Babesia bovis merozoite surface antigen 2 (MSA-2) are proteins encoded by msa multigene family and implicated in erythrocyte recognition and invasion, yet their roles remain only partially understood. We investigated spatiotemporal differences of three MSA-2 paralogs (MSA-2a1, MSA-2a2, and MSA-2b) by using epitope-specific rabbit antisera. Alignment of amino acid sequences of B. bovis T2Bo strain and Texas strain C1 clone showed that MSA-2a1 is highly conserved (99.6% identity) among the parasites, whereas MSA-2a2 (84%) and MSA-2b (94%) are more variable; notably, the least conserved paralog, MSA-2a2, harbored a contiguous 24 amino acid deletion. Indirect immunofluorescence assays using the antisera revealed apical localization of all three proteins in intraerythrocytic parasite. Following merozoite purification and brief incubation, localization of MSA-2a1 was observed on the merozoite surface before the attachment, whereas MSA-2a2 and MSA-2b remained at the apical end. Moreover, growth-inhibition assays, antiserum Ab-1 recognizing MSA-2a2 and MSA-2b inhibited 90.4% at 48 h incubation, whereas Ab-2 recognizing MSA-2a1 and MSA-2a2 inhibited only 13.1%. These results suggest functional difference among MSA-2 paralogs: MSA-2a1 was secreted to the surface before attachment, while MSA-2a2 and MSA-2b remain inside the merozoite at the onset of attachment. Taken together, MSA-2a1, MSA-2a2, and MSA-2b appear structurally similar, yet functionally distinct proteins that may contribute to the coordinated control of erythrocyte invasion.

Bats are recognized reservoirs for numerous pathogens, including zoonotic agents. They harbor a variety of ectoparasites, such as ticks, which pose a potential risk for cross-species pathogen spillover. Borrelia, a genus of spirochete bacteria responsible for several vector-borne diseases, represents a significant public health concern due to its global distribution. However, its genetic diversity and circulation in Southeast Asia remain poorly understood. To address this knowledge gap, this study investigated the occurrence and genetic diversity of Borrelia spp. in Thai bats and the ticks associated with them based on PCR and sequencing, without bacterial isolation or clinical case data. A total of 1031 bats were sampled across nine provinces in Thailand between February 2018 and February 2023. Morphological and molecular analysis-based methods identified and confirmed 96 Reticulinasus sp. ticks infesting Craseonycteris thonglongyai and Eonycteris spelaea. Molecular screening of 48 pooled tick samples (from 96 individual ticks) detected four Borrelia-positive pools originating from three individual C. thonglongyai bats. PCR screening of 814 bats targeting the flaB gene revealed 13 positive individuals (1.6%; 95% CI: 0.9-2.7%), comprising Taphozous melanopogon, Hipposideros gentilis, and C. thonglongyai. Multilocus sequence analysis of the 16S rRNA, flaB, and glpQ genes revealed three distinct Borrelia lineages. Two clustered within the relapsing fever Borrelia group, highlighting the need for further investigation of their transmission potential and epidemiological significance in other animal hosts. The third lineage formed a highly divergent, bat-associated clade, suggesting possible cryptic speciation based on sequence divergence across the three loci; however, formal taxonomic classification will require broader sampling and additional genetic markers. This study presents the first molecular-level evidence of Borrelia in Thai bats and bat-infesting ticks.

