Development of Three PCR Assays for the Differentiation between Echinococcus shiquicus, E. granulosus (G1 genotype), and E. multilocularis DNA in the Co-Endemic Region of Qinghai-Tibet plateau, China

Belgees Boufana Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, Salford, Greater Manchester, United Kingdom; French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Nancy Rabies and Wildlife Laboratory, Technopole Agricole et Vétérinaire, Malzéville, France; Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China; Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Sidi Thabet, Tunisia; School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia

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Gérald Umhang Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, Salford, Greater Manchester, United Kingdom; French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Nancy Rabies and Wildlife Laboratory, Technopole Agricole et Vétérinaire, Malzéville, France; Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China; Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Sidi Thabet, Tunisia; School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia

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Jiamin Qiu Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, Salford, Greater Manchester, United Kingdom; French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Nancy Rabies and Wildlife Laboratory, Technopole Agricole et Vétérinaire, Malzéville, France; Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China; Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Sidi Thabet, Tunisia; School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia

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Xingwang Chen Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, Salford, Greater Manchester, United Kingdom; French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Nancy Rabies and Wildlife Laboratory, Technopole Agricole et Vétérinaire, Malzéville, France; Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China; Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Sidi Thabet, Tunisia; School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia

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Samia Lahmar Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, Salford, Greater Manchester, United Kingdom; French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Nancy Rabies and Wildlife Laboratory, Technopole Agricole et Vétérinaire, Malzéville, France; Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China; Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Sidi Thabet, Tunisia; School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia

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Franck Boué Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, Salford, Greater Manchester, United Kingdom; French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Nancy Rabies and Wildlife Laboratory, Technopole Agricole et Vétérinaire, Malzéville, France; Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China; Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Sidi Thabet, Tunisia; School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia

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David Jenkins Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, Salford, Greater Manchester, United Kingdom; French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Nancy Rabies and Wildlife Laboratory, Technopole Agricole et Vétérinaire, Malzéville, France; Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China; Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Sidi Thabet, Tunisia; School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia

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Philip Craig Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, Salford, Greater Manchester, United Kingdom; French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Nancy Rabies and Wildlife Laboratory, Technopole Agricole et Vétérinaire, Malzéville, France; Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China; Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Sidi Thabet, Tunisia; School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia

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To investigate echinococcosis in co-endemic regions, three polymerase chain reaction (PCR) assays based on the amplification of a fragment within the NADH dehydrogenase subunit 1 (ND1) mitochondrial gene were optimized for the detection of Echinococcus shiquicus, Echinococcus granulosus G1, and Echinococcus multilocularis DNA derived from parasite tissue or canid fecal samples. Specificity using parasite tissue-derived DNA was found to be 100% except for E. shiquicus primers that faintly detected E. equinus DNA. Sensitivity of the three assays for DNA detection was between 2 and 10 pg. Ethanol precipitation of negative PCR fecal samples was used to eliminate false negatives and served to increase sensitivity as exemplified by an increase in detection from 0% to 89% of E. shiquicus coproDNA using necropsy-positive fox samples.

Author Notes

* Address correspondence to Belgees Boufana, Cestode Zoonoses Research Group, School of Enviroment and Life Sciences, University of Salford, United Kingdom. E-mail: B.Boufana@salford.ac.uk

Financial support: The research described here was supported by a grant (RO1 TW001565) from the National Institutes of Health and Natural Science Foundation (USA) Ecology of Infectious Diseases Program and by the Sichuan Centre for Disease Control, Chengdu, China.

Authors' addresses: Belgees Boufana and Philip Craig, Cestode Zoonoses Research Group, School of Environment and Life Sciences, University of Salford, United Kingdom, E-mails: B.Boufana@salford.ac.uk and P.S.Craig@salford.ac.uk. Gérald Umhang and Franck Boué, Anses Rabies and Wildlife Laboratory, Technopole Agricole et Veterinaire, Malzeville, France, E-mails: Gerald.UMHANG@anses.fr and Frank.BOUE@anses.fr. Jiamin Qiu and Xingwang Chen, Institute of Parasitic Diseases, Sichuan Centre for Disease Control and Prevention, Chengdu, Sichuan, China, E-mails: qiujiamin45@163.com and chenxingwang028@sohu.com. Samia Lahmar, Service de Parasitologie, Ecole Nationale de Medecine Veterinaire, Thabet, Tunisia, E-mail: drlsamia@yahoo.fr. David Jenkins, School of Animal and Veterinary Sciences, Charles Sturt University, New South Wales, Australia, E-mail: djjenkins@csu.edu.au.

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