CN108719204A - A kind of drug resistance Acinetobacter bauamnnii infects method for building up and its application of Caenorhabditis elegans medicaments sifting model - Google Patents
A kind of drug resistance Acinetobacter bauamnnii infects method for building up and its application of Caenorhabditis elegans medicaments sifting model Download PDFInfo
- Publication number
- CN108719204A CN108719204A CN201710576315.4A CN201710576315A CN108719204A CN 108719204 A CN108719204 A CN 108719204A CN 201710576315 A CN201710576315 A CN 201710576315A CN 108719204 A CN108719204 A CN 108719204A
- Authority
- CN
- China
- Prior art keywords
- drug
- acinetobacter bauamnnii
- drug resistance
- caenorhabditis elegans
- infection
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 239000003814 drug Substances 0.000 title claims abstract description 94
- 241000589291 Acinetobacter Species 0.000 title claims abstract description 53
- 206010059866 Drug resistance Diseases 0.000 title claims abstract description 41
- 241000244203 Caenorhabditis elegans Species 0.000 title claims abstract description 36
- 238000000034 method Methods 0.000 title claims abstract description 26
- 229940079593 drug Drugs 0.000 claims abstract description 65
- 241000244206 Nematoda Species 0.000 claims abstract description 41
- 208000015181 infectious disease Diseases 0.000 claims abstract description 40
- 241000588724 Escherichia coli Species 0.000 claims abstract description 5
- 102000002322 Egg Proteins Human genes 0.000 claims abstract description 4
- 108010000912 Egg Proteins Proteins 0.000 claims abstract description 4
- 210000004681 ovum Anatomy 0.000 claims abstract description 4
- 230000035935 pregnancy Effects 0.000 claims abstract description 4
- 230000001360 synchronised effect Effects 0.000 claims abstract description 4
- 239000007853 buffer solution Substances 0.000 claims description 19
- 241000894006 Bacteria Species 0.000 claims description 17
- 230000004083 survival effect Effects 0.000 claims description 16
- 239000013641 positive control Substances 0.000 claims description 7
- 208000000291 Nematode infections Diseases 0.000 claims description 6
- 239000013642 negative control Substances 0.000 claims description 6
- 229920001817 Agar Polymers 0.000 claims description 5
- 239000001888 Peptone Substances 0.000 claims description 5
- 108010080698 Peptones Proteins 0.000 claims description 5
- 239000008272 agar Substances 0.000 claims description 5
- 235000019319 peptone Nutrition 0.000 claims description 5
- 239000006228 supernatant Substances 0.000 claims description 5
- 235000015278 beef Nutrition 0.000 claims description 3
- 238000007877 drug screening Methods 0.000 claims description 2
- 238000012360 testing method Methods 0.000 abstract description 22
- 238000000338 in vitro Methods 0.000 abstract description 18
- 238000002474 experimental method Methods 0.000 abstract description 11
- 230000035945 sensitivity Effects 0.000 abstract description 11
- 238000012216 screening Methods 0.000 abstract description 7
- 238000001727 in vivo Methods 0.000 abstract description 5
- 230000012447 hatching Effects 0.000 abstract 1
- 238000011282 treatment Methods 0.000 description 9
- 230000001225 therapeutic effect Effects 0.000 description 8
- 230000000844 anti-bacterial effect Effects 0.000 description 7
- 229940088710 antibiotic agent Drugs 0.000 description 6
- 230000003115 biocidal effect Effects 0.000 description 6
- MYSWGUAQZAJSOK-UHFFFAOYSA-N ciprofloxacin Chemical compound C12=CC(N3CCNCC3)=C(F)C=C2C(=O)C(C(=O)O)=CN1C1CC1 MYSWGUAQZAJSOK-UHFFFAOYSA-N 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- FFTVPQUHLQBXQZ-KVUCHLLUSA-N (4s,4as,5ar,12ar)-4,7-bis(dimethylamino)-1,10,11,12a-tetrahydroxy-3,12-dioxo-4a,5,5a,6-tetrahydro-4h-tetracene-2-carboxamide Chemical compound C1C2=C(N(C)C)C=CC(O)=C2C(O)=C2[C@@H]1C[C@H]1[C@H](N(C)C)C(=O)C(C(N)=O)=C(O)[C@@]1(O)C2=O FFTVPQUHLQBXQZ-KVUCHLLUSA-N 0.000 description 5
- 235000013305 food Nutrition 0.000 description 5
- 229960004023 minocycline Drugs 0.000 description 5
- WKDDRNSBRWANNC-UHFFFAOYSA-N Thienamycin Natural products C1C(SCCN)=C(C(O)=O)N2C(=O)C(C(O)C)C21 WKDDRNSBRWANNC-UHFFFAOYSA-N 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 4
- 230000001580 bacterial effect Effects 0.000 description 4
- 229960002182 imipenem Drugs 0.000 description 4
- ZSKVGTPCRGIANV-ZXFLCMHBSA-N imipenem Chemical compound C1C(SCC\N=C\N)=C(C(O)=O)N2C(=O)[C@H]([C@H](O)C)[C@H]21 ZSKVGTPCRGIANV-ZXFLCMHBSA-N 0.000 description 4
- 229960002260 meropenem Drugs 0.000 description 4
- DMJNNHOOLUXYBV-PQTSNVLCSA-N meropenem Chemical compound C=1([C@H](C)[C@@H]2[C@H](C(N2C=1C(O)=O)=O)[C@H](O)C)S[C@@H]1CN[C@H](C(=O)N(C)C)C1 DMJNNHOOLUXYBV-PQTSNVLCSA-N 0.000 description 4
- 238000002360 preparation method Methods 0.000 description 4