Hemotropic mycoplasmas, particularly Mycoplasma wenyonii and “Candidatus Mycoplasma haematobovis”, are increasingly recognized as emerging vector-borne pathogens of veterinary importance in cattle and buffalo. Due to their complex epidemiology and the limited understanding of their pathogenic mechanisms, hemoplasma infections in large ruminants are often underdiagnosed and underreported, especially in resource-limited settings. This study investigated the genetic diversity and phylogenetic relationships of hemotropic Mycoplasma species in blood samples collected from cattle and buffalo across seven provinces in Thailand between 2016 and 2023. A total of 422 samples (214 from cattle and 208 from buffalo) were analyzed, with 373 testing positive for hemotropic Mycoplasma DNA. Phylogenetic analyses of the 16S and 23S rRNA gene sequences revealed two distinct clades corresponding to M. wenyonii and “Ca. M. haematobovis”. Most sequences clustered with M. wenyonii strains from multiple countries, while a smaller subset grouped with “Ca. M. haematobovis” strains. Intraspecific sequence similarity among M. wenyonii 16S rRNA gene sequences ranged from 97.29% to 100%, whereas “Ca. M. haematobovis” sequences showed similarities ranging from 99.75% to 100%. Interspecific similarity between the two species ranged from 83.48% to 84.34%. For the 23S rRNA gene sequences, M. wenyonii exhibited intraspecific similarity levels ranging from 90.94% to 100%, while “Ca. M. haematobovis” sequences demonstrated 99.53% similarity; interspecific similarity for this gene ranged from 76.69% to 77.34%. We identified 11 nucleotide sequence types (ntSTs) for M. wenyonii, with ntST10 being the most widely distributed, shared among isolates from Cuba, Thailand, Japan, and Brazil. The single “Ca. M. haematobovis” sequence type detected in this study clustered within ntST7, alongside sequences from Cuba, Malaysia, Brazil, Turkey, Kyrgyzstan, and Japan. These findings highlight the substantial genetic diversity and broad geographical distribution of hemotropic Mycoplasma species in Thailand and emphasize the critical importance of molecular screening for managing the risk of pathogen transmission within livestock populations.

Abstract Background Second only to mosquitoes, ticks (Acari: Ixodida) are significant blood-feeding ectoparasites and vectors of numerous pathogens affecting both animals and humans. Despite bats serving as hosts to various tick species, they remain relatively understudied due to their nocturnal behavior and laborious capture procedures. Soft ticks in particular display diverse ecological behaviors, inhabiting bat roosts, caves, and occasionally human dwellings. This overlap in habitats suggests soft ticks may play a critical role as vectors of zoonotic pathogens. In Southeast Asia, research on soft ticks has primarily focused on island nations, with limited studies on bat-associated ticks in Thailand. This study aimed to examine the identity and distribution of bat ticks in Thailand. Methods Bats were captured across ten provinces in Thailand between 2018 and 2023. Ticks were removed from the bats’ skin and identified through morphological examination using a stereomicroscope, with molecular confirmation. Scanning electron micrographs were recorded. Prevalence, mean abundance, and mean intensity of tick infestations were calculated. The mitochondrial genomes of the ticks were sequenced, annotated, and subjected to phylogenetic analysis. Results A total of 1031 bats, representing 7 families, 11 genera, and 28 species, were captured. Tick infestations were found in 34 bats (3.30%), specifically in two species: Craseonycteris thonglongyai (33/139, 23.74%) and Eonycteris spelaea (1/2, 50%). All ticks were in the larval stage. Basic local alignment search tool for nucleotide (BLASTN) searches using 16S rRNA (425 bp) and COI (825 bp) sequences, along with Barcode of Life Database (BOLD) database queries, revealed the highest similarity to tick in the genus Reticulinasus found on bats in Zambia. The mitochondrial genomes of ticks collected from C. thonglongyai and E. spelaea were 14,433 bp and 14,439 bp in length, respectively, and contained 13 protein-coding genes, 22 tRNA genes, and 2 rRNA genes. Phylogenetic analysis placed these ticks within the Reticulinasus clade, with strong support indicated by high bootstrap values. Conclusions This study identified Reticulinasus sp. infestations on C. thonglongyai and E. spelaea bats, marking the first report of soft ticks in bats from Thailand, with potential implications for zoonotic disease transmission. Graphical Abstract

