The importance of social and spatial structuring of wildlife populations for disease spread, though widely recognized, is still poorly understood in many host-pathogen systems. In particular, system specific kin relationships among hosts can create contact heterogeneities and differential disease transmission rates. Here, we investigate how distance-dependent infection risk is influenced by genetic relatedness in a novel wild boar ( Sus scrofa) - African swine fever (ASF) system. We hypothesized that the infection risk would correlate positively with proximity and relatedness to ASF-infected individuals but expected those relationships to weaken with distance between individuals due to decay in contact rates and genetic similarity. ASF infection risk was shaped by the number of infected animals throughout the zone of potential contact (0-10 km) but not beyond it. This effect was the strongest at close distances (0-2 km) and weakened further on (2-10 km), consistent with decreasing probability of contact. Overall, there was a positive association between genetic relatedness to infectees and infection risk within the contact zone but this effect varied in space. In the high-contact zone (0-2 km), infection risk was not influenced by relatedness when controlled for the number of ASF-positive animals. However, infections were more frequent among close relatives indicating that familial relationships could have played a role in ASF transmission. In the medium-contact zone (2-5 km), infection risk and frequency of paired infections were associated with relatedness. Relatedness did not predict infection risk in low- and no-contact zones (5-10 and >10 km, respectively). Together, our results indicate that the number of nearby infected individuals overrides the effect of relatedness in shaping ASF transmission rates which nevertheless can be higher among close relatives. Highly localized transmission highlights the possibility to control the disease if containment measures are employed quickly and efficiently.

Classical swine fever (CSF) is caused by classical swine fever virus (CSFV) and has led to huge ecnomic losses for the pig industry worldwide. Although vaccination and other control measures have been carried out, it is essential to establish a rapid and valid method for CSF vaccination monitoring and clinical diagnosis. CSFV E2 protein has been well-known as a major antigen for antibody detection. It is significant to improve affinity between E2 protein and CSFV antibody for a better performance of detection method. In this study, a recombinant E2 extracellular protein (aa 1-331), which has a native homodimer conformation and has a high affinity with anti-CSFV-E2 monoclonal antibody WH303, was expressed using Bac-to-Bac baculovirus expression system. A novel immunochromatographic test strip based on the recombinant CSFV E2 protein was developed for CSFV antibody detection. The sensitivity of this strip for detecting CSFV standard positive serum was 1:102400, 4 times higher than that of the previously developed CnC2 test strip. No cross reaction with antibodies of other swine viruses was observed. The detection of clinical swine serum samples (n=138) demonstrated that the agreements of this E2 test strip with three commercial ELISA kits were 88.40% (122/138), 86.23% (119/138), and 96.38% (133/138), respectively. Our data indicated that a novel E2 test strip with higher sensitivity has been developed and can be applied for clinical sample detections, providing a new powerful and simple approach for CSFV antibody monitoring.

Leptospirosis is a zoonotic neglected disease of worldwide public health concern. Leptospira species can infect a wide range of wild and domestic mammals and can lead to a spectrum of disease, including severe and fatal forms. Herein, we report for the first time a fatal Leptospira interrogans infection in a free-ranging nonhuman primate (NHP), a black-tufted marmoset. Icterus, pulmonary hemorrhage, interstitial nephritis and hepatocellular dissociation were the main findings raising the suspicion of leptospirosis. Diagnostic confirmation was based on specific immunohistochemical and PCR assays for Leptospira species. Immunolocalization of leptospiral antigens and identification of pathogenic species ( L. interrogans species) were important for better understanding the pathogenesis of disease. One Health related implications of free-ranging NHPs in anthropized areas and transmission dynamics of human and animal leptospirosis are discussed.

