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Reagent kit and screening method for screening carbapenem resistant enterobacteria from excrement and urine sample

A technology for screening carbapenems and enterobacteriaceae, applied in the fields of molecular biology and medical microbiology, to achieve convenient and quick use, increase positive rate, and reduce fungal interference

Inactive Publication Date: 2019-04-02
王汉宇
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] At present, the most common screening method for CRE in feces reported in the literature is the method recommended by the US CDC, that is, sampling ertapenem discs and adding 5ml of broth, picking an appropriate amount of feces, penem for 18-20 hours, and the next day Transfer to MacConkey or China blue plates, and obtain CRE according to the growth of colonies on the plates. This method is relatively simple to operate, but the problem is that there are a large number of bacteria and fungi in the feces. In the presence of the antibacterial drug Ertapenem Under conditions, the fungus is naturally resistant to the antibiotic and thus grows in large numbers

Method used

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  • Reagent kit and screening method for screening carbapenem resistant enterobacteria from excrement and urine sample
  • Reagent kit and screening method for screening carbapenem resistant enterobacteria from excrement and urine sample
  • Reagent kit and screening method for screening carbapenem resistant enterobacteria from excrement and urine sample

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Experimental program
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Effect test

Embodiment 1

[0020] Embodiment 1 (broth enrichment method)

[0021] 1 Sample inoculation and enrichment for the first time: pick an appropriate amount of feces (about the size of a matchhead) with a cotton swab and inoculate into 5ml of feces enrichment broth, and incubate in a 35°C incubator for 8-12 hours.

[0022] 2. The second drug-containing plate screening of the sample: pick a loop of the enrichment solution after the first broth enrichment and inoculate it on a Chinese blue plate with 0.3ug / ml meropenem, and incubate in a 35°C incubator for 8- 12h.

[0023] 3. From the colonies grown on the plate containing the drug, determine whether there is CRE by the following methods:

[0024] 3.1 Bacterial identification: Pick a single colony from the drug-containing plate and spread it evenly on the target plate for mass spectrometry. After drying, add 1 μl of matrix solution to cover the target point and let it dry naturally. Put the target plate into the mass spectrometer to start the id...

Embodiment 2

[0027] Embodiment 2 (CDC method)

[0028] 1 Sample inoculation: Pick up an appropriate amount of feces (about the size of a matchhead) with a cotton swab and inoculate into 5ml of fecal enrichment broth, add a piece of ertapenem disc (10ug / ml), and incubate in a 35°C incubator for 8-12h.

[0029] 2. Transfer of the enrichment solution: Pick a loop of the enrichment solution after the first enrichment of the broth containing the drug and inoculate it on a Chinese blue plate commonly used in clinical practice (the Chinese blue plate in this example does not contain meropenem), 35 Incubate in an incubator for 8-12 hours.

[0030] 3. From the colonies grown on the plate containing the drug, determine whether there is CRE by the following methods:

[0031] 3.1 Bacterial identification: Pick a single colony from the drug-containing plate and spread it evenly on the target plate for mass spectrometry. After drying, add 1 μl of matrix solution to cover the target point and let it dry...

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Abstract

The invention discloses a reagent kit for screening carbapenem resistant enterobacteria (CRE) from an excrement and urine sample and a method for screening the carbapenem resistant enterobacteria (CRE). The reagent kit comprises 5ml of excrement and urine and broth bacterium increasing fluid and a medicine-containing plate, wherein the excrement and urine and broth bacterium increasing fluid comprises 0.5wt% of sodium chloride, 0.5wt% of yeast powder and 1wt% of phytone, a substrate is bacteria-free water, the PH is 7.0, and high-pressure sterilization is performed at 121 DEG C for 15min; andthe medicine-containing plate is a China blue plate which comprises meropenem, and the concentration of the meropenem is 0.3 [mug] / m. A two-step method is used for screening, firstly, bacterium increasing is performed for the first time to obtain medicine-resistance bacteria, and secondly, screening is performed with a medicine plate to obtain medicine-resistance single colonies. The method is convenient to use, the interference of infectious microbes of fungi is greatly reduced, the screening positive rate is high, and the method can be widely applied to clinical laboratories and can meet CRErapid diagnosis requirements of clinical treatment.

Description

technical field [0001] The method belongs to the technical fields of molecular biology and medical microbiology, and in particular relates to a kit and a screening method for screening carbapenem-resistant Enterobacteriaceae (CRE) in stool samples. Background technique [0002] Since the first Klebsiella pneumoniae strain producing KPC-type carbapenemase was reported in the United States in 2001, Enterobacteriaceae bacteria producing KPC enzyme have been reported in various countries and regions around the world, even in Israel, Greece, Italy and some Epidemics or outbreaks have occurred in South American countries. Among them, carbapenem-resistant Klebsiellapneumoniae (CRKP) is particularly prominent. According to the historical data of CHINET bacterial resistance monitoring in my country, the resistance rate of Enterobacteriaceae bacteria to carbapenem antibiotics is increasing year by year, especially the resistance rate of Klebsiella pneumoniae to carbapenem antibiotics...

Claims

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Application Information

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IPC IPC(8): C12Q1/10
CPCC12Q1/10Y02A50/30
Inventor 王汉宇李佳萍黄子衔胡洁孙巧玲舒玲斌张嵘
Owner 王汉宇
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