Haemosporidian Parasites of White-Breasted Waterhens (Amaurornis phoenicurus), with a Report and Molecular Characterization of Haemoproteus gallinulae in Thailand.

Haemosporidian parasites are vector-borne parasites infecting terrestrial vertebrates as well as avian species, such as the White-breasted Waterhen, a Gruiformes waterbird found in lowlands near wetlands and distributed throughout Thailand. However, information regarding haemosporidia infection in this species is lacking. To establish regional information, 17 blood samples were collected from White-breasted Waterhens. Four haemoparasite lineages were identified in six blood samples: Haemoproteus gallinulae, Plasmodium collidatum, Plasmodium elongatum, and an unidentified Plasmodium species. H. gallinulae was characterized with morphological features in White-breasted Waterhens for the first time; the morphological characteristics were consistent with previous descriptions. H. gallinulae was more closely related to Haemoproteus species of Passeriformes birds than to those of Gruiformes birds. The Plasmodium parasites infecting these White-breasted Waterhens previously caused severe avian malaria in other host species. The unidentified Plasmodium species had rarely been documented, although it was reported in the Culex vector and was possibly associated with specialist parasites either as host or habitat. Our findings reveal multiple haemosporidian species reflecting the role of this avian host as a carrier of haemosporidians. This study offers species records and molecular materials that may provide critical information for further targeted research into these haemosporidia.

Baseline study of the morphological and genetic characteristics of Haemoproteus parasites in wild pigeons (Columba livia) from paddy fields in Thailand.

Haemoproteus columbae is a common haemosporidian parasite of wild pigeons (Columba livia) reported worldwide. In Thailand, the wild pigeon population is increasing due to paddy field monoculture. However, there are limited reports on the presence of H. columbae in these pigeon populations. The aim of the study was to characterize H. columbae in wild pigeons. A total of 87 wild pigeons were examined using microscopic and molecular methods. Haemoproteus columbae was detected in approximately 27.6% of pigeons and their morphological characteristics were described. The partial cytochrome b (cyt b) gene sequence of H. columbae was then characterized into three common lineages (HAECOL1, COLIV03, and COQUI05). By highlighting the morphologic and genetic characteristics of H. columbae commonly found in this population of pigeons, this study provides essential regional knowledge about haemosporidian parasites that could benefit future taxonomic and phylogeographic studies.

Performance of the onstructural 1 Antigen Rapid Test for detecting all four DENV serotypes in clinical specimens from Bangkok, Thailand.

BACKGROUND: Dengue is an arboviral disease that has a large effect on public health in subtropical and tropical countries. Rapid and accurate detection of dengue infection is necessary for diagnosis and disease management. We previously developed highly sensitive immunochromatographic devices, the TKK 1st and TKK 2nd kits, based on dengue virus (DENV) nonstructural protein 1 detection. However, these TKK kits were evaluated mainly using DENV type 2 clinical specimens collected in Bangladesh, and further validation using clinical specimens of other serotypes was needed. METHODS: In the present study, one of the TKK kits, TKK 2nd, was evaluated using 10 DENV-1, 10 DENV-2, 4 DENV-3, 16 DENV-4, and 10 zika virus-infected clinical specimens collected in Bangkok, Thailand. RESULTS: The TKK 2nd kit successfully detected all four DENV serotypes in patient serum specimens and did not show any cross-reactivities against zika virus serum specimens. The IgM and/or IgG anti-DENV antibodies were detected in seven serum specimens, but did not seem to affect the results of antigen detection in the TKK 2nd kit. CONCLUSION: The results showed that the TKK 2nd kit successfully detected all four DENV serotypes in clinical specimens and confirmed the potential of the kit for dengue diagnosis in endemic countries.

Overview of avian influenza virus in urban feral pigeons in Bangkok, Thailand.

This survey assessed the presence of avian influenza virus (AIV) in urban feral pigeons (UFPs) in Bangkok, Thailand. A total of 485 UFPs were collected from eight study sites, and blood, tracheal, and cloacal samples were collected from each bird. Virus isolation and molecular methods did not detect AIV in any of the birds tested. A hemagglutination inhibition test was used to test for antibodies to high and low pathogenicity AIV subtypes. AIV subtype H9 antibodies were the only antibodies detected. The overall seroprevalence of AIV subtype H9 antibodies was 6.9%, and subtype H9 antibodies were found in UFPs at all eight sites. The overall geometric mean titer was 11.07 (range: 8-64). These results reveal that UFPs in Bangkok do not currently pose a risk of transmitting AIV to humans. However, monitoring of AIV in UFPs is necessary for disease control and to minimize the possibility of influenza outbreaks.