Mongolia and Eastern Siberia, Russia are border regions in Asia with high incidence of tuberculosis (TB). In this study, we aimed to investigate MDR -TB transborder transmission with a focus on endemic and epidemic Mycobacterium tuberculosis clones and drug resistance patterns. M. tuberculosis strains (291 from Mongolia and 754 from Russia) were collected within cross-sectional population-based surveys in 2010-2016. DNA was genotyped in 24 MIRU-VNTR loci and by PCR testing of the key SNP markers to discriminate within Beijing genotype. In total, 1045 isolates were divided into 435 MIRU-types that were assigned to Lineage 2 (Beijing isolates) and Lineage 4 (Ural, Haarlem, Latin-American-Mediterranean [LAM], S, and unclassified isolates). Beijing genotype was dominant in both countries, but most of Russian and all Mongolian Beijing strains belonged to different subtypes of the modern Beijing sublineage with only negligible overlap between the two countries. In particular, the Beijing types #342-32, #3819-32, #1773-32 (Asian African 2 group) were found only in Mongolia. LAM was the most common non-Beijing genotype (11.0% in Mongolia and 14.7% in Russia) and its isolates mostly belonged to LAM-RUS branch. MDR rate was higher in Russia compared to Mongolia among newly diagnosed patients: 29.4% versus 4.2% (p < 0.001) but similar and high in the retreatment subgroups (65.8% and 67.4%, respectively). In Russian collection, a higher MDR rate was observed in (i) Beijing compared to non-Beijing (47.5% versus 38.8%, p = 0.03), (ii) Beijing B0/W148 subtype compared to Beijing Central Asian/Russian subtype (64.5% versus 39.3%, p <0.001). In Mongolia, MDR rate was similar in Beijing (29.7%) and non-Beijing (27.5%) genotypes. In conclusion, population structures of the Beijing genotype in Mongolia and Russian borderline regions differ significantly including specific patterns of drug resistance. In contrast, largely overlapping LAM subtypes may correlate with historical endemic circulation of the LAM-RUS branch in Northern Eurasia.

Rabbit Haemorrhagic Disease (RHD) is a significant viral disease affecting lagomorphs. The first documented cases of RHD in Singapore occurred in adult pet European rabbits in September 2020. Singapore subsequently declared the outbreak resolved in December 2020. Epidemiological investigations ruled out introductions via importation of infected rabbits and contaminated feed. The source could not be definitively determined. However, the findings suggest that the incident involved both inter- and intra-household transmission and veterinary clinic-household transmission. This incident demonstrated the importance of sustained application of biosecurity measures, epidemiological investigations, and control, including active case finding, expedient vaccine dissemination, and risk communications. It shows that Singapore, an urbanised city-state, without a significant lagomorph population, could still encounter emerging diseases such as rabbit haemorrhagic disease. Given its social impact on rabbit owners, the National Parks Board Singapore and the private veterinarians worked together to communicate and urge the adoption of biosecurity measures and assuage the concerns of rabbit owners.

This study aimed to compare the infection dynamics of three genotype II African swine fever viruses (ASFV) circulating in Europe. Eighteen domestic pigs divided into three groups were infected intramuscularly or by direct contact with two haemadsorbent ASFVs (HAD) from Poland (Pol16/DP/ OUT21) and Estonia (Est16/WB/Viru8), and with the Latvian non-HAD ASFV (Lv17/WB/Rie1). Parameters such as symptoms, pathogenicity, and distribution of the virus in tissues, humoral immune response, and dissemination of the virus by blood, oropharyngeal and rectal routes were investigated. The Polish ASFV caused a case of rapidly developing fatal acute disease, while the Estonian ASFV caused acute to subacute infections in the presence of surviving animals. In contrast, animals infected with the ASFV from Latvia developed a more subtle, mild, or even subclinical disease. Oral excretion was sporadic or even absent in the attenuated group, whereas in animals that developed an acute or subacute form of ASF, oral excretion began at the same time as in the blood, or even 3 days earlier, and persisted up to 22 days. Regardless of virulence, blood was the main route of transmission of ASFV and infectious virus was isolated from persistently infected animals for at least 19 days in the attenuated group and up to 44 days in the group of moderate virulence. Rectal excretion was limited to the acute phase of infection. In terms of diagnostics, the ASFV genome was detected in contact pigs from oropharyngeal samples earlier than in blood, independently of virulence and, together with blood, both samples could cover a longer range to detect ASFV infection. The results presented here provide quantitative data on the spread and excretion of ASFV strains of different virulence among domestic pigs that can help to better focus surveillance activities and thus increase the ability to detect ASF introductions earlier.

