WO2024058680A1 - Ensemble d'amorces, composition du mélange réactionnel et procédé de détection des types a et b du virus respiratoire syncytial humain (rsva et rsvb) - Google Patents

Ensemble d'amorces, composition du mélange réactionnel et procédé de détection des types a et b du virus respiratoire syncytial humain (rsva et rsvb) Download PDF

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WO2024058680A1
WO2024058680A1 PCT/PL2023/050074 PL2023050074W WO2024058680A1 WO 2024058680 A1 WO2024058680 A1 WO 2024058680A1 PL 2023050074 W PL2023050074 W PL 2023050074W WO 2024058680 A1 WO2024058680 A1 WO 2024058680A1
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sequence
rsva
rsvb
primers
reverse
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Matylda CZOSNYKOWSKA-ŁUKACKA
Miron TOKARSKI
Małgorzata MAŁODOBRA-MAZUR
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Genomtec S.A.
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/70—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/70—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
    • C12Q1/701—Specific hybridization probes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6844—Nucleic acid amplification reactions
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2537/00—Reactions characterised by the reaction format or use of a specific feature
    • C12Q2537/10—Reactions characterised by the reaction format or use of a specific feature the purpose or use of
    • C12Q2537/143—Multiplexing, i.e. use of multiple primers or probes in a single reaction, usually for simultaneously analyse of multiple analysis
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2563/00—Nucleic acid detection characterized by the use of physical, structural and functional properties
    • C12Q2563/107—Nucleic acid detection characterized by the use of physical, structural and functional properties fluorescence
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/16—Primer sets for multiplex assays

Definitions

  • the invention relates to a set of primers for detecting human respiratory syncytial virus types A and B (RSVA and RSVB), a method for detecting RSVA and RSVB using the set of primers, and an application of the set of primers for detecting RSVA and RSVB.
  • the invention is applicable in medical diagnostics.
  • RSV is an RNA virus. It belongs to the Paramyxoviridae family, Pneumovirus genus. A distinction is currently made between 2 antigenic types of the virus (A and B). RSV is the main pathogen responsible for respiratory tract infections in older children and the most common cause of lower respiratory tract disease in infants. During outbreaks, the incidence rate is more than 50% (in children's communities such as nurseries, it reaches 100%) and is the cause of 75% of hospitalisations of children with bronchiolitis and 25% with pneumonia. It is also the second leading cause of infant mortality after malaria. In adolescents and adults, the disease is mild, characterised by a high prevalence again in the elderly, especially when residing in community settings such as nursing homes and in immunocompromised individuals.
  • RSV infection may refer to the lower respiratory tract and progress into bronchiolitis and pneumonia, less commonly as tracheobronchitis. Upper respiratory tract infections follow a similar course to the common cold, with cough being the main symptom. Very rarely (less than 1%), the first RSV infection is asymptomatic. Without appropriate diagnostics, clinical diagnosis of infection with the described pathogen is not possible, as similar symptoms are observed with infections with other respiratory viruses and atypical bacteria.
  • RSV virus isolation is a method for detecting RSV virus antigens. This is the most commonly used detection method and is based on immunofluorescence or the enzyme immunoassay technique (EIA). Detection of viral RNA by RT-PCR in epithelial cells of the upper respiratory tract, with nasopharyngeal lavage, posterior pharyngeal wall swabs or tracheal secretions being the appropriate material for these determinations.
  • IA enzyme immunoassay technique
  • NAAT methods Nucleic Acid Amplification Tests
  • the most commonly used tests in NAAT technology are assays based on Real-Time RT-PCR method.
  • Many different tests using the Real-Time PCR technique are available on the market, but despite the fierce competition, these methods are still relatively expensive.
  • they require highly specialized personnel, expensive devices, and the extraction of genetic material from the patient's sample.
  • cyclic heating and cooling of the reaction mixture is necessary, this method is time-consuming, and the equipment used consumes relatively large amounts of energy to perform the diagnostic process.