Serologic evidence of pandemic (H1N1) 2009 virus infection in camel and Eld's deer, Thailand.

BACKGROUND AND AIM: The pandemic (H1N1) 2009 influenza (H1N1pdm09) virus has affected both human and animal populations worldwide. The transmission of the H1N1pdm09 virus from humans to animals is increasingly more evident. Captive animals, particularly zoo animals, are at risk of H1N1pdm09 virus infection through close contact with humans. Evidence of exposure to the H1N1pdm09 virus has been reported in several species of animals in captivity. However, there is limited information on the H1N1pdm09 virus infection and circulation in captive animals. To extend the body of knowledge on exposure to the H1N1pdm09 virus among captive animals in Thailand, our study investigated the presence of antibodies against the H1N1pdm09 virus in two captive animals: Camelids and Eld's deer. MATERIALS AND METHODS: We investigated H1N1pdm09 virus infection among four domestic camelid species and wild Eld's deer that were kept in different zoos in Thailand. In total, 72 archival serum samples from camelid species and Eld's deer collected between 2013 and 2014 in seven provinces in Thailand were analyzed for influenza antibodies using hemagglutination inhibition (HI), microneutralization, and western blotting (WB) assays. RESULTS: The presence of antibodies against the H1N1pdm09 virus was detected in 2.4% (1/42) of dromedary camel serum samples and 15.4% (2/13) of Eld's deer serum samples. No antibodies were detected in the rest of the serum samples derived from other investigated camelids, including Bactrian camels (0/3), alpacas (0/5), and llamas (0/9). The three positive serum samples showed HI antibody titers of 80, whereas the neutralization titers were in the range of 320-640. Antibodies specific to HA and NP proteins in the H1N1pdm09 virus were detected in positive camel serum samples using WB. Conversely, the presence of the specific antibodies in the positive Eld's deer serum samples could not be determined using WB due to the lack of commercially labeled secondary antibodies. CONCLUSION: The present study provided evidence of H1N1pdm09 virus infection in the captive dromedary camel and Eld's deer in Thailand. Our findings highlight the need for continuous surveillance for influenza A virus in the population of dromedary camels and Eld's deer. The susceptible animal populations in close contact with humans should be closely monitored. Further study is warranted to determine whether Eld's deer are indeed a competent reservoir for human influenza virus.

The occurrence of elephant endotheliotropic herpesvirus infection in wild and captive Asian elephants in Thailand: Investigation based on viral DNA and host antibody.

BACKGROUND AND AIM: Elephant endotheliotropic herpesvirus (EEHV) is a serious disease, threatening the life of young elephants. Many elephants have been infected with no clinical signs and may serve as carriers spreading this disease. It is important to monitor the disease through clinical signs and molecular diagnosis. In this study we investigated the occurrence of EEHV and the efficiency of different techniques used to monitor EEHV infection in various samples and populations of Asian elephants. MATERIALS AND METHODS: Blood and trunk swabs were collected from live elephants, while visceral organs (lung, digestive tract, spleen, lymph nodes, and kidney) were collected from dead elephants. EEHV was detected by polymerase chain reaction (PCR) in whole blood, trunk swabs, and visceral organs as samples, while elephant anti-EEHV immunoglobulin G (IgG) in serum was detected by enzyme-linked immunosorbent assay (ELISA). A total of 162 samples were analyzed in this study: 129 from healthy, 26 from dead, and 7 from sick elephants. RESULTS: The present study showed that the overall incidence of EEHV was 40.1% (n=65/162). Approximately 46.2% (n=12/26) and 85.7% (n=6/7) of dead and sick elephants were positive for EEHV by PCR, respectively. All sick elephants that were young and affected by EEHV clinical disease tested negative for the IgG antibody ELISA, suggesting primary EEHV infection in this group. In addition, 2.3% (n=3/129) of subclinical infections were detected using PCR, and trunk swab samples showed slightly higher sensitivity (5.3%, n=2/38) to detect EEHV than whole blood (1.2%, n=1/84). As many as, 48.4% (n=44/91) of healthy elephants were EEHV seropositive (ELISA-positive), suggesting that many elephants in Thailand had previously been infected. Overall, 30% of dead wild elephants had been infected with EEHV (n=3/10). Moreover, statistical analysis revealed no significant differences in the EEHV detection rate between different age groups or sexes (p>0.05). CONCLUSION: PCR is better than ELISA to detect EEHV active infection in dead/sick elephants and to monitor EEHV in young elephants. ELISA is suitable for detecting previous EEHV infection and carriers, particularly adults. Theoretically, we could use both PCR and ELISA to increase the sensitivity of testing, along with observing abnormal behavior to efficiently monitor this disease. Identification of EEHV carriers within elephant populations is important to prevent transmission to healthy individuals, especially young elephants with high mortality from EEHV. This is the first report from Thailand regarding EEHV infection in wild elephants, showing the importance of preventing disease transmission between captive and wild elephants.