Bacteriophage is considered an alternative to antibiotics and environmentally friendly approach to tackle antimicrobial resistance (AMR) in aquaculture. Here, we reported isolation, morphology and genomic characterizations of a newly isolated lytic bacteriophage, designated pAh6.2TG. Host range and stability of pAh6.2TG in different environmental conditions, and protective efficacy against a pathogenic multidrug-resistant (MDR) Aeromonas hydrophila in Nile tilapia were subsequently evaluated. The results showed that pAh6.2TG is a member of the family Myoviridae which has genome size of 51,780 bp, encoding 65 putative open reading frames (ORFs), and is most closely related to Aeromonas phage PVN02 (99.33% nucleotide identity). The pAh6.2TG was highly specific to A. hydrophila and infected 83.3% tested strains of MDR A. hydrophila (10 out of 12) with relative stability at pH 7 ­ 9, temperature 0 ­ 40 °C and salinity 0 ­ 40 ppt. In experimental challenge, pAh6.2TG treatments significantly improved survivability of Nile tilapia exposed to a lethal dose of the pathogenic MDR A. hydrophila, with relative percent survival (RPS) of 73.3% and 50% for phage multiplicity of infection (MOI) 1.0 and 0.1, respectively. Significant reduction of bacterial counts in rearing water at 3 h (6.7 ± 0.5 to 18.1 ± 6.98 folds) and in fish liver at 48 h post-treatment (2.7 ± 0.24 to 34.08 ± 26.4 folds) was observed in phage treatment groups while opposite pattern for bacterial counts was observed in untreated control. Interestingly, the surviving fish provoked specific antibody (IgM) against the challenged A. hydrophila. These results might explain the higher survival in phage treatment groups. In summary, the findings suggested that the lytic bacteriophage pAh6.2TG is an effective alternative to antibiotics to control MDR A. hydrophila in tilapia and possibly other freshwater fish.

This systematic review and meta-analysis aimed to recalculate the efficacy of these two vaccine strains, and to discuss the main variables associated with controlled trials to evaluate bovine brucellosis vaccines efficacy. The most used vaccine strain was S19, at the dose of 10 10 colony forming units (CFU), followed by the vaccine strain RB51 at 10 10 CFU. The most used challenge strain was B. abortus 2308, at the dose of 10 7 CFU by intraconjunctival route. For the meta-analysis, trials were grouped according to the vaccine strain and dose to recalculate protection against abortion (four groups) or infection (five groups), using pooled risk ratio (RR) and vaccine efficacy (VE). For protection against abortion (n = 15 trials), S19 vaccine at 10 9 CFU exhibited the highest protection rate (RR = 0.25, 95% CI: 0.12 to 0.52; VE = 75.09%, 95% CI: 48.08 – 88.05), followed by RB51 10 10 (RR = 0.31, 95% CI: 0.16 to 0.61; VE = 69.25%, 95% CI: 39.48 – 84.38). For protection against infection (n = 23 trials), only two subgroups exhibited significant protection: S19 at 10 9 CFU (RR = 0.28, 95% CI: 0.14 to 0.55; VE = 72.03%, 95% CI: 57.70 – 81.50) and RB51 at 10 10 CFU dose (RR = 0.43, 95% CI: 0.22 to 0.84; VE = 57.05%, 95% CI: 30.90 – 73.30). In conclusion, our results suggest that the dose of 10 9 CFU for S19 and 10 10 CFU for RB51 are the most suitable for the prevention of abortion and infection caused by B. abortus.

Introduction: The Coronavirus Disease 2019 (COVID-19) pandemic has had a variable worldwide impact, likely related to country-level characteristics. In this ecological study, we explored the association of COVID-19 case rates (per 100,000 people) and death rates (per 100,000 people) with country-level population health characteristics, economic and human development indicators, and habitat-related variables. Methods: To calculate country-level COVID-19 case and death rates, the number of cases and deaths were extracted from the Johns Hopkins Coronavirus Resource Center for 2020. Country-level population health characteristics, economic and human development indicators, and habitat-related variables were extracted from several publicly available online sources of international organizations. Results were tabulated according to world zones and country economies. Univariate and multivariable linear regression analyses were performed to examine determinants of COVID-19 case rates and death rates. Results: A total of 187 countries and territories were analyzed, with an aggregate COVID-19 case rate of 779 per 100,000 people, a death rate of 19 per 100,000 people, and a case-fatality rate of 2.4%. For country-level population health characteristics, a higher percentage rate of adults with obesity and a higher percentage rate of adults with high blood pressure was independently associated with a higher COVID-19 case rate, and a higher percentage rate of adults with obesity was associated with a higher COVID-19 death rate. For country-level economic and human development indicators, only a higher gross domestic product percentage rate spent on total health expenditure and a higher human development index was independently associated with a higher COVID-19 case rate and death rate. A higher percentage of urban population was independently associated with a higher COVID-19 death rate, whereas a higher income per capita was independently associated with a lower COVID-19 death rate. For country-level habitat-related variables, a higher average household size and a higher percentage rate of population with primary reliance on polluting fuels and technologies was independently associated with a lower COVID-19 case rate and death rate whereas a higher percentage rate of households with at least one-member age 65 years or over was associated with a higher case rate and death rates. Conclusion: This ecological study informs the need to develop country-specific public health interventions to better target populations at high risk for COVID-19, and test environmental interventions to prevent indoor transmission of SARS-CoV-2, taking into consideration population health characteristics, economic and human development indicators, and habitat-related variables that are unique to each country.

Peste des petits ruminants (PPR) is a viral transboundary disease of small ruminants that causes significant damage to agriculture. This disease has not been previously registered in the Republic of Kazakhstan (RK). This paper presents an assessment of the susceptibility of the RK’s territory to the spread of the disease in the event of its importation from infected countries. The Generalized Linear Negative Binomial regression model that was trained on the PPR outbreaks in China was used to rank municipal districts in the RK in terms of PPR spread risk. The outbreaks count per administrative district was used as a risk indicator, while a number of socio-economic, landscape and climatic factors were considered as explanatory variables. Summary road length, altitude, the density of small ruminants, the maximum green vegetation fraction, cattle density and the Engel coefficient were the most significant factors. The model demonstrated a good performance in training data (R 2 = 0.69) and was transferred to the RK, suggesting a significantly lower susceptibility of this country to the spread of PPR. Hot Spot analysis identified three clusters of districts at the highest risk, located in the western, eastern and southern parts of Kazakhstan. As part of the study, a countrywide survey was conducted to collect data on the distribution of livestock populations, which resulted in the compilation of a complete geo-database of small ruminant holdings in the RK. The research results may be used to formulate a national strategy for preventing the importation and spread of PPR in Kazakhstan through targeted monitoring in high-risk areas.

Foot-and-mouth disease (FMD) is endemic in India, where circulation of serotypes O, A and Asia 1 is frequent. In the past two decades, many of the most widespread and significant FMD lineages globally have emerged from the South Asia region. Here, we provide an epidemiological assessment of the ongoing mass vaccination programs in regard to post-vaccination monitoring and outbreak occurrence. The objective of this study was to quantify the spatiotemporal dynamics of FMD outbreaks and to assess the impact of the mass vaccination program between 2008 to 2016 with available antibody titer data from the vaccination monitoring program, alongside other risk factors that facilitate FMD spread in the country. We first conducted a descriptive analysis of epidemiological outcomes of governmental vaccination programs in India, focusing on antibody titer data from >1 million animals sampled as part of pre- and post-vaccination monitoring and estimates of standardized incidence ratios calculated from reported outbreaks per state/administrative unit. The percent of animals with inferred immunological protection (based on ELISA) was highly variable across states, but there was a general increase in the overall percent of animals with inferred protection through time. In addition, the number of outbreaks in a state was negatively correlated with the percent of animals with inferred protection. Because standardized incidence ratios of outbreaks were heterogeneously distributed over the course of eight years, we analyzed the distribution of reported FMD outbreaks using a Bayesian space-time model to map high-risk areas. This model demonstrated a ~50% reduction in the relative risk of outbreaks in states that were part of the vaccination program. In addition, states that did not have an international border experienced reduced risk of FMD outbreaks. These findings help inform risk-based control strategies for India as the country progresses towards reducing reported clinical disease.

