Distribution and Genetic Diversity of Cystic Echinococcosis in Busia and Bungoma Counties, Western Kenya

Abstract

Cystic Echinococcosis (CE) is a zoonosis caused by infection with the larval stages of the taeniid cestodes of the species complex Echinococcus granulosus sensu lato. It is prevalent among transhumant communities in East Africa, including those residing in northern Kenya. The distribution of the disease is enhanced by several factors, including livestock trade, which may facilitate the spread of CE to non-endemic areas such as Western Kenya. Poor condemnation of offal at the slaughterhouses potentially creates an indirect risk of disease spill-over to humans through dogs. This study: assessed the possible establishment of the CE life cycle by screening for the presence of human CE using ultrasound, examined dogs’ infection levels with taeniid eggs and their potential role in contaminating the environment with intestinal parasites. The study also described intraspecific variance of E. granulosus s. s. and the parasites’ haplotypes in livestock slaughtered in Western Kenya and conducted economic losses analysis through condemned offal. Eight sentinel sites in Bungoma County were purposively selected and a portable ultrasound scanner was used to screen for the presence of human CE. In the livestock package, a cross-sectional survey was conducted at slaughterhouses in Bungoma and Busia counties. A post-mortem visit was conducted on livestock sampled at the slaughterhouse, to screen carcasses for CE infection and the organ(s) infected. The DNA was extracted from the protoscoleces and nested PCR performed targeting NADH dehydrogenase subunit I (NAD1) and Cytochrome c oxidase I genes followed by sequencing. In the dog work package, sixteen ruminant slaughterhouses were selected in Busia and Bungoma Counties, and around each slaughterhouse ten homesteads owning free-roaming dogs were identified. A questionnaire was administered on dog management practices to the homestead owner and a faecal sample was collected from the dog’s rectum. In Busia County the sampled dog were collared with a GPS tracker to assess their movement patterns. The faecal samples were examined microscopically following the zinc-chloride sieving-floatation technique for the presence of taeniid eggs and other canine intestinal parasites. Polymerase Chain Reaction – Restriction Fragment Length Polymorphism of NADH dehydrogenase subunit 1 gene and sequencing were used to confirm taeniid eggs identified during microscopy. Additionally, the Coproantigen-ELISA was used to detect the presence of taeniid antigen in a sub-set of the faecal samples. Economic losses were established by assessing the losses incurred from condemned offal at the slaughterhouses. In humans, 1002 participants were screened and sixty-seven participants (6.7%) had abnormal ultrasound findings and, of these, 7 (1.1%) had simple liver cysts/CL, as per WHO classification. In Livestock, a total of 153 E. granulosus s. s. cysts were sampled from livestock. In total, 135/153 cysts were genotyped as E. granulosus s. s. based on Restriction fragment length polymorphism (RFLP) of the Nad1 gene. Upon sequencing 11 nad1 and 19 cox1 haplotypes were identified in 120 and 122 sequences, respectively. This work serves as the first report of E. granulosus s. s. haplotypes in East Africa and will enhance the understanding of the genetic variability of the cestode sub-region. In dogs, helminths were detected in the 155 dogs sampled. They included hookworms (n=92; 59.4%), ascarids (n=15; 9.7%), and taeniids (n=1; 0.6%). Through Copro-PCR, 13 eggs extracted from the sample of the only taeniid infected dog were sequenced and identified as E. canadensis (G6/7) [n=1], Taenia multiceps [n=1], and Taenia serialis [n=6]; the remaining were indeterminate. Of the 77 faecal samples tested for E. granulosus sensu lato (s. l.) with the Copro-ELISA, 64 (83.1%) were negative, 12 (15.6%) were positive, while 1 (1.3%) was suspicious. The human data results contribute to CE baseline data while providing insights on the implementation of ultrasound diagnosis in the field, as recommended by the WHO for targeted control of echinococcosis by 2030. The livestock data results serve as first report of E. granulosus s. s. haplotypes in East Africa and will enhance the understanding of the genetic variability of the cestode in the region. The dog data results indicate a relatively high carriage of zoonotic parasites by free-roaming domestic dogs in Western Kenya, which poses a risk to human and livestock populations. This study report for the first time a domestic lifecycle of Echinococcus canadensis and Taenia multiceps in Western Kenya, as well as a presumptive sylvatic cycle of coenurosis by T. serialis. The human study recommends routine screening of the population for CE in the health units as well as deployment of Ultrasound as a diagnostic tool to aid epidemiological investigations. The livestock study recommends sustained assessment of the significance of haplotypes variation in the control of CE in the region. Whereas, the measures to control infection by the parasite will require more understanding of its genetic diversity and how it affects transmission and pathogenicity. The dog study recommends development of an incountry Copro-antigen capacity to aid in surveillance programmes, a broader assessment of dog parasites with zoonotic potential, adherence to slaughterhouse management practices, and dog-ownership programmes to highlight the importance of deworming and restricted dog movements.