Seroprevalence of anti-SARS-CoV-2 antibodies and associated factors among household contacts of COVID-19 confirmed cases in Bangkok, Thailand.

Background: High COVID-19 transmission among household (HH) contacts of infected cases were reported with seroprevalence varying from 5.5% to 57.2% worldwide. Data on seroprevalence among HH contacts and factors associated with seropositivity in Thailand are limited. Objectives: To determine the seroprevalence and factors associated with anti-SARS-CoV-2 antibodies among HH contacts of COVID-19 confirmed cases. Materials and methods: Data on confirmed COVID-19 cases (primary cases) in Bangkok from March 2020-July 2021 were retrieved from Institute for Urban Disease Control and Prevention. Primary cases were contacted within 14 days of testing positive for permission to contact their HH contacts via telephone. HH contacts were then recruited to complete questionnaires about demographics, and risk factors and blood was collected and tested for total immunoglobulin antibody against SARS-CoV-2 spike S1 protein. Factors associated with seropositivity were analysed by logistic regression. Results: Eligible participants of 452 HH contacts of infected cases in Bangkok were contacted. Seroprevalence was 20.5% among HH contacts. Factors associated with seropositivity after multivariate analysis were relationship to index case (being other relatives to index case (other than close relatives/spouse) [aOR 4.04, 95% CI; 1.15, 14.14, p.029] and being a co-worker to index cases [aOR 0.16, 95% CI; 0.045, 0.60, p.006]), always staying in the same room with index case [aOR 5.64, 95% CI; 1.95, 16.34, p.001], sharing utensil [aOR 0.25, 95% CI; 0.074, 0.82, p.023], and participation in leisure activities together with index case [aOR 4.77, 95% CI; 1.47, 15.51, p.009]. Conclusion: Serological investigation can be used in detecting COVID-19 infection in conjunction with other molecular techniques. It is a useful tool for studies on seroprevalence in a population as well as seroconversion after a vaccination campaign. Sharing living environments are associated with seropositivity in HH contacts. Nevertheless, individual practices can be affected by awareness, cultural differences, and control measures implemented by each country.

COVID-19 Transmission among Healthcare Workers at a Quarantine Facility in Thailand: Genomic and Outbreak Investigations.

During the COVID-19 pandemic, Thailand implemented a quarantine program at approved quarantine facilities for every international traveler. Here, we report an epidemiological and genomic investigation of a COVID-19 cluster consisting of seven healthcare workers (HCWs) at a quarantine facility and its partnered hospital in Thailand. Outbreak investigations were implemented to obtain contact tracing data and to establish chains of transmission. Genomic sequencing of SARS-CoV-2 with samples within the cohort was performed. Investigations of 951 HCWs and staff with quarantined travelers were implemented to determine the chain of transmission. Genomic and outbreak investigations identified the international travelers infected with the B.1.1.31 SARS-CoV-2 lineage as the source of this outbreak. The genomic data and the investigated timeline revealed a putative transmission chain among HCWs, pointing toward the transmission via the use of common living quarters at the investigated quarantine site. The evaluation of this cohort has led to a policy recommendation on quarantine facility management. International travel quarantine is an important strategy to contain importation of COVID-19 cases. However, a quarantine facility is likely to become a potential hotspot, requiring thorough preventive measures. Reducing the exposure risk by providing private living quarters and scheduling clinical duties at a quarantine site separated from the conventional healthcare workforce have been implemented.

Molecular epidemiology of the first wave of severe acute respiratory syndrome coronavirus 2 infection in Thailand in 2020.

The coronavirus disease 2019 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a major global concern. Several SARS-CoV-2 gene mutations have been reported. In the current study associations between SARS-CoV-2 gene variation and exposure history during the first wave of the outbreak in Thailand between January and May 2020 were investigated. Forty samples were collected at different time points during the outbreak, and parts of the SARS-CoV-2 genome sequence were used to assess genomic variation patterns. The phylogenetics of the 40 samples were clustered into L, GH, GR, O and T types. T types were predominant in Bangkok during the first local outbreak centered at a boxing stadium and entertainment venues in March 2020. Imported cases were infected with various types, including L, GH, GR and O. In southern Thailand introductions of different genotypes were identified at different times. No clinical parameters were significantly associated with differences in genotype. The results indicated local transmission (type T, Spike protein (A829T)) and imported cases (types L, GH, GR and O) during the first wave in Thailand. Genetic and epidemiological data may contribute to national policy formulation, transmission tracking and the implementation of measures to control viral spread.

Detection of Babesia canis vogeli and Hepatozoon canis in canine blood by a single-tube real-time fluorescence resonance energy transfer polymerase chain reaction assay and melting curve analysis.

