| PrefacesStaphylococcus aureus (S. aureus), which is widely distributed in the nature and the surface of skin and mucous membrane in human and animals, can generate a variety of toxins and toxin enzyme. S. aureus is one of the main pathogens which can lead to human opportunistic infection and nosocomial infection. Since MRSA (methicillin-resistant Staphylococccus aureus) was firstly discovered in Europe and the US in the 1960's, The proportion of MRSA from clinical isolates of S. aureus has been increasing. It has become an important pathogen of nosocomial infections. The results showed that the proportion of MRSA infections in China was up to more than 25% in nosocomial infections, and the proportion of MRSA detection from S. aureus infections in hospital was more than 74%. However, MRSA infections are no longer just a hospital-acquired diseases. The community-acquired MRSA (CA-MRSA) infections and frequent reports of its epidemic outbreaks are also common. In 1998-1999, the proportion of CA-MRSA infections from the infections of S. aureus was up to 21.84% in nine big cities of China. In 2002-2003, the infection rate of CA-MRSA was up to 37%, and it had been higher more and more. Researchers believe that CA-MRSA infection may become a threat to human health. Therefore, it is necessary to understand the nasal colonization status and drug resistance of S. aureus in the healthy students from the university, in order to prevent the formation of more resistant strains, and to guide the rational use of antibiotics.Methods1. The Germiculture of the nasal swab specimentsThe bacteria were obtained in the nasal cavity from healthy students of China Medical University and cultured in LB medium in order to increase the amount of bacteria. The collections were divided into two groups, A group and B group. A group of 320 specimens were collected by the students themselves. B group of 210 specimens were collected by the researcher. The following steps of A group were done by the students and the researcher respectively and that of B group were done by the researcher.2. Gram stainingThe samples which had been proliferated were stained with the method of gram stainming:smear-desiccation-fixation-staining-microscopic examination.3. Isolating the samples of positive and spherical bacteria with high-salt mannitol mediumThe samples of positive and spherical bacteria from the LB medium were cultured with high-salt mannitol medium in the incubator of 37℃for 36 hours.4. Coagulase testA clean glass slide was marked three circles with the number 1-3 on its back; Take 50μl NS onto the 2,3-marked circle; Take 50μl rabbit plasma onto the 1,3-marked circle; Finally, take 50μl spherical bacteria from the LB onto the 1,2-marked circle. The results were observed within 15s after shanking.5. Test of drug sensitivityThe samples of the double-positive S. aureus were daubed repeatedly on the ordinary agarplate by the sterile glass rod, in order to make the bacteria grow equably. Then the drug papers(penicillin G, ampicillin, erythromycin, streptomycin, gentamicin) were put on the plate medium and pressed tightly by the sterilized tweezers. Finally, the agarplates were cultured in the incubator of 37℃for 17 hours.6. PCR amplification for mecA gene detectionThe 5μl bacterium of the double-positive S. aureus are broken with the template; The PCR started at 94℃for 5 min, and the condition of 40 cycles is at 94℃30s,50℃30s,72℃1min, and nextly extend to 5 min at 72℃. After amplification,5μl products was taken to make electrophoresis of 10g/L agarose gel (with EB). Finally, the result was observed and photographed by the system of computer camera. 7. Statistical treatmentThe statistical analysis of the obtained data was made with the software SPSS13.5.Results1. A group:Among 320 samples,56 of which have showed spherical bacteria with gram staining positive; Among these 56 cases,26 cases are yellow colony in high-salt mannitol, and 27cases are positive in coagulase test, and 24 cases are double-positive S. aureus; In these 24 cases,5 cases have resistance to penicillin G, and 4 cases have resistance to ampicillin, and 3 cases have resistance to erythromycin, and 4 cases are positive with mecA gene by PCR amplification.2. B group:Among 210 samples,123 of which have showed spherical bacteria with gram staining positive; Among these 123 cases,27 cases are yellow colony in high-salt mannitol, and 24cases are positive in coagulase test, and 22 cases are double-positive S. aureus; In these 22 cases,12 cases have resistance to penicillin G, and 10 cases have resistance to ampicillin, and 3cases have resistance to erythromycin, and 11 cases are positive with mecA gene by PCR amplification.Conclusion1. A group:There are 24 cases where the staphylococcus aureus exist in 320 cases of healthy students, it is 7.5%, and 4 cases have mecA gene positive, it is 1.3%.4 cases of positive mecA gene have resistant to penicillin G (100%), ampicillin (100%) and erythromycin (75%), sensitivity to streptomycin (100%) and gentamicin (100%).2. B group:There are 22 cases where the staphylococcus aureus exist in 210 cases of healthy students, it is 10.5%, and 11 cases have mecA gene positive, it is 5.2%.11 cases of positive mecA gene have resistant to penicillin G (100%), ampicillin (90.9%) and erythromycin (27.3%), sensitivity to streptomycin (100%) and gentamicin (100%). |
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