Identification and Genetic Diversity Analysis of Rickettsia in Dermacentor Nuttalli Within Inner Mongolia, China

Background The pathogen genus Rickettsia contains the linages spotted fever group, typhus group, transitional group, and the ancestral group, of which the spotted fever group Rickettsia (SFGR) is transmitted by ticks. Dermacentor nuttalli is considered the main vector carrying SFGR. Studying the genetic diversity and population structure of Rickettsia is essential for developing effective control strategies and predicting evolutionary trends of the pathogens. Methods We collected 408 Dermacentor nuttalli in the Inner Mongolia Autonomous region in 2019, detected Rickettsia infection, and characterized the haplotypes. The extracted Rickettsia DNA of the gltA and ompA genes were amplified and sequenced. Result In this study, 10 haplotypes of the gltA gene and 22 haplotypes of the ompA gene were obtained. In the two resulting phylogenetic trees, the haplotypes G1-G7 and G9 of the gltA gene clustered with Rickettsia raoultii, while G8 and G10 clustered with Rickettsia sibirica. Haplotypes O1-O15, O18 and O20-O22 of the ompA gene clustered with Rickettsia raoultii, while O16 and O19 clustered with Rickettsia sibirica. The average haplotype diversity was 0.3 for gltA and 0.7 for ompA, while the average nucleotide diversity was greater than 0.05. Neutrality tests were insignificant for Tajima’s D results and Fu’s Fs results. The fixation index values (FST) showed that the degree of genetic differentiation between most sampled populations was small (FST<0.05), while others were medium (FST>0.05) and large (FST>0.15). Analysis of molecular variance (AMOVA) revealed that the variation within populations was greater than that between populations. The mismatch analysis of Rickettsia showed double peaks. Conclusion We found two genotypes of Rickettsia: Rickettsia raoultii and Rickettsia sibirica. The high genetic diversity of Rickettsia allows for easy adaption to different environments; furthermore, genetic differentiation between populations is small and Rickettsia populations do not show a pedigree geographical structure. The high rates of retention and infestation of Rickettsia in Dermacentor nuttalli together with the animal husbandry exchange in China gradually lead to the genetic characteristics of Rickettsia harmonizing across various regions. Overall, the significant genetic diversity and geographic structure of Rickettsia in Dermacentor nuttalli are critical for SFGR control.

CLINICAL AND LABORATORY CHARACTERISTICS OF TICK-BORNED RICKETTSIOSIS RELATED TO RICKETTSIA SIBIRICA AND "CANDIDATUS RICKETTSIA TARASEVICHIAE"

The Siberian tick-borne typhus (STT) is the most common tick-borne rickettsiosis (TBR) in Russia, registered in 17 administrative territories of the Southern Siberia and the Far East. The aim of this study was to describe clinical picture, pathological data and the results of laboratory diagnostics during fatal mixed infection caused by two rickettsia species most common in Russia - pathogenic Rickettsia sibirica and poorly examined Candidatus R. tarasevichiae, including identification of its etiological agents.A four-year-old girl in the Krasnoyarsk Territory in a hyperendemic focus of the Siberian tick-borne typhus after tick sucking revealed typical TBR symptoms (scab at the site of tick suction, fever, spotted rash, myalgia) and meningeal syndrome, which is not typical for the Siberian tick-borne typhus. The child died on the seventh day of the illness. Autopsy data (hepatosplenomegaly; cerebral edema, which was the immediate cause of death) and the results of histological examination (productive vasculitis of the brain, spinal cord and skin, polymorphic cell perivascular infiltrates in the liver and lungs, serous meningitis, myeloid hyperplasia of the spleen and lymph nodes, interstitial lymphoid infiltration in the myocardium) confirmed the clinical diagnosis of tick-borne rickettsiosis.The patient's blood and brain samples were tested for a wide range of tick-borne pathogens and enteric viruses that cause brain damage using PCR followed by sequencing of the positive samples.The DNA of Rickettsia sibirica and "Candidatus Rickettsia tarasevichiae" was found in both blood and brain samples. R. sibirica was identified by the nucleotide sequences of gene fragments gltA, OmpA and ompB, and "Candidatus R. tarasevichiae" - the gltA and ompB genes using nested PCR and sequencing. All amplified fragments were sequenced in both directions; the obtained sequences were deposited in the GenBank database under the inventory numbers: MK048467-MK048475. We have not identified other tick-borne pathogens or intestinal viruses in the patient samples able to result in meningeal syndrome.The area where the child was sucked by the tick belongs to the regions with a high incidence of STT. Several species of mites coexist in this area; of these, Haemaphisalis concinna, the carrier of R. sibirica, dominates the populations of Ixodes mites, while Ixodes persulcatus, the main reservoir of "Candidatus R. tarasevichiae", is less common.As a result of the conducted studies, for the first time in the Russian Federation, a verified case of a lethal infection associated with two species of rickettsiae - Rickettsia sibirica and "Candidatus R. tarasevichiae" was identified and described.

