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.

Four Tick-Borne Microorganisms and Their Prevalence in Hyalomma Ticks Collected from Livestock in United Arab Emirates

Ticks and associated tick-borne diseases in livestock remain a major threat to the health of animals and people worldwide. However, in the United Arab Emirates (UAE), very few studies have been conducted on tick-borne microorganisms thus far. The purpose of this cross-sectional DNA-based study was to assess the presence and prevalence of tick-borne Francisella sp., Rickettsia sp., and piroplasmids in ticks infesting livestock, and to estimate their infection rates. A total of 562 tick samples were collected from camels, cows, sheep, and goats in the Emirates of Abu Dhabi, Dubai, and Sharjah from 24 locations. DNA was extracted from ticks and PCR was conducted. We found that Hyalomma dromedarii ticks collected from camels had Francisella sp. (5.81%) and SFG Rickettsia (1.36%), which was 99% similar to Candidatus Rickettsia andeanae and uncultured Rickettsia sp. In addition, Hyalomma anatolicum ticks collected from cows were found to be positive for Theileria annulata (4.55%), whereas H. anatolicum collected from goats were positive for Theileria ovis (10%). The widespread abundance of Francisella of unknown pathogenicity and the presence of Rickettsia are a matter of concern. The discovery of T. ovis from relatively few samples from goats indicates the overall need for more surveillance. Increasing sampling efforts over a wider geographical range within the UAE could reveal the true extent of tick-borne diseases in livestock. Moreover, achieving successful tick-borne disease control requires more research and targeted studies evaluating the pathogenicity and infection rates of many microbial species.

Development of a Multiplex Loop-Mediated Isothermal Amplification (LAMP) Method for Simultaneous Detection of Spotted Fever Group Rickettsiae and Malaria Parasites by Dipstick DNA Chromatography

Spotted fever group (SFG) rickettsiae causes febrile illness in humans worldwide. Since SFG rickettsiosis’s clinical presentation is nonspecific, it is frequently misdiagnosed as other febrile diseases, especially malaria, and complicates proper treatment. Aiming at rapid, simple, and simultaneous detection of SFG Rickettsia spp. and Plasmodium spp., we developed a novel multiple pathogen detection system by combining a loop-mediated isothermal amplification (LAMP) method and dipstick DNA chromatography technology. Two primer sets detecting SFG Rickettsia spp. and Plasmodium spp. were mixed, and amplified products were visualized by hybridizing to dipstick DNA chromatography. The multiplex LAMP with dipstick DNA chromatography distinguished amplified Rickettsia and Plasmodium targeted genes simultaneously. The determined sensitivity using synthetic nucleotides was 1000 copies per reaction for mixed Rickettsia and Plasmodium genes. When genomic DNA from in vitro cultured organisms was used, the sensitivity was 100 and 10 genome equivalents per reaction for Rickettsia monacensis and Plasmodium falciparum, respectively. Although further improvement will be required for more sensitive detection, our developed simultaneous diagnosis technique will contribute to the differential diagnosis of undifferentiated febrile illness caused by either SFG Rickettsia spp. or Plasmodium spp. in resource-limited endemic areas. Importantly, this scheme is potentially versatile for the simultaneous detection of diverse infectious diseases.

Identification and Characterization of “Candidatus Rickettsia Thierseensis”, a Novel Spotted Fever Group Rickettsia Species Detected in Austria

Rickettsia spp. are the second most common pathogens detected in Ixodes ricinus ticks in Austria after Borrelia burgdorferi sensu lato. Species belonging to the spotted fever group (SFG) are the causative agents for tick-borne rickettsiosis across the world. So far, only four SFG Rickettsia spp. were detected in Austria, namely R. helvetica, R. raoultii, R. monacensis and R. slovaca. Here, we describe the identification of a new SFG Rickettsia species detected in an I. ricinus tick. Sequencing of various rickettsial genes revealed a nucleotide sequence similarity of 99.6%, 98.5%, 97.3% and 98.5% to the gltA, ompA, ompB, and sca4 genes, respectively, of known and validated species. Additionally, sequencing of the htrA gene and 23S-5S intergenic spacer region also only showed 99.6% and 99.2%, respectively, similarity to known species. Therefore, and in accordance with current criteria for Rickettsia species discrimination, we hereby describe a new species of the SFG with putative pathogenic potential. We propose the name “Candidatus Rickettsia thierseensis” based on the village Thiersee in the Austrian province of Tyrol, where the carrying tick was found.

Human Spotted Fever Group Rickettsia Infecting Yaks (Bos grunniens) in the Qinghai-Tibetan Plateau Area

The Qinghai-Tibetan Plateau Area (QTPA) is a plateau with the highest average altitude, located in Northwestern China. There is a risk for interspecies disease transmission, such as spotted fever rickettsioses. However, information on the molecular characteristics of the spotted fever group (SFG) Rickettsia spp. in the area is limited. This study performed screenings, and detected the DNA of human pathogen, SFG Rickettsia spp., with 11.3% (25/222) infection rates in yaks (Bos grunniens). BLASTn analysis revealed that the Rickettsia sequences obtained shared 94.3–100% identity with isolates of Rickettsia spp. from ticks in China. One Rickettsia sequence (MN536161) had 100% nucleotide identity to two R. raoultii isolates from Chinese Homo sapiens, and one isolate from Qinghai Dermacentor silvarum. Meanwhile, another Rickettsia sequence (MN536157) shared 99.1–99.5% identity to one isolate from Dermacentor spp. in China. Furthermore, the phylogenetic analysis of SFG Rickettsia spp. ompA gene revealed that these two sequences obtained from yaks in the present study grouped with the R. slovaca and R. raoultii clades with isolates identified from Dermacentor spp. and Homo sapiens. Our findings showed the first evidence of human pathogen DNA, SFG Rickettsia spp., from animals, in the QTPA.

Molecular Investigation of Tick-Borne Pathogens in Ticks removed from Tick-Bitten Humans in the Republic of Korea

Background Tick-borne infections are continuously increasing due to climate change, increased outdoor activities and increased travel between countries. This study was to investigate the presence of tick-borne pathogens in ticks removed from tick-bitten humans in southwest provinces of Republic of Korea (ROK). Methods Ticks were obtained from those tick-bitten humans between May 2014 and September 2017 in Jeollanam provinces and Gwangju metropolitan city in ROK. The presence of the tick-borne pathogens in ticks removed from tick-bitten humans was analyzed using pathogen-specific polymerase chain reaction (PCR). Results We identified 33 ticks from three tick species, namely Amblyomma testudinarium (60.6%), Haemaphysalis longicornis (27.3%), and Ixodes nipponensis (12.1%) in order of occurrence by morphology and 16S rDNA-targeting PCR. Tick-borne pathogens were found in 16 ticks using pathogen-specific PCR. From the results, 12 ticks (36.4%) tested positive for spotted fever group (SFG) Rickettsia: Rickettsia monacensis (1/12), R. tamurae (8/12), and Candidatus Rickettsia jingxinensis (3/12). Three ticks (9.1%) were positive for Anaplasma phagocytophilum . In addition, three ticks (9.1%) tested positive for Babesia gibsoni (1/3) and B. microti (2/3). Conclusions In conclusion, we identified three tick species; the most common species was A. testudinarium followed by H. longicornis and I. nipponensis . SFG Rickettsia , A. phagocytophilum , and Babesia spp. were the most frequently detected pathogens in ticks removed from tick-bitten humans. R. tamurae and Ca. R. jingxinensis were firstly detected in Korea. The present results will contribute to the understanding of tick-borne infections in animals and humans in the ROK.

Diversity of Rickettsia in Ticks Collected in Abruzzi and Molise Regions (Central Italy)

Rickettsiae have worldwide occurrence and rickettsiosis are widely recognized as emerging infections in several parts of the world. For decades, it was thought that a single pathogenic tick-borne spotted fever group (SFG), Rickettsia, occurred in each continent. Nowadays, thanks to molecular biology, new species of Rickettsia responsible for disease in humans are continuously identified worldwide. In a framework of diagnostic activities of the Istituto Zooprofilattico Sperimentale dell’Abruzzo e del Molise “G. Gaporale” and considering some reports of suspected human clinical cases of rickettsiosis, a survey on ticks collected form animals and humans was carried out with the aim to identify the Rickettsia species circulating in Abruzzi and Molise regions. A total of 603 ticks, previously identified at species level by morphology, pooled into 178 tick samples, were tested by pan-Rickettsia RealTime PCR. DNA from specimens positive for Rickettsia spp. was then sequenced in order to identify the Rickettsia species involved. The highest infection rate was detected in Dermacentor marginatus followed by Ixodes ricinus. The selected targets for this purpose were OmpA and gltA. Rickettsia slovaca, Rickettsia monacensis, Rickettsia massiliae, Rickettsia conorii, Rickettsia aeschlimannii, Rickettsia helvetica, Rickettsia raoultii, and Rickettsia felis – like organisms were identified in this study. These are the first data available in the literature for the circulation of SFG Rickettsia species in the selected geographical area. Results made evidence of high rate of infection in ticks. All Rickettsia species detected have been previously involved in human infection. The diversity of Rickettsia detected, and tick species collected reflects the biodiversity in term of wildlife and environment of the area. An association between Rickettsia species, ticks, and the relationships with vertebrate host species are discussed. Due to the peculiar eco-biology of each Rickettsia species, the use of diagnostic tools able to identify Rickettsia at the species level is thus recommended in order to assess the risk for humans and to elucidate more precise etiological diagnosis in clinical cases.

RETROSPECTIVE SEROLOGICAL DIAGNOSTICS OF SPOTTED FEVER GROUP RIKKETSIOSES IN HIGH RISK AREAS OF RICKETTSIA CONORII SUBSP. CASPIA INFECTION

723 blood sera from 537 patients of Regional Infectious Clinical Hospital, Astrakhan were obtained during high activity period of Rhipicephalus ticks (May-September 2015) and retrospectively studied for IgG/IgM to antigen of spotted fever group (SFG) Rickettsia. IgG and/or IgM to Rickettsia conorii were detected in 145 sera from 130 patients, and antibodies to R. sibirica (group-specific) were detected in 143 sera from 145. Antibodies to R. conorii were detected for 71,4% patients with Astrakhan spotted fever (ASF), for 28,4% patients with acute respiratory viral infection, for 19,1% patients with infection of unspecified etiology and for 40% patients having symptoms of a adenovirus infection. Acute rickettsiosis, provably ASF, is serologically validated for 71 patients. Dynamic of IgM/IgG to R. conorii in sera of patients having different preliminary diagnoses is discussed. IgM to R. conorii in sera of patients having adenovirus infection symptoms were detected at a later time as compared with others. For regions of high risk of R. conorii subsp. caspia infection the differentiation of diagnostic and anamnestic specific antibodies is very important. The absence of serological and molecular biological markers in third of patients with ASF symptoms is necessary to study. Preparations and algorithms for diagnosis of SFG rickettsioses are needed to improve.

Serological Evidence of Anaplasma phagocytophilum and Spotted Fever Group Rickettsia spp. Exposure in Horses from Central Italy

Anaplasma phagocytophilum and Rickettsia spp. are tick-borne bacteria of veterinary and human concern. In view of the One-Health concept, the present study wanted to evaluate the spreading of these pathogens in horses living in central Italy. In particular, the aim of the investigation was to verify the exposure to A. phagocytophilum in order to update the prevalence of this pathogen in the equine population from this area, and to spotted fever group (SFG) Rickettsia spp. to evaluate a possible role of horses in the epidemiology of rickettsiosis. Indirect immunofluorescent assay was carried out to detect antibodies against A. phagocytophilum and SFG (spotted fever group) Rickettsia spp. in blood serum samples collected from 479 grazing horses living in central Italy during the period from 2013 to 2018. One hundred and nine (22.75%) horses were positive for A. phagocytophilum, 72 (15.03%) for SFG Rickettsia spp., and 19 (3.96%) for both antigens. The obtained results confirm the occurrence of A. phagocytophilum in equine populations, and also suggest the involvement of horses in the epidemiology of SFG rickettsiosis. In both cases, in view of the zoonotic aspect of these pathogens and the frequent contact between horses and humans, the monitoring of equine populations could be useful for indication about the spreading of the tick-borne pathogens in a certain geographic area.

Algorithms of Rickettsiae’s Detection and Laboratory Diagnosis of Rickettsioses of Spotted Fever Group in Russia

An analysis of modern trends of Rickettsiae’s detection and laboratory diagnostics for spotted fever group rickettsioses is present. Due to the sharp decline in the range of manufactured products and increase the spectrum of rickettsial species identified in Russia, new approaches required to laboratory verification of diagnoses. IFA and ELISA with antigens of relevant species of Rickettsia can be recommended to detect antibodies to the SFG rickettsiae. PCR-restriction analysis and sequencing of PCR products most appropriate for the detection and identification of SFG Rickettsia, biological methods are necessary for studying of pathogenic species of Rickettsia.