The Study On Sanhuang Capsule To Eliminate R Plasmid Of Neisseria Gonorrhoeae

objective: Bacterial plasmid included the F factor (fertility factor),R factor(resistance factor), Col factor (coliconogenic factor), Ti factor (tumor inducing plasmid), huge plasmid (mega plasmid) and the plasmid of degradation. R factor which was also known as the R plasmid carried resistance genes, giving certain bacterial resistance to antibiotics. The past 30 years, a number of researchers both at home and abroad actively sought an effective way to eliminate resistance plasmids in order to reverse bacterial resistance. Gonorrhea was one of the most common sexually transmitted diseases;with the widespread use of antibiotics, the resistance of Neisseria gonorrhoeae became more and more serious. The study found that many more drug-resistant Neisseria gonorrhoeae was mediated by the plasmid. Internal used the traditional Chinese medicine of clearing away heat and toxic types for elimination of G-bacteria plasmid, and this medicine was precisely the prescription of Chinese medicine treatment of Gonorrhea. So we could speculate that the base of traditional Chinese medicine treatment of Gonorrhea was a certain degree with the elimination of plasmid. This experiment planed to study whether the Sanhuang could eliminate R plasmid of Neisseria gonorrhoeae,whether Sanhuang could reforce the effect of its elimination results with the extending time. Methods : 1.Extraction of Neisseria gonorrhoeae Plasmid: 10 Neisseria gonorrhoeae was selected from Sichuan Institute of Dermatology out-patient in 2008 after identification, and first recoveried on the non-selective TM, 24h latter was switched to the new 10 TM mediums until 24h ,then respectively scratched Neisseria gonorrhoeae to 10 Ep tubes filled with 1ml of normal saline and marked .Follewed the AxyPrep plasmid extraction kit and extracted plasmid .Observed the plasmid after electrophoresis. 2.Determination of Sanhuang sub-MIC: Prepared 105/ml concentration of Neisseria gonorrhea. Neisseria gonorrhea would be inoculated on different amount (6.25ul, 12.5ul, 25ul, 50ul, 100ul, 200ul) of Sanhuang TM medium (the medium in each dish of 15ml) , 24h latter observed the growth in order to determine sub-MIC. 3.Elimination experiment of Neisseria gonorrhoeae plasmid: the above 10 Neisseria Gonorrhoeae recoveried the same way again, and the species was transferred on the Sanhuang TM, 24h, 48h, 72h of elimination,then observed . The operation was as follows: The transferred was inoculated on Sanhuang TM for 24h,then extracted and observed(compared to pre-elimination);at the same time transferred on Sanhuang TM for 24h another ,then extracted and observed(compared to pre-elimination); at the same time transferred on Sanhuang TM for 24h another ,then extracted and observed(compared to pre-elimination);at last the conversion rates of 24h,48h,72h were compared. Results: All of selected Neisseria gonorrhoeae growth was good, and not be contaminated. 1. Extraction of plasmid: nine of the 10 gonorrhoeae's plasmid could be seen more clearly, and their molecular weights were 7.4Kb both, PPNG strain of the Asian type. 2. Neisseria gonorrhoeae could grow on the Sanhuang TM medium of different amounts(6.25ul, 12.5ul, 25ul, 50ul, 100ul, 200ul),so Sanhuang sub-MIC (ie, Neisseria gonorrhoeae can grow under the highest drug concentration) identified as 200ul / dish 3. 10 Neisseria gonorrhoeae grew on the Sanhuang TM medium, respectively, 24h, 48h, 72h, the Neisseria gonorrhoeae plasmids of three time points were extracted and observed after electrophoresis (compared to pre-elimination ): two bands reduction after 24h , three bands reduction after 48h,eight bands reduction after 72h. The conversion rates of 24h and 48h were no significant differences from that of pre-elimination (P>0.05). The conversion rate of 72h was significant difference from that of pre- elimination (P<0.05).The conversion rates of 24h,48h,72h had significant differences(P<0.05). Conclusion: In this experiment ,the result showed that Sanhuang could eliminate the plasmid of Neisseria gonorrhoeae, and the effect of the elimination became more and more obvious with extending time,72h highest.