Behavioral-biological surveillance of emerging infectious diseases among a dynamic cohort in Thailand.

BACKGROUND: Interactions between humans and animals are the key elements of zoonotic spillover leading to zoonotic disease emergence. Research to understand the high-risk behaviors associated with disease transmission at the human-animal interface is limited, and few consider regional and local contexts. OBJECTIVE: This study employed an integrated behavioral-biological surveillance approach for the early detection of novel and known zoonotic viruses in potentially high-risk populations, in an effort to identify risk factors for spillover and to determine potential foci for risk-mitigation measures. METHOD: Participants were enrolled at two community-based sites (n = 472) in eastern and western Thailand and two hospital (clinical) sites (n = 206) in northeastern and central Thailand. A behavioral questionnaire was administered to understand participants' demographics, living conditions, health history, and animal-contact behaviors and attitudes. Biological specimens were tested for coronaviruses, filoviruses, flaviviruses, influenza viruses, and paramyxoviruses using pan (consensus) RNA Virus assays. RESULTS: Overall 61/678 (9%) of participants tested positive for the viral families screened which included influenza viruses (75%), paramyxoviruses (15%), human coronaviruses (3%), flaviviruses (3%), and enteroviruses (3%). The most salient predictors of reporting unusual symptoms (i.e., any illness or sickness that is not known or recognized in the community or diagnosed by medical providers) in the past year were having other household members who had unusual symptoms and being scratched or bitten by animals in the same year. Many participants reported raising and handling poultry (10.3% and 24.2%), swine (2%, 14.6%), and cattle (4.9%, 7.8%) and several participants also reported eating raw or undercooked meat of these animals (2.2%, 5.5%, 10.3% respectively). Twenty four participants (3.5%) reported handling bats or having bats in the house roof. Gender, age, and livelihood activities were shown to be significantly associated with participants' interactions with animals. Participants' knowledge of risks influenced their health-seeking behavior. CONCLUSION: The results suggest that there is a high level of interaction between humans, livestock, and wild animals in communities at sites we investigated in Thailand. This study highlights important differences among demographic and occupational risk factors as they relate to animal contact and zoonotic disease risk, which can be used by policymakers and local public health programs to build more effective surveillance strategies and behavior-focused interventions.

An Efficient DNA Extraction for a Blue Xestospongia sp. Sponge and Its Associated Microorganisms Containing Cytotoxic Substances.

Extraction of high quantity and quality DNAs from marine sponges, which contain diverse and abundant microbial communities, is important to molecular biology techniques for the analysis of nucleic acids. Several marine sponges and their associated microorganisms have been known to produce cytotoxic natural products on several cancer cell lines via DNA damage mechanisms. These marine cytotoxic substances might be one of the factors that cause the low quantity and quality of DNAs during the DNA extraction from its living origin. Therefore, the extraction of DNA of a Thai blue marine sponge Xestospongia sp. with sufficient purity and quantity for molecular study can be challenging. In this study, we developed an efficient extraction method to prepare DNAs from a Thai blue marine sponge Xestospongia sp. which accumulated a highly potent cytotoxic alkaloid with DNA-damaging activity, named Renieramycin M (RM), as a major constituent in high quantity. We demonstrated that removal of RM from the sponge samples by a simple methanolic extraction before DNA extraction dramatically increased the yield and purity of DNAs compared to the RM-unremoved sponge samples. High molecular weight (HMW) genomic DNA was obtained from sponge samples with 8 times of RM elimination by using modified NaOAc salting-out extraction method. The quantity and quality of the prepared DNAs were comparatively determined via spectrophotometry, electrophoresis, and 16S rRNA gene amplification. Our result suggests that the removal of DNA-damaging constituents from the samples is a crucial step and must be seriously taken as the necessary consideration for the practical protocol of DNA extraction.

Detection of influenza virus in rectal swabs of patients admitted in hospital for febrile illnesses in Thailand.

