Cross-correlation and time series analysis of rabies in different animal species in Nepal from 2005 to 2018.

Rabies is a fatal zoonotic disease, resulting in human and livestock deaths. In Nepal, animal rabies has posed a significant challenge to public health. Because animals are the primary source of rabies in humans, a better understanding of rabies epidemiology in animals is necessary. The objectives of this study were to determine the correlation between rabies occurrences in dogs and livestock animals and to detect the trends and change points of the disease using longitudinal data. The nationwide rabies dataset from 2005 to 2018 was analyzed using cross-correlation, multiple change points, and time series methods. Autoregressive Integrated Moving Average (ARIMA) and Neural Network Autoregression (NNAR) were applied to the time series data. The results show a positive correlation between canine rabies and livestock rabies occurrences. Three significant change points were detected in the time series data, demonstrating that the occurrences were high in the initial years but stabilized before peaking to an upward trend in the final years of the study period. Nonetheless, there was no seasonality pattern in rabies occurrences. The most suitable models were ARIMA (2,1,2) and NNAR (5,1,4) (12). Based on the study findings, both locals and tourists in Nepal need to have enhanced awareness of the potential dangers posed by rabies in canines and livestock. This study offers much-needed insight into the patterns and epidemiology of animal rabies which will be helpful for policymakers in drafting rabies control plans for Nepal.

Impacts of the Pandemic, Animal Source Food Retailers' and Consumers' Knowledge and Attitudes toward COVID-19, and Their Food Safety Practices in Chiang Mai, Thailand.

The COVID-19 pandemic affected the food supply chain, retailers, and consumers owing to infection awareness. This study evaluated the impacts COVID-19 on ASF retailers' businesses and consumers' livelihoods, as well as their knowledge toward the disease, attitudes, and food safety practices to prevent infections. The study includes a cross-sectional component that was conducted in urban/peri-urban (U/PU) and rural areas in Chiang Mai province. In another part of the study, a structured questionnaire was developed for animal source food (ASF) retailers and consumers, with three primary parts for data analysis: general information, COVID-19 impacts, and knowledge, attitudes, and practices (KAP) assessment. Data corresponding to three periods of interest (before the COVID-19 outbreak, during partial lockdown, and present) were gathered and analyzed. In this study, 155 retailers and 150 consumers participated, of which the majority of the respondents were female (70.3% and 82.7%, respectively) with average ages of 47.4 and 44.9 years, respectively. The most noticeable effect of COVID-19 was a decline in income for retailers and consumers. The KAP scores of consumers in both areas were not significantly different, whereas the retailer attitudes toward COVID-19 prevention and food safety practices scored more highly in rural areas than in U/PU. During the partial lockdown, food safety practices significantly improved relative to the time preceding the outbreak, and these practices have remained constant to the present day. The results revealed that gender, age group, business type, and type of ASF retailers were associated with the KAP of the retailers, whereas gender, age group, education, number of family members, and occupation were associated with the KAP of the consumers. Our findings provide in-depth information about the effects of COVID-19 on ASF retailers and consumers, as well as their KAP regarding the outbreak and food safety, which may serve as support in developing policies for improved health and food safety.

A Cutoff Determination of Real-Time Loop-Mediated Isothermal Amplification (LAMP) for End-Point Detection of Campylobacter jejuni in Chicken Meat.

Campylobacter jejuni is one of the leading causes of foodborne illness worldwide. C. jejuni is commonly found in poultry. It is the most frequent cause of contamination and thus resulting in not only public health concerns but also economic impacts. To test for this bacterial contamination in food processing plants, this study attempted to employ a simple and rapid detection assay called loop-mediated isothermal amplification (LAMP). The best cutoff value for the positive determination of C. jejuni calculated using real-time LAMP quantification cycle (Cq) was derived from the receiver operating characteristic (ROC) curve modeling. The model showed an area under curve (AUC) of 0.936 (95% Wald CI: 0.903-0.970). Based on Youden's J statistic, the optimal cutoff value which had the highest sensitivity and specificity from the model was calculated as 18.07. The LAMP assay had 96.9% sensitivity, 95.8% specificity, and 93.9 and 97.9% positive and negative predictive values, respectively, compared to a standard culture approach for C. jejuni identification. Among all non-C. jejuni strains, the LAMP assay gave each of 12.5% false-positive results to C. coli and E. coli (1 out of 8 samples). The assay can detect C. jejuni at the lowest concentration of 103 CFU/mL. Our results suggest a preliminary indicator for the application of end-point LAMP assays, such as turbidity and UV fluorescence tests, to detect C. jejuni in field operations. The LAMP assay is an alternative screening test for C. jejuni contamination in food samples. The method provides a rapid detection, which requires only 9 min with a cutoff value of Cq. We performed the extraction of DNA from pure cultures and the detection of C. jejuni using the LAMP assay within 3 h. However, we were not able to reduce the time for the process of enrichment involved in our study. Therefore, we suggest that alternative enrichment media and rapid DNA extraction methods should be considered for further study. Compared to other traditional methods, our proposed assay requires less equipment and time, which is applicable at any processing steps in the food production chain.

