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Multi-drug resistance Gram-negative bacteria combined drug sensitivity testing card and preparation method thereof

A technique for Gram-negative bacteria and drug susceptibility testing, which is applied to biochemical equipment and methods, and microbial measurement/inspection, can solve the problems of poor timeliness of results, high cost, and long time consumption, so as to facilitate popularization and use, reduce Effects of test cost and shortened test time

Pending Publication Date: 2019-10-25
长沙市第三医院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the current clinical commercial drug sensitivity test cards are only for a single drug variety, and none of them can provide combined drug sensitivity results. When encountering multi-drug resistant bacterial infections, physicians often can only choose combination drugs based on their own clinical experience. Species and doses, it is impossible to formulate a reasonable and effective treatment plan through effective combined drug susceptibility test results, so that the failure rate of clinical treatment increases significantly
[0005] In order to solve the problem of combined drug susceptibility, some laboratories use the disk diffusion method to conduct combined drug susceptibility tests of different drugs to investigate whether different drugs have synergistic effects so as to guide drug use, but this method can only The change of the inhibition zone presumes whether there is a synergistic effect between the two, which cannot provide accurate MIC results and thus cannot guide clinical precision medication
Some scholars have used the manual broth dilution method to conduct combined drug susceptibility studies of different drugs. For medication, Zhou Menglan, Wang Yao, etc. have studied cefoperazone, ceftazidime, imipenem, tigecycline, colistin single drug and its combination with sulbactam, but the above researches have the following problems: (1) The preparation and testing of the test card by manual method takes a long time, the timeliness of the result is poor, the cost is high, and the prepared test card cannot be stored for a long time, so it cannot be effectively promoted in the clinic; (2) The drug concentration design adopts conventional doubling dilution Design, the concentration of some drugs does not fully meet the precise drug use requirements of the drug

Method used

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  • Multi-drug resistance Gram-negative bacteria combined drug sensitivity testing card and preparation method thereof
  • Multi-drug resistance Gram-negative bacteria combined drug sensitivity testing card and preparation method thereof
  • Multi-drug resistance Gram-negative bacteria combined drug sensitivity testing card and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Test card layout: Taking the 144-hole test card as an example, four 144-hole test cards are used for layout. The design method is as follows Figure 1-4 shown or shown in Table 1-4.

[0036] Table 1 Four 144-hole test cards 1

[0037]

[0038] Table 2 Four 144-hole test cards 2

[0039]

[0040] Table 3 Four 144-hole test cards 3

[0041]

[0042] Table 4 Four 144-hole test cards 4

[0043]

[0044] Note: 1. MP-meropenem; CL-colistin; TG-tigecycline; SU-sulbactam; FM-fosfomycin; AK-amikacin; DX-doxycycline; LE- Levofloxacin; CZ-Ceftazidime; C+ means positive control well

[0045] 2. For example: CL0.25+TG0.125 means that the well is coated with 0.25ug / ml colistin and 0.125ug / ml tigecycline.

Embodiment 2

[0047] Test card preparation, the steps are as follows: (1) prepare MH broth + 0.0001% polysorbate 80 solution (PH7.2-7.4), adopt 121 ℃, 15min to carry out autoclave sterilization, prepare dilution liquid; (2) make the The required antibiotics are dissolved with the required solvent and filtered with a 0.22um filter membrane; (3) according to the layout of the drug in Example 1, the antibiotic solution is prepared by diluting the antibiotic mother liquor with a sterilized diluent; (4) the prepared Add the completed antibiotic solution according to the layout in Example 1 to a 144-well card, 50ul per well; (5) Dry by freeze-drying; (6) Finally, use aluminum film bags for vacuum packaging and store at 2-8°C .

[0048] Test card test, the steps are as follows: (1) take fresh bacterial colonies with a bacterial age of 16-24 hours as the strain to be tested; (2) use sterile saline to adjust the bacteria to be tested to 0.5 McFarland; (3) take the required amount 0.5 of McFly's liq...

Embodiment 3

[0050] Clinical case 1: The patient, male, 74 years old, had severe pneumonia complicated with sepsis. The lower respiratory tract secretions and blood cultures showed all-drug-resistant Acinetobacter baumannii. The preliminary test of the commercial drug susceptibility system showed that there was no alternative Single drugs, including colistin, meropenem, fosfomycin, amikacin, sulbactam, doxycycline, levofloxacin, and ceftazidime, were all resistant to high concentrations. The combination drug susceptibility test card designed in the present invention was used to test, and the results showed that only meropenem combined with fosfomycin had a synergistic effect, and the specific test results are shown in Table 5 below.

[0051] Table 5 Clinical case 1 using combined drug susceptibility test card test results

[0052]

[0053]

[0054] The above results show that the concentration of meropenem at the conventional resistance breakpoint concentration (8 μg / ml) combined wit...

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Abstract

The invention relates to the field of microbial in-vitro diagnosis and discloses a multi-drug resistance Gram-negative bacteria combined drug sensitivity testing card and a preparation method thereof.Combined drug sensitivity combinations comprise colistin+meropenem, colistin+fosfomycin, colistin+tigecycline, colistin+ceftazidime, tigecycline+amikacin, tigecycline+meropenem, tigecycline+fosfomycin, sulbactam+meropenem, sulbactam+doxycycline, sulbactam+tigecycline, amikacin+fosfomycin, meropenem+amikacin, meropenem+fosfomycin and meropenem+levofloxacin. The combined drug sensitivity testing card simultaneously provides 9 single drugs and 14 groups of combined drug sensitivity results, obviously improves the test efficiency, reduces the test workload and the test cost, provides MIC resultsmeeting clinical demands and offers bases to precise clinical medicine application in multi-drug resistance bacteria.

Description

technical field [0001] The invention relates to the field of in vitro diagnosis of microorganisms, in particular to a multidrug-resistant Gram-negative bacteria combined drug sensitivity test card and a preparation method thereof. Background technique [0002] The susceptibility test of antibacterial drugs (referred to as drug susceptibility test) refers to the use of in vitro methods to determine the ability of antibacterial drugs to inhibit the growth of pathogenic microorganisms. Provide a basis for the selection of drugs. Clinical microbiology laboratories use a variety of experimental methods, including disk diffusion method, agar dilution method, micro broth dilution method or various commercial detection systems, to test the drug susceptibility of clinical isolates. Different bacteria require different types of antimicrobial drugs for drug susceptibility testing. The selection of antimicrobial drug types and the determination of drug sensitivity results for various b...

Claims

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Application Information

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IPC IPC(8): C12Q1/18
CPCC12Q1/18
Inventor 李昕徐兵何开大
Owner 长沙市第三医院
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