- 238000011160 research Methods 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- KMEGBUCIGMEPME-LQYKFRDPSA-N (2s,5r,6r)-6-[[(2r)-2-amino-2-phenylacetyl]amino]-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxylic acid;(1r,4s)-3,3-dimethyl-2,2,6-trioxo-2$l^{6}-thiabicyclo[3.2.0]heptane-4-carboxylic acid Chemical compound O=S1(=O)C(C)(C)[C@H](C(O)=O)C2C(=O)C[C@H]21.C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 KMEGBUCIGMEPME-LQYKFRDPSA-N 0.000 description 3
- 241000588624 Acinetobacter calcoaceticus Species 0.000 description 3
- APKFDSVGJQXUKY-KKGHZKTASA-N Amphotericin-B Natural products O[C@H]1[C@@H](N)[C@H](O)[C@@H](C)O[C@H]1O[C@H]1C=CC=CC=CC=CC=CC=CC=C[C@H](C)[C@@H](O)[C@@H](C)[C@H](C)OC(=O)C[C@H](O)C[C@H](O)CC[C@@H](O)[C@H](O)C[C@H](O)C[C@](O)(C[C@H](O)[C@H]2C(O)=O)O[C@H]2C1 APKFDSVGJQXUKY-KKGHZKTASA-N 0.000 description 3
- 229960004821 amikacin Drugs 0.000 description 3
- LKCWBDHBTVXHDL-RMDFUYIESA-N amikacin Chemical compound O([C@@H]1[C@@H](N)C[C@H]([C@@H]([C@H]1O)O[C@@H]1[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O1)O)NC(=O)[C@@H](O)CCN)[C@H]1O[C@H](CN)[C@@H](O)[C@H](O)[C@H]1O LKCWBDHBTVXHDL-RMDFUYIESA-N 0.000 description 3
- APKFDSVGJQXUKY-INPOYWNPSA-N amphotericin B Chemical compound O[C@H]1[C@@H](N)[C@H](O)[C@@H](C)O[C@H]1O[C@H]1/C=C/C=C/C=C/C=C/C=C/C=C/C=C/[C@H](C)[C@@H](O)[C@@H](C)[C@H](C)OC(=O)C[C@H](O)C[C@H](O)CC[C@@H](O)[C@H](O)C[C@H](O)C[C@](O)(C[C@H](O)[C@H]2C(O)=O)O[C@H]2C1 APKFDSVGJQXUKY-INPOYWNPSA-N 0.000 description 3
- 229960003942 amphotericin b Drugs 0.000 description 3
- 230000002924 anti-infective effect Effects 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 229960003405 ciprofloxacin Drugs 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 229960001699 ofloxacin Drugs 0.000 description 3
- 244000052769 pathogen Species 0.000 description 3
- 230000001717 pathogenic effect Effects 0.000 description 3
- 238000004393 prognosis Methods 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 229960004261 cefotaxime Drugs 0.000 description 2
- GPRBEKHLDVQUJE-VINNURBNSA-N cefotaxime Chemical compound N([C@@H]1C(N2C(=C(COC(C)=O)CS[C@@H]21)C(O)=O)=O)C(=O)/C(=N/OC)C1=CSC(N)=N1 GPRBEKHLDVQUJE-VINNURBNSA-N 0.000 description 2
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 2
- 238000004140 cleaning Methods 0.000 description 2
- 238000010219 correlation analysis Methods 0.000 description 2
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical class [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 229940000406 drug candidate Drugs 0.000 description 2
- 235000013399 edible fruits Nutrition 0.000 description 2
- 230000002458 infectious effect Effects 0.000 description 2
- 239000003112 inhibitor Substances 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 238000011369 optimal treatment Methods 0.000 description 2
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 238000004659 sterilization and disinfection Methods 0.000 description 2
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 1
- 241000304886 Bacilli Species 0.000 description 1
- 206010060976 Bacillus infection Diseases 0.000 description 1
- 208000035143 Bacterial infection Diseases 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- 206010011409 Cross infection Diseases 0.000 description 1
- 241001269238 Data Species 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 239000007836 KH2PO4 Substances 0.000 description 1
- 206010029803 Nosocomial infection Diseases 0.000 description 1
- 206010034133 Pathogen resistance Diseases 0.000 description 1
- -1 Sulbactam compound Chemical class 0.000 description 1
- 239000004098 Tetracycline Substances 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 150000001336 alkenes Chemical class 0.000 description 1
- 230000002421 anti-septic effect Effects 0.000 description 1
- 230000001363 autoimmune Effects 0.000 description 1
- 208000022362 bacterial infectious disease Diseases 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000001124 body fluid Anatomy 0.000 description 1
- 239000010839 body fluid Substances 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- HVFLCNVBZFFHBT-ZKDACBOMSA-N cefepime Chemical compound S([C@@H]1[C@@H](C(N1C=1C([O-])=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1C[N+]1(C)CCCC1 HVFLCNVBZFFHBT-ZKDACBOMSA-N 0.000 description 1
- 229960002100 cefepime Drugs 0.000 description 1
- 229960004682 cefoperazone Drugs 0.000 description 1
- GCFBRXLSHGKWDP-XCGNWRKASA-N cefoperazone Chemical compound O=C1C(=O)N(CC)CCN1C(=O)N[C@H](C=1C=CC(O)=CC=1)C(=O)N[C@@H]1C(=O)N2C(C(O)=O)=C(CSC=3N(N=NN=3)C)CS[C@@H]21 GCFBRXLSHGKWDP-XCGNWRKASA-N 0.000 description 1
- 229960004755 ceftriaxone Drugs 0.000 description 1
- VAAUVRVFOQPIGI-SPQHTLEESA-N ceftriaxone Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1CSC1=NC(=O)C(=O)NN1C VAAUVRVFOQPIGI-SPQHTLEESA-N 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 235000012000 cholesterol Nutrition 0.000 description 1
- 238000002648 combination therapy Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000003113 dilution method Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 235000019441 ethanol Nutrition 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000011010 flushing procedure Methods 0.000 description 1