  • Isothermal methods including the LAMP (Loop-mediated isothermal amplification) method, are methods that allow to accelerate the diagnostic process and reduce the cost of energy and reagents needed to perform the analysis. Moreover, according to the literature data, these methods are characterized by higher sensitivity and specificity than the aforementioned Real-Time PCR technique, they are also much faster. Their isothermal course does not require specialized equipment.
  • LAMP Loop-mediated isothermal amplification
  • isothermal methods are an ideal diagnostic solution both for primary care units (POCT - point-of-care testing), where the test can be performed in the practice of a general practitioner or specialist doctor during the patient's first office visit.
  • POCT - point-of-care testing Such a solution enables a rapid diagnostic test (in less than 15 minutes), allowing targeted therapy to be selected at the first visit.
  • This is particularly important in the case of RSV infection, due to the rapid progression of the infection, which quickly leads to an immediately life-threatening condition. Delayed diagnosis of RSV infection increases the patient's risk of death, as well as the risk of developing serious complications.
  • freeze-dried reagents allows the tests to be stored at room temperature, without the need to freeze the diagnostic tests.
  • the use of primers in the LAMP method for the diagnosis of RSV is known from the patent applications published so far: CN109762942A, CN104419713A, CN104419713A; CN107099619A; KR20220040073A.
  • the LAMP method is disclosed, for example, in patent applications WO0028082, WO0224902.
  • the detection method is not quantitative, and it is an end-point detection with the use of agarose gel or other markers based on the colour change of the reaction mixture upon a positive amplification.
  • the first subject of the invention is a set of primers for amplifying the nucleotide sequence of the N (nucleocapsid) gene of RSVA, characterized in that it comprises a set of internal primers with the following nucleotide sequences a) and b), as well as a set of external primers containing the following nucleotide sequences c) and d) specific for a selected fragment the N gene of RSVA: a) 5' GCACACTAGCATGTCCTAACATAAT 3'- (nucleic sequence SEQ ID NO:
  • the primer set comprises a set of loop primer sequences comprising nucleic sequences contained in or complementary to the RSVA N gene SEQ ID NO:7 5' TGATTTTGCTAAGACTCCCC3A'C and SEQ ID NO: 8 5' ATGCCCAAAAATTGGGTGGAG 3' or sequences reverse and complementary thereto.
  • the second subject of the invention is a set of primers for amplifying the nucleotide sequence of the N (nucleocapsid) gene of RSVB, characterized in that it comprises a set of internal primers with the following nucleotide sequences a) and b), as well as a set of external primers containing the following nucleotide sequences c) and d) specific for a selected fragment the N gene of RSVB: a) 5' GGACACTAGCATGTCCTAGCATG 3'- (nucleic sequence SEQ ID NO: 13 or its reverse and complementary sequence), linked from the 3' end, preferably by a TTTT bridge, to the sequence 5' GTAATGCTAAGATGGGGAGTT 3'- (nucleic sequence SEQ ID NO: 11 or its reverse and complementary sequence) b) 5' GGAGCAAGTTGTGGAAGTCTATGA 3'- (nucleic sequence SEQ ID NO: 14 or its reverse and complementary sequence), linked at the 3'
  • the primer set comprises a set of loop primer sequences comprising nucleic sequences contained in or complementary to the RSVB N gene SEQ ID NO: 15: 5' GCACAGAAGTTGGGAGGAGAAGC 3' or its reverse and complementary sequence.
  • the third subject of the invention is a method for detecting RSVA and RSVB viruses, characterised in that selected regions of the nucleic sequence of the RSVA genome (N - nucleocapsid gene fragment) and the RSVB genome (N - nucleocapsid gene fragment) are amplified using a mixture of primer sets as defined in the first and second subjects of the invention, the amplification method being the RT-LAMP method.
  • the amplification is carried out with a temperature profile: 63°C, 40 min.
  • the end- point reaction is carried out with an additional temperature profile of 80°C, 5 min carried out after the amplification step.
  • the fourth subject of the invention is a method for detecting an infection caused by RSVA and/or RSVB characterised in that it comprises the detection method defined in the third subject of the invention.