Arthropod vectors play a crucial role in the transmission of hemotropic mycoplasmas, small bacteria that infect red blood cells in a wide range of animals and humans globally, leading to intravascular infections. Traditional Giemsa-stained thin blood smears, used for diagnosing hemotropic mycoplasmas through microscopic examination, have low sensitivity and are effective only when bacteremia levels are high. This study aimed to employ molecular methods to detect and genetically characterize hemotropic mycoplasmas in goats as well as investigate the potential role of fleas as vectors. Blood and flea samples were collected concurrently from goats on 16 farms across seven provinces in Thailand from January 2017 to October 2023. The 16 S rRNA, 23 S rRNA, and rnpB genes of hemoplasmas were amplified and sequenced. All fleas were identified morphologically and molecularly through DNA barcoding of the cytochrome oxidase I gene. A total of 78 out of 500 goats (15.6%), three pooled flea samples (3/6, 50%), and one individual flea (1/49, 2.04%) tested positive for hemoplasmas and all fleas were identified as Ctenocephalides orientis. BLASTN searches utilizing the three genetic markers revealed that the hemoplasmas detected in this study showed 97.81-100% similarity to Mycoplasma ovis and Candidatus Mycoplasma haemovis, which have been previously reported in sheep, goats, and humans, suggesting their zoonotic potential. The sequences were grouped into 28 unique nucleotide sequence types (ntSTs) based on minor variations in the 16 S rRNA gene. Hemotropic mycoplasma infection was significantly associated with farm locations and seasonality of sample collection (p < 0.0001), indicating that farm management practices or environmental conditions may play a critical role in the epidemiology of these infections. This study represents the first report of hemotropic mycoplasmas in goats in Thailand, confirms their presence in fleas, and provides valuable insights for farm management, such as guiding the rational use of insecticides and antibiotics.

Virulent species or strains of hematophagous borne pathogens such as Anaplasma spp., Babesia spp., Theileria spp., and Trypanosoma spp., are lethal to susceptible animals or reduce their productivity on a global scale. Nonetheless, efforts to diagnose the causative agents and assess the genotypic profiles as well as quantify the parasite burden of aforementioned parasites across seasons remain limited. Therefore, the present investigation sought to elucidate the genotypic composition of Anaplasma spp., Babesia spp., Theileria spp., and Trypanosoma spp. The findings revealed heightened infection rates during the summer, manifesting a correlation between Trypanosoma spp. infection and seasonal fluctuations. Among the identified pathogens, Anaplasma marginale emerged as the most dominant species, while the occurrence of Anaplasma platys in Thai cattle was confirmed via the sequencing of the groEL gene. Moreover, the study successfully identified two lineages of Trypanosoma theileri. The findings of this investigation offer valuable insights that can inform the development of preventive strategies for vector-borne diseases, such as considering the appropriate use of insect repellent, mosquito or insect nets, or eliminating breeding places for insects in each season.

BackgroundSand flies, belonging to the Psychodidae family, represent small, hairy insects that serve as significant vectors in various important medical and veterinary diseases. Despite being recognized by the World Health Organization as an endemic area for leishmaniasis, Southeast Asia lacks comprehensive information on the species composition and biology of sand flies. To address this, the current study aimed to survey sand fly biodiversity.MethodsSand flies from six provinces in Southern Vietnam were collected using CDC light traps. Sand flies were subsequently identified morphologically and confirmed molecularly using mitochondrial cytochrome oxidase c subunit I (COI) and cytochrome b (cytb) sequences. BLASTN searches were conducted, and the species identity of sand flies was further confirmed through a Barcode of Life Database (BOLD) search utilizing COI sequences. Subsequently, nucleotide sequences were subjected to a panel of analyses including intraspecific variation, phylogenetic relationships and haplotype network. The average densities of collected sand flies (sand flies/trap/night) and species richness were also recorded.ResultsA total of 753 sand flies were collected. After excluding damaged specimens, six sand fly species, namely Phlebotomus stantoni, Sergentomyia khawi, Se. silvatica, Se. barraudi, Se. bailyi and Grassomyia indica, were identified. All conspecific sand fly sequences, including Ph. stantoni, Se. barraudi, Gr. indica, Se. bailyi, Se. khawi and Se. silvatica, clustered with their reference sequences, corroborating the results of morphology-based identification, BLASTN analysis and BOLD search. For intraspecific variation of sand flies obtained from the current study, COI diversity indices were consistently higher than those of cytb.ConclusionsThis study provides the first updates on morphological and molecular characterization of sand flies in Southern Vietnam. This acquired knowledge on sand fly species composition is essential for controlling sand fly-borne diseases in this potentially endemic region.