- 230000004907 flux Effects 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 238000007403 mPCR Methods 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 238000005374 membrane filtration Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 230000036457 multidrug resistance Effects 0.000 description 1
- 238000011017 operating method Methods 0.000 description 1
- 229960002292 piperacillin Drugs 0.000 description 1
- IVBHGBMCVLDMKU-GXNBUGAJSA-N piperacillin Chemical compound O=C1C(=O)N(CC)CCN1C(=O)N[C@H](C=1C=CC=CC=1)C(=O)N[C@@H]1C(=O)N2[C@@H](C(O)=O)C(C)(C)S[C@@H]21 IVBHGBMCVLDMKU-GXNBUGAJSA-N 0.000 description 1
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 230000005180 public health Effects 0.000 description 1
- 238000013139 quantization Methods 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 229960005256 sulbactam Drugs 0.000 description 1
- 235000019364 tetracycline Nutrition 0.000 description 1
- 150000003522 tetracyclines Chemical class 0.000 description 1
- 229940040944 tetracyclines Drugs 0.000 description 1
- 239000011534 wash buffer Substances 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/033—Rearing or breeding invertebrates; New breeds of invertebrates
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Environmental Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Zoology (AREA)
- Animal Husbandry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The present invention provides a kind of method for building up of drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model, includes the following steps：Step 1：Isolate general drug resistance Acinetobacter bauamnnii；Step 2：Worm's ovum is detached from pregnancy adult, larva is transferred to continued growth in the incubator on E.coli lawns, after eluting polypide, polypide is transferred on the lawn of general drug resistance Acinetobacter bauamnnii by hatching, cultivates 8h~infected for 24 hours in the incubator；Step 3：Polypide is washed down, is placed it on the culture plate containing 5-FU, is incubated overnight, synchronized Caenorhabditis elegans is obtained, medicaments sifting model, which is established, to be completed.It is experimental subjects that the present invention, which selects nematode, carries out the drug sensitive experiment of general drug resistant Acinetobacter bauamnnii in vivo, and result is compared to each other with drug sensitivity testing in vitro result, has clinical reference value, can make up the omission in traditional In vitro chemo-drug sensitive test screening.
Description
Technical field
The present invention relates to a kind of drug resistance Acinetobacter bauamnnii infect Caenorhabditis elegans medicaments sifting model method for building up,
It is obtained the invention further relates to the method for building up of drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model is used
The application of medicaments sifting model.
Background technology
Infectious diseases is always one of the major public health problem in China, due to widely applying wide spectrum antibiosis for a long time
Element so that many pathogens produce antibiotic surprising drug resistance.Nosocomial infection is in rising trend in recent years, in some institutes
Infection is in intractable, and is multidrug resistant, seriously threatens the health and lives safety of patient.Acinetobacter bauamnnii has become respectively
One of most common gram negative bacilli in the infection of large hospital inpatient, in recent years the recall rate of Bao Man not levers it is continuous on
It rises, prodigious difficulty is caused to the treatment of infection.The resistant rate of drug resistance Acinetobacter bauamnnii has the tendency that constantly increasing, and often
There is multidrug resistance and general drug resistance phenomenon.For general drug resistant Difficult infection, clinician often rule of thumb carries out antibacterial
Combination therapies or escalated dose achieve the purpose that control drug-fast bacteria infection.However in empirical treatment, Bao Man is motionless
Lapsing to for bacillus infection patient is different, the state of an illness of some patients, which can be eased, even cures, and some patients occur
It is dead.Therefore, it is quickly timely to screen drug and formation for the optimal treatment of specific drug resistant Acinetobacter bauamnnii infection
Anti-infective therapeutic schemes, this is to ensure that patient has the Critical policies of more preferable prognosis.Therefore needle can quickly be screened by providing one kind
It is the powerful guarantee of this strategy to the model of property drug.
Invention content
The purpose of the present invention is to provide a kind of drug resistance Acinetobacter bauamnniis to infect Caenorhabditis elegans medicaments sifting model
Method for building up and its application, with achieve the purpose that quickly screen for drug resistance Acinetobacter bauamnnii drug.