Extended-spectrum beta-lactamase-producing Escherichia coli from pork in Muang district, Chiang Mai Province, Thailand.

Background and Aim: Contaminated pork is one of the transmission routes for pathogens. Extended-spectrum beta-lactamases (ESBL)-producing Escherichia coli is one of the critical threats to global public health. This study aimed to examine pork from different types of markets in Muang district, Chiangmai Province, Thailand, for a proportion of ESBL-producing E. coli, antibiotic resistance of ESBL-producing E. coli and ESBL-producing E. coli genotypes. Materials and Methods: Samples were collected from different market types; fresh markets, pork stores, and supermarkets, enriched and inoculated on selective media. Extended-spectrum beta-lactamases-producing E. coli was identified using double-disk diffusion method according to Clinical and Laboratory Standards Institute 2016. Antibiotic susceptibility test was performed through VITEK® System and ESBL-encoding genes were detected using a multiplex polymerase chain reaction. Results: About 69% of the samples were positive to ESBL-producing E. coli and showed high rates of resistance for ampicillin (100%), piperacillin (100%), cefalexin (100%), cefpodoxime (100%), cefovecin (100%) and ceftiofur (100%), gentamycin (89.86%), and tetracycline (TE) (84.06%). All isolates were multiple drug resistant; resistance patterns of beta-lactams, aminoglycosides, TEs, nitrobenzene derivatives, and sulfonamide groups were observed. The ESBL-producing E. coli-positive isolates carried blaCTX-M groups (100%), blaTEM (98.55%), and blaSHV (1.45%). None of blaOXA was found in this study. Conclusion: Extended-spectrum beta-lactamases-producing E. coli was found in various types of markets; all isolates were detected as multidrug-resistant. The dissemination of such strains can conceivably cause concerning public health, implying that supervised antimicrobial use in pork production and sanitary food preparation is recommended.

A Knowledge, Attitudes and Practices (KAP) survey on canine rabies prevention and control in four rural areas of Sri Lanka.

Rabies is a fatal viral disease that affects all mammals. It causes an estimated 59,000 human deaths worldwide annually. Dogs are the main reservoir and transmitter of rabies to human in Sri Lanka. Prevention and control measures include the mass vaccination of dogs and human post-exposure treatment. While these measures appear to be homogeneous across the country, there is a need to identify community-level gaps due to the decentralization of regional health and veterinary services in conducting such activities. A community-based questionnaire survey was conducted to identify potential risk factors and gaps related to knowledge, attitudes and practices on rabies prevention and control in the rural regions of Sri Lanka. Lower knowledge scores were associated with respondents between 18 and 38 years old, had no experience of being bitten, and own semi-independent dogs. Despite associated fear of stray dogs with health issues, some communities provide shelter and/or food for the stray dogs. Uptake of population control and vaccination of dogs across the grama niladhari divisions (GNDs), which are the smallest administrative units, differed. More than 80% of dogs were not desexed and community knowledge on vaccination delivery, booster and vaccination schedules were also variable. Even though 69% of respondents identified stray/community dogs as potential carriers' owners allow their dogs to roam the neighbourhood. Most of the respondents were able to identify the need for post-exposure treatment after a dog bite. However, post-exposure prophylaxis uptake by the local victims ranged between 83.00% and 87.50%. Low participation was reported in health education in the communities. Lack of knowledge on other potential carriers was similar to previous survey studies in the last decade. Collaborations between local communities, veterinary and medical services are necessary to scale up the control programme in order to achieve the global target of zero rabies deaths by 2030. Further community-level studies are recommended.

Evaluation of strategies using simulation model to control a potential outbreak of highly pathogenic avian influenza among poultry farms in Central Luzon, Philippines.

