A novel Rickettsia, Candidatus Rickettsia takensis, and the first record of Candidatus Rickettsia laoensis in Dermacentor from Northwestern Thailand.

Three hundred and forty-four tick samples were collected from vegetation at Taksin Maharat National Park, Tak province, northwestern Thailand. They were morphologically identified and molecularly confirmed by 16S rRNA and COI genes as Dermacentor laothaiensis (n = 105), D. steini (n = 139), and D. auratus (n = 100). These ticks were examined for the spotted fever group rickettsiae (SFGRs) using PCR and DNA sequencing of six genes; 17-kDa, gltA, 16S rRNA, ompA, ompB, and sca4. Of these ticks, 6.10% (21/344) gave positive results for the presence of SFGRs. Phylogenetic analyses of the SFGRs clearly indicated that a novel genotype assigned as Candidatus Rickettsia takensis was detected in D. laothaiensis (19/105) and at lesser frequency in D. steini (1/139). Furthermore, Candidatus Rickettsia laoensis was also found at a low frequency in D. auratus (1/100), the first record in Thailand. Although, the pathogenicities of these SFGRs remain unknown, our findings suggest potential risks of SFGRs being transmitted via ticks near the border between Thailand and Myanmar, a gateway of daily migrations of local people and visitors both legal and illegal.

Spotted fever group Rickettsia, Anaplasma and Coxiella-like endosymbiont in Haemaphysalis ticks from mammals in Thailand.

Ticks are ectoparasites of vertebrates and vectors of various pathogenic microorganisms. In this study, the presence of bacteria and protozoa was evaluated by PCR and DNA sequencing in 233 mammal ticks collected from 8 provinces in Thailand. Sequence and phylogenetic analyses of partial rickettsial ompA, ompB, sca4 and partial Coxiella 16S rRNA, GroEL, rpoB genes clearly revealed, for the first time, a co-infection of SFG Rickettsia belonging to R. massiliae subgroup and Coxiella-like endosymbiont (CLE), Cox-hein, in a male of Haemaphysalis heinrichi tick infesting Burmese ferret-badger in Loei province. Moreover, a male of H. hystricis tick infesting the same host was infected with another CLE, Cox-hys. Based on the 16S rRNA gene sequence, Anaplasma sp., closely related to Anaplasma bovis was also detected in a male of H. heinrichi infesting the same Burmese ferret-badger. In addition, the third CLE, Cox-asia, found in H. asiatica collected from Asian palm civet in Chiang Rai province, was different from both Cox-hein and Cox-hys. This study provided important data and broadened our knowledge on tick-borne pathogens and endosymbionts in Thailand and Southeast Asia.

Phylogenetic Studies of Coxiella-Like Bacteria and Spotted Fever Group Rickettsiae in Ticks Collected From Vegetation in Chaiyaphum Province, Thailand.

Ticks can transmit a wide variety of pathogens, including bacteria. Here, we report the detection of tick-associated bacteria in Chaiyaphum Province, northeastern Thailand. There have been few reports of tick-borne bacterial pathogens in the study areas, which are evergreen forests dominated by plateaus at elevations of approximately 1,000 m. In total, 94 ticks were collected from vegetation. They were screened for the presence of Coxiella, Francisella, Rickettsia, and Borrelia bacteria using PCR assays. In this study, we found ticks from two genera, Haemaphysalis and Amblyomma, that were positive for Coxiella-like bacteria (CLB) and Rickettsia. Francisella and Borrelia spp. were not detected in these two tick genera. The results revealed the evolutionary relationships of CLB in Amblyomma testudinarium, Haemaphysalis lagrangei, and Haemaphysalis obesa ticks using the 16S rRNA and rpoB markers, which clustered together with known isolates of ticks from the same genera. In contrast, the groEL marker showed different results. On the basis of the groEL phylogenetic analysis and BLAST results, three groups of CLB were found: (1) CLB from A. testudinarium grouped as a sister clade to CLB from Ixodes ricinus; (2) CLB from Haemaphysalis lagrangei was distantly related to CLB from Haemaphysalis wellingtoni; and (3) CLB from A. testudinarium grouped as sister clade to CLB from Amblyomma from French Guiana and Brazil. For Rickettsia studies, phylogenetic trees of the gltA, ompB, and sca4 genes revealed two groups of Spotted Fever Group (SFG) Rickettsiae: (1) SFG Rickettsiae that formed a sister clade with Rickettsia tamurae AT-1 (belong to the Rickettsia helvetica subgroup) in A. testudinarium and (2) SFG Rickettsiae that formed a distantly related group to Rickettsia rhipicephali 3-7-female6-CWPP (belong to the Rickettsia massiliae subgroup) in A. testudinarium. This study expanded our knowledge of the diversity of tick-borne Coxiella and Rickettsia bacteria. The pathogenic roles of these bacteria also need to be investigated further.