Rickettsiae in red fox (Vulpes vulpes), marbled polecat (Vormela peregusna) and their ticks in northwestern China

Background Previously, twelve Rickettsia species were identified in ticks, fleas, sheep keds (Melophagus ovinus), bats (Pipistrellus pipistrellus) and a tick-bitten patient in the Xinjiang Uygur Autonomous Region (XUAR) in northwestern China. Here we aimed to molecularly detect rickettsial agents in red fox (Vulpes vulpes), marbled polecat (Vormela peregusna) and their ticks. Methods During 2018–2019, 12 red foxes, one marbled polecat and their ticks were sampled in two counties and a city of the XUAR. The heart, liver, spleen, lung and kidney of these 13 carnivores were dissected, followed by DNA extraction. Hard ticks were identified both morphologically and molecularly. All samples were examined for the presence of rickettsiae by amplifying four genetic markers (17-kDa, gltA, ompA, sca1). Results A total of 26 adult ticks and 28 nymphs (38 Ixodes canisuga, nine Ixodes kaiseri, six Haemaphysalis erinacei and one Dermacentor marginatus) were collected from red foxes, and four Ha. erinacei ticks were removed from the marbled polecat. Analysis of cytochrome c oxidase subunit I (COI) gene sequences indicated that 2–32 nucleotides differed between I. canisuga, I. kaiseri and Ha. erinacei from northwestern China and Europe. Rickettsia raoultii was detected in three red foxes, Candidatus Rickettsia barbariae in a red fox, Rickettsia sibirica in a red fox and a marbled polecat, and R. raoultii in two tick species (I. canisuga and D. marginatus). Conclusions To the best of our knowledge, I. canisuga and I. kaiseri have not been previously reported from red foxes in China. The DNA of R. sibirica and R. raoultii was detected for the first time in the organs of red foxes, and R. sibirica in the organs of a marbled polecat. This is also the first molecular evidence for the presence of R. raoultii in I. canisuga. Our findings expand the range of tick-borne pathogens in wildlife species and associated ticks in China.

Rickettsiae in Red Fox (Vulpes Vulpes), Marbled Polecat (Vormela Peregusna) and Their Ticks in Northwestern China

BackgroundPreviously, twelve Rickettsia species were found in ticks, fleas, sheep keds (Melophagus ovinus), bats (common pipistrelle: Pipistrellus pipistrellus) and a tick-bitten patient in Xinjiang Uygur Autonomous Region (XUAR), northwestern China. Here we aimed to molecularly detect rickettsial agents in red fox (Vulpes vulpes), marbled polecat (Vormela peregusna) and their ticks.MethodsDuring 2018-2019, 12 red foxes, 1 marbled polecat and their ticks were sampled in two counties and a city of Xinjiang Uygur Autonomous Region (northwestern China). The heart, liver, spleen, lung and kidney of these 13 carnivores were dissected, followed by DNA extraction. Hard ticks were identified both morphologically and molecularly. All samples were examined for the presence of rickettsiae by amplifying four genetic markers.ResultsA total of 26 adult ticks and 28 nymphs (38 Ixodes canisuga, nine Ixodes kaiseri, six Haemaphysalis erinacei and one Dermacentor marginatus) were collected from red foxes, and four H. erinacei ticks were removed from a marbled polecat. Analysis of cytochrome c oxidase subunit I (COI) gene sequences indicated that 2-32 nucleotides differed between I. canisuga, I. kaiseri and H. erinacei from northwestern China and Europe. Rickettsia raoultii was detected in three red foxes, Candidatus Rickettsia barbariae in a red fox, Rickettsia sibirica in a red fox and a marbled polecat, and R. raoultii in two tick species (I. canisuga and D. marginatus).ConclusionsTo the best of our knowledge, I. canisuga and I. kaiseri have not been previously reported from red foxes in China. The DNA of R. sibirica and R. raoultii was detected for the first time in organs of red foxes, and R. sibirica in organs of marbled polecat. This is also the first molecular evidence for the presence of R. raoultii in I. canisuga. Our findings add to the range of tick-borne pathogens in wildlife species and associated ticks in China.