The Philippines confirmed its first epidemic of Highly Pathogenic Avian Influenza (HPAI) on August 11, 2017. It ended in November of 2017. Despite the successful management of the epidemic, reemergence is a continuous threat. The aim of this study was to conduct a mathematical model to assess the spatial transmission of HPAI among poultry farms in Central Luzon. Different control strategies and the current government protocol of 1 km radius pre-emptive culling (PEC) from infected farms were evaluated. The alternative strategies include 0.5km PEC, 1.5km PEC, 2 km PEC, 2.5 km PEC, and 3 km PEC, no pre-emptive culling (NPEC). The NPEC scenario was further modeled with a time of government notification set at 24hours, 48 hours, and 72 hours after the detection. Disease spread scenarios under each strategy were generated using an SEIR (susceptible-exposed-infectious-removed) stochastic model. A spatial transmission kernel was calculated and used to represent all potential routes of infection between farms. We assumed that the latent period occurs between 1-2 days, disease detection at 5-7 days post-infection, notification of authorities at 5-7 days post-detection and start of culling at 1-3 days post notification. The epidemic scenarios were compared based on the number of infected farms, the total number of culled farms, and the duration of the epidemic. Our results revealed that the current protocol is the most appropriate option compared with the other alternative interventions considered among farms with reproductive ratio (Ri) > 1. Shortening the culling radius to 0.5 km increased the duration of the epidemic. Further increase in the PEC zone decreased the duration of the epidemic but may not justify the increased number of farms to be culled. Nonetheless, the no-pre-emptive culling (NPEC) strategy can be an effective alternative to the current protocol if farm managers inform the government immediately within 24 hours of observation of the presence of HPAI in their farms. Moreover, if notification is made on days 1-3 after the detection, the scale and length of the outbreak have been significantly reduced. In conclusion, this study provided a comparison of various control measures for confronting the spread of HPAI infection using the simulation model. Policy makers can use this information to enhance the effectiveness of the current control strategy.

Identification of High-Risk Areas for the Spread of Highly Pathogenic Avian Influenza in Central Luzon, Philippines.

Highly pathogenic avian influenza virus (HPAIV) is a major problem in the poultry industry. It is highly contagious and is associated with a high mortality rate. The Philippines experienced an outbreak of avian influenza (AI) in 2017. As there is always a risk of re-emergence, efforts to manage disease outbreaks should be optimal. Linked to this is the need for an effective surveillance procedure to capture disease outbreaks at their early stage. Risk-based surveillance is the most effective and economical approach to outbreak management. This study evaluated the potential of commercial poultry farms in Central Luzon to transmit HPAI by calculating their respective reproductive ratios (R0). The reproductive number for each farm is based on the spatial kernel and the infectious period. A risk map has been created based on the calculated R0. There were 882 (76.63%) farms with R0 < 1. Farms with R0 ≥ 1 were all located in Pampanga Province. These farms were concentrated in the towns of San Luis (n = 12) and Candaba (n = 257). This study demonstrates the utility of mapping farm-level R0 estimates for informing HPAI risk management activities.

Qualitative risk assessment of transmission pathways of highly pathogenic avian influenza (HPAI) virus at live poultry markets in Dhaka city, Bangladesh.

Analysis of environmental samples obtained from the Live Poultry Markets (LPMs) of Dhaka City, Bangladesh, has revealed that the highest degree of prevalence of highly pathogenic avian influenza A (HPAI, H5N1), besides other subtypes of the LPAI virus, poses the plausible risk of transmission of these viruses between human and poultry species. The present study was conducted using the OIE risk analysis framework to assess the risk level of each pathway successively. The estimated risk parameters were integrated towards to obtain the overall risk level for each specific HPAI transmission pathway using the matrix adapted by Cristobel Zepeda accompanying other expert consultations. The relevant data obtained from published and unpublished sources, together with survey data of field observations, were used to formulate and confirm the risk pathways and their associated risks. The results revealed that the risk of the release of the HPAI virus was medium when exposure was high. Additionally, the consequence would be considered very high with a medium degree of uncertainty for all parameters. Ultimately, the overall risk for transmission was estimated as medium with a medium degree of uncertainty. The findings of this study reveal that there is a significant threat that HPAI virus transmission could occur among poultry and humans and effectively sustain within the environment of the LPMs. Our findings are primarily focused on public health considerations, the hygienic slaughter of poultry and the relevant cleaning and sanitation practices conducted in the LPMs to support evidence-based decision-making processes. The findings of the study have the potential to be used to formulate effective risk reduction measures and can be further adapted in low-resource settings without major infrastructural changes required of the LPMs. All of which would reduce the risk of HPAI virus release and further lessen the degree of exposure and transmission in established LPMs.

Detection of virus-neutralising antibodies and associated factors against rabies in the vaccinated household dogs of Kathmandu Valley, Nepal.