Novel phlebovirus-like-AYUT and Stenotrophomonas maltophilia bacterial co-infection in a Rhipicephalus sanguineus s.l. tick.

Tick-borne viruses and bacteria that can cause diseases of animals and humans have high impact and are of concern as significant threats to human health worldwide. In this research, we screened microorganisms related to those pathogens in ticks from dogs, a cat, and a cow. The techniques used were PCR, DNA sequencing and phylogenetic analysis to detect and classify the microorganisms [Flavivirus, severe fever with thrombocytopenia syndrome virus (SFTSV), Phlebovirus, Coronavirus, Canine Parvovirus, eubacteria, Coxiella and Rickettsia]. A novel virus named Phlebovirus-like-AYUT and Stenotrophomonas maltophilia bacteria were found in one individual tick (Rhipicephalus sanguineus s.l.) from a dog. All tick samples were negative for Rickettsia, while 9/21 (42.9 %) were positive for Coxiella bacteria. The novel virus "Phlebovirus-like-AYUT" (the name derives from Phra Nakhon Si Ayutthaya Province in Thailand) was resolved by phylogenetic analysis of the partial L segment by maximum likelihood (ML) method using MEGA X. The phylogenetic tree also indicated that the virus was related to Phlebovirus in brown dog ticks reported in Trinidad and Tobago. In contrast, Phlebovirus-like-AYUT was in a distinct clade from Lihan tick Phlebovirus-Thailand (LTPV), which was previously found in cow ticks, Rhipicephalus microplus, in Nan Province, Thailand. This study reports the Stenotrophomonas maltophilia bacterium with a novel Phlebovirus-like-AYUT in a brown dog tick. The roles of this bacterium in a virus-positive tick or in viral transmission from animal host requires further investigation.

Reinstatement of Dermacentor tricuspis (Schulze, 1933) n. comb., n. stat. (Acari: Ixodidae) as a valid species, synonymization of D. atrosignatus Neumann, 1906 and description of a new species from Indonesia, Malaysia and Thailand.

Re-examination of the holotype of Dermacentor atrosignatus Neumann, 1906 (Acari: Ixodidae) stored in the Natural History Museum (London, UK) revealed that this taxon is identical with D. auratus Supino, 1897 and should be treated as a junior synonym of the latter species. A correct name for the distinct species previously identified as D. atrosignatus Neumann, 1906 sensu Wassef & Hoogstraal, 1984 should be D. tricuspis (Schulze, 1933) n. comb., n. stat. Adults of D. tricuspis are redescribed here. Re-examination of extensive holdings of Oriental Dermacentor Koch, 1844 ticks stored in the United States National Tick Collection revealed that a morphologically distinct new species of this genus, namely D. falsosteini D. Apanaskevich, M. Apanaskevich & Nooma n. sp. should be recognized. Adults of D. tricuspis and D. falsosteini n. sp. can be distinguished from other species of Oriental Dermacentor and each other by the colour pattern of the conscutum and scutum, the pattern of punctations on the pseudoscutum and scutum, the shape of female genital structures and spurs on coxa I. Dermacentor tricuspis is recorded from Indonesia, Malaysia, the Philippines and Thailand where the adults were mostly collected from various species of wild pigs (Artiodactyla: Suidae) and vegetation; few adults were available from other mammals (Artiodactyla: Bovidae; Carnivora: Canidae, Felidae, Ursidae; Pholidota: Manidae), as well as humans and reptiles (Squamata: Elapidae, Varanidae). One male was reared from a nymph collected on a rodent (Rodentia: Muridae). Dermacentor falsosteini n. sp. is found in Indonesia, Malaysia and Thailand where the adults were collected from bearded pig, Sus barbatus Müller, wild boar, S. scrofa Linnaeus, unidentified wild pig, Sus sp. (Artiodactyla: Suidae), Malayan tapir, Tapirus indicus Desmarest (Perissodactyla: Tapiridae), human and vegetation.

