One Health Approach to Rickettsiosis: A Five-Year Study on Spotted Fever Group Rickettsiae in Ticks Collected from Humans, Animals and Environment

The spotted fever group of Rickettsiae is a heterogeneous group of Rickettsiae transmitted by ticks, causing similar diseases in humans (spotted fever). Until recently, it was supposed that a single pathogenic tick-borne SFG Rickettsia circulated in each different geographic area and that R. conorii subsp. conorii was the SFG Rickettsiae circulating in Italy, but in the last decade, thanks to molecular diagnostic, several different Rickettsia species, previously not considered pathogenic for decades, have been isolated from ticks and definitively associated to human disease, also in Italy. The present survey was carried out with the aim of investigating the presence of different SFG Rickettsia species in a geographic area where no information was available. Ticks collected from animals submitted to necropsy, removed from humans in local hospitals and collected from the environment were identified and tested by PCR for Rickettsia spp. based on the gltA gene, and positive PCR products were sequenced. A total of 3286 ticks were collected. Fifteen tick species were recognized, the most represented (79.52%) species in the collection was Ixodes ricinus, followed by Rhipicephalus sanguineus (9.13%). The overall prevalence of Rickettsia infection was 7.58%. Eight species of Rickettsia were identified, the most frequent was R. monacensis (56%), followed by R. helvetica (25.50%). Noteworthy, is the detection in the present study of Rrhipicephali, detected only twice in Italy. These are the first data available on SFG Rickettsiae circulation in the study area and they can be considered as starting point to assess the possible risk for humans.

Molecular Detection of Bartonella Species in Rodents Residing in Urban and Suburban Areas of Central Thailand

Bartonella spp. are Gram-negative zoonotic bacteria transmitted to humans via various blood-sucking arthropods. Rodents have been identified as reservoir hosts of several zoonotic pathogens, including Bartonella spp. In Thailand, studies of Bartonella spp. in rodents from urban areas are limited; thus, a study in this area is necessary. The objectives of this study were to detect Bartonella spp. in rodents in Thailand and to compare the species’ distribution across different areas. In total, 70 blood samples from rodents in urban and suburban areas were tested for Bartonella spp. using a conventional polymerase chain reaction that targeted the citrate synthase (gltA) gene. All Bartonella-positive sequences were analyzed using polymorphism in order to build a phylogenetic tree. Approximately 38% of the rodents studied contained Bartonella DNA. Both Rattus exulans (Pacific rat) and R. tanezumi (Asian house rat) contained Bartonella spp. Four species of Bartonella were detected in blood samples: B. tribocorum, B. phoceensis, B. grahamii, and B. rattimassiliensis. In addition, eight Pacific rats contained the B. kosoyi–B. tribocorum complex. Bartonella phoceensis and B. tribocorum–B. kosoyi complexes were found in a specific habitat (p < 0.05). Interestingly, only seven haplotypes were identified in the sequences analyzed, and only haplotype A was found in both rodent species. Finally, a monitoring program for zoonotic Bartonella infection, especially the B. kosoyi–B. tribocorum complex, B. phoceensis, B. grahamii, and B. rattimassiliensis should be established, especially in high-risk areas.

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.

Molecular Survey of Anaplasmataceae Agents and Coxiellaceae in Non-Hematophagous Bats and Associated Ectoparasites from Brazil

The Anaplasmataceae family (order Rickettsiales) encompasses obligately intracellular bacteria of the genera Anaplasma, Ehrlichia, and Neorickettsia. Together with Coxiella burnetii (Coxiellaceae family, order Legionellales), these bacteria represent important causative agents of diseases in humans and animals. The scarcity of studies that investigated the occurrence of these agents in bats and their associated ectoparasites, emphasizes the need to achieve a better understanding of the role of these animals in the maintenance of such bacteria. Herein, 418 samples (133 blood, 135 spleen, and 150 ectoparasites) are collected from 135 non-hematophagous bats belonging to 12 species in a periurban area of Campo Grande city, Mato Grosso do Sul state, midwestern Brazil. In the results, 1.65% (7/418), 12.04% (50/418), and 13.63% (57/418) of samples are positive in PCR assays for Anaplasma spp. (16S rRNA gene), Ehrlichia spp. (dsb gene), and Neorickettsia spp. (16S rRNA gene), respectively. Anaplasma spp. and Neorickettsia spp. are detected in one (5.26%) Ornithodoros hasei tick larva. Ehrlichia spp. is detected in 14% of bat flies (represented by Megistopoda aranea, Trichobius costalimai, and Strebla hertigi), 6% of tick larvae (O. hasei), 12% of Spinturnicidae mites (represented by Periglischrus sp., P. torrealbai, and P. acutisternus), and 38% of Macronyssidae mites (Steatonyssuss sp.). The obtained sequences are observed to be similar to Anaplasma phagocytophilum (97.42–97.6% identified), Ehrlichia minasensis (96.73–100% identified), Neorickettsia risticii (96.7–100% identified), and Neorickettsia findlayensis (95.07–100% identified) by BLASTn analyses, and closely related to Ehrlichia ruminantium by phylogenetic analyses based on the gltA gene. No bat samples (blood/spleen) are positive in the qPCR assay for C. burnetii based on the IS1111 gene. The present work shows, for the first time, the occurrence of Anaplasmataceae in bats and associated ectoparasites (ticks, mites, and bat flies) from Brazil.

Ixodes tropicalis (Acari: Ixodidae) infesting a human and molecular detection of Rickettsia bellii, Colombia

Introduction: Ixodes tropicalis is a little-known tick species reported parasitizing wild rodents only in Colombia and Perú.Objective: To report a case of I. tropicalis infesting a human in the south of the metropolitan area of the Valle de Aburrá, Antioquia, Colombia, and to report the molecular detection of Rickettsia bellii in this species.Materials and methods: The tick was identified using a morphological key and sequencing of tick mitochondrial 16S rRNA. Additionally, bacterial and protozoa pathogens were evaluated using PCR for the detection of Rickettsia spp., family Anaplasmataceae, Borrelia spp., and piroplasmid.Results: We identified the tick as an I. tropicalis female according to Kohls, 1956, description and to partial 16S rRNA sequences showing a minimum of 5% divergencies compared to Ixodes sequences. We also detected the gltA gene of R. bellii in the tick with 99.87% of identity.Conclusion: This is the first report in Colombia of a species of the Ixodes genus parasitizing a human and the first report of the detection of R. bellii in this tick species.

Molecular Detection and Genetic Identification of Rickettsia Infection in Ixodes granulatus Ticks, an Incriminated Vector for Geographical Transmission in Taiwan

Tick-borne Rickettsia pathogens have become an emerging source of zoonotic infections and have a major impact on human health worldwide. In this study, the prevalence and genetic identity of Rickettsia infections in Ixodes granulatus ticks was firstly determined in Kinmen Island of Taiwan. A total of 247 I. granulatus ticks were examined for Rickettsia infection by nested-PCR assay targeting the citrate synthase (gltA) gene of Rickettsia. The Rickettsia infection was detected with a general infection rate of 4.86%, and was detected in nymph, male and female stages with an infection rate of 3.81%, 0% and 6.84%, respectively. Phylogenetic relationships were analyzed by comparing the gltA sequences obtained from four Taiwan strains and 19 other strains representing 13 genospecies of Rickettsia. Phylogenetic analyses reveal that all Taiwan strains were genetically affiliated to the genospecies of spotted fever (R. parkeri) and transitional (R. felis) groups of Rickettsia. Our findings reveal the first detection of R. parkeri-like and R. felis in I. granulatus ticks from Kinmen Island. As a tourist island between Taiwan and mainland China, these results demonstrate the epidemiological significance of diverse Rickettsia species existed in I. granulatus ticks and highlight the potential threat of geographical transmission among humans in the Taiwan area.

Serological Evidence of Exposure to Spotted Fever Group and Typhus Group Rickettsiae in Australian Wildlife Rehabilitators

Rickettsioses are arthropod-borne zoonotic diseases, several of which occur in Australia. This study aimed to assess the exposure levels and risk factors for Rickettsia spp. among Australian wildlife rehabilitators (AWRs) using serology, PCR and a questionnaire. Antibody titres against Spotted Fever Group (SFG), Typhus Group (TG) and Scrub Typhus Group (STG) antigens were determined using an immunofluorescence assay. PCR targeting the gltA gene was performed on DNA extracts from whole blood and serum. Logistic regression was used to identify risk factors associated with seropositivity. Of the 27 (22.1%; 27/122) seropositive participants all were seropositive for SFG, with 5/27 (4.1%) also positive for TG. Of the 27 positive sera, 14.8% (4/27) were further classified as exposure to R. australis, 3.7% (1/27) to R. honei, 3.7% (1/27) to R. felis and 77.8% (21/27) were classified as ‘indeterminate’—most of which (85.7%; 18/21) were indeterminate R. australis/R. honei exposures. Rickettsia DNA was not detected in whole blood or serum. Rehabilitators were more likely to be seropositive if more than one household member rehabilitated wildlife, were older than 50 years or had occupational animal contact. These findings suggest that AWRs are at increased risk of contracting Rickettsia-related illnesses, however the source of the increased seropositivity remains unclear.