BACKGROUND: Rabies is a vaccine-preventable neglected tropical viral zoonosis. It occurs worldwide, creating a very heavy burden in many developing countries, including Nepal. Dogs are the principle vector for the transmission of this disease in urban areas. Vaccination is the most important preventive measure in areas where dogs are the principle source of infection. This study was conducted with the aim of detecting virus-neutralising antibodies and associated factors against rabies in vaccinated household dogs of Kathmandu valley. METHODS: Blood samples were collected from 110 vaccinated pet dogs in Kathmandu, Bhaktapur, and Lalitpur districts of Nepal. The samples were taken to the laboratory of the National Zoonosis and Food Hygiene Research Center where serum was separated. An indirect immune-enzymatic assay (PlateliaTM Rabies II kit ad usum Veterinarium, Biorad, China) was used for the detection of rabies virus anti-glycoprotein antibodies in the dog serum samples following the manufacturer's recommendations and instructions. Optical density values for unknown samples were compared with the positive sera titers in quantification tests obtained after a direct reading on the standard curve. Results were expressed as equivalent units per ml (EU/ml). FINDINGS: Of the total samples, 89.09% exceeded the required seroconversion level (≥ 0.5 EU/ml); another 9.09% did not reach the seroconversion level (0.125-0.5 EU/ml); and 1.81% had undetectable seroconversion levels (<0.125 EU/ml) suggesting that the animal had not seroconverted according to the PLATELIA™ RABIES II test. Only one factor, the condition under which the dog was kept, was significantly associated with the antibody titer level. No association was found for any of the other factors included in the study. INTERPRETATION: Vaccination is the most effective measure for prevention and control of rabies. The locally manufactured brand of vaccine, which is available in Nepal, is potent enough to generate a sufficient amount of protective antibodies, equal to international brands.

Genetic Characterization of Cambodian Fasciola gigantica and Dispersal Direction of the Species in Asia.

Fasciola gigantica and hybrid Fasciola are distributed throughout Asia. Herein, we investigated the species of the Fasciola fluke distributed in three hotspots of fascioliasis in Cambodia. A total of 92 flukes collected from 21 slaughtered cattle from Kandal (44), Battambang (41), and Kratie (7) Provinces were identified as F. gigantica using multiplex PCR for a nuclear phosphoenolpyruvate carboxykinase (PEPCK) gene. The overall prevalence of F. gigantica infestation was 7.14% (21/294). Phylogenetic as well as population genetics analyses were performed using the mitochondrial NADH dehydrogenase subunit 1 (ND1). The 19 ND1 haplotypes were identified from Cambodian F. gigantica (haplotype diversity, 0.83). All of the haplotypes were classified into F. gigantica haplogroup C, which includes ND1 haplotypes detected from Thailand, Vietnam, Indonesia, Myanmar, and China. Among haplogroup C, novel and unique haplotypes of Cambodia were found in the Battambang and Kandal Provinces, and the nucleotide diversity of the Cambodian population (0.00532) was the highest. Pairwise fixation indices among the F. gigantica populations from these countries indicated that the Cambodian and Thailand populations were related to each other. The highest genetic diversity in the Cambodian population suggests that F. gigantica in Cambodia may be the ancestor of the populations in Southeast Asian countries. Most likely, livestock movement, including Zebu cattle, played an important role in the transmission of F. gigantica. In this study, the hybrid Fasciola flukes that are commonly found in neighboring countries, were not found in Cambodia. Further comprehensive investigations of Fasciola prevalence should be conducted by analyzing a wider range of hosts throughout Cambodia to reach a more solid conclusion about the absence of hybrid flukes.

A Participatory System for Preventing Pandemics of Animal Origins: Pilot Study of the Participatory One Health Disease Detection (PODD) System.

