Foot-and-mouth disease (FMD) is widely distributed in Sudan where outbreaks occur on an annual basis especially during the winter months (December-February). This study aimed to increase our understanding of the epidemiological patterns of FMD in Sudan and connections to neighbouring countries by characterising the genetic sequences of FMD viruses (FMDV) collected from seven Sudanese states over a 10-year period (between 2009 and 2018). FMDV was detected in 91 of the 265 samples using an antigen-detection ELISA. Three serotypes were detected: O (46.2%), A (34.1%), and SAT 2 (19.8%). Fifty-three of these samples were submitted for sequence analyses, generating sequences that were characterised as belonging to O/EA-3 (n=18), A/AFRICA/G-IV (n=23) and SAT 2/VII/Alx-12 (n=12) viral lineages. Phylogenetic analyses provided evidence that FMDV lineages were maintained within Sudan, and also highlighted epidemiological connections to FMD outbreaks reported in neighbouring countries in East and North Africa (such as Ethiopia and Egypt). This study motivates continued FMD surveillance in Sudan to monitor the circulating viral lineages and broader initiatives to improve our understanding of the epidemiological risks in the region.

Livestock industry supports the livelihood of around 1.3 billion people in the world, with swine industry contributing with 30 % of total livestock production worldwide. To maintain and guarantee this production, a pivotal point according to the OIE is addressing potential biohazards. To control them, permanent sero-surveillance is crucial to achieve more focused veterinary public health intervention and prevention strategies, to break the chains of transmission, and to enable fast responses against outbreaks. Within this context, multiplex assays are powerful tools with the potential to simplify surveillance programs, since they reduce time, labour, and variability within analysis. In the present work, we developed a multiplex bead-based assay for the detection of specific antibodies to six relevant pathogens affecting swine: ASFV, CSFV, PRRSV, SIV, TB, and HEV. The most immunogenic target antigen of each pathogen was selected as the target protein to coat different microsphere regions in order to develop this multiplex assay. A total of 1544 serum samples from experimental infections as well as field samples were included in the analysis. The 6plex assay exhibited credible diagnostic parameters with sensitivities ranging from 87.0 % to 97.5 % and specificities ranging from 87.9 % to 100.0 %, demonstrating it to be a potential high throughput tool for surveillance of infectious diseases in swine.

Animals like mink, cats and dogs are susceptible to SARS-CoV-2 infection. In the Netherlands, 69 out of 127 mink farms were infected with SARS-CoV-2 between April and November 2020 and all mink on infected farms were culled after SARS-CoV-2 infection to prevent further spread of the virus. On some farms, (feral) cats and dogs were present. This study provides insight into the prevalence of SARS-CoV-2 positive cats and dogs in ten infected mink farms and their possible role in transmission of the virus. Throat and rectal swabs of 101 cats (12 domestic and 89 feral cats) and 13 dogs of ten farms were tested for SARS-CoV-2 using PCR. Serological assays were performed on serum samples from 62 adult cats and all 13 dogs. Whole Genome Sequencing was performed on one cat sample. Cat-to-mink transmission parameters were estimated using data from all ten farms. This study shows evidence of SARS-CoV-2 infection in twelve feral cats and two dogs. Eleven cats (19%) and two dogs (15%) tested serologically positive. Three feral cats (3%) and one dog (8%) tested PCR-positive. The sequence generated from the cat throat swab clustered with mink sequences from the same farm. The calculated rate of mink-to-cat transmission showed that cats on average had a chance of 12% (95%CI 10% to 18%) of becoming infected by mink, assuming no cat-to-cat transmission. As only feral cats were infected it is most likely that infections in cats were initiated by mink, not by humans. Whether both dogs were infected by mink or humans remains inconclusive. This study presents one of the first reports of interspecies transmission of SARS-CoV-2 that does not involve humans, namely mink-to-cat transmission, which should also be considered as a potential risk for spread of SARS-CoV-2.

In South Vietnam, live bird markets (LBMs) are key in the value chain of poultry products and spread of avian influenza virus (AIV) although they may not be the sole factor to determine avian influenza (AI) prevalence in the southern part. Therefore, a risk analysis of AIV spread was conducted by including all possible value chain factors. A cross-sectional study was performed in backyard farms, high-biosecurity farms (bio-farms), LBMs, and poultry delivery stations (PDSs) in the four districts of Vinh Long Province in December 2016 and August 2017. A total of 3 597 swab samples were collected from individual poultry at 101 backyard farms, 50 bio-farms, 58 sellers in LBMs, and 17 traders in PDSs and then investigated for AIV isolation. Concurrently, information related to participants and birds was collected and used to identify the fixed and random effects of factors in AIV infection. A total of 274 birds were positive for virus isolation, with a prevalence of 7.6% (95% confidence interval [CI]: 6.8–8.5) at the individual poultry level, and the adjusted prevalence based on the sampling weight was 7.9% (95% CI: 7.6–8.2). The significantly higher prevalence in PDSs (20.7%) and LBMs (14.2%) compared to backyard farms (3.0%) and bio-farms (0.6%) suggested that PDSs are another hot spot for AIV circulation. The high diversity in the seller and trader population characteristics was revealed using multiple-correspondence analysis to analyze the participants’ demographic factors in LBM and PDS. The mixed-effect logistic regression model revealed that keeping duck at the sampling time and the owner’s older age should be risk factors of AIV infection in PDS. Therefore, functional AI control efforts to monitor the PDS system should be emphasized to minimize AIV circulation risk in Vietnam.