  • the fifth subject of the invention is a kit for detecting an infection caused by RSVA and/or RSVB characterised in that it comprises a set of primers as defined in the first and second subjects of the invention.
  • the infection detection kit comprises 5.0 ⁇ L of WarmStart® LAMP (DNA & RNA) Master Mix.
  • the kit comprises individual amplification primers as defined in the first and second subjects of the invention, the primers having the following concentrations: 0.13 ⁇ M F3RSVA, 0.13 ⁇ M B3RSVA, 1.06 ⁇ M FIPRSVA, 1.06 ⁇ M BIPRSVA, 0.26 ⁇ M LoopFRSVA, 0.26 ⁇ M LoopBRSVA, 0.13 ⁇ M F3RSVB, 0.13 ⁇ M B3RSVB, 1.06 ⁇ M FIPRSVB, 1.06 ⁇ M BIPRSVB, 0.26 ⁇ M LoopBRSVB; D-(+)-Trehalose dihydrate - 6%; mannitol - 1.25%; fluorescent marker interacting with double- stranded DNA - EvaGreen ⁇ 1X (Biotium) or Fluorescent Dye (New England Biolabs) in the amount of dl ⁇ L or Syto-13 ⁇ 16 ⁇ M (ThermoFisher Scientific) or SYTO-82 ⁇ 16
  • primer sets of the invention for detecting of RSVA and RSVB are the possibility of using them in medical diagnostics at the point of care (POCT) in the target application with a portable genetic analyser. Freeze-drying of the reaction mixtures of the invention allows the diagnostic kits to be stored at room temperature without reducing the diagnostic parameters of the tests.
  • POCT point of care
  • the use of a fluorescent dye to detect the amplification product increases the sensitivity of the method, allows to lower the detection limit (down to 320 genome copies/reaction for the RSVA virus and down to 195 genome copies/reaction for the RSVB virus), as well as it enables the measurement of viral load in the test sample.
  • FIG. 1 shows the sensitivity characteristics of the method, where a specific signal was obtained with the template: AMPLIRUN® Respiratory syncytial Virus (subtype A) RNA Control - Vircell and RNA AMPLIRUN® Respiratory Syncytial Virus (subtype B) RNA Control over the range of 1000-320 copies/ ⁇ L, but there was no product in NTC;
  • fig. 2 shows the sensitivity characteristics of the method, where a specific signal was obtained with the template: AMPLIRUN® Respiratory syncytial Virus (subtype B) RNA Control - Vircell over the range of 1000- 195 copies/ ⁇ L, but there was no product in NTC.
  • lane 1 mass marker (Quick-Load® Purple 100 bp DNA Ladder, NewEngland Biolabs); lane 2: 2000 copies of RSVA; lane 3: 1000 copies of RSVA; lane 4: 500 copies of RSVA; lane 5: 300 copies of RSVA; and lane 6: NTC.
  • Fig. 2 lane 1: mass marker (Quick-Load® Purple 100 bp DNA Ladder, NewEngland Biolabs); lane 2: 2000 copies of RSVB; lane 3: 1000 copies of RSVB; lane 4: 500 copies of RSVB; lane 5: 320 copies of RSVB; lane 6: NTC; and Fig. 3 shows the sensitivity of the method of the invention as measured by assaying a serial dilution of the AMPLIRUN® Respiratory syncytial Virus (subtype
  • RNA Control standard over a range of 1000-320 copies/reaction of the RNA standard, where the product amplification was measured in real time.
  • the results of the real-time RSVA detection are presented in Table 1, providing the minimum time required to detect the fluorescence signal; and
  • Fig. 4 shows the sensitivity of the method of the invention as measured by assaying a serial dilution of the AMPLIRUN® Respiratory syncytial Virus (subtype
  • RNA Control standard over a range of 1000-195 copies/reaction of the RNA standard, where the product amplification was measured in real time.