Present invention employs following technical solutions：
A kind of method for building up of drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model, feature exist
In including the following steps：
Step 1：Isolate general drug resistance Acinetobacter bauamnnii；
Step 2：Caenorhabditis elegans infects drug resistance Acinetobacter bauamnnii：From pregnancy adult detach worm's ovum, incubated overnight,
L1 phase larvas are obtained, larva is transferred on the E.coli lawns of NGM culture mediums continued growth 48h in the incubator, obtains L4
Polypide is transferred to the general drug resistance Bao of plain agar tablet by young adult respectively after eluting polypide and cleaning time with M9 buffer solutions
On the lawn of graceful acinetobacter calcoaceticus, 8h~infected for 24 hours is cultivated in the incubator, is transferred to after infection in M9 buffer solutions and continues to train
It supports, observes and records daily death condition；
Step 3：Nematode infections synchronize：The good tablet of nematode growth is collected, is washed down polypide with M9 buffer solutions, from
The heart takes supernatant, and washing places it on the culture plate containing 5-FU, is incubated overnight, obtains synchronized Caenorhabditis elegans,
Medicaments sifting model, which is established, to be completed.
Preferably, the foundation side of the drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model of the present invention
Method, it is characterised in that：In step 3, centrifugal condition 20S takes supernatant, is repeated 3 times.
Preferably, the foundation side of the drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model of the present invention
Method can also have the feature that：By percentage to the quality, M9 buffer solutions include：Peptone 2%, beef extract 4%.
Preferably, the foundation side of the drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model of the present invention
Method can also have the feature that：In step 2, infection time is 16 hours.
Preferably, the foundation side of the drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model of the present invention
Method can also have the feature that：In step 2, nematode becomes linear type, touches reactionless be judged to without movement, with platinum wire
It is dead.
The present invention also provides a kind of drug resistance Acinetobacter bauamnniis established using the above method to infect Caenorhabditis elegans mould
Application of the type in drug screening.
Preferably, application of the invention, includes the following steps：
Step 1：Negative control is set；
Step 2：Positive control is set；
Step 3：Establish the concentration gradient of each drug；
Step 4：Drug resistance Acinetobacter bauamnnii is added into the drug of each concentration gradient and infects Caenorhabditis elegans model,
To the nematodal accounting of survival after culture, nematode survival quantity is at most optimum medicine, and nematode is deposited in each drug
The optimum concentration of the most a concentration of drug of quantity living.
Preferably, the application of the present invention, can also have the feature that：The negative control is M9 buffer solutions, institute
Stating positive control is：The nematode of Acinetobacter bauamnnii but non-administration is infected.
Preferably, the application of the present invention, can also have the feature that：In step 3, drug concentration gradient difference
For 1000 μ gml-1, 500 μ gml-1, 250 μ gml-1, 125 μ gml-1, 62.5 μ gml-1, 31.25 μ gml-1,
15.625μg·ml-1, 7.8125 μ gml-1。
Preferably, the application of the present invention, can also have the feature that：Wherein, it is resistance to that 10-30 items are added in each concentration
Medicine Acinetobacter bauamnnii infects Caenorhabditis elegans model.
Advantageous effect of the invention
It is experimental subjects that the present invention, which selects nematode, carries out the drug sensitive experiment of general drug resistant Acinetobacter bauamnnii in vivo, knot
Fruit is compared to each other with drug sensitivity testing in vitro result, has clinical reference value, can make up traditional In vitro chemo-drug sensitive test screening
In omission.This will largely improve disease prognosis, reduce the death rate and hospital stays, Economy type medicine during being hospitalized
Resource reduces medical treatment cost.
Description of the drawings
Fig. 1 is nematode state diagram in 96 orifice plates under microscope；
Specific implementation mode
Illustrate the specific implementation mode of the present invention below.
1. materials and methods
The identification and collection of 1.1 bacterial strains：It is clinically separated out from the humoral specimen of drug-fast bacteria infection patient for being included in research
General drug resistance Acinetobacter bauamnnii carries out the discriminating of strain by Multiplex PCR and colloidal gold method, uses direct Sequencing when necessary
Method carries out precise Identification to bacterial strain.Culture is collected to bacterial strain to preserve.N2 Wild-type C. elegans (C.elegans,
Bristolstrain N2) and E.coli OP50 given by Genetics and Developmental Biology Institute of the Chinese Academy of Sciences.
1.2 drug sensitivity testing in vitro：Patient's drug sensitivity testing in vitro uses paper disk method and dilution method, the use of drug is conventional Bao
Graceful acinetobacter calcoaceticus drug sensitivity testing in vitro scheme.
The preparation of 1.3 instruments, reagent, culture medium
Reagent：NGM culture mediums, M9 buffer solutions, agar powder, peptone, Sodium azide (Sigma companies), agents useful for same are logical
It crosses commercial sources and buys or further use conventional reagent and prepare to obtain.The use of drug is susceptibility examination outside specific pathogen thalline
It tests routine and uses drug and other drug candidates, be formulated as a concentration of 1mg/mL, 100 μ g/ according to drug requirement respectively
ML, 50 μ g/mL set refrigerator and save backup with a diameter of 0.45 μm of membrane filtration after the completion of preparing.