The Cambodian government is attempting to mobilise government, donor and private sector funding to implement a coordinated FMD vaccination program (FMDVP). A necessary first step is to convince the farmers of the benefits of participating in and potentially financially supporting this program. Information was collected from 300 farmers in order to estimate the on-farm benefits and costs of their participation in an FMDVP. Implementing a successful vaccination program is difficult, and farmers understand from previous experience that there may be institutional, social, technical and financial constraints which limit its success. A benefit-cost analysis needs to take into account that outbreaks do not occur every year, not all cattle will be successfully vaccinated, not all sick animals successfully treated and sometimes sick animals simply sold. This study sensitises these variables in order to give a realistic estimation of the farmer participation benefits in an FMDVP. A general result is that it is worthwhile for farmers to participate in the FMDVP if there are average annual outbreaks, or at least two major outbreaks, in the ensuing five years. However, the results are influenced by the interaction of vaccination success and treatment success and coverage. Ineffective coverage and poor treatment of sick animals reduce the benefits of an FMDVP. It is also important that farmers do not sell sick stock and, if they do, that they are able to breed replacements rather than purchase replacements. There are many factors in the smallholder cattle farming system that will influence the success of an FMDVP; farmers will only choose to participate if they can be convinced of the short and long-term economic benefits.

Brucellosis is an endemic disease in many developing countries and ranked by the World Health Organization among the top seven “neglected zoonoses”. Although a Palestinian brucellosis control program was launched in 1998, the disease reemerged after 2012. Interestingly, a similar reemerging pattern was reported in the neighboring Israeli regions. The aim of this work was to characterize the reemerging strains and delineate their genetic relatedness. During 2015-2017, blood samples from 1324 suspected patients were analyzed using two serological tests. Seropositive samples were cultured, and their DNAs were analyzed by different genetic markers to determine the involved Brucella species and rule out any possible involvement of the Rev.1 vaccine strain. The rpoB gene was sequenced from 9 isolates to screen for rifampicin-resistance mutations. Multi Locus VNTR Analysis (MLVA-16) was used for genotyping the isolates. The molecular analysis showed that all isolates were B. melitensis strains unrelated to the Rev.1 vaccine. The rpoB gene sequences showed four single nucleotide variations (SNVs) not associated with rifampicin resistance. MLVA-16 analysis clustered the isolates into 22 unique genotypes that belong to the East Mediterranean lineage. Altogether, our findings show that the reemergence of brucellosis was due to B. melitensis strains of local origin, the Palestinian and Israeli control programs’ weaknesses could be a major factor behind the reemergence of the disease. However, other socioeconomic and environmental factors must be investigated. Moreover, strengthening brucellosis control programs and enhancing cooperation between all stakeholders is essential to ensure long-term program outcomes to fight brucellosis.

Food-and-mouth disease (FMD) is endemic in Cambodia. The control program for FMD has relied on vaccination, with poor vaccination uptake by smallholder farmers an increasing concern. A study to improve the understanding of farmer knowledge, attitudes and practices of FMD and FMD vaccination was conducted in two Cambodian provinces. The aim was to identify opportunities to improve the disease control programs provided by both the government and private sectors. The survey comprised 300 smallholder farmers using a one-on-one interview technique. Results identified that over two-thirds of the respondent farmers had not vaccinated their cattle over two years. Of those who did, most cattle were vaccinated either once a year or once every three years. A booster had never been administered. FMD outbreaks occurred every year during the study period, with a morbidity rate of over 30%. Isolation of first infected cattle from the household herd was not practiced, with treatment identified as the first preference intervention. Farmers often assisted other farmers to restrain and treat infected cattle both before (57%) and after (43%) their own cattle were infected. This indicated that most farmers did not practice basic biosecurity measures and chose to report FMD outbreaks to the village animal health workers (VAHW), friends, neighbors, and relatives in preference to government officials. It was concluded that poor knowledge of disease transmission and biosecurity, with low FMD vaccination coverage and a focus on treatment, contribute to regular FMD outbreaks in these communities. Improvement of FMD control requires the cooperation of villagers, VAHWs, and village leaders in disease reporting, with either improved funding of government vaccination services or private FMD vaccination service. Training programs for farmers on disease transmission, and the importance of biosecurity and vaccination, including information on the cost-benefits of treatment versus full fee bi-annual FMD vaccination, are required.

