CN103525735A - Salmonella separation selection and detection method - Google Patents
Salmonella separation selection and detection method Download PDFInfo
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- CN103525735A CN103525735A CN201310493238.8A CN201310493238A CN103525735A CN 103525735 A CN103525735 A CN 103525735A CN 201310493238 A CN201310493238 A CN 201310493238A CN 103525735 A CN103525735 A CN 103525735A
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Abstract
The invention discloses a salmonella separation selection and detection method which comprises the steps of enrichment, isolated culture, picking of suspected bacterial colony, biochemical test and serological identification, wherein the picking of suspected bacterial colony comprises the steps of stirring the suspected bacterial colony and picking the broken bacterial colony. According to the detection method, the salmonella is quickly identified in a large-scale salmonella screening work, so that the workload is remarkably reduced and the working efficiency is improved.
Description
Technical field
The separation that the present invention relates to a kind of salmonella is chosen and detection method.
Background technology
Salmonella is important enteric pathogenic bacteria, is one of important pathogen of food poisoning and diarrhoea.
At present, detect salmonella and mainly adopt the several different methods such as traditional conventional sense method, molecular biology method, immunological method.Tradition conventional sense method: gather after ight soil carries out selective enrichment with food samples and use selective separation dull and stereotyped (WS, SS, BS, DHL, HE etc.) to carry out separated, on separating plate, generally need to select a plurality of doubtful bacterium colonies, then it is carried out to biochemistry and serological evaluation.Adopt automatic bacterial analyser to detect: bacterium micro biochemical identification systems are connected with computer, and the qualification result of bacterium is processed by computer, and automatically reports the result, and is exactly microbiological analysis instrument.After sample to be checked increases bacterium, culture is processed through specific, gets a certain amount for the treatment of solution injection instrument and detects, and detected data are calculated through MICROCOMPUTER PROCESSING, the data of contrast computer database storage, then judged result.Immunological method has euzymelinked immunosorbent assay (ELISA) (ELISA): the method is utilized antigen---the specificity of antibody response, adopt the micro plate that has been coated with in advance Salmonellas (A-E group) monoclonal antibody, add the sample of processing through increasing bacterium, after reaction, add again developer, after effect, by microplate reader, measure OD value and carry out result of determination.ELISA method detects the limit range of Salmonellas 10
5-10
6cfu/ml, therefore, draw reliable result, detects sample and first must increase in advance bacterium, selective enrichment.As euzymelinked immunosorbent assay (ELISA) (ELISA), gene chips, polymerase chain method (PCR), Transia salmonella card detection method etc., these methods preferably resolve specificity and the sensitive question that salmonella detects, but all need expensive plant and instrument, and testing cost is higher, and be not suitable for the examination of large sample.
Although traditional cellar culture isolation technique much time power, no matter be that medical diagnosis on disease or hygienical evaluation remain indispensable important method up to now, is that the checks such as Protocols in Molecular Biology are irreplaceable.But in the method, when separating plate is selected doubtful bacterium colony, mixing of a large amount of doubtful bacterium colonies needs follow-up biochemistry and serological identification, wastes time and energy, and therefore how reducing in institute's choosing colony to the quantity of the similar bacterium colony of salmonella of target detect is need to deal with problems at present.
Summary of the invention
For a difficult problem for above-mentioned prior art, the object of this invention is to provide a kind of detection method of salmonella, reduce in institute's choosing colony the quantity to the similar bacterium colony of salmonella of target detect.
Technical scheme of the present invention is such:
A separated choosing method, it is characterized in that, stir the broken bacterium colony of doubtful bacterium colony and picking.
A detection method for salmonella, comprises and increases bacterium, separation and Culture, the doubtful bacterium colony of picking, biochemical test and serological identification, and it is characterized in that, the doubtful bacterium colony of described picking comprises step: stir the broken bacterium colony of doubtful bacterium colony and picking.
It is described that to stir doubtful bacterium colony be to stir the bacterium colony that translucent center is black.
Described increasing bacterium be by specimen inoculation in selenite enrichment medium, cultivate 18～24h for 35 ℃～37 ℃.
Described separation and Culture is 35 ℃～37 ℃ cultivation 18～24h on WS agar plate.
Technical scheme provided by the present invention, has improved the recognition rate of the doubtful bacterium colony of salmonella on normal isolation culture base WS flat board, to reduce false positive colony counts to reduce follow-up unnecessary loaded down with trivial details evaluation work.In salmonella testing, a large amount of false positives are abandoned in pick, are conducive to accurately detect real salmonella.Present method, on traditional substratum WS flat board, is used that specific identification method selects that Suspicious Salmonella feature is obvious, method simple, easily identification, specificity be high.In extensive salmonella screening operation, identify fast salmonella, significantly reduce workload, increase work efficiency, Rapid Detection salmonella.The salmonella of carrying disease germs enteron aisle detects from traditional 4-7 days, reduces to 2-3 days.