Partial DnaK protein expression from Coxiella-like endosymbiont of Rhipicephalus annulatus tick.

Q fever is one of the most important zoonotic diseases caused by the obligate intracellular bacteria, Coxiella burnetii. This bacterial infection has been frequently reported in both humans and animals, especially ruminants. Ticks are important ectoparasite and serve as reservoir hosts of Coxiella-like endosymbionts (CLEs). In this study, we have attempted to express chaperone-coding genes from CLEs of Rhipicephalus annulatus ticks collected fromcow path. The partial DnaK coding sequence has been amplified and expressed by Escherichia coli. Amino acid sequences have been analyzed by MS-MS spectrometry and the UniProt database. Despites nucleotide sequences indicating high nucleotide variation and diversity, many nucleotide substitutions are synonymous. In addition, amino acid substitutions compensate for the physicochemical properties of the original amino acids. Immune Epitope Database and Analysis Resource (IEDB-AR) was employed to indicate the antigenicity of the partial DnaK protein and predict the epitopes of B-and T-cells. Interestingly, some predicted HLA-A and B alleles of the MHC-I and HLA-DR alleles belonging to MHC-II were similar to T-cell responses to C. burnetii in Q fever patients. Therefore, the partial DnaK protein of CLE from R. annulatus could be considered a vaccine candidate and immunogenic marker with future prospects.

Roles of RcsA, an AhpD Family Protein, in Reactive Chlorine Stress Resistance and Virulence in Pseudomonas aeruginosa.

Reactive chlorine species (RCS), particularly hypochlorous acid (HOCl), are powerful antimicrobial oxidants generated by biological pathways and chemical syntheses. Pseudomonas aeruginosa is an important opportunistic pathogen that has adapted mechanisms for protection and survival in harsh environments, including RCS exposure. Based on previous transcriptomic studies of HOCl exposure in P. aeruginosa, we found that the expression of PA0565, or rcsA, which encodes an alkyl hydroperoxidase D-like protein, exhibited the highest induction among the RCS-induced genes. In this study, rcsA expression was dominant under HOCl stress and greatly increased under HOCl-related stress conditions. Functional analysis of RcsA showed that the distinguishing core amino acid residues Cys60, Cys63, and His67 were required for the degradation of sodium hypochlorite (NaOCl), suggesting an extended motif in the AhpD family. After allelic exchange mutagenesis in the P. aeruginosarcsA, the P. aeruginosarcsA deletion mutant showed significantly decreased HOCl resistance. Ectopic expression of P. aeruginosarcsA led to significantly increased NaOCl resistance in Escherichia coli Moreover, the pathogenicity of the rcsA mutant decreased dramatically in both Caenorhabditis elegans and Drosophila melanogaster host model systems compared to the wild type (WT). Finally, the Cys60, Cys63, and His67 variants of RcsA were unsuccessful at complementing phenotypes of the rcsA mutant. Overall, our data indicate the importance of P. aeruginosa RcsA in defense against HOCl stress under disinfections and during infections of hosts, which involves the catalytic Cys60, Cys63, and His67 residues.IMPORTANCEPseudomonas aeruginosa is a common pathogen that is a major cause of serious infections in many hosts. Hypochlorous acid (HOCl) is a potent antimicrobial agent found in household bleach and is a widely used disinfectant. P. aeruginosa has evolved adaptive mechanisms for protection and survival during HOCl exposure. We identified P. aeruginosarcsA as a HOCl-responsive gene encoding an antioxidant protein that may be involved in HOCl degradation. RcsA has a distinguishing core motif containing functional Cys60, Cys63, and His67 residues. P. aeruginosarcsA plays an important role in bleach tolerance, with expression of P. aeruginosarcsA in Escherichia coli also conferring HOCl resistance. Interestingly, RcsA is required for full virulence in worm and fruit fly infection models, indicating a correlation between mechanisms of bleach toxicity and host immunity during infection. This provides new insights into the mechanisms used by P. aeruginosa to persist in harsh environments such as hospitals.

Description of a new species of Dermacentor Koch, 1844 (Acari: Ixodidae) from the mountains of Laos and Thailand.

Dermacentor pasteuri n. sp. (Acari: Ixodidae) is described based on adults ex wild boar and vegetation from Laos and Thailand. Adults of D. pasteuri n. sp. are similar to those of D. compactus Neumann, 1901, but can be distinguished by the shape of conscutum in the male, development of dorsal cornua in the female, size and shape of spurs on coxae and coloration of leg segments in both sexes.

Reptile-associated Borrelia spp. In Amblyomma ticks, Thailand.

A total of 127 Amblyomma ticks (A. helvolum, A. varanense and A. geoemydae) were collected from reptiles: water monitors (Varanus salvator), Bengal monitors (Varanus bengalensis), Burmese pythons (Python bivittatus), yellow-spotted keelbacks (Xenochrophis flavipunctatus), keeled rat snakes (Ptyas carinata) and elongated tortoises (Indotestudo elongata) from nine provinces in Thailand. The presence of Borrelia spp. of the 16S rRNA, flaB, glpQ, groEL and gyrB genes was examined by conventional, semi-nested and nested PCR. Phylogenetic analyses using maximum likelihood method of housekeeping genes showed that most sequences of Borrelia spp. in these Amblyomma ticks belonged to the clade of reptile-associated (REP) borreliae. Interestingly, one Borrelia sp. in an A. geoemydae tick collected from an elongated tortoise clustered in the same clade as a Borrelia sp. detected from an A. geoemydae-infested turtle in Japan (it may belong to the same species given the identical sequences of their 16S rRNA, flaB and glpQ genes) and formed the same group with tick-borne relapsing fever (RF) borreliae of B. miyamotoi and B. theileri. Our findings are the first report on the presence of Borrelia spp. in A. helvolum and A. geoemydae ticks from reptiles in Thailand adding to the geographic distribution of Borrelia spp. in Asia.

Description of a new species of Dermacentor Koch, 1844 (Acari: Ixodidae) from Laos and Thailand.

Dermacentor laothaiensis n. sp. (Acari: Ixodidae) is described based on adults ex wild boar and vegetation from Laos and Thailand. Adults of D. laothaiensis n. sp. are similar to those of D. bellulus (Schulze, 1935) and D. steini (Schulze, 1933) but can be distinguished by the conscutum shape, colour pattern of the conscutum and scutum, the density of punctations on the pseudoscutum and scutum and the shape of female genital structures.

Coxiella-like bacteria in fowl ticks from Thailand.

BACKGROUND: Coxiella bacteria were identified from various tick species across the world. Q fever is a zoonotic disease caused by the bacteria Coxiella burnetii that most commonly infects a variety of mammals. Non-mammalian hosts, such as birds, have also been reported to be infected with the pathogenic form of "Candidatus Coxiella avium". This research increases the list of tick species that have been found with Coxiella-like bacteria in Thailand. METHODS: A total of 69 ticks were collected from 27 domestic fowl (Gallus gallus domesticus), 2 jungle fowl (Gallus gallus) and 3 Siamese firebacks (Lophura diardi) at 10 locations (provinces) in Thailand. Ticks were identified and PCR was used to amplify Coxiella bacteria 16S rRNA, groEL and rpoB genes from the extracted tick DNA. MEGA6 was used to construct phylogenetic trees via a Maximum Likelihood method. RESULTS: The phylogenetic analysis based on the 16S rRNA gene showed that the Coxiella sequences detected in this study grouped in the same clade with Coxiella sequences from the same tick genus (or species) reported previously. In contrast, rpoB gene of the Coxiella bacteria detected in this study did not cluster together with the same tick genus reported previously. Instead, they clustered by geographical distribution (Thai cluster and Malaysian cluster). In addition, phylogenetic analysis of the groEL gene (the chaperonin family) showed that all Coxiella bacteria found in this study were grouped in the same clade (three sister groups). CONCLUSIONS: To our knowledge, we found for the first time rpoB genes of Coxiella-like bacteria in Haemaphysalis wellingtoni ticks forming two distinct clades by phylogenetic analysis. This may be indicative of a horizontal gene transfer event.

Pseudomonas aeruginosa glutathione biosynthesis genes play multiple roles in stress protection, bacterial virulence and biofilm formation.

Pseudomonas aeruginosa PAO1 contains gshA and gshB genes, which encode enzymes involved in glutathione (GSH) biosynthesis. Challenging P. aeruginosa with hydrogen peroxide, cumene hydroperoxide, and t-butyl hydroperoxide increased the expression of gshA and gshB. The physiological roles of these genes in P. aeruginosa oxidative stress, bacterial virulence, and biofilm formation were examined using P. aeruginosa ΔgshA, ΔgshB, and double ΔgshAΔgshB mutant strains. These mutants exhibited significantly increased susceptibility to methyl viologen, thiol-depleting agent, and methylglyoxal compared to PAO1. Expression of functional gshA, gshB or exogenous supplementation with GSH complemented these phenotypes, which indicates that the observed mutant phenotypes arose from their inability to produce GSH. Virulence assays using a Drosophila melanogaster model revealed that the ΔgshA, ΔgshB and double ΔgshAΔgshB mutants exhibited attenuated virulence phenotypes. An analysis of virulence factors, including pyocyanin, pyoverdine, and cell motility (swimming and twitching), showed that these levels were reduced in these gsh mutants compared to PAO1. In contrast, biofilm formation increased in mutants. These data indicate that the GSH product and the genes responsible for GSH synthesis play multiple crucial roles in oxidative stress protection, bacterial virulence and biofilm formation in P. aeruginosa.

Phylogenetic studies of bacteria (Rickettsia, Coxiella, and Anaplasma) in Amblyomma and Dermacentor ticks in Thailand and their co-infection.

In this study, we attempted to detect Rickettsia, Coxiella and Anaplasma bacteria in one hundred and fourteen-Dermacentor and thirty three-Amblyomma unfed adult ticks that were collected from under leaves along animal trails at different places across Thailand. PCR amplification was used to identify bacterial infection with general conserved sequences of bacteria. The results revealed single infection in Amblyomma testudinarium ticks with Rickettsia (24%) and Coxiella (6%). Anaplasma bacteria were often detected in Dermacentor auratus ticks (32%). Coxiella spp. were detected in Dermacentor atrosignatus (6%) and D. auratus ticks (3%) in this study. Moreover, we found co-infection by Coxiella and Rickettsia bacteria (39%) in Am. testudinarium. In contrast, D. atrosignatus ticks were co-infected with Coxiella and Anaplasma bacteria (3%) and Dermacentor compactus ticks were co-infected with Rickettsia and Anaplasma spp. (25%). Interestingly, Am. testudinarium ticks (12%) were found for the first time to exhibit triple infection by these three bacteria. Phylogenetic studies showed the rickettsiae from ticks causing both single and multiple infections had sequence similarity with spotted fever group rickettsial strains, including Rickettsia massilliae, R. raoultii and R. tamurae. In addition, the phylogenetic analysis of the 16S rRNA gene of Coxiella bacteria showed that they were closely grouped with Coxiella endosymbionts in both Dermacentor and Amblyomma. Moreover, the Anaplasma identified in a D. auratus tick was grouped in the same clade with the pathogenic bacterium Anaplasma phagocytophilum. Bacterial co-infections in Dermacentor and Amblyomma ticks may cause co-transmission of some tick-borne microorganisms (pathogen and endosymbiont, whether enhance or reduce) in humans and animals and they could affect medical and veterinary health.

The FinR-regulated essential gene fprA, encoding ferredoxin NADP+ reductase: Roles in superoxide-mediated stress protection and virulence of Pseudomonas aeruginosa.

Pseudomonas aeruginosa has two genes encoding ferredoxin NADP(+) reductases, denoted fprA and fprB. We show here that P. aeruginosa fprA is an essential gene. However, the ΔfprA mutant could only be successfully constructed in PAO1 strains containing an extra copy of fprA on a mini-Tn7 vector integrated into the chromosome or carrying it on a temperature-sensitive plasmid. The strain containing an extra copy of the ferredoxin gene (fdx1) could suppress the essentiality of FprA. Other ferredoxin genes could not suppress the requirement for FprA, suggesting that Fdx1 mediates the essentiality of FprA. The expression of fprA was highly induced in response to treatments with a superoxide generator, paraquat, or sodium hypochlorite (NaOCl). The induction of fprA by these treatments depended on FinR, a LysR-family transcription regulator. In vivo and in vitro analysis suggested that oxidized FinR acted as a transcriptional activator of fprA expression by binding to its regulatory box, located 20 bases upstream of the fprA -35 promoter motif. This location of the FinR box also placed it between the -35 and -10 motifs of the finR promoter, where the reduced regulator functions as a repressor. Under uninduced conditions, binding of FinR repressed its own transcription but had no effect on fprA expression. Exposure to paraquat or NaOCl converted FinR to a transcriptional activator, leading to the expression of both fprA and finR. The ΔfinR mutant showed an increased paraquat sensitivity phenotype and attenuated virulence in the Drosophila melanogaster host model. These phenotypes could be complemented by high expression of fprA, indicating that the observed phenotypes of the ΔfinR mutant arose from the inability to up-regulate fprA expression. In addition, increased expression of fprB was unable to rescue essentiality of fprA or the superoxide-sensitive phenotype of the ΔfinR mutant, suggesting distinct mechanisms of the FprA and FprB enzymes.

FRANCISELLA-LIKE ENDOSYMBIONT IN A TICK COLLECTED FROM A CHICKEN IN SOUTHERN THAILAND.

Francisella is a genus of bacterial pathogens potentially lethal to humans. We report here for the first time a novel Francisella-like endosymbiont discovered in a hard-tick (Rhipicephalus sanguineus s.l.) obtained from a chicken (Gallus domesticus) in Thailand. The phylogenetic results indicate the 16S rDNA sequences of this Francisella bacterium form a unique clade with the Francisella-like endosymbiont of the tick species, Amblyomma varanense and Amblyomma helvolum, that have previously been found on snakes in Thailand. This species of Francisella is in a different group from the other Francisella-like endosymbionts previously reported from other countries. No Francisella was detected in Haemaphysalis wellingtoni ticks obtained from chickens in this study.

Borrelia sp. phylogenetically different from Lyme disease- and relapsing fever-related Borrelia spp. in Amblyomma varanense from Python reticulatus.

BACKGROUND: Species of the genus Borrelia are causative agents of Lyme disease and relapsing fever. Lyme disease is the most commonly reported vector-borne disease in the northern hemisphere. However, in some parts of the world Lyme borreliosis and relapsing fever may be caused by novel Borrelia genotypes. Herein, we report the presence of a Borrelia sp. in an Amblyomma varanense collected from Python reticulatus. METHODS: Ticks were collected from snakes, identified to species level and examined by PCR for the presence of Borrelia spp. flaB and 16S rRNA genes. Phylogenetic trees were constructed using the neighbour-joining method. RESULTS: Three A. varanense ticks collected from P. reticulatus were positive for a unique Borrelia sp., which was phylogenetically divergent from both Lyme disease- and relapsing fever-associated Borrelia spp. CONCLUSION: The results of this study suggest for the first time that there is a Borrelia sp. in A. varanense tick in the snake P. reticulatus that might be novel.

Molecular detection of Rickettsia, Anaplasma, Coxiella and Francisella bacteria in ticks collected from Artiodactyla in Thailand.