Rickettsiae exposure related to habitats of the oriental house rat (Rattus tanezumi, Temminck, 1844) in Salaya suburb, Thailand.

Rickettsial zoonotic diseases, in particular scrub typhus, murine typhus, and tick typhus, are caused by Orientia tsutsugamushi, Rickettsia typhi, and Rickettsia honei infections. Rickettsiae exposure is typically related to a rodent host in various habitats of marginal regions, including between rural areas and communities such as the Salaya suburb. This allows the oriental house rat (OHR), a highly adaptive species, to live in close proximity to the community and possibly introduce rickettsial diseases. To understand rickettsial exposure in the OHR from different habitats, knowledge of disease exposure can serve as baseline information for disease management and prevention. A total of 185 OHRs from three unrelated habitats were assessed using a standard indirect immunofluorescence assay (IFA) for specific antibody reaction to O. tsutsugamushi, R. typhi, and R. honei. The presence of antibody in the OHR to rickettsiae, either scrub or murine typhus, was associated with the habitat, whereas tick typhus had general exposure. This finding shows the OHR to be a potential reservoir host for rickettsial diseases along the borders of geographic regions in the suburban environment.

Antibody reaction of leptospirosis in asymptomatic feral boars, Thailand.

AIMS: This study aimed to determine the proportion of exposure to leptospirosis and evaluate the degree of serovar antibody reaction in feral boars. MATERIALS AND METHODS: A total of 58 sera obtained from feral boars in Khao Prathab Chang Wildlife Breeding Center, Ratchaburi, Thailand, were screened for leptospirosis exposure by microscopic agglutination test, conducted with a reference panel of 23 pathogenic serovars and a non-pathogenic serovar. RESULTS: Overall exposure rate of 62.07% was found in the studied population. An antibody reaction presented in 18 of 24 leptospiral serovars. Among the seropositive, Ballum serovar showed predominant exposure in the feral boar population. CONCLUSION: The results show a relatively high exposure to leptospirosis and the predominant serovar was Ballum followed by Canicola, the first finding in feral boars in Thailand. It has been revealed that feral boars act as a natural reservoir host of leptospirosis. There should be more concern about public health problems in leptospirosis arising where feral boars appear.

A LONG-TERM SEROSURVEY OF AVIAN INFLUENZA H5 AMONG WILD BIRDS IN NAKHON SAWAN PROVINCE, THAILAND.

An outbreak of HPAIV H5N1 in Nakhon Sawan province, Thailand, in 2004 caused sporadic deaths of Asian openbill storks ( Anastomus oscitans). An investigation was undertaken to determine if this virus occurs and circulates in wild birds in Nakhon Sawan province. Following the outbreak, a widespread serosurvey was conducted using the hemagglutination inhibition assay and microneutralization assay to detect antibodies against AIV H5. From 2007 to 2014, blood was collected from a total of 753 wild birds, representing 10 orders and 44 species. The results reveal that 10 serum samples were positive for AIV H5 antibodies. These seropositive results, found in the orders Ciconiiformes and Anseriformes, demonstrate that waterfowl serve as a reservoir host of AIV. Moreover, the seroprevalences in streak-eared bulbul showed habitat sharing with waterfowl or duck.

Detection and characterization of Chlamydophila psittaci in asymptomatic feral pigeons (Columba livia domestica) in central Thailand.