Embodiment
Below in conjunction with embodiment, the invention will be further described, but not as a limitation of the invention.
Get stool sample 0.5～1g and be inoculated in 35 ℃～37 ℃ cultivation 18～24h of 10ml selenite enrichment medium (SF enrichment liquid), with transfering loop picking enrichment liquid, be inoculated on selective medium (WS agar plate), cultivate 18～24h for 35 ℃～37 ℃, from WS agar plate, with inoculating needle, stir the suspicious bacterium colony of central black, and the bacterium colony that picking is stirred rear fragmentation carries out biochemical test and serological identification.Biochemical test: picking can bacterium colony culture transferring in Ke Shi disaccharide, and do oxydase, nitrate, KCN, Methionin test; Or culture transferring is in salmonella biochemical identification plate; Or use microorganism automatic identifying system to identify.What meet salmonella biochemical reaction does serological test again; Serological identification determines group according to salmonella O serum, then uses H factor serum the 1st phase and its affiliated serotype of the 2nd phase study result synthetic determination.Inventive point of the present invention is after WS agar plate separation and Culture, during picking colony, first stir the suspicious bacterium colony that meets salmonella colony characteristics, then in these suspicious bacterium colonies of picking, be vaccinated pin and stir broken bacterium colony, obtain the suspicious bacterium colony of salmonella of less false positive bacterium colony, reduce the workload of follow-up biochemical test and serological identification.
13278 intestines inspection samples, adopt the inventive method to detect 102 strains, and conventional method detects 101 strains, susceptibility contrast there was no significant difference (x2=0.116, p>0.05); Specificity contrast the inventive method 99.98%, conventional method 90.33%, has significant differences (P<0.005).Ke Majia CAS color developing culture medium, owing to containing special substance that show color, without by any instrument, only can with the naked eye judge at 24-48 hour by observing colony colour.Foreign scholar detects and isolates 20 strain salmonella 508 parts of clinical samples, and sensitivity is 100%, and specificity is 88.9%.Dull and stereotyped (CAS) simultaneous test of the inventive method and import colour developing: by 936 parts of health check-up sample detection, result detects 6 strains, and dull and stereotyped (CAS) susceptibility of colour developing is 100%, and specificity is 96.1%; The inventive method susceptibility is 100%, and specificity is 100%.
Claims (5)
1. a separated choosing method for salmonella, is characterized in that, stirs the broken bacterium colony of doubtful bacterium colony and picking.
2. a detection method for salmonella, comprises and increases bacterium, separation and Culture, the doubtful bacterium colony of picking, biochemical test and serological identification, and it is characterized in that, the doubtful bacterium colony of described picking comprises step: stir the broken bacterium colony of doubtful bacterium colony and picking.
3. the detection method of salmonella according to claim 2, is characterized in that: described in to stir doubtful bacterium colony be to stir the bacterium colony that translucent center is black.
4. the detection method of salmonella according to claim 2, is characterized in that: described increasing bacterium for by specimen inoculation in selenite enrichment medium, cultivate 18～24h for 35 ℃～37 ℃.
5. the detection method of salmonella according to claim 2, is characterized in that: described separation and Culture is 35 ℃～37 ℃ cultivation 18～24h on WS agar plate.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN201310493238.8A CN103525735A (en) | 2013-10-21 | 2013-10-21 | Salmonella separation selection and detection method |
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| CN201310493238.8A CN103525735A (en) | 2013-10-21 | 2013-10-21 | Salmonella separation selection and detection method |
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2013
- 2013-10-21 CN CN201310493238.8A patent/CN103525735A/en active Pending
Non-Patent Citations (5)
| Title |
|---|
| 刘继倩等: "两种培养基在健康人群中沙门菌检测的比较", 《中国实用医药》 * |
| 刘继倩等: "两种培养基在健康人群中沙门菌检测的比较", 《中国实用医药》, vol. 6, no. 31, 10 November 2011 (2011-11-10) * |
| 张缪伟: "利用WS琼脂特殊菌落挑选沙门氏菌", 《中国卫生检验杂志》 * |
| 罗蓓等: "两种检测方法在沙门菌初筛中的应用比较", 《上海预防医学杂志》 * |
| 胡雪明等: "沙门菌分离技术关键点的研究和评价", 《中国人兽共患病学报》 * |
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Application publication date: 20140122 |