A total of 79 ticks collected from Sambar deer (Cervus unicolor), Barking deer (Muntiacus muntjak) and Wild boar (Sus scrofa) were examined by PCR for the presence of Rickettsia, Anaplasma, Coxiella, and Francisella bacteria. Of the 79 ticks, 13% tested positive for Rickettsia, 15% tested positive for Anaplasma, 4% tested positive for Coxiella, and 3% tested positive for Francisella. Interestingly, triple infection with Anaplasma, Rickettsia and Francisella was determined in a Dermacentor auratus tick. Moreover, another triple infection with Rickettsia, Anaplasma, and Coxiella was found in a Haemaphysalis lagrangei tick. Double infection of Rickettsia with Coxiella was also detected in another H. lagrangei tick. From the phylogenetic analyses, we found a Rickettsia sp. with a close evolutionary relationship to Rickettsia bellii in the H. lagrangei tick. We also found the first evidence of a Rickettsia sp. that is closely related to Rickettsia tamurae in Rhipicephalus (Boophilus) microplus ticks from Thailand. H. lagrangei and Haemaphysalis obesa ticks collected from Sambar deer tested positive for Anaplasma species form the same clade with Anaplasma bovis. In contrast, other H. lagrangei ticks collected from Sambar deer and D. auratus ticks collected from Wild boar were also reported for the first time to be infected with an Anaplasma species that is closely related to Anaplasma platys. The phylogenetic analysis of the 16S rRNA gene of Coxiella bacteria revealed that Coxiella symbionts from H. lagrangei formed a distinctly different lineage from Coxiella burnetii (a human pathogen). Additionally, Francisella bacteria identified in D. auratus ticks were found to be distantly related to a group of pathogenic Francisella species. The identification of these bacteria in several feeding ticks suggests the risk of various emerging tick-borne diseases and endosymbionts in humans, wildlife, and domestic animals in Thailand.

Detection of Rickettsia and Anaplasma from hard ticks in Thailand.

We collected a total of 169 adult hard ticks and 120 nymphs from under the leaves of plants located along tourist nature trails in ten localities. The results present data examining the vector competence of ticks of different genera and the presence of Rickettsia and Anaplasma species. The ticks belonged to three genera, Amblyomma, Dermacentor, and Haemaphysalis, comprising 11 species. Rickettsia bacteria were detected at three collection sites, while Anaplasma bacteria were detected at only one site. Phylogenetic analysis revealed new rickettsia genotypes from Thailand that were closely related to Rickettsia tamurae, Rickettsia monacensis, and Rickettsia montana. This study was also the first to show that Anaplasma bacteria are found in Haemaphysalis shimoga ticks and are closely related evolutionarily to Anaplasma bovis. These results provide additional information for the geographical distribution of tick species and tick-borne bacteria in Thailand and can therefore be applied for ecotourism management.

Hepatozoon and Theileria species detected in ticks collected from mammals and snakes in Thailand.

We report the detection of Hepatozoon and Theileria in 103 ticks from mammals and snakes in Thailand. By using a genus-specific 18S rRNA PCR, Hepatozoon and Theileria spp. were detected in 8% and 18%, respectively, of ticks (n=79) removed from mammals. Of the ticks removed from snakes (n=24), 96% were infected with Hepatozoon spp., but none were infected with Theileria. Phylogenetic analysis revealed that Hepatozoon spp. detected from Dermacentor astrosignatus and Dermacentor auratus ticks from Wild boar (Sus scrofa) formed a phylogenetic group with many isolates of Hepatozoon felis that were distantly related to a species group containing Hepatozoon canis and Hepatozoon americanum. In contrast, a phylogenetic analysis of the Hepatozoon sequences of snake ticks revealed that Hepatozoon spp. from Amblyomma varanense from King cobra (Ophiophagus hannah) and Amblyomma helvolum ticks from Indochinese rat snake (Ptyas korros), and Asiatic water snake (Xenochrophis piscator) are grouped with Hepatozoon spp. recently isolated from Monocellate cobras, Reticulated pythons and Burmese pythons, all of Thai origin, and with Hepatozoon sp. 774c that has been detected from a tick species obtained from Argus monitors in Australia. A phylogenetic analysis demonstrated that Theileria spp. from Rhipicephalus (Boophilus) microplus, Haemaphysalis obesa, and Haemaphysalis lagrangei ticks from Sambar deer (Cervus unicolor) cluster with the Theileria cervi isolates WU11 and 239, and Theileria sp. Iwate 141. We report for the first time a Hepatozoon species that shares genetic similarity with Hepatozoon felis found in Dermacentor astrosignatus and Dermacentor auratus ticks collected from Wild boars in Thailand. In addition, we found the presence of a Theileria cervi-like sp. which suggests the potential role of Haemaphysalis lagrangei as a Theileria vector in Thailand.