Sero-epidemiology and molecular characterization of Rickettsiae infecting humans, selected animals and arthropod vectors in Asembo, western Kenya, 2007-2010

Abstract

Rickettsiae are gram negative obligate intracellular bacteria in the class α- proteobacteria. They are transmitted by arthropod vectors, including fleas, ticks, mites, andbody lice. The prevalence of rickettsial infections in humans, animals and arthropod vectors in Kenya is undefined.Thisstudy was conducted to investigate rickettsial infections in western Kenya. To assess previous exposure to Rickettsia, a randomly selected subset of sera from patients attending Lwak Hospital in western Kenya in 2007-8 were tested for IgG antibodies against Rickettsia(n=359) with indirect fluorescence antibody (IFA) test. To detect acute rickettsial infections, blood specimens from patients with acute febrile illness (n=699) and from asymptomatic individuals (n=236) were tested using four monoplex qPCR assays that target gene fragments of gltA,17-kDa and ompB. Buffy coats,spleen specimens collected from peri-domestic small mammals, ticks and fleas were tested for Rickettsia using gltAqPCR.Selected positive samples were further characterized by sequencing genes encoding the 17-kDa, ompA, ompB and R. felis plasmids.Further testing of the unique Rickettsia species was done using multi-locus sequence typing of rrs, gltA, ompA, sca4 and ompB genes. Immunoglobulin G against all rickettsiae were detected in 57.4% of the patients (n = 357), including 56% positive for spotted fever group (SFG)Rickettsiae and and 14.5% positive for typhus group (TG) xxii rickettsiae.Seropositivity for SFG antibodies increased significantly with age (p <0.001). Febrile patients had 2.2 times higher odds of testing positive for rickettsial infection by PCR than asymptomatic individuals (7.15% vs 3.39%, odds ratio 2.20, CI; 1.03-4.70). From animals in the same region, Rickettsia DNA were detected by PCR in 3.68% (n=299) of the dogs and 7.69% (n=26) of the cats. No rickettsia was detected in cattle, sheep and goats. Partial sequences from 17-kDa gene showed 97% homology with specimens from cats, dogs and R. felis URRWXCal2. Of the ticks obtained from cattle and dogs, 96.9% (n=162 pools) and 20.34% (n=59) were positive for Rickettsia, respectively. Nucleotide sequences of ompA and ompB gene showed ≥ 98% similarity with tick specimens and Rickettsia africae. Of the flea specimens tested, 59.7% (n=134) were positive for Rickettsiae. Partial sequences of human isolates and one flea specimen were 100% homologous to Rickettsia felis, while11 other fleas were ≤93% homologous to R. felis. MLST of the 6/11 specimens determined the flea specimens to be unique and genetically similar to Rickettsia RF2125, a previously-described rickettsial agent of fleas and mites.This study provides evidence of R. felis-associated illness in febrile patients in western Kenya and demonstrates at least one novel rickettsial agent“CandidatusRickettsia asemboensis”. Further work is needed to elucidate the biological characteristics and pathogenecity of the novel Rickettsia species.