Marek’s disease (MD) is a re-emerging viral disease of chicken and a serious economic threat to poultry industry worldwide. Continuous surveillance with molecular investigation is mandatory to monitor the emergence of virulent MDV strains and to devise any appropriate vaccination strategy and implement bio-security programs. In the present study, we investigated the cases of MD outbreaks in vaccinated poultry flocks. The MD outbreak was confirmed through necropsy (majorily visceral tumors), histopathology and viral gene specific PCR. The pathotypes of the field MDV strains were assessed by molecular analysis of three oncogenes -Meq, pp38 and vIL-8. The Meq sequence of the field strains analyzed in this study lacked the 59 aa unique to mild strains indicating that they are virulent strains. Mutation at position 71 and presence of five proline rich repeats in the transactivation domain, both associated with virulence were observed in these strains, however, the signature sequences specific to very virulent plus strains were absent. Phylogenetic analysis of Meq gene sequences revealed clustering of the field strains with North Indian strains and with a very virulent plus ATE 2539 strain from Hungary. Analyses of pp38 protein at positions 107 and 109 and vIL-8 protein at positions 4 and 31 showed signatures of virulence. Sequence and phylogenetic analysis of oncogenes from field MDVs from vaccinated flock indicated these strains possessing molecular features of very virulent strains. Our data shows here that Meq, vIL-8 and pp38 genes can be used as markers for molecular analysis to decipher the pathotype of MDV strains. Our present study suggests evolution of virulent MDV induced by vaccination.

We report a COVID-19 case with unprecedented viral complexity. In the first severe episode, two different SARS-CoV-2 strains (superinfection) were identified within a week. Three months after discharge, patient was readmitted and was infected in a nosocomial outbreak with a different strain, suffering a second milder COVID-19 episode.

Bovine leptospirosis is a bacterial disease that affects cattle herds, causing economic losses due to reproductive problems, which require expensive treatments. The main source of transmission for cattle is still uncertain, but it has been described that rodents and bats can play an important role in the transmission cycle by being maintenance hosts for the pathogenic species of the bacterium and spreading it through urine. In this study, we characterize possible risk areas for bovine leptospirosis exposure in the state of Veracruz, Mexico, based on the geographical distribution of flying (bats) and terrestrial (rodents and opossums) wild hosts of Leptospira sp. reported in Mexico in addition with climatic, geographic, soil characteristics, land use and human activities variables (environmental variables). We used a generalized linear regression model (GLM) to understand the association between the frequency of anti- Leptospira sp. antibodies (a proxy of exposure to) in cattle herds exposed to Leptospira, the favorability of wild hosts of Leptospira as well as the environmental variables. The parameterized model explains 12.3% of the variance. The frequency of anti- Leptospira sp. antibodies exposoure in cattle herds was associated with elevation, geographic longitude, pH of the soil surface and environmental favorability for the presence of rodents, opossums, and bats. The variation in exposure is mainly explained by a longitudinal gradient (6.4% of the variance) and the favorability-based indices for wild hosts (9.6 % of the variance). Describing the possible risks for exposure to Leptospira in an important and neglected livestock geographical region, we provide valuable information to the selection of areas for diagnosis and prevention of this relevant disease.

Influenza viruses have been posing a great threat to public health and animal industry. The developed vaccines have been widely used to reduce the risk of potential pandemic; however, the ongoing antigenic drift makes influenza virus escape from host immune response and hampers vaccine efficacy. Until now, the genetic basis of antigenic variation remains largely unknown. In this study, we used A/swine/Guangxi/18/2011 (GX/18) and A/swine/Guangdong/104/2013 (GD/104) as models to explore the molecular determinant for antigenic variation of Eurasian avian-like H1N1 (EA H1N1) swine influenza viruses (SIVs), and found that the GD/104 virus exhibited 32~64-fold lower antigenic cross-reactivity with antibodies against GX/18 virus. Therefore, we generated polyclonal antibodies against GX/18 or GD/104 virus and a monoclonal antibody (mAb), named mAb102-95, targeted to the hemagglutinin (HA) protein of GX/18 virus, and found that a single amino acid substitution at position 158 in HA protein substantially altered the antigenicity of virus. The reactivity of GX/18 virus containing G158E mutation with the mAb102-95 decreased 8-fold than that of the parental strain. Contrarily, the reactivity of GD/104 virus bearing E158G mutation with the mAb102-95 increased by 32 times as compared with that of the parental virus. Structural analysis showed that the amino acid mutation from G to E was accompanied with the R group changing from -H to -(CH 2) 2-COOH. The induced steric effect and increased hydrophilicity of HA protein surface jointly contributed to the antigenic drift of EA H1N1 SIVs. Our study provides experimental evidence that G158E mutation in HA protein affects the antigenic property of EA H1N1 SIVs, and widens our horizon on the antigenic drift of influenza virus.

