Isolation and identification of Yersinia pestis phages from squirrels in Yunnan and their epidemiological significance / 中华地方病学杂志

ABSTRACT

Objective:To investigate whether the squirrels in Yunnan Province carried Yersinia pestis phages and their epidemiological significance. Methods:From 2015 to 2018, plague host animals were investigated in five of Yunnan plague foci and non-plague foci. The spleen, liver and intestinal specimens of the squirrels captured in the investigation were taken and stored at low temperature for later use. Intestinal specimens with PBS solution, were filtered by 0.22 μm and added to LB liquid medium containing 100 μl suspension of plague vaccine strain (EV76) and then oscillated in a constant temperature gas bath at 28 ℃ and 220 r/min for 18 to 24 h. The double-layer plate method was used to isolate and observe the growth of plaque. The morphology and structure of Yersinia pestis phages were observed under electron microscope. Meanwhile, spleen, liver and intestinal specimens were taken for detection of Yersinia pestis specific marker gene caf1. Results:A total of 10 squirrels were captured (8 Callosciurus erythraeus and 2 Dremomys pernyi), and four Yersinia pestis phages were isolated (2 in Callosciurus erythraeus and 2 in Dremomys pernyi). Two were isolated from non-plague foci (Yongshan County), two from house rats plague foci (Mile County and Xinping County), and none was isolated from wild radents plague foci (Jianchuan County and Eryuan County). By naked eye observation, two bacteriophages from the plague foci produced transparent plaques and grew well, while two bacteriophages from non-plague foci produced translucent plaques and with poor growth. By electron microscopy, these Yersinia pestis phages were of typical Myoviridae family, their head diameter was about 40 nm, muscle tail was about 120 nm, and tail filament cluster was slightly visible at the end of muscle tail. And all the 10 samples of squirrels were negative of plague-specific caf1 gene. Conclusions:The proportion of plague phages carried by Yunnan squirrels is relatively high. Although the detection of caf1 is negative. Squirrels may be a carrier of plague transmission due to the existence of Yersinia pestis phages.