Molecular Prevalence and Identification of Ehrlichia canis and Anaplasma platys from Dogs in Nay Pyi Taw Area, Myanmar

Ticks are vectors of different types of viruses, protozoans, and other microorganisms, which include Gram-negative prokaryotes of the genera Rickettsiales, Ehrlichia, Anaplasma, and Borrelia. Canine monocytic ehrlichiosis caused by Ehrlichia canis and canine cyclic thrombocytopenia caused by Anaplasma platys are of veterinary importance worldwide. In Myanmar, there is limited information concerning tick-borne pathogens, Ehrlichia and Anaplasma spp., as well as genetic characterization of these species. We performed nested PCR for the gltA gene of the genus Ehrlichia spp. and the 16S rRNA gene of the genus Anaplasma spp. with blood samples from 400 apparently healthy dogs in Nay Pyi Taw area. These amplicon sequences were compared with other sequences from GenBank. Among the 400 blood samples from dogs, 3 (0.75%) were positive for E. canis and 1 (0.25%) was positive for A. platys. The partial sequences of the E. canis gltA and A. platys 16SrRNA genes obtained were highly similar to E. canis and A. platys isolated from different other countries.

Bartonella spp. in Small Mammals and Their Fleas in Differently Structured Habitats From Germany

Most Bartonella spp. are transmitted by fleas and harbored by small mammals which serve as reservoirs. However, little is known about the composition of fleas and their Bartonella spp. from small mammals in Central Europe. Therefore, the aims of this study were to investigate flea communities on small mammals from three differently structured sites (urban, sylvatic, renatured) in Germany as well as the prevalence of Bartonella spp. in small mammals and their parasitizing fleas. In total, 623 small mammals belonging to 10 different species (the majority were Myodes glareolus and Apodemus flavicollis) were available. Fleas were removed from the small mammals' fur, morphologically identified and DNA was extracted. To detect Bartonella spp., two conventional PCRs targeting the gltA gene and the 16S−23S rRNA intergenic spacer were carried out followed by sequencing. Obtained sequences were compared to those in GenBank. In total, 1,156 fleas were collected from 456 small mammals. Altogether, 12 different flea species (the majority were Ctenophthalmus agyrtes, Nosopsyllus fasciatus, and Megabothris turbidus) were detected. At the urban site mostly Leptopsylla segnis and N. fasciatus were collected which may be vectors of zoonotic pathogens to companion animals. The overall prevalence for Bartonella in small mammals was 43.3% and in fleas 49.1%. Five different Bartonella spp. were detected in small mammals namely B. grahamii, B. taylorii, B. doshiae, Bartonella sp. N40 and uncultured Bartonella sp. whereas in fleas four Bartonella spp. were found which were with the exception of B. doshiae identical to the Bartonella species detected in their small mammal hosts. While B. grahamii was the only zoonotic Bartonella sp. most Bartonella strains found in fleas and small mammals belonged to uncultured Bartonella spp. with unknown zoonotic potential. This study showed a high diversity of flea species on small mammals from Germany. Further, high prevalence rates of Bartonella species were detected both in fleas and in their mammalian hosts. Several different Bartonella species with a high genetic variability were discovered. Especially at the urban study sites, this may pose a risk for Bartonella transmission to companion animals and humans.

Short Communication: Molecular characterization and blood hematology profile of dogs infected by Ehrlichia canis in Yogyakarta, Indonesia

Wuhan YOP, Haryanto A, Tjahajati I. 2020. Short Communication: Molecular characterization and blood hematology profile of dogs infected by Ehrlichia canis in Yogyakarta, Indonesia. Biodiversitas 21: 3242-3248. Ehrlichia canis is Gram-negative intracellular obligate bacteria that cause ehrlichiosis, a companion vector-borne disease is a potentially fatal disease that attacks dogs. The purpose of this study was to molecular characterize and determine the features of Ehrlichia-infected blood based on the amplification of the gltA gene in Ehrlichia infected dogs from Yogyakarta, Indonesia. Blood samples were collected from 51 dog patients from the Prof. Dr. Soeparwi Animal Hospital, animal clinics, and pet shops based on the anamnesis, clinical sign, and physical examination, followed by microscopic examination, routine hematology, PCR amplification, and DNA sequencing were carried out on the blood samples. Based on positive PCR amplification and blood hematology profile examination ehrlichiosis-positive in dogs showed that thrombocytopenia case was 82.3%, anemia was 70.5%, eosinopenia was 70.5%, neutropenia was 29.4%, monocytopenia was 23%, leukopenia was 17% and lymphopenia was 11.7%. Morulae of Ehrlichia sp.was not found in microscopic examination. Molecularly, detected of E. canis using the gltA gene showed that 34% of samples were positive results. Then 5 of positive Ehrlichia samples were DNA sequenced, they showed a high homology of 100% with Hat Yai isolates (KU765199.1). There was no genetic diversity between E. canis samples in Yogyakarta.