  • the results of the real-time RSVB detection are presented in Table 2, giving the minimum time required to detect the fluorescence signal; while Fig. 5 shows the specificity of the product obtained after RSVA detection as measured by the melting curve of the amplification product using the AMPLIRUN® Respiratory syncytial Virus (subtype A) RNA Control standard over the range 1000-320 copies/reaction by real-time fluorescence measurement, with a target melting temperature (Tm) of 81.5°C for a specific RSVA reaction product; while Fig.
  • Tm target melting temperature
  • FIG. 6 shows the specificity of the product obtained after RSVB detection as measured by the melting curve of the amplification product using the AMPLIRUN® Respiratory syncytial Virus (subtype B) RNA Control standard over the range 1000-195 copies/reaction by real-time fluorescence measurement, with a target dissociation temperature (Tm) of 84°C for a specific RSVB reaction product; and Figs 7, 8 and 9 show the specificity of the method of the invention with standard materials of a number of pathogens potentially present in the tested biological material as natural physiological flora, those which may result from co-infections or those which share similar genomic sequences; and Fig.
  • Tm target dissociation temperature
  • lane 1 mass marker (Quick-Load® Purple 100 bp DNA Ladder, NewEngland Biolabs); lanes 2 and 3: methicillin-sensitive Staphylococcus aureus (MSSA); lanes 4 and 5: Influenza B virus; lanes 6 and 7: Influenza A virus (H1N1); lanes 8 and 9: Influenza A virus (H3N2); lanes 10 and 11: Mycoplasma genitalium; lanes 12 and 13: HPV 16; lanes 14 and 15: Klebsiella pneumoniae; lanes 16 and 17: Bordetella pertussis; lanes 18 and 19: Streptococcus pyogenes; lanes 20 and 21: Staphylococcus aureus (MRSA); lanes 22 and 23: Enterococcus faecalis; lanes 24 and 25: Enterococcus faecium; lanes 26 and 27: Pseudomonas aeruginosa; lanes 28 and
  • lane 1 mass marker (Quick-Load® Purple 100 bp DNA Ladder, NewEngland Biolabs); lanes 2 and 3: Homo sapiens; lanes 4 and 5: SARS CoV-2 Frankfurt 1; lanes 6 and 7: SARS CoV-2 USA-WA12020; lanes 8 and 9: SARS CoV-2 Isolate USA-WI/202; lanes 10 and 11: SARS CoV-2 Isolate Hong Kong; lanes 12 and 13: SARS CoV-2 Isolate Italy-INMIl; lanes 14 and 15: NTC, and Fig. 9.
  • lane 1 mass marker (Quick-Load® Purple 100 bp DNA Ladder, NewEnglandBiolabs); lanes 2 and 3:Human Coronavirus, OC43; lanes 4 and 5:SARS-Related Coronavirus 2, Isolate Germany/BavPatl/2020; lanes6 and 7: SARS CoV-2 NR 52726 Human Coronavirus 229E; lanes 8 and9: Amplirun SARS-CoV-2 B.1.351 RNA Control; lanes 10 and 11:SARS CoV-2 Isolate USA-CA1/CA2/CA3/2020; lanes 12 and 13: SARSCoV-2 Isolate New York-PVO8410/2020; lanes 14 and 15: SARS CoV-2 Isolate USA-IL1/2020; lane 16:mass marker (Quick-Load® Purple100 bp DNA Ladder, NewEngland Biolabs); lanes 17 and 18: SARSCoV-2 Isolate Chile/Santiago_op4dl/2020; la
  • Example 1 Primer sequences The sequences of the specific oligonucleotides used for thedetection of RSVA and RSVB genetic materialusing LAMP technologyare presented and characterised below.
  • the RSVA NB3 oligonucleotide sequence:5' GTGAGGAAATTGAGTCAAAGA 3' is a complementary fragment of theRSVA N gene (5'-3' strand) 179 nucleotides away from the 3' endof the oligonucleotide 1. 3.
  • the RSVA NF2 oligonucleotide sequence:5' CAGGGCAAGTGATGTTACG 3' is a sequence identical to the RSVA Ngene (5'-3' strand) immediately adjacent to the 3' end of theoligonucleotide 1. 4.
