPV).
83. 83. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 155 to 160, and at least one Label Extender probe is selected from SEQ ID NO. 161 to 172, and optionally at least one blocking label extender is selected from SEQ ID NO. 173 to 180 for the detection of a high risk type 45 human papilloma virus (HPV).
84. 84. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 181 to 187, and at least one

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    Label Extender is selected from SEQ ID NO. 188 to 201, and optionally at least one blocking label extender is selected from SEQ ID NO. 202 to 206 for the detection of a high risk type 51 human papilloma virus (HPV).
85. 85. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 207 to 212, and at least one Label Extender is selected from SEQ ID NO. 213 to 224, and optionally at least one blocking label extender is selected from SEQ ID NO. 225 to 238 for the detection of a high risk type 52. human papilloma virus (HPV).
86. 86. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 239 to 245, and at least one Label Extender is selected from SEQ ID NO. 246 to 257, and optionally at least one blocking label extender is selected from SEQ ID NO. 258 to 262 for the detection of a high risk type 56 human papilloma virus (HPV).
87. 87. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 263 to 269, and at least one Label Extender is selected from SEQ ID NO. 2.70 to 2.81, and optionally at least one blocking label extender is selected from SEQ ID NO. 282 to 286 for the detection of a high risk type 58 human papilloma virus (HPV).
88. 88. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 287 to 292, and at least one Label Extender is selected from SEQ ID NO. 293 to 304, and optionally at least one blocking label extender is selected from SEQ ID NO. 305 to 317 for the detection of a high risk type 59 human papilloma virus (HPV).
89. 89. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 318 to 324, and at least one Label Extender is selected from SEQ ID NO. 325 to 338, and optionally at least one blocking label extender is selected from SEQ ID NO. 339 to 347 for the detection of a high risk type 68 human papilloma virus (HPV).
90. 90. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender probe is selected from SEQ ID NO. 348 to 354, and at least one Label Extender probe is selected from SEQ ID NO. 355 to 368 and optionally at least one blocking label extender is selected from SEQ ID NO.

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    369 to 375 for the detection of a low risk type-6 human papilloma virus (HPV).
91. 91. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender probe is selected from SEQ ID NO. 376 to 382, and at least one Label Extender probe is selected from SEQ ID NO. 383 to 398, and optionally at least one blocking label extender is selected from SEQ ID NO. 399 to 403 for the detection of a low risk type-11 human papilloma virus (HPV).
92. 92. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender is selected from SEQ ID NO. 404 to 410, and at least one Label Extender is selected from SEQ ID NO. 411 to 424, and optionally at least one blocking label extender is selected from SEQ ID NO. 425 to 432 for the detection of a low risk type-40 human papilloma virus (HPV).
93. 93. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender probe is selected from SEQ ID NO. 433 to 439, and at least one Label Extender probe is selected from SEQ ID NO. 440 to 451, and optionally at least one blocking label extender is selected from SEQ ID NO. 452 to 462 for the detection of a low risk type-42 human papilloma vims (HPV).
94. 94. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender probe is selected from SEQ ID NO. 463 to 468, and at least one Label Extender probe is selected from SEQ ID NO. 469 to 482, and optionally at least, one blocking label extender is selected from SEQ ID NO. 483 to 488 for the detection of a low risk type-43 human papilloma virus (HPV).
95. 95. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender probe is selected from SEQ ID NO. 489 to 494, and at least one Label Extender probe is selected from SEQ ID NO. 495 to 506, and optionally at least one blocking label extender is selected from SEQ ID NO. 507 to 510 for the detection of a low risk type-44 human papilloma vims (HPV).

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96. 96. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 511 to 519, and at least one Label Extender selected from SEQ ID NO. 520 to 535, and optionally at least one blocking label extender is selected from SEQ ID NO. 536 to 553 for the detection of HBV.
97. 97. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 708 to 714, and at least one Label Extender selected from SEQ ID NO. 715 to 727, and optionally at least one blocking label extender is selected from SEQ ID NO. 728 to 732, for the detection of HCV.
98. 98. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 554 to 558, and at least one Label Extender selected from SEQ ID NO. 559 to 567, and optionally at least one blocking label extender is selected from SEQ ID NO. 568 to 571, for the detection of HIV-A.
99. 99. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 572 to 577, and at least one Label Extender selected from SEQ ID NO. 578 to 590, and optionally at least one blocking label extender is selected from SEQ ID NO. 591 to 595, for the detection of HIV-B.
100. 100. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 596 to 600, and at least one Label Extender selected from SEQ ID NO. 601 to 608, and optionally at least one blocking label extender is selected from SEQ ID NO. 609, for the detection of HIV-F.
101. 101. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 610 to 629, and at least one Label Extender selected from SEQ ID NO. 630 to 689, and optionally at least one blocking label extender is selected from SEQ ID NO. 690 to 707, for the detection of orthomyxovirus (flu virus).
102. 102. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 733 to 738, and at least one

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     Label Extender selected from SEQ ID NO. 739 to 750, and optionally at least one blocking label extender is selected from SEQ ID NO. 751, for the detection of SARS virus.
103. 103. The kit of any one of claims 70 ίο 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 752 to 756, and at least one Label Extender selected from SEQ ID NO. 757 to 770, and optionally at least one blocking label extender is selected from SEQ ID NO. 771 to 773, for the detection of mycobacterium.
104. 104. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 774 to 779, and at least one Label Extender selected from SEQ ID NO. 780 to 791, and optionally at least one blocking label extender is selected from SEQ ID NO. 792, for the detection of syphilis.
105. 105. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 793 to 798, and at least one Label Extender selected from SEQ ID NO. 799 to 810, and optionally at least one blocking label extender is selected from SEQ ID NO. 811 to 813, for the detection of Aspergillus.
106. 106. The kit of any one of claims 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 814 to 819, and at least one Label Extender selected from SEQ ID NO. 820 to 831, and optionally at least one blocking label extender is selected from SEQ ID NO. 832 to 833, for the detection of Candida.
107. 107. The kit of any one of 70 to 77, wherein the kit comprises at least one Capture Extender selected from SEQ ID NO. 834 to 839, and at least one Label Extender selected from SEQ ID NO. 840 to 851, and optionally at least one blocking label extender is selected from SEQ ID NO. 852 to 854, for the detection of cyrptococcus.