The total impact of the worldwide COVID-19 pandemic is still emerging, changing all relationships as a result, including those with pet animals. In the infection process, the use of Angiotensin-converting enzyme 2 (ACE2) as a cellular receptor to the spike protein of the new coronavirus is a fundamental step. In this sense, understanding which residue plays what role in the interaction between SARS-CoV-2 spike glycoprotein and ACE2 from cats, dogs, and ferrets is an important guide for helping to choose which animal model can be used to study the pathology of COVID-19 and if there are differences between these interactions and those occurring in the human system. Hence, trying to help to answer these questions, we performed classical molecular dynamics simulations to evaluate, from an atomistic point of view, the interactions in these systems. Our results show that there are significant differences in the interacting residues between the systems from different animal species, and the role of ACE2 key residues are different in each system and can assist in the search for different inhibitors for each animal.

Toxoplasma gondii was initially classified in three main lineages related to its virulence: Types I, II and III. The recombination of genes during sexual cycle in felids gut led to more than 200 genotypes, found in ToxoDB database, using 11 RFLP markers. Free-range chickens are good bioindicators of soil contamination with T. gondii oocysts. In this sense, there are systematic reviews regarding data of genetic characterization of this parasite in felines and ruminants, but not in chickens heretofore, what makes this work necessary. A systematic review in the literature was performed with papers published prior to September 21st, 2020. The main inclusion criteria was the presence of T. gondii genotypes, isolated strictly from free-range chickens, in experimental works. Initially, a total of 1,343 studies related to the terms were identified on databases and 30 studies were selected to be systematically reviewed. A total of 561 isolates of T. gondii from 6,356 free-range chickens were analyzed for genotyping, revealing 190 genotypes. ToxoDB #59 and #2 were the most frequent in America, #1 was the most frequent in Africa and 3 atypical isolates from genotype ToxoDB #9 were found in Asia. There is not data from Europe and Oceania. The majority of studies were Brazilian (16/30). A total of 68 RFLP genotypes were recognized among the 561 isolates’ DNAs analyzed from the 30 studies. Some studies show new genotypes never described before, which reinforces the idea that some years from now, even more new genotypes will be isolated, due to progressive genetic recombination. The large amount of undefined genotypes makes it necessary to perform Nested PCR technique when genotyping. Moreover, the lack of data in Continents such as Europe, Asia and Oceania makes it necessary to perform new isolating and genotyping studies in these places.

Anthroponotic cutaneous leishmaniasis (ACL) due to Leishmania tropica is spreading to new areas. Exposure to the vector, Phlebotomus sergenti, is the only proven risk factor. Our objective was to compare the densities and genetic characteristics of P. sergenti populations in two nearby localities in Morocco, one within an ACL endemic area (El Borouj) and another undamaged (Sidi Hajjaj). Statistically significant differences were detected between P. sergenti densities with a higher density of P. sergenti in the endemic town (p≤ 0.032). A different main P. sergenti mitochondrial lineage was evidenced in each one of the 2 localities, and for the first time, the lineage of P. sergenti specimens that are acting as a vector of L. tropica has been identified. Bioclimatic differences were detected between both localities. In conclusion, between an ACL endemic locality and another ACL free there are differences in both the density of P. sergenti and the mitochondrial lineage that may explain the different epidemiological situation. Given that the density of P. sergenti in the locality without ACL cases seems sufficient to allow transmission, the main factor that would justify its ACL undamaged character could be the absence of P. sergenti Lineage IV, which seems to prefer warmer and drier climates.

Worldwide, wild birds are frequently suspected to be involved in the occurrence of outbreaks in captive-bred birds although proofs are lacking and most of the dedicated studies are insufficiently conclusive to confirm or characterize the roles of wild birds in such outbreaks. The aim of this study was to assess and compare, for the most prevalent peridomestic wild birds, the different exposure routes for Avian Influenza and Newcastle disease viruses in conservation breeding sites of Houbara bustards in the United Arab Emirates. To do so, we considered all of the potential pathways by which captive bustards could be exposed to Avian Influenza and Newcastle disease viruses by wild birds, and ran a comparative study of the likelihood of exposure via each of the pathways considered. We merged data from an ecological study dedicated to local wild bird communities with an analysis of the contacts between wild birds and captive bustards and with a prevalence survey of AIV and NDV in wild bird populations. We also extracted data from an extensive review of the scientific literature and by the elicitation of expert opinion. Overall, this analysis highlighted that captive bustards had a high risk of being exposed to pathogens by wild birds. This risk was higher for Newcastle disease virus than Avian influenza virus, and House sparrows represented the riskiest species for the transmission of both viruses through indirect exposure from consumption of water contaminated from the faeces of an infectious bird that got inside the aviary. Thus, this analysis reveals that wild peridomestic birds may play a role in the transmission of avian pathogens to captive bred birds. These results also reaffirm the need to implement sanitary measures to limit contacts between wild and captive birds and highlight priority targets for a thoughtful and efficient sanitary management strategy.

Numerous studies have unsuccessfully tried to unravel the definitive host of the coccidian parasite Besnoitia besnoiti. Cattle infections by B. