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PCR detection of Yersinia pestis in fleas: comparison with mouse inoculation
D M Engelthaler1, K L Gage, J A Montenieri
1Division of Vector-Borne Infectious Diseases, Centers for Disease Control and Prevention, Public Health Service, U.S. Department of Health and Human Services, Fort Collins, Colorado 80522, USA.
Abstract:
The "gold standard" for identifying Yersinia pestis-infected fleas has been inoculation of mice with pooled flea material. Inoculated mice are monitored for 21 days, and those that die are further analyzed for Y. pestis infection by fluorescent-antibody assay and/or culture. PCR may provide a more rapid and sensitive alternative for identifying Y. pestis in fleas. To compare these assays, samples were prepared from 381 field-collected fleas. Each flea was analyzed individually by both PCR and mouse inoculation. Sixty of the 381 flea samples were positive for Y. pestis by PCR; 48 of these PCR-positive samples caused death in mice (80.0% agreement). None of the 321 PCR-negative samples caused death. Among the 12 mice that survived inoculation with PCR-positive samples, 10 were later demonstrated by serology or culture to have been infected with Y. pestis. This suggests that death of inoculated mice is less reliable than PCR as an indicator of the presence of Y. pestis in flea samples. Mouse inoculation assays produce results that are comparable to PCR only when surviving as well as dead mice are analyzed for infection. The rapidity and sensitivity (10 to 100 CFU of Y. pestis) of PCR suggest that it could serve as a useful alternative to mouse inoculation for routine plague surveillance and outbreak investigations.
Insights
Polymerase chain reaction (PCR) offers a faster and more sensitive method for detecting Yersinia pestis in fleas compared to traditional mouse inoculation. This study highlights PCR as a valuable tool for plague surveillance and outbreak investigations.
Area of Science:
- Veterinary Entomology
- Microbiology
- Infectious Diseases
Background:
- The traditional method for detecting Yersinia pestis in fleas involves mouse inoculation, a process that is time-consuming and potentially less sensitive.
- Polymerase chain reaction (PCR) has emerged as a potentially more rapid and sensitive diagnostic tool for bacterial identification.
Purpose of the Study:
- To compare the efficacy of PCR versus traditional mouse inoculation for identifying Yersinia pestis in field-collected fleas.
- To evaluate the reliability and sensitivity of each method for plague surveillance.
Main Methods:
- 381 field-collected fleas were individually tested using both PCR and mouse inoculation.
- Mouse inoculation involved monitoring mice for 21 days, with dead mice analyzed for Y. pestis.
- PCR analysis was performed on flea samples, and results were compared with mouse inoculation outcomes.
Main Results:
- PCR identified 60 out of 381 fleas as positive for Yersinia pestis.
- 80% agreement was observed between PCR-positive samples and those causing mouse mortality.
- PCR demonstrated higher sensitivity, detecting as few as 10-100 CFU of Y. pestis, and identified infections in mice that survived inoculation.
Conclusions:
- PCR is a more reliable and sensitive method than mouse inoculation for detecting Yersinia pestis in fleas.
- PCR's speed and sensitivity make it a suitable alternative for routine plague surveillance and outbreak investigations.
- Analyzing both dead and surviving mice is crucial for comparable results with PCR in mouse inoculation assays.