A real-time fluorescence resonance energy transfer polymerase chain reaction (qFRET PCR) coupled with melting curve analysis was developed for detection of Babesia canis vogeli and Hepatozoon canis infections in canine blood samples in a single tube assay. The target of the assay was a region within the 18S ribosomal RNA gene amplified in either species by a single pair of primers. Following amplification from the DNA of infected dog blood, a fluorescence melting curve analysis was done. The 2 species, B. canis vogeli and H. canis, could be detected and differentiated in infected dog blood samples (n = 37) with high sensitivity (100%). The detection limit for B. canis vogeli was 15 copies of a positive control plasmid, and for H. canis, it was 150 copies of a positive control plasmid. The assay could simultaneously distinguish the DNA of both parasites from the DNA of controls. Blood samples from 5 noninfected dogs were negative, indicating high specificity. Several samples can be run at the same time. The assay can reduce misdiagnosis and the time associated with microscopic examination, and is not prone to the carryover contamination associated with the agarose gel electrophoresis step of conventional PCR. In addition, this qFRET PCR method would be useful to accurately determine the range of endemic areas or to discover those areas where the 2 parasites co-circulate.

Detection of Ehrlichia canis in canine blood samples by real-time fluorescence resonance energy transfer (FRET) PCR and melting curve analysis.

Ehrlichia canis is a small pleomorphic gram-negative, coccoid, obligatory intracellular bacterium and the cause of canine monocytic ehrlichiosis. A real-time fluorescence resonance energy transfer polymerase chain reaction (real-time FRET PCR) coupled with melting curve analysis was established for detection of E. canis infection in canine blood samples. The VirB9 gene was amplified using one pair of primers and the melting curve analysis was generated by heating the hybridizing probes and amplified products. Eight E. canis-infected dog blood samples were initially identified using the Giemsa staining/microscopic method followed by conventional PCR (cPCR)/Sanger sequencing for confirmation. The sensitivity and specificity of the real-time FRET PCR detection were 87.5% and 100%, respectively and the limit of detection was 6.6 x 10(3) copies of positive E. canis control plasmids. The real-time FRET PCR with melting curve analysis reported here is better than microscopic visualization or cPCR because the method is not affected by the false bias inherent in the microscopic method. Furthermore, many samples can be processed rapidly at the same time. This convenient tool is beneficial as an alternative assay for the epidemiologic study of canine ehrlichiosis as well as for eradication of these organisms in prevention and control programs in endemic areas.

High throughput pyrosequencing technology for molecular differential detection of Babesia vogeli, Hepatozoon canis, Ehrlichia canis and Anaplasma platys in canine blood samples.

Canine babesiosis, hepatozoonosis, ehrlichiosis, and anaplasmosis are tick-borne diseases caused by different hemopathogens. These diseases are causes of morbidity and mortality in dogs. The classic method for parasite detection and differentiation is based on microscopic observation of blood smears. The limitations of the microscopic method are that its performance requires a specially qualified person with professional competence, and it is ineffective in differentiating closely related species. This study applied PCR amplification with high throughput pyrosequencing for molecular differential detection of the following 4 hemoparasites common to tropical areas in dog blood samples: Babesia vogeli, Hepatozoon canis, Ehrlichia canis, and Anaplasma platys. PCR was initially used to amplify specific target regions of the ribosomal RNA genes of each parasite using 2 primer pairs that included 18S rRNA for protozoa (B. vogeli and H. canis) and 16S rRNA for rickettsia (E. canis and A. platys). Babesia vogeli and H. canis were discriminated using 9 nucleotide positions out of 30 base pairs, whereas E. canis and A. platys were differentiated using 15 nucleotide positions out of 34 base pairs that were determined from regions adjacent to 3' ends of the sequencing primers. This method provides a challenging alternative for a rapid diagnosis and surveillance of these tick-borne diseases in canines.

Molecular differentiation of Schistosoma japonicum and Schistosoma mekongi by real-time PCR with high resolution melting analysis.

Human schistosomiasis caused by Schistosoma japonicum and Schistosoma mekongi is a chronic and debilitating helminthic disease still prevalent in several countries of Asia. Due to morphological similarities of cercariae and eggs of these 2 species, microscopic differentiation is difficult. High resolution melting (HRM) real-time PCR is developed as an alternative tool for the detection and differentiation of these 2 species. A primer pair was designed for targeting the 18S ribosomal RNA gene to generate PCR products of 156 base pairs for both species. The melting points of S. japonicum and S. mekongi PCR products were 84.5±0.07℃ and 85.7±0.07℃, respectively. The method permits amplification from a single cercaria or an egg. The HRM real-time PCR is a rapid and simple tool for differentiation of S. japonicum and S. mekongi in the intermediate and final hosts.