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Objective:To assess the prevalence ofLeptospira detected in wildlife and domesticated animals in Jiangxi Province, China, in2009.Methods:Urine samples from 28 buffaloes and kidney samples from50 pigs,50 dogs and38 rats were collected from Fuliang and Shangrao County, Jiangxi Province, China, in October2009. Polymerase chain reaction(PCR) and culture analyses were used to detectLeptospira. The cultured isolates were typed using the microscopic agglutination test (MAT).Results:The results showed that rats potentially serve as the main reservoir of leptospiral infection, followed by dogs. Although16% of rats (6/38) were positive using culture analysis, PCR analysis using the diagnostic primersG1/G2 and B64I/B64II or lipL32 showed identification as50% and24%, respectively, of the rat samples as positive for the presence of leptospiralDNA.Conclusions:PCR-based detection of leptospiralDNA in infected kidney tissues of reservoirs is more efficient when usingG1/G2 primers thanlipL32 primers. However, the latter primers have a potential application for detection in urine samples. The alarmingly high prevalence of leptospiralDNAin the wild rat population near human habitation underscores the utility of routineLeptospira surveillance, preferably usingPCR methods, which are more sensitive than traditional culture-based methods.