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Method and kit for rapidly detecting drug resistance and virulence of staphylococcus aureus

A staphylococcus, golden yellow technology, applied in the field of clinical testing, can solve life-threatening, tedious, time-consuming and labor-intensive detection problems, and achieve the effect of reducing the possibility and improving the survival rate

Inactive Publication Date: 2017-03-29
JIANGSU UNIV
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  • Claims
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AI Technical Summary

Problems solved by technology

Although Staphylococcus aureus is traditionally regarded as hospital-acquired bacteria, Staphylococcus aureus and MRSA infections, which are prevalent in the community and family wards, can cause severe pneumonia and life-threatening symptoms in patients, and a simple and rapid method is urgently needed Detect MRSA bacteria and assess bacterial virulence
[0003] The existing identification method of Staphylococcus aureus includes spreading the specimen to be tested on a blood plate to separate a single colony, picking a single colony for biochemical reaction identification, and performing drug sensitivity testing on the identified Staphylococcus aureus. The whole process is at least It takes 3 to 5 days, and the existing methods are cumbersome, time-consuming and labor-intensive, and cannot meet the requirements for rapid detection of MRSA bacteria in communities and family wards
In addition, the existing identification methods for Staphylococcus aureus can only detect bacterial drug resistance, but cannot detect bacterial virulence

Method used

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  • Method and kit for rapidly detecting drug resistance and virulence of staphylococcus aureus
  • Method and kit for rapidly detecting drug resistance and virulence of staphylococcus aureus

Examples

Experimental program
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Effect test

Embodiment 1

[0021] Embodiment 1: Multiplex PCR method detects that Staphylococcus aureus carries spa gene, mecA gene and pvl gene

[0022] Randomly selected 17 clinical isolates of Staphylococcus aureus (from the Affiliated Hospital of Jiangsu University), including 10 strains of MRSA bacteria and 7 strains of non-MRSA bacteria. The types and drug resistance of the strains were identified by the VITEK 2-Compact automatic bacterial identification and drug susceptibility instrument of the French company BioMérieux.

[0023] The Staphylococcus aureus used in the experiment was inoculated on a blood agar plate, cultivated at 35°C for 16 hours to form a single colony, and then picked a single colony and inoculated it on MH medium, cultured with shaking at 37°C for 24 hours, sucked 200 μL of fresh saturated bacterial solution, 12000 Centrifuge at rpm for 2 min to obtain bacterial pellet for bacterial genomic DNA extraction.

[0024] Staphylococcus aureus genomic DNA extraction steps:

[0025]...

Embodiment 2

[0044] Embodiment 2: multiplex PCR method specific detection

[0045] Extract Escherichia coli (American Type Culture Collection, ATCC 25922), Pseudomonas aeruginosa (American Type Culture Collection, ATCC 27853), Klebsiella pneumoniae (American Type Culture Collection, ATCC 700603) , Streptococcus pneumoniae (American Type Culture Collection, ATCC 496), the above bacteria were purchased. Genomic DNA was extracted from the pure culture of the above bacteria with a bacterial genomic DNA extraction kit (Beijing Tiangen Biochemical Co., Ltd., DP302), and the extracted DNA was directly used for multiplex PCR amplification.

[0046] The primer sequences for the spa gene, mecA gene and pvl gene of Staphylococcus aureus are as described in Example 1.

[0047] The total PCR reaction system is 50 μL, and 25 μL of 2×PCR Mix premix solution, 0.25 μM of spa gene primers, 0.5 μM of mecA gene primers, and 1 μM of pvl gene primers are added in turn to the PCR reaction tube, and bacterial ge...

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Abstract

The invention belongs to the field of clinical examination and diagnosis, and relates to a method and kit for rapidly detecting the drug resistance and virulence of staphylococcus aureus. The kit comprises three pairs of multiple PCR amplification primers, 2*PCR premix system, and sterilized double distilled water, wherein the multiple PCR amplification primers are spa gene upstream / downstream primers, mecA gene amplification upstream / downstream primers, and pvl gene amplification upstream / downstream primers. The kit comprises three pairs of PCR primers and thus can simultaneously amplify the spa, mecA, and pvl genes of staphylococcus aureus. The kit can be used to detect staphylococcus aureus, which is resistant to methicillin, in a clinical sample, can detect staphylococcus aureus with high virulence at the same time, and provides references for the clinical treatment on staphylococcus aureus infection. The invention establishes a method for rapidly detecting drug resistance and virulence of staphylococcus aureus, and the method is simple, rapid, and cheap and has an important meaning for the clinical treatment on staphylococcus aureus infection.

Description

technical field [0001] The invention belongs to the technical field of clinical testing, and relates to a rapid detection method and kit for staphylococcus aureus drug resistance and virulence. Background technique [0002] Staphylococcus aureus ( Staphylococcus aureus , Sa) is one of the clinically important pathogenic bacteria, which can produce more virulence factors and cause food poisoning, pseudomembranous enteritis, scalded skin syndrome, toxic shock syndrome, suppurative inflammation and sepsis, etc. , causing great damage to human health. Methicillin-resistant Staphylococcus aureus (Multiple-resistant Staphylococcus aureus , MRSA) is a unique strain of Staphylococcus aureus that is resistant to all penicillins, including methicillin and other β-lactamase-resistant penicillins. MRSA was first discovered in the United Kingdom in 1961, and has now spread widely, becoming one of the most serious drug-resistant bacteria in clinical practice. Although Staphylococcus a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/14C12N15/11
CPCC12Q1/689C12Q2600/16
Inventor 陈盛霞吴亮阴晴姜旭淦孙晓春
Owner JIANGSU UNIV
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