INTRODUCTION: Influenza virus favours the respiratory tract as its primary site of host entry and replication, and it is transmitted mainly via respiratory secretions. Nasopharyngeal swab is the gold standard specimen type for influenza detection, but several studies have also suggested that the virus replicates in the human gastrointestinal tract. METHODS: A retrospective study was conducted on all patients positive for influenza virus and initially recruited as part of the PREDICT project from 2017 to 2018. The objectives of the study were to investigate whether rectal swab could aid in improving influenza detection, and if there was any correlation between gastrointestinal disturbances and severity of infection, using length of hospital stay as an indicator of severity. RESULTS: Of the 51 influenza-positive patients, 12 had detectable influenza virus in their rectal swab. Among these 12 rectal swab positive patients, influenza virus was not detected in the nasopharyngeal swab of three of them. Gastrointestinal symptoms were observed for 28.2% patients with a negative rectal swab negative and 25.0% patients with a positive rectal swab. Average length of hospital stay was 4.2 days for rectal swab positive group and 3.7 days for rectal swab negative group. This difference was not statistically significant (p = 0.288). CONCLUSIONS: There is no correlation between influenza virus detection in rectal swab and gastrointestinal disturbances or disease severity, and there is currently insufficient evidence to support replicative ability in the gastrointestinal tract.

First Complete Genome Sequence of Human Coronavirus HKU1 from a Nonill Bat Guano Miner in Thailand.

Human coronavirus HKU1 (HCoV-HKU1) was first detected in a patient with viral pneumonia from Hong Kong in 2004. Here, we report the first complete genome sequence of HCoV-HKU1 from Thailand, obtained from a nonill person who worked in a bat cave. Phylogenetic tree analysis revealed it as a group B HCoV-HKU1.

Full-length genome analysis of two genetically distinct variants of porcine epidemic diarrhea virus in Thailand.

Porcine epidemic diarrhea virus (PEDV) has continued to cause sporadic outbreaks in Thailand since 2007 and a pandemic variant containing an insertion and deletion in the spike gene was responsible for outbreaks. In 2014, there were further outbreaks of the disease occurring within four months of each other. In this study, the full-length genome sequences of two genetically distinct PEDV isolates from the outbreaks were characterized. The two PEDV isolates, CBR1/2014 and EAS1/2014, were 28,039 and 28,033 nucleotides in length and showed 96.2% and 93.6% similarities at nucleotide and amino acid levels respectively. In total, we have observed 1048 nucleotide substitutions throughout the genome. Compared to EAS1/2014, CBR1/2014 has 2 insertions of 4 ((56)GENQ(59)) and 1 ((140)N) amino acid positions 56-59 and 140, and 2 deletions of 2 ((160)DG(161)) and 1 ((1199)Y) amino acid positions 160-161 and 1199. The phylogenetic analysis based on full-length genome of CBR1/2014 isolate has grouped the virus with the pandemic variants. In contrast, EAS1/2014 isolate was grouped with CV777, LZC and SM98, a classical variant. Our findings demonstrated the emergence of EAS1/2014, a classical variant which is novel to Thailand and genetically distinct from the currently circulating endemic variants. This study warrants further investigations into molecular epidemiology and genetic evolution of the PEDV in Thailand.

Complete Genome Sequences of Two Genetically Distinct Variants of Porcine Epidemic Diarrhea Virus in the Eastern Region of Thailand.

Porcine epidemic diarrhea virus (PEDV) has continued to cause sporadic outbreaks in Thailand since 2007. Previously, PEDV in Thailand was a new variant containing an insertion and deletion in the spike gene. Herein, full-length genome sequences are reported for two variants of PEDV isolates from pigs displaying diarrhea in Thailand.

Replacement of a quinone by a 5-O-acetylhydroquinone abolishes the accidental necrosis inducing effect while preserving the apoptosis-inducing effect of renieramycin M on lung cancer cells.

Renieramycin M (1), a bistetrahydroisoquinolinequinone alkaloid isolated from the marine sponge Xestospongia sp., has been reported to possess promising anticancer effects. However, its accidental necrosis inducing effect has limited further development due to concerns of unwanted toxicity. The presence of two quinone moieties in its structure was demonstrated to induce accidental necrosis and increase reactive oxygen species (ROS) levels. Therefore, one quinone of 1 was modified to produce the 5-O-acetylated hydroquinone derivative (2), and 2 dramatically reduced the accidental necrosis inducing effect while preserving the apoptosis-inducing effect of parent 1 on lung cancer H23 cells. Addition of the antioxidant N-acetylcysteine suppressed the accidental necrosis mediated by 1, suggesting that its accidental necrosis inducing effect was ROS-dependent. The fluorescent probe dihydroethidium revealed that the accidental necrosis mediated by 1 was due to its ability to generate intracellular superoxide anions. Interestingly, the remaining quinone in 2 was required for its cytotoxicity, as the 5,8,15,18-O-tetraacetylated bishydroquinone derivative (3) exhibited weak cytotoxicity compared to 1 and 2. The present study demonstrates a simple way to eliminate the undesired accidental necrosis inducing effect of substances that may be developed as improved anticancer drug candidates.