The Composition of a Selective Media for Trichomonas vaginalis Isolation

Yang, Nam Woong; Lim, Yong; Shin, Sung Heui; Park, Jong Hun; Lee, Kang Kil; Ra, Yun Jung; Jang, Ha Jong

Abstract

BACKGROUND
Modified Diamond medium (MDM) supplemented with 10% heat-inactivated horse serum, streptomycin, penicillin G, and mycostatin is commonly used for the isolation of Trichomonas vaginalis from vaginal swab. But, judging from our experience, the above usual MDM antibiotic composition was frequently contaminated with facultative anaerobes, and isolation rate of T. vaginalis was no more than 12% in 142 korean woman patients whose chief complaints were foul odored, increased vaginal discharge. This isolation rate is low in comparison with reports of another countries including U.S.A (about 15~30%) and could be attributed to the prevalence of antibiotic resistance in Korea. So, we exploited more selective antibiotic compositions in modified Diamond medium for pure isolation of T. vaginalis.
METHODS
We used new self-devised anaerobic pack for sample maintenance and tested several antibacterial and antifungal agent combinations in modified Diamond medium supplemented with 5% human erythrocyte lysate and 5% heat-inactivated human serum in the place of 10% horse serum with the object of increased and pure isolation of T. vaginalis. Several drugs and chemicals were tested to fourteen wild strains isolated in a local clinic, in the hope of finding the agents that have no effect on T. vaginalis growth in high drug concentrations. Anaerobic jar was used for culture of T. vaginalis and cell count performed in the improved Neubauer's haemocytometer.
RESULTS
Strains of T. vaginalis grew batter in modified Diamond medium supplemented with 5% human erythrocyte lysate and 5% heat-inactivated human serum (mean 1.166X106, about 5.83 fold) than 10% horse serum (mean 2.0X105 after 48 hours culture), and their growth rate in the former was more rapid than the latter in early grow phase. On the basis of this results, we examined selectivity of modified Diamond media supplemented with several antibacterial and antifungal combinations by a double blind test. Isolation rate in the conventional modified Diamond's medium (combination A; 10% horse serum, streptomycin 1,200 microgram/mL, penicillin G 1,500 unit/mL, mycostatin 37.5 microgram/mL, pH 6.2) was 9/73 (12.3%) while in modified Diamond medium supplemented with 5% human erythrocyte lysate and 5% heat-inactivated human serum, isolation rates in various drug combinations were as follows; Combination B (cefazolin 100 microgram/mL, streptomycin 1,200 microgram/mL, clindamycin 150 microgram/mL, pH 6.5), combination C (bacitracin 14.6 unit/mL, streptomycin 1,200 microgram/mL, clindamycin 150 microgram/mL pH 6.5) and combination D (vancomycin 100 microgram/mL, streptomycin 1,200 microgram/mL, clindamycin 150 microgram/mL pH 6.5) were all 11/73 (15.0%). Combination D allowed the least bacterial growth rate.
CONCLUSION
We consider that a new modified Diamond medium supplemented with 5% human erythrocyte lysate, 5% heat-inactivated human serum and combination D might be provide the highest selection for Trichomonas vaginalis pure isolation from vaginal swabs.