Nematode growth media (NGM), for maintaining nematode.The preparation of nematode culture medium：Total volume 1L:3g NaCl,
17g agar, 2.5g peptone, 1ml cholesterol (5mg/ml is dissolved in 95% ethyl alcohol), 975ml H2O, high pressure sterilization,
Then the following solution for passing through high pressure steam sterilization is added:1ml 1M CaCl2, 1ml 1M MgSO4, 25ml 1M dipotassium hydrogen phosphates
MgSO is being added in order to avoid precipitation in pH 64Mixing is wanted when with dipotassium hydrogen phosphate.
The preparation of M9 buffer solutions：The M9 buffer solutions of 1000ml contain 15.12 gNa2HPO4·12H2O, 5gNaCl, 3g
KH2PO4, 0.25gMgSO4·7H2O。
1.5 antibacterials store the preparation of liquid：The use of drug is that specific pathogen bacterium drug sensitivity testing in vitro routinely uses drug,
And other drug candidates.
1.6 Caenorhabditis elegans bacterium infection methods：Worm's ovum is detached from pregnancy adult, incubated overnight obtains L1 phase larvas,
Larva is transferred on the E.coli lawns of NGM culture mediums the continued growth 48h in 25 DEG C of incubators, obtains L4 youth adult,
After eluting polypide with M9 buffer solutions and cleaning 3~4 times repeatedly, polypide is transferred to the general drug resistance Bao Man of plain agar tablet respectively
On acinetobacter calcoaceticus lawn, culture 8h~24 hour are infected in 25 DEG C of incubators, and the M9 that 96 orifice plates are transferred to after infection is slow
(buffer solution peptone content is 2%, beef extract content is 4%) continues to cultivate in fliud flushing, observes and records daily dead feelings
Condition.Nematode become linear type, without movement, touched with platinum wire and reactionless be judged to death.
1.7 nematode infections synchronize：The good tablet of nematode growth is collected, is washed down polypide with M9 buffer solutions, centrifuges, takes
Supernatant, washing, places it on the culture plate containing 5-FU (5-FU), is incubated overnight, obtain synchronized nematode.From
Heart condition will influence to collect the synchronization degree of nematode, centrifuge 20s, take supernatant, be repeated 3 times.
The statistical disposition of 1.8 clinical datas：This project statistics carries out statistical using 5.01 softwares of GraphPad Prism
Analysis.Survival analysis is mainly carried out using Kaplan-Meier methods, log-rank methods of inspection data between group carry out significant difference
Analysis, P < 0.01 have significant difference.
Drug sensitive test in 1.9 bodies：Drug sensitive test in nematode culture body is carried out in 96 orifice plates, and medicine 20 is added per hole
μ l, each drug multiple holes 4, including negative control and positive control.After the synchronization of M9 wash buffers is added in per hole
Nematode solution makes every hole nematode 20 or so.96 orifice plates are completed, nematode is counted, count living nematodes every 12h later, count 7
It.
2. experimental result
2.1 synchronize and infect the determination of scheme
Acinetobacter bauamnnii is not the conventional food of nematode, some researches show that nematode can distinguish the smell of different bacterium to
It avoids its harmful food and food refusal, therefore is different from nematode conventional food, the bacterium after drug sensitivity testing in vitro culture is directly by line
Worm is added thereto, and can avoid infectious bacteria to avoid nematode in this way is gathered in white space, and infection purpose is not achieved.In selected infection
On the basis of bacterium condition, infectious effect of the different infection times to nematode is demonstrated, is nematode infections Acinetobacter bauamnnii respectively
8,12,16,24h.Continue to cultivate with M9 buffer solutions after washing, observes the time-to-live of nematode, and the journey of comparison nematode infections
Degree, combined treatment process select most suitable gradient of infection to infect 16h optimums overnight for nematode, have both reached certain infection,
The time for working and observing there are treatment again.
The determination of 2.2 optimum drug concentrations：It is screened with 96 orifice plates, the 1st is classified as negative control, 8 hole of each column, injection
100 μ l of M9 buffer solutions；2nd~11 is classified as medicine sample, often arranges 8 holes, and 100 μ l of M9 buffer solutions are first added per hole, then exist
100 μ l of mother liquid medicine are added toward the 1st hole, drawing 100 μ l from the 1st hole after mixing is added to the 2nd hole, repeats the above steps, from the
100 μ l are drawn when 8 hole to discard, it is respectively 1000,500,250,125,62.5,31.25,15.625 to obtain drug concentration in this way,
7.8125μg·ml-1Serial solution；12nd is classified as positive control, each drug do two it is parallel.Positive control refers to infection
The nematode of Acinetobacter bauamnnii but non-administration.By in the metainfective polypide adding hole of Acinetobacter bauamnnii, per hole 10-30 items.
It sets in 25 DEG C of incubators and cultivates 5 days, incubator relative humidity is between 80%~85%.Just each medicine in two plates is sifted out after 5 days
The most nematode concentration of object survival are optimum drug concentration.As a result it see the table below 1.