  • the RSVA NQ2 oligonucleotide sequence:5' GGTTGTTCAATATATGGTAGAATCC 3' is a complementary fragment ofthe RSVA N gene (5'-3' strand) 137 nucleotides away from the 3'end of the oligonucleotide 1. 5.
  • the RSVA NFlc oligonucleotide sequence: 5' GCACACTAGCATGTCCTAACATAAT 3' is a complementary fragment of the RSVA N gene (5'-3' strand) 50 nucleotides away from the 3' end of the oligonucleotide 1.
  • the RSVA NBlc oligonucleotide sequence: 5' TGGAACAAGTTGTTGAGGTTTATGA 3' is a sequence identical to the RSVA N gene (5'-3' strand) 84 nucleotides away from the 3' end of the oligonucleotide 1.
  • the RSVB NF3 oligonucleotide sequence: 5' TGAATGCCTATGGTTCA3G'G is a sequence identical to the RSVB N gene (5'-3' strand).
  • the RSVB NB3 oligonucleotide sequence: 5' TTGAGTTAATGACAGCAATGA 3' is a complementary fragment of the RSVB N gene (5'-3' strand) 166 nucleotides away from the 3' end of the oligonucleotide 9.
  • the RSVB NF2 oligonucleotide sequence: 5' GTAATGCTAAGATGGGGAGTT 3' is a sequence identical to the RSVB N gene (5'-3' strand) 4 nucleotides away from the 3’ end of the oligonucleotide 9.
  • the RSVB NB2 oligonucleotide sequence: 5' GATTGTTCAATATATGGTAGAATCC 3' is a complementary fragment of the RSVA N gene (5'-3' strand) 133 nucleotides away from the 3' end of the oligonucleotide 9.
  • the RSVB NF1C oligonucleotide sequence: 5' GGACACTAGCATGTCCTAGCATG 3' is a complementary fragment of the RSVB N gene (5'-3' strand) 48 nucleotides away from the3' end of the oligonucleotide 9.
  • the RSVB NB1c oligonucleotide sequence: 5' GGAGCAAGTTGTGGAAGTCTATGA 3' is a sequence identical to the RSVA N gene (5'-3' strand) 81 nucleotides away from the 3' end of the oligonucleotide 9.
  • sequences of the Flc and F2 oligonucleotides have preferably been linked by a TTTT bridge and used as FTP.
  • sequences of the Bic and B2 oligonucleotides have preferably been linked by a TTTT bridge and used as BIP.
  • the RSVB NLoopB oligonucleotide sequence 5' GCACAGAAGTTGGGAGGAGAAGC 3'.
  • reaction components were mixed according to the composition described in Example 2 and Example 3, except the template RNA, to a total volume of 10 ⁇ L.
  • the mixture was transferred to 0.2 mL tubes and subjected to the freeze-drying process according to the parameters below.
  • the mixture placed in test tubes was pre-cooled to -80°C for 2 hours. Then the freeze-drying process was carried out at the temperature of -25°C for 18 hours under the pressure of 5 -2 mBar and a further 3 hours 30 minutes at the temperature of 25°C under the same pressure.
  • the sensitivity was determined by assaying serial dilutions of the Respiratory syncytial virus (subtype A) RNA Control standard and Respiratory syncytial virus (subtype B) RNA Control standard with a minimum amount of 320 copies for RSVA and a minimum amount of 195 copies for RSVB per reaction mixture, where the product amplification was measured in real time - Figure 3 and Figure 4 (Real-Time LAMP for serial dilutions) along with recording the dissociation temperature of 81.5°C ( Figure 5) for RSVA and 84°C ( Figure 6) for RSVB.
  • the characterized primers allow for the detection of RSVA and RSVB by detecting the N gene fragment at a minimum number of 320 genome copies/reaction mixture for RSVA and a minimum number of 195 genome copies/reaction mixture for RSVB.