Ticks and tick-borne pathogens (TTBP) pose a serious threat to animal and human health globally. Anaplasma bovis, an obligatory intracellular bacterium, is one of the more recent species of the Family Anaplasmaceae to be formally described. Owing to its diminutive size, microscopic detection presents a formidable challenge, leading to it being overlooked in laboratory settings lacking advanced equipment or resources, as observed in various regions, including Thailand. This study aimed to undertake a genetic analysis of A. bovis and determine its prevalence in goats and ticks utilizing three genetic markers (16S rRNA, gltA, groEL). A total of 601 goat blood and 118 tick samples were collected from 12 sampling sites throughout Thailand. Two tick species, Haemaphysalis bispinosa (n = 109), and Rhipicephalus microplus (n = 9) were identified. The results herein showed that 13.8% (83/601) of goats at several farms and 5% (1/20) of ticks were infected with A. bovis. Among infected ticks, A. bovis and an uncultured Anaplasma sp. which are closely related to A. phagocytophilum-like 1, were detected in each of H. bispinosa ticks. The remaining R. microplus ticks tested positive for the Anaplasma genus. A nucleotide sequence type network showed that A. bovis originated from Nan and Narathiwat were positioned within the same cluster and closely related to China isolates. This observation suggests the potential dispersal of A. bovis over considerable distances, likely facilitated by activities such as live animal trade or the transportation of infected ticks via migratory birds. The authors believe that the findings from this study will provide valuable information about TTBP in animals.

Trypanosomiasis caused Trypanosoma evansi is current public health concern especially, in south Asia and Southeast Asia. Although polymerase chain reaction is currently used as a standard method, the techniques required skilled personnel, were performed in multiple steps, and required expensive instruments. Fundamental microscopic approach also has limitation in use by facing both inter- and intra-variability of interpretation by examiners. New automatic tool with the microscopic examination is needed. The study aimed to develop the mobile application-based YOLO neural network algorithms to predict T. evansi blood stages from thin-blood film examination. YOLO v4 tiny model is outperformed to localize and classify unseen images with the best performance at 95% of sensitivity, specificity, precision, accuracy and F1 score, respectively, with less misclassification rate than 5%. Simulation implementation platform, calling CiRA bot, give the empirical result and reliably comparable to that from the computational experiment studied with the area under ROC and precision-recall curves as 0.964 and 0.962, respectively. The result obtained from the CIRA bot platform is good enough for further distribution in field site. In the future, the study could contribute human and animal public health staff to simply identify the unicellular parasitic flagellate infection and also benefit them for designing the strategy in prevention and treatment of the disease.

Tick-borne pathogens of the genus Theileria and Anaplasma has been widely distributed and infect small ruminant in tropical and subtropical countries. Several species are considered highly pathogenic, while some cause subclinical infections in small ruminants. The distribution data of theileriosis and anaplasmosis in Samigaluh, Kulon Progo remain scarce and poorly understood. Therefore, further investigations are required to control this tick-borne disease. This study aimed to investigate the occurrence of theileriosis and anaplasmosis in domestic goats in Samigaluh, Kulon Progo, Western Yogyakarta. A total of 53 blood samples were collected from domestic goats. Blood sample was drawn from the jugular vein individually and kept in an EDTA tube. Thin blood smear was stained by Giemsa and observed under microscope for genus-level identification. As a result, it was found that theileriosis 12/53 (23%), anaplasmosis 25/53 (47%) and mixed infection 6/53 (11%) were detected in the present study.The conclusion of this study is that the incidence of Anaplasma sp. infection in goats is greater than Theileria sp. infection in Samigaluh, Kulon Progo.Although all domestic goats were asymptomatic, the finding of this study may shed light on the distribution of theileriosis and anaplasmosis infecting domestic goats in Samigaluh, Kulon Progo.