The optimal treatment concentration for the caenorhabditis elegan that 1 different pharmaceutical of table infects Acinetobacter bauamnnii
| Drug | Concentration (μ gml-1) | Drug | Concentration (μ gml-1) |
| Piperacillin | 250 | Cefotaxime | 125 |
| Cefotaxime | 250 | Cefoperazone | 250 |
| Ampicillin and sulbactam | 125 | Amikacin | 62.5 |
| Imipenem | 125 | Lavo-ofloxacin | 125 |
| Ceftriaxone | 250 | Ciprofloxacin | 62.5 |
| Cefepime | 125 | Minocycline | 31.25 |
| Meropenem | 125 | Amphotericin B | 62.5 |
The result of 2.3 different antibiosis extract for treating infection drug-fast bacteria nematodes
According to the above operating procedure, nematode amount of survival is recorded, and draw its survival curve.Internal drug sensitive experiment screening
As a result such as the following table 2.As figure shows, Meropenem, Imipenem, ampicillin and sulbactam, Minocycline In The Treatment effect is preferable,
In addition lavo-ofloxacin, Ciprofloxacin, amikacin, amphotericin B also have different degrees of therapeutic effect.Table 2 is drug pair
The therapeutic effect of the caenorhabditis elegan of Acinetobacter bauamnnii infection.
Table 2：The survival rate of each drug different time points
2.3 with the comparison of drug sensitivity testing in vitro
It is general drug-fast bacteria that screening, which obtains 3 plants of Acinetobacter bauamnniis,.Experiment in vitro such as the following table 1.Drug sensitive experiment in comparison body
As a result, it can be seen that Imipenem and the antibiotic of Meropenem carbon blueness enzyme alkenes still have more apparent effect, with external susceptibility
As a result there is gap；It is also sensitive, therefore external in vivo that Tetracyclines minocycline, which also has therapeutic effect, experiment in vitro minocycline,
Drug sensitive test result is consistent, separately has Sulbactam compound formulation inside and outside experimental result also slightly different, Vitro Experimental Results only have
An example is quick in being, and cylinder therapeutic effect is obvious.The drug resistance situation to all kinds of antibacterials and show by Acinetobacter bauamnnii
Survival condition after beautiful nematode infections, carry out correlation analysis the results show that Acinetobacter bauamnnii to all kinds of antibacterials
Drug resistance situation is with the metainfective survival condition of caenorhabditis elegan without notable positive correlation (P > 0.05, as a result do not show).
The In vitro chemo-drug sensitive test result of the not homophyletic drug resistance Acinetobacter bauamnnii of table 3
The external drug sensitivity tests of this research are also shown, and have general drug resistant trend between the different strains of Acinetobacter bauamnnii,
It is shown in Table 3.However, to drugs such as lefofloxacin, minocyclines, the drug sensitivity testing in vitro susceptibilitys of different strains each not phase
Together.Therefore the dosage regimen that is obtained according to traditional In vitro chemo-drug sensitive test, may to cross resistance that bacterium itself has,
Bacterial Drug Resistance of Patients variation lags behind situations such as antibacterials dosage variation, and shortage considers.The present invention utilizes Bao Man not levers
The Caenorhabditis elegans of bacterium is model, is treated to it with antibiotic, observation treatment nematode survival state, will in vivo and experiment in vitro
It is compared.In this experiment, the incubation time and Infection Action of nematode are convenient, and the bacterium infection degree of nematode can pass through control
Infection time is adjusted, and establishes the model of Caenorhabditis elegans food antiseptic by preliminary experiment condition optimizing.And pass through this
The screening of 3 drug-fast bacterias of model pair obtains, Meropenem, Imipenem, ampicillin and sulbactam, Minocycline In The Treatment effect
More apparent, in addition, lavo-ofloxacin, the therapeutic effect of Ciprofloxacin, amikacin, amphotericin B does not have significant difference (P
> 0.05).
This experiment infects caenorhabditis elegan using the general drug resistant Acinetobacter bauamnnii that patient body fluid sample is isolated, and is resisted
The high flux screening of infection medicine and research establish optimal operation parameter, and have studied different antibiotic to nematode survival
The influence of time, obtained the general drug resistant Acinetobacter bauamnnii gradient of infection of nematode can quantization operation conclusion.To online polypide
Inside there is the drug of anti-infection activity, compares extracorporeal bacteria inhibitor test result and patient clinical therapeutic effect, carried out correlation analysis.
By Acinetobacter bauamnnii to the drug resistance situation and the metainfective survival condition of caenorhabditis elegan of all kinds of antibacterials, correlation point is carried out
Analysis the results show that Acinetobacter bauamnnii to drug resistance situation and the metainfective survival condition of caenorhabditis elegan to all kinds of antibacterials
Without notable positive correlation (P > 0.05).This result reflects that some sensitive medicaments may pass through enhancing by acting on host
The mechanism such as host autoimmune ability resist pathogenic bacterial infection.Therefore, to there is the drug of anti-infection activity in online polypide,
Control extracorporeal bacteria inhibitor test result and patient clinical therapeutic effect are also needed to, comprehensive evaluation is carried out.
It is experimental subjects that the present invention, which selects nematode, carries out the drug sensitive experiment of general drug resistant Acinetobacter bauamnnii in vivo, knot
Fruit is compared to each other with drug sensitivity testing in vitro result, has clinical reference value, can make up traditional In vitro chemo-drug sensitive test screening
In omission.This will largely improve disease prognosis, reduce the death rate and hospital stays, Economy type medicine during being hospitalized
Resource reduces medical treatment cost.
Claims (10)
1. a kind of method for building up of drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model, which is characterized in that
Include the following steps：
Step 1：Isolate general drug resistance Acinetobacter bauamnnii；
Step 2：Caenorhabditis elegans infects drug resistance Acinetobacter bauamnnii：Worm's ovum is detached from pregnancy adult, incubated overnight obtains
Larva is transferred on the E.coli lawns of NGM culture mediums continued growth 48h in the incubator by L1 phase larvas, obtains L4 youth
Polypide is transferred to the general drug resistance Bao Man of plain agar tablet not lever by adult respectively after being eluted polypide with buffer solution and cleaned
On the lawn of bacterium, 8h~infected for 24 hours is cultivated in the incubator；
Step 3：Nematode infections synchronize：The good tablet of nematode growth is collected, is washed down polypide with buffer solution, centrifuges, takes
Clearly, it washs, places it on the culture plate containing 5-FU, be incubated overnight, obtain synchronized Caenorhabditis elegans, drug screening
Model foundation is completed.
2. the foundation side of drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model as described in claim 1
Method, it is characterised in that：
In step 3, centrifugal condition 20S takes supernatant, is repeated 3 times.
3. the foundation side of drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model as described in claim 1
Method, it is characterised in that：
By percentage to the quality, M9 buffer solutions include：Peptone 2%, beef extract 4%.
4. the foundation side of drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model as described in claim 1
Method, it is characterised in that：
In step 2, infection time is 16 hours.
5. the foundation side of drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans medicaments sifting model as described in claim 1
Method, it is characterised in that：
Further include being transferred to the step of continuing to cultivate, observe and record daily death condition in buffer solution after infecting in step 2.
6. the drug resistance Acinetobacter bauamnnii that the method as described in claim 1 is established infects Caenorhabditis elegans model in drug sieve
The application chosen.
7. application as claimed in claim 6, which is characterized in that include the following steps：
Step 1：Negative control is set；
Step 2：Positive control is set；
Step 3：Establish the concentration gradient of each drug；
Step 4：Drug resistance Acinetobacter bauamnnii is added into the drug of each concentration gradient and infects Caenorhabditis elegans model, culture
Afterwards to the nematodal accounting of survival, nematode survival quantity is at most optimum medicine, nematode survival number in each drug
The optimum concentration of the most a concentration of drug of amount.
8. the use as claimed in claim 7, it is characterised in that：
The negative control is M9 buffer solutions, and the positive control is：The nematode of Acinetobacter bauamnnii but non-administration is infected.
9. the use as claimed in claim 7, it is characterised in that：
In step 3, drug concentration gradient is respectively 1000,500,250,125,62.5,31.25,15.625,7.8125 μ g
ml-1。
10. the use as claimed in claim 7, it is characterised in that：
Wherein, 10-30 drug resistance Acinetobacter bauamnnii infection Caenorhabditis elegans model is added in each concentration.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710576315.4A CN108719204A (en) | 2017-07-14 | 2017-07-14 | A kind of drug resistance Acinetobacter bauamnnii infects method for building up and its application of Caenorhabditis elegans medicaments sifting model |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710576315.4A CN108719204A (en) | 2017-07-14 | 2017-07-14 | A kind of drug resistance Acinetobacter bauamnnii infects method for building up and its application of Caenorhabditis elegans medicaments sifting model |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN108719204A true CN108719204A (en) | 2018-11-02 |
Family
ID=63940407
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710576315.4A Pending CN108719204A (en) | 2017-07-14 | 2017-07-14 | A kind of drug resistance Acinetobacter bauamnnii infects method for building up and its application of Caenorhabditis elegans medicaments sifting model |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108719204A (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110269923A (en) * | 2019-06-06 | 2019-09-24 | 中国人民解放军南部战区总医院 | Turmeric P.E is preparing the application in overriding resistance Acinetobacter bauamnnii drug |
| CN110302308A (en) * | 2019-06-06 | 2019-10-08 | 中国人民解放军南部战区总医院 | Pseudobulbus Cremastrae seu Pleiones extract is preparing the application in overriding resistance Acinetobacter bauamnnii drug |
| CN110302219A (en) * | 2019-06-06 | 2019-10-08 | 中国人民解放军南部战区总医院 | Cortex Pseudolaricis extract is preparing the application in overriding resistance Acinetobacter bauamnnii drug |
| CN111378715A (en) * | 2020-02-29 | 2020-07-07 | 广西壮族自治区兽医研究所 | Caenorhabditis elegans model for identifying bovine escherichia coli virulence |
| CN111398573A (en) * | 2019-01-02 | 2020-07-10 | 广东省第二人民医院 | Drug screening method based on anti-lung cancer drug resistance effect of caenorhabditis elegans |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104945329A (en) * | 2014-03-24 | 2015-09-30 | 中国医学科学院医药生物技术研究所 | Antituberculosis drug and screening method thereof |
| CN106562943A (en) * | 2016-11-09 | 2017-04-19 | 济南市儿童医院 | Memantine and application thereof in preparation of medicines for therapy of acinetobacter baumannii infection |
-
2017
- 2017-07-14 CN CN201710576315.4A patent/CN108719204A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104945329A (en) * | 2014-03-24 | 2015-09-30 | 中国医学科学院医药生物技术研究所 | Antituberculosis drug and screening method thereof |
| CN106562943A (en) * | 2016-11-09 | 2017-04-19 | 济南市儿童医院 | Memantine and application thereof in preparation of medicines for therapy of acinetobacter baumannii infection |
Non-Patent Citations (1)
| Title |
|---|
| 欧阳妮等: "秀丽隐杆线虫－多重耐药鲍曼不动杆菌感染模型的建立", 《广东医学》 * |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111398573A (en) * | 2019-01-02 | 2020-07-10 | 广东省第二人民医院 | Drug screening method based on anti-lung cancer drug resistance effect of caenorhabditis elegans |
| CN111398573B (en) * | 2019-01-02 | 2024-04-30 | 广东省第二人民医院 | Method for screening medicines with anti-lung cancer drug resistance based on caenorhabditis elegans |
| CN110269923A (en) * | 2019-06-06 | 2019-09-24 | 中国人民解放军南部战区总医院 | Turmeric P.E is preparing the application in overriding resistance Acinetobacter bauamnnii drug |
| CN110302308A (en) * | 2019-06-06 | 2019-10-08 | 中国人民解放军南部战区总医院 | Pseudobulbus Cremastrae seu Pleiones extract is preparing the application in overriding resistance Acinetobacter bauamnnii drug |
| CN110302219A (en) * | 2019-06-06 | 2019-10-08 | 中国人民解放军南部战区总医院 | Cortex Pseudolaricis extract is preparing the application in overriding resistance Acinetobacter bauamnnii drug |
| CN110302219B (en) * | 2019-06-06 | 2023-10-03 | 中国人民解放军南部战区总医院 | Application of cortex pseudolaricis extract in preparation of drug-resistant Acinetobacter baumannii drug |
| CN110269923B (en) * | 2019-06-06 | 2023-10-03 | 中国人民解放军南部战区总医院 | Application of turmeric extract in preparation of drug-resistant Acinetobacter baumannii drug |
| CN110302308B (en) * | 2019-06-06 | 2023-10-03 | 中国人民解放军南部战区总医院 | Application of Indian iphigenia bulb extract in preparation of drug-resistant acinetobacter baumannii drug |
| CN111378715A (en) * | 2020-02-29 | 2020-07-07 | 广西壮族自治区兽医研究所 | Caenorhabditis elegans model for identifying bovine escherichia coli virulence |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN108719204A (en) | A kind of drug resistance Acinetobacter bauamnnii infects method for building up and its application of Caenorhabditis elegans medicaments sifting model | |
| Beaudoin et al. | Staphylococcus aureus interaction with Pseudomonas aeruginosa biofilm enhances tobramycin resistance | |
| Arciola et al. | Detection of biofilm-forming strains of Staphylococcus epidermidis and S. aureus | |
| JP2020143119A (en) | Phage therapy of pseudomonas infections | |
| CN107751089A (en) | It is a kind of that the method for resisting general resistance Acinetobacter bauamnnii medicine is screened using Caenorhabditis elegans | |
| TW201130980A (en) | A new lytic phage specific to Klebsiella pneumoniae | |
| Alp et al. | The role of hemagglutination and effect of exopolysaccharide production on bifidobacteria adhesion to Caco‐2 cells in vitro | |
| Lopez et al. | Management of the infected total hip arthroplasty | |
| Müller-Schulte et al. | Peptoniphilus asaccharolyticus–Commensal, pathogen or synergist? Two case reports on invasive Peptoniphilus asaccharolyticus infection | |
| CN104651473B (en) | The method for determining the sub-lethal dose staphylococcus aureus resistance to the action of a drug and drug resistance simultaneously | |
| CN101177668B (en) | Novel neisseria gonorrhoeae culture medium and method for making same | |
| CN102373213B (en) | Mycobacterium tuberculosis surface lipolysaccharide-antistatic nucleic acid aptamer and application thereof | |
| KR20140077930A (en) | Biomarkers for respiratory infection | |
| Alexander et al. | Experimental investigations as a basis for treatment of type b Hemophilus influenzae meningitis in infants and children | |
| CN113975317A (en) | Application of coptis chinensis in preparation of product for promoting proliferation of beneficial bacteria in intestinal tract | |
| RU2593725C1 (en) | METHOD FOR PRODUCING ex vivo ALVEOLAR MACROPHAGE CULTURES OF THE OPERATIONAL MATERIAL OF PATIENTS SUFFERING PULMONARY TUBERCULOSIS AND METHOD FOR ESTIMATING VIRULENCE OF Mycobacterium tuberculosis USING THE OBTAINED ex vivo ALVEOLAR MACROPHAGE CULTURES | |
| Krag et al. | Serious human infections due to bacilli of the Arizona group | |
| Sibil et al. | A Rare Case of Burkholderia Pseudomallie Presenting as Brain Abscess | |
| De Seta et al. | Preclinical evaluation of pharmaceutical compositions and their components for urinary health | |
| CN105079289B (en) | Fermented Chinese gall herb and tea leaves are preparing the application in Anti-helicobacter pylori drugs | |
| CN110302308A (en) | Pseudobulbus Cremastrae seu Pleiones extract is preparing the application in overriding resistance Acinetobacter bauamnnii drug | |
| CN107875374A (en) | The method that Caenorhabditis elegans infection model is precisely quickly established based on orthogonal optimization | |
| CN113999792B (en) | Bifidobacterium proliferation promoter and preparation method and application thereof | |
| Meshram et al. | Pulmonary scedosporiosis–A rare entity | |
| Al-Najar et al. | Effect of alcoholic extracts of Punica grantum and Trigonella foenum on E. coli isolated from UTI |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20181102 |
|
| RJ01 | Rejection of invention patent application after publication |