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Abstract

Le premier objet de l'invention est un ensemble d'amorces permettant d'amplifier la séquence nucléotidique du gène N du RSVA. Le deuxième objet de l'invention est un ensemble d'amorces permettant d'amplifier la séquence nucléotidique du gène N du RSVB. Le troisième objet de l'invention est un procédé de détection du RSVA et du RSVB. Un autre objet de l'invention est un procédé permettant de détecter une infection causée par le RSVA ou le RSVB. Le cinquième objet de l'invention est un kit de détection d'une infection causée par le RSVA ou le RSVB.
PCT/PL2023/050074 2022-09-12 2023-09-11 Ensemble d'amorces, composition du mélange réactionnel et procédé de détection des types a et b du virus respiratoire syncytial humain (rsva et rsvb) WO2024058680A1 (fr)

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PL442248A PL442248A1 (pl) 2022-09-12 2022-09-12 Zestaw starterów, skład mieszaniny reakcyjnej oraz metoda wykrywania ludzkiego syncytialnego wirusa oddechowego (ang. Human Respiratory Syncital Virus) typ A i typ B (RSVA) i (RSVB)

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Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
HART D., VAN ZYL G.: "Loop mediated isothermal amplification to detect respiratory syncytial virus in respiratory specimens", INTERNATIONAL JOURNAL OF INFECTIOUS DISEASES, INTERNATIONAL SOCIETY FOR INFECTIOUS DISEASES, HAMILTON, CA, vol. 21, 1 April 2014 (2014-04-01), CA , pages 328, XP093151083, ISSN: 1201-9712, DOI: 10.1016/j.ijid.2014.03.1097 *
MAHONY JAMES, CHONG SYLVIA, BULIR DAVID, RUYTER ALEXANDRA, MWAWASI KEN, WALTHO DANIEL: "Development of a Sensitive Loop-Mediated Isothermal Amplification Assay That Provides Specimen-to-Result Diagnosis of Respiratory Syncytial Virus Infection in 30 Minutes", JOURNAL OF CLINICAL MICROBIOLOGY, AMERICAN SOCIETY FOR MICROBIOLOGY, US, vol. 51, no. 8, 12 June 2013 (2013-06-12), US , pages 2696 - 2701, XP093151076, ISSN: 0095-1137, DOI: 10.1128/JCM.00662-13 *
SHIRATO, K. NISHIMURA, H. SAIJO, M. OKAMOTO, M. NODA, M. TASHIRO, M. TAGUCHI, F.: "Diagnosis of human respiratory syncytial virus infection using reverse transcription loop-mediated isothermal amplification", JOURNAL OF VIROLOGICAL METHODS, ELSEVIER BV, NL, vol. 139, no. 1, 6 December 2006 (2006-12-06), NL , pages 78 - 84, XP005794349, ISSN: 0166-0934, DOI: 10.1016/j.jviromet.2006.09.014 *
TAKAYAMA IKUYO; NAKAUCHI MINA; TAKAHASHI HITOSHI; OBA KUNIHIRO; SEMBA SHOHEI; KAIDA ATSUSHI; KUBO HIDEYUKI; SAITO SHINJI; NAGATA S: "Development of real-time fluorescent reverse transcription loop-mediated isothermal amplification assay with quenching primer for influenza virus and respiratory syncytial virus", JOURNAL OF VIROLOGICAL METHODS, ELSEVIER BV, NL, vol. 267, 1 January 1900 (1900-01-01), NL , pages 53 - 58, XP085650988, ISSN: 0166-0934, DOI: 10.1016/j.jviromet.2019.02.010 *
USHIO MASANOBU, YUI IKUKO, YOSHIDA NAOKO, FUJINO MOTOKO, YONEKAWA TOSHIHIRO, OTA YOSHINORI, NOTOMI TSUGUNORI, NAKAYAMA TETSUO: "Detection of respiratory syncytial virus genome by subgroups‐A, B specific reverse transcription loop‐mediated isothermal amplification (RT‐LAMP)", JOURNAL OF MEDICAL VIROLOGY, JOHN WILEY & SONS, INC., US, vol. 77, no. 1, 1 September 2005 (2005-09-01), US , pages 121 - 127, XP093151071, ISSN: 0146-6615, DOI: 10.1002/jmv.20424 *

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