Metagenomic Approach to Characterizing Disease Epidemiology in a Disease-Endemic Environment in Northern Thailand

Abstract

In this study, we used a metagenomic approach to analyze bacterial communities from diverse populations (humans, animals, and vectors) to investigate the role of these microorganisms as causative agents of disease in human and animal populations. Wild rodents and ectoparasites were collected from 2014 to 2018 in Nan province, Thailand where scrub typhus is highly endemic. Samples from undifferentiated febrile illness (UFI) patients were obtained from a local hospital. A total of 200 UFI patient samples were obtained and 309 rodents and 420 pools of ectoparasites were collected from rodents ( n = 285) and domestic animals ( n = 135). The bacterial 16S rRNA gene was amplified and sequenced with the Illumina. Real-time PCR and Sanger sequencing were used to confirm the next-generation sequencing (NGS) results and to characterize pathogen species. Several pathogens were detected by NGS in all populations studied and the most common pathogens identified included Bartonella spp., Rickettsia spp., Leptospira spp., and Orientia tsutsugamushi . Interestingly, Anaplasma spp. was detected in patient, rodent and tick populations, although they were not previously known to cause human disease from this region. Candidatus Neoehrlichia, Neorickettsia spp., Borrelia spp., and Ehrlichia spp. were detected in rodents and their associated ectoparasites. The same O. tsutsugamushi genotypes were shared among UFI patients, rodents, and chiggers in a single district indicating that the chiggers found on rodents were also likely responsible for transmitting to people. Serological testing using immunofluorescence assays in UFI samples showed high prevalence (IgM/IgG) of Rickettsia and Orientia pathogens, most notably among samples collected during September–November. Additionally, a higher number of seropositive samples belonged to patients in the working age population (20–60 years old). The results presented in this study demonstrate that the increased risk of human infection or exposure to chiggers and their associated pathogen ( O. tsutsugamushi ) resulted in part from two important factors; working age group and seasons for rice cultivation and harvesting. Evidence of pathogen exposure was shown to occur as there was seropositivity (IgG) in UFI patients for bartonellosis as well as for anaplasmosis. Using a metagenomic approach, this study demonstrated the circulation and transmission of several pathogens in the environment, some of which are known causative agents of illness in human populations.

Bartonella spp. Prevalence (Serology, Culture, and PCR) in Sanitary Workers in La Rioja Spain

“…Subsequently the BAPGM platform has been used diagnostically to assess bloodstream infection in dogs, other animal species, [37,38] and in humans [19,20,[39][40][41]. It is clear that the developmental and microbiological utilization of more sensitive or robust microbiological techniques such as MALDI TOF, Next Generation Sequencing and specialized enrichment/PCR techniques enhance the detection of microorganisms in various patient samples [42][43][44].…”

Detection of Anaplasma spp. and Bartonella spp. from wild‐caught rodents and their ectoparasites in Nakhon Ratchasima Province, Thailand

“…Since Haemaphysalis ticks are three-host ticks, there is a possibility that our fed tick specimens may have fed on a non-infected rodent before contacting an infected rodent. A recent report by Takhampunya et al (2019) found that the A. bovis pathogen was detected in H. bandicota ticks that were removed from B. indica and Rattus spp. rodents collected in northern Thailand.…”

A Host-Specific Blocking Primer Combined with Optimal DNA Extraction Improves the Detection Capability of a Metabarcoding Protocol for Canine Vector-Borne Bacteria

“…Thorough characterisation and monitoring of CVBD is crucial to protect the health of dogs and humans, especially in regions of Asia where there has been little explorative research done into established, emerging and novel CVBDs [2,5]. To address these knowledge gaps, state-of-the-art methodologies could be employed such as next-generation sequencing (NGS) based 16S ribosomal RNA (16S rRNA) metabarcoding that can supersede conventional PCR (cPCR) techniques with regard to their ability to detect and discover rare and/or novel organisms [18,19]. 16S rRNA metabarcoding is better able to elucidate diversity in explorative research by not relying on likely pathogen prevalence in a region nor on a priori knowledge of a pathogen's target genetic sequences, whilst also being better able to characterise coinfection [18,20].…”

“…To address these knowledge gaps, state-of-the-art methodologies could be employed such as next-generation sequencing (NGS) based 16S ribosomal RNA (16S rRNA) metabarcoding that can supersede conventional PCR (cPCR) techniques with regard to their ability to detect and discover rare and/or novel organisms [18,19]. 16S rRNA metabarcoding is better able to elucidate diversity in explorative research by not relying on likely pathogen prevalence in a region nor on a priori knowledge of a pathogen's target genetic sequences, whilst also being better able to characterise coinfection [18,20]. Previous work has already developed a novel 16S and 18S rRNA metabarcoding methodology to characterise the range of blood-borne bacterial, apicomplexan and kinetoplastid organisms infecting canine hosts in Thailand, a country of substantial CVBD diversity [4,8,[21][22][23].…”