Bovine babesiosis is one of the most economically important tick-borne diseases in tropical and subtropical countries. A conventional microscopic diagnosis is typically used because it is inexpensive and expeditious. However, it is highly dependent on well-trained microscopists and tends to be incapable of detecting subpatent and chronic infections. Here, we developed a novel nucleic acid-based amplification method using loop-mediated isothermal amplification (LAMP) in conjunction with a colori-fluorometric dual indicator for the rapid and accurate detection of Babesia bovis based on the mitochondrial cytochrome b gene. We aimed to improve the thermostability, sensitivity, specificity, and alternative visualization of LAMP-based methods. We assessed its diagnostic performance compared to two conventional PCR agarose gel electrophoresis (PCR-AGE) methods. The thermostability of LAMP reaction mixtures and DNA templates in variable conditions was also assessed. In addition, we evaluated alternative visualization methods using different light sources including neon, LED, and UV lights. We found that the LAMP-neon was ten times more sensitive than the PCR-AGE, while the LAMP-LED and LAMP-UV were 1,000 times more sensitive. The current LAMP method showed no cross-amplification with uninfected cattle DNA or other common blood parasites in cattle, including Babesia bigemina, Theileria orientalis, Anaplasma marginale, and Trypanosoma evansi. In addition, the developed LAMP method has good thermostability and the potential for on-site utility as B. bovis DNA could still be detected up to 72 h after initial preparation. Our findings suggested that the developed LAMP method provides an alternative approach for B. bovis detection with sensitivity higher than PCR-AGE diagnostics, high specificity, and the flexibility to use neon, LED, and UV light sources for positive signal observations.

In recent phylogenetic studies, bat Polychromophilus and ungulate Plasmodium, two relatively understudied haemosporidian parasites within the Apicomplexa phylum, have often been overlooked. Instead, the focus has been primarily on haemosporidian parasites in primates, rodents, and birds. Several phylogenetic analyses of bat Polychromophilus have relied on limited datasets and short informative DNA sequences. As a result of these inherent limitations, the substantiation of their evolutionary stance has encountered a diminished degree of robust validation. This study successfully obtained complete mitochondrial genome sequences from 11 Polychromophilus parasites originating from Hipposideros gentilis and Myotis siligoensis bats for the first time. Additionally, the authors have sequenced the apicoplast caseinolytic protease C genes from Polychromophilus murinus and a potentially new Polychromophilus species. These mitochondrial genomes range in length from 5994 to 6001 bp and consist of three protein-coding genes (PCGs), seven small subunit ribosomal RNA genes (SSU rRNA), 12 large subunit ribosomal RNA genes (LSU rRNA), and seven miscellaneous RNA genes. Phylogenetic analyses using Bayesian Inference and Maximum Likelihood methods indicated robust support for the grouping of ungulate Plasmodium and bat Polychromophilus in a single clade separate from other Plasmodium spp., confirming previous reports, albeit with stronger evidence in this study. The divergence between Polychromophilus in bats and Plasmodium in ungulates occurred approximately 29.61 to 55.77 million years ago (Mya), with a node age estimated at 40.63 Mya. These findings highlight that the genus Plasmodium, which includes species found in ungulates, birds, reptiles, and other mammals, does not form a monophyletic group. By incorporating Polychromophilus in bats and Plasmodium in ungulates, this study contributes significantly to understanding the phylogenetic relationships within the Haemosporida order. It provides valuable insights into the evolutionary history and interconnections among these diverse parasites, thereby expanding knowledge in this field.