The incidence of bovine tuberculosis (TB, caused by Mycobacterium bovis) in cattle has been associated with TB in badgers ( Meles meles) in parts of England. The aim was to identify badger associated M. bovis reservoirs in the Edge Area, between the High and Low Risk Areas for cattle TB. Data from badger TB surveys were sparse. Therefore, a definition for a local M. bovis reservoir potentially shared by cattle and badgers was developed using cattle TB surveillance data. The performance of the definition was estimated through Latent Class Analysis using badger TB survey data. Spatial units (25 km 2 ) in the Edge Area were classified as having a reservoir if they had i) at least one OTF-W (Officially Tuberculosis Free – Withdrawn) incident in a cattle herd not attributed to cattle movement in the previous two years, ii) more OTF-W incidents than Officially Tuberculosis Free – Suspended (OTF-S) incidents in the previous two years and iii) at least one TB incident (OTF-S or OTF-W) in at least three of the previous seven years. Approximately twenty percent of the Edge Area was classified as having a local M. bovis reservoir using the cattle-based definition. Assuming 15% TB prevalence in Edge Area badgers, sensitivity for the local M. bovis reservoir definition varied from 25.7% (95% Credible Interval (CrI) 10.7 to 85.1 %) to 64.8 % (95% CrI 48.1 to 88.0 %). Specificity was 91.9% (CrI 83.6 to 97.4 %). Over ninety percent of the local reservoir was in stable endemic TB areas identified through previous work and its spatial distribution was largely consistent with local veterinary knowledge. Uncertainty in the reservoir spatial distribution was explored through its recalculation in spatial units shifted in different directions. We recommend that the definition is re-evaluated as further data on badger infection with M. bovis becomes available.

African Swine Fever Virus (ASFV) causes a deadly disease of pigs which spread through southeast Asia in 2019. We investigated one of the first outbreaks of ASFV in Lao Peoples Democratic Republic amongst smallholder villages of Thapangtong District, Savannakhet Province. In this study, two ASFV affected villages were compared to two unaffected villages. Evidence of ASFV-like clinical signs appeared in pig herds as early as May 2019, with median epidemic days on 1 and 18 June in the two villages, respectively. Using participatory epidemiology mapping techniques, we found statistically significant spatial clustering in both outbreaks (P < 0.001). Villagers reported known risk factors for ASFV transmission − such as free-ranging management systems and wild boar access − in all four villages. The villagers reported increased pig trader activity from Vietnam before the outbreaks; however, the survey did not determine a single outbreak source. The outbreak caused substantial household financial losses with an average of 9 pigs lost to the disease, and Monte Carlo analysis estimated this to be USD 215 per household. ASFV poses a significant threat to food and financial security in smallholder communities such as Thapangtong, where 40.6% of the district’s population are affected by poverty. This study shows ASFV management in the region will require increased local government resources, knowledge of informal trader activity and wild boar monitoring alongside education and support to address intra-village risk factors such as free-ranging, incorrect waste disposal and swill feeding.

During the current COVID-19 pandemic, different methods have been used to evaluate patients suspected with infection of SARS-CoV-2. In this study, we evaluate the longevity of saliva and dry swab samples to retain SARS-CoV-2 for storage and transport at different environmental settings. Our results show that at ambient temperature of 20°C, SARS-CoV-2 RNA remains stable for up to 9 days giving a long span of time for transport and storage without compromising clinical results. Additionally, this study demonstrates that sali­­­va and dry swabs specimens can also be stored at -20°C and +4°C for up to 26 days without affecting RT-qPCR results. Our data is relevant for low-and middle-income countries, which have limited access to rapid refrigerated transport and storage of samples representing an economical alternative. Finally, our study demonstrates that dry swabs provide clear advantages over using transport medium.