BACKGROUND: Aiming for early disease detection and prompt outbreak control, digital technology with a participatory One Health approach was used to create a novel disease surveillance system called Participatory One Health Disease Detection (PODD). PODD is a community-owned surveillance system that collects data from volunteer reporters; identifies disease outbreak automatically; and notifies the local governments (LGs), surrounding villages, and relevant authorities. This system provides a direct and immediate benefit to the communities by empowering them to protect themselves. OBJECTIVE: The objective of this study was to determine the effectiveness of the PODD system for the rapid detection and control of disease outbreaks. METHODS: The system was piloted in 74 LGs in Chiang Mai, Thailand, with the participation of 296 volunteer reporters. The volunteers and LGs were key participants in the piloting of the PODD system. Volunteers monitored animal and human diseases, as well as environmental problems, in their communities and reported these events via the PODD mobile phone app. LGs were responsible for outbreak control and provided support to the volunteers. Outcome mapping was used to evaluate the performance of the LGs and volunteers. RESULTS: LGs were categorized into one of the 3 groups based on performance: A (good), B (fair), and C (poor), with the majority (46%,34/74) categorized into group B. Volunteers were similarly categorized into 4 performance groups (A-D), again with group A showing the best performance, with the majority categorized into groups B and C. After 16 months of implementation, 1029 abnormal events had been reported and confirmed to be true reports. The majority of abnormal reports were sick or dead animals (404/1029, 39.26%), followed by zoonoses and other human diseases (129/1029, 12.54%). Many potentially devastating animal disease outbreaks were detected and successfully controlled, including 26 chicken high mortality outbreaks, 4 cattle disease outbreaks, 3 pig disease outbreaks, and 3 fish disease outbreaks. In all cases, the communities and animal authorities cooperated to apply community contingency plans to control these outbreaks, and community volunteers continued to monitor the abnormal events for 3 weeks after each outbreak was controlled. CONCLUSIONS: By design, PODD initially targeted only animal diseases that potentially could emerge into human pandemics (eg, avian influenza) and then, in response to community needs, expanded to cover human health and environmental health issues.

Virulence characterization of Campylobacter jejuni isolated from resident wild birds in Tokachi area, Japan.

The prevalence of Campylobacter jejuni in wild birds is a potential hazard for human and animal health. The aim of this study was to establish the prevalence of C. jejuni in wild birds in Tokachi area, Hokkaido, Japan and investigate their virulence in vitro. In total, 173 cloacal swabs from individual wild birds were collected for the detection of Campylobacter spp. Thirty four samples (19.7%) were positive for Campylobacter of which 94.1% (32/34 samples) were C. jejuni. Additionally, one C. coli and one C. fetus were isolated. Seven C. jejuni isolates (one from crows and the other from pigeons) had important virulence genes including all three CDT genes (cdtA, cdtB and cdtC) and flaA, flaB, ciaB and cadF, and the other isolates were lacking cdtA gene. Further studies on in vitro virulence-associated phenotypes, such as motility assay on soft agar and invasion assay in Caco-2 cells, were performed. The wild bird C. jejuni isolates adhered and invaded human cells. Although the numbers of viable intracellular bacteria of wild bird isolates were lower than a type strain NCTC11168, they persisted at 48-hr and underwent replication in host cells.

Prevalence and antimicrobial susceptibility of Listeria monocytogenes on chicken carcasses in Bandung, Indonesia.

This study was conducted to determine the prevalence and quantify the number of Listeria monocytogenes in fresh chicken carcasses sold in traditional markets and supermarkets in Bandung, West Java, Indonesia, and to determine the antimicrobial resistance patterns of the isolated L. monocytogenes strains. The overall prevalence of L. monocytogenes in chicken carcasses was 15.8% (29/184). When comparing samples from traditional markets and supermarkets, no significant difference in the L. monocytogenes prevalence was detectable (15.2 versus 16.3%). Of the samples, 97.3% had L. monocytogenes counts <100 CFU/g, 2.2% had L. monocytogenes counts between 101 and 1,000 CFU/g, and 0.5% had L. monocytogenes counts of 1,001 to 10,000 CFU/g. Of the isolates, 27.6% were resistant to at least one of the 10 antimicrobials tested, with the major resistant phenotypes to penicillin (17.2%), ampicillin (6.9%), and erythromycin (6.9%). All 29 isolates recovered in this study were grouped into the molecular serogroup IIb, comprising the serovars 1/2b, 3b, and 7.

Physiological characterization of Campylobacter jejuni under cold stresses conditions: its potential for public threat.

Campylobacter jejuni is the major cause of human gastroenteritis worldwide. Under stress conditions, C. jejuni can enter a viable but non-culturable (VBNC) state. We found that the C. jejuni was able to enter a VBNC state by prolonged incubation at 4°C. The standard isolation methods using pre-enrichment steps in Bolton broth or Preston broth could not detect the VBNC cells in spiked chicken meat. The transcription levels of virulence-associated genes (flaA, flaB, cadF, ciaB, cdtA, cdtB and cdtC) were expressed in VBNC cells but in low levels. The VBNC cells retained the ability to invade Caco-2 human intestinal epithelial cells in vitro. In most cases, VBNC cells failed to resuscitate in Caco-2 cells, but in some experiments, they formed colonies after co-incubation with host cells. Collectively, C. jejuni enters into a VBNC state at 4°C and the VBNC C. jejuni remains virulent which may possibly lead to disease in humans. C. jejuni in VBNC state is a potential concern for food safety.