Bovine anaplasmosis is a serious tick-borne disease that is responsible for economic loss worldwide. The major surface proteins (MSPs), encoded by msp1 to msp5 genes of Anaplasma marginale, play an important role in host-pathogen and tick-pathogen interactions. These markers have been used for genetic characterization and phylogenetic studies. Despite domestic reports concerning suspected outbreaks of anaplasmosis in Thailand, genetic analysis of A. marginale in the country remains largely limited. Therefore, we aim to investigate the infection rate of the rickettsia organism in the Anaplasmataceae family throughout five regions of Thailand and to further characterize the key genetic markers: msp1a, msp2, and msp5 of A. marginale. From 2016 to 2021, we collected a total of 384 cattle blood samples across 18 provinces. Overall, the infection rate of the rickettsia organism in the Anaplasmataceae family was 46.1%. Over 65% of the positive samples were confirmed as A. marginale. We successfully obtained a total of 138 A. marginale msp1a (38), msp2 (79), and msp5 (21) sequences from all regions of the country. The msp1a and msp2 genes exhibit a high degree of genetic diversity, while the msp5 gene is highly conserved among the Thai isolates. Our findings regarding msp1a corroborated the genetic heterogeneity of A. marginale strains in endemic regions worldwide. Additionally, we found multiple novel variants for the first time in the current nationwide survey. We found 45 tandem repeat characters of the msp1a sequence. Among them, 24 characters were not shared with other countries. Collectively, we expanded the extent of genetic diversity in key markers; msp1a and msp2 genes, and further confirmed the previous finding that msp5 was highly conserved. The msp1a and msp2 genes could be useful for the surveillance of newly introduced strains. The current data may also be useful in designing a vaccine containing potential epitopes of different antigens in the future.

Ungulate malaria parasites and their vectors are among the least studied when compared to other medically important species. As a result, a thorough understanding of ungulate malaria parasites, hosts, and mosquito vectors has been lacking, necessitating additional research efforts. This study aimed to identify the vector(s) of Plasmodium bubalis . A total of 187 female mosquitoes (133 Anopheles spp., 24 Culex spp., 24 Aedes spp., and 6 Mansonia spp. collected from a buffalo farm in Thailand where concurrently collected water buffalo samples were examined and we found only Anopheles spp. samples were P. bubalis positive. Molecular identification of anopheline mosquito species was conducted by sequencing of the PCR products targeting cytochrome c oxidase subunit 1 ( cox1 ), cytochrome c oxidase subunit 2 ( cox2 ), and internal transcribed spacer 2 ( ITS2 ) markers. We observed 5 distinct groups of anopheline mosquitoes: Barbirostris, Hyrcanus, Ludlowae, Funestus, and Jamesii groups. The Barbirostris group ( Anopheles wejchoochotei or Anopheles campestris ) and the Hyrcanus group ( Anopheles peditaeniatus ) were positive for P. bubalis . Thus, for the first time, our study implicated these anopheline mosquito species as probable vectors of P. bubalis in Thailand.

The vast majority of trypanosome species is vector-borne parasites, with some of them being medically and veterinary important (such as Trypanosoma cruzi and Trypanosoma brucei) and capable of causing serious illness in vertebrate hosts. The discovery of trypanosomes in bats emphasizes the importance of bats as an important reservoir. Interestingly, there is a hypothesis that bats are ancestral hosts of T. cruzi. Trypanosome diversity has never been investigated in bats in Thailand, despite being in a biodiversity hot spot. To gain a better understanding of the diversity and evolutionary relationship of trypanosomes, polymerase chain reaction-based surveys were carried out from 2018 to 2020 in 17 sites. A total of 576 bats were captured, representing 23 species. A total of 38 (6.6%) positive samples was detected in ten bat species. Trypanosoma dionisii and Trypanosoma noyesi were identified from Myotis siligorensis and Megaderma spasma, respectively. The remaining 18S rRNA sequences of trypanosomes were related to other trypanosomes previously reported elsewhere. The sequences in the current study showed nucleotide identity as low as 90.74% compared to those of trypanosomes in the GenBank database, indicating the possibility of new species. All bat trypanosomes identified in the current study fall within the T. cruzi clade. The current study adds to evidence linking T. noyesi to a bat trypanosome and further supports the bat host origin of the T. cruzi clade. To the best of authors' knowledge, this is the first study on bat trypanosomes in Thailand and their phylogenetic relationships with global isolates.

The study of bacterial zoonoses has been under-pursued despite the fact that bacteria cause the majority of zoonotic diseases, of which 70% have a wildlife origin. More Bartonella species are being identified as the cause of human diseases, and several of them have been linked to domestic and wild animals. Bats are outstanding reservoirs for Bartonella species because of their wide distribution, mobility, roosting behaviour, and long life span. Here, we carried out a PCR-based survey on bats that were collected from 19 sampling sites in eight provinces of Thailand from February 2018 to April 2021. Bartonella infection was investigated in a total of 459 bats that belong to 24 different bat species (21 species of which had never been previously studied in Thailand). PCR diagnostics revealed that 115 out of 459 (25.5%) blood samples tested positive for Bartonella. The nucleotide identities of the Bartonella 16S rRNA sequences in this study were between 95.78-99.66% identical to those of known zoonotic species (Bartonella ancashensis, Bartonella henselae, Bartonella bacilliformis and Bartonella australis) as well as to an unidentified Bartonella spp. In addition, the citrate synthase (gltA) and RNA polymerase-beta subunit (rpoB) genes of Bartonella were sequenced and analyzed in positive samples. The gltA and rpoB gene sequences from Hipposideros gentilis and Rhinolophus coelophyllus bat samples showed low nucleotide identity (<95%) compared to those of the currently deposited sequences in the GenBank database, indicating the possibility of new Bartonella species. The phylogenetic inference and genetic diversity were generated and indicated a close relationship with other Bartonella species previously discovered in Asian bats. Overall, the current study demonstrates the primary evidence pointing to a potential novel Bartonella species in bats. This discovery also contributes to our current understanding of the geographical distribution, genetic diversity, and host ranges of bat-related Bartonella.

Malaria parasites in the phylum Apicomplexa (Order: Haemosporida) infect diverse vertebrates and invertebrate hosts. At least seven genera of haemosporidian parasites have been described to exclusively infect bats. Most of these parasites remain enigmatic with a poorly known host range. Here, we investigated 271 bats belonging to 21 species and seven families from six provinces of Thailand. Overall, 124 out of 271 bats (45.8%) were positive for haemosporidian parasites, while none had Plasmodium, based on microscopic examination of blood smears and PCR amplification. We obtained 19 distinct cytochrome b (cytb) nucleotide haplotypes of Hepatocystis from seven bat species (families: Craseonycteridae, Hipposideridae, Pteropodidae, and Rhinolophidae). Nycteria was found in four bat species (Craseonycteridae, Emballonuridae, Megadermatidae, and Pteropodidae) and Polychromophilus in two species (Emballonuridae, Vespertilionidae). Phylogenetic analysis inferred from cytb sequences placed Hepatocystis into 2 different clades. Most Hepatocystis infections were found in insectivorous bats and clustered together with a sequence from Hipposideros larvatus in Cambodia (in subclade 1a). A single sequence of Hepatocystis obtained from a frugivorous bat, Cynopterus brachyotis, was placed in the same clade with Hepatocystis from the same bat species previously reported in Malaysia (clade 2). Nycteria in these Thai bats were clearly separated from the African isolates previously reported in bats in the family Rhinolophidae. Polychromophilus murinus from Myotis siligorensis was placed in a distinct clade (clade 2) from Polychromophilus melanipherus isolated from Taphozous melanopogon (clade 1). These results confirmed that at least two distinct species of Polychromophilus are found in Thailand. Collectively, Hepatocystis presented no host specificity. Although Megaderma spasma seemed to be infected by only Nycteria, its respective parasite does not show specificity to only a single bat host. Polychromophilus murinus and P. melanipherus seem to infect a narrower host range or are somehow restricted to bats in the families Vespertilionidae and Emballonuridae, respectively.