OBJECTIVE: To detect and characterize Chlamydophila psittaci (C. psittaci) in asymptomatic feral pigeons in central Thailand. METHODS: A total 814 swabs from the trachea and cloacae of 407 non-clinical feral pigeons in central Thailand were collected and tested for the presence of C. psittaci. RESULTS: A 10.8% of feral pigeons in the sample group were positive as determined by nested PCR primer specific to C. psittaci. The outer membrane protein A (ompA) gene of positive samples exhibited amino acid identity of C. psittaci ranging from 71 to 100% and were grouped in genotype B. Exceptionally, BF1676-56 isolate was closely related to Chlamydia avium with 99% identification of the 16S ribosomal (r) RNA gene. CONCLUSIONS: This is the first report on C. psittaci isolated from asymptomatic feral pigeons in Thailand, which provides knowledge for the disease status in pigeon populations in Thailand.

Detection and phylogenetic characterization of hepatitis E virus genotype 3 in a captive wild boar in Thailand.

Hepatitis E virus (HEV) was studied in different types of wild boar captive settings in Thailand, including a wildlife breeding research station, zoo, and commercial wild boar farm, which were located in different locations of Thailand. Fifty-one fecal samples were collected and screened for HEV RNA and then analyzed. One sample obtained from a wildlife breeding research station in Ratchaburi province was HEV positive. Phylogenetic characterization revealed that the virus was HEV genotype 3 and belongs to subgroup 3e, which is closely related to HEV recently isolated from domestic pigs and humans in the country. It was hypothesized that HEV is shared among wild boars, domestic pigs, and humans in Thailand.

Genetic analysis of beak and feather disease virus isolated from captive psittacine birds in Thailand.

Beak and feather disease virus (BFDV) is a causative agent of psittacine beak and feather disease. Genome sequences of BFDVs isolated from Thailand have not hitherto been reported. The whole genomes of 17 BFDV isolates, obtained from 12 psittacine genera, were amplified and subjected to direct sequencing revealing a length ranging from 1,990 to 2,015 nucleotides. The predicted open reading frames (ORFs) in the viral genome varied from four to six. Only ORF1, ORF2, and ORF5 were found in all isolates. Deduced amino acid sequences of BFDV ORF2 were used to construct a phylogenetic tree. The phylogram grouped BFDV into ten clusters, which showed either host species relationship or regional restriction. The Thai isolates, were grouped into three clusters, cluster I, II, and V. Cluster I and II showed restricted geographical region to Thailand, and cluster II also showed a close relationship with BFDV isolated from Australia. Cluster V demonstrated neither restricted region nor species specificity of birds. In this cluster, there was an insertion of 16 nucleotides at non coding region of all BFDV isolates. The genetic information obtained from this study can be used to help understand BFDV diversity and evolution in Thailand.

Comparison of outbreaks of H5N1 highly pathogenic avian influenza in wild birds and poultry in Thailand.

Wild bird surveillance for highly pathogenic avian influenza (HPAI) H5N1 virus from 2004 to 2007 in Thailand indicated that the prevalence of infection with avian influenza H5N1 virus in wild birds was low (1.0%, 95% confidence interval [CI]: 0.7-1.2, 60/6,263 pooled samples). However, the annual prevalence varied considerably over this period, with a peak of 2.7% (95% CI: 1.4, 4.1) in 2004. Prevalence dropped to 0.5% (95% CI: 0.3, 0.8]) and 0.6% (95% CI: 0.3, 1.0) in 2005 and 2006, respectively, and then increased to 1.8% (95% CI: 1.0, 2.6) in 2007. During this period, 16 species from 12 families of wild birds tested positive for H5N1 virus infection. All samples from juvenile birds were negative for H5N1 virus, whereas 0.6% (95% CI: 0.4, 0.9) of pooled samples from adult birds were positive. Most positive samples originated from peridomestic resident species. Infected wild bird samples were only found in provinces where poultry outbreaks had occurred. Detection of H5N1 virus infection in wild birds was reported up to 3 yr after eradication of the poultry outbreaks in those provinces. As observed with outbreaks in poultry, the frequencies of H5N1 outbreaks in wild birds were significantly higher in winter. Further understanding of the mechanisms of persistence and ongoing HPAI H5N1 transmission between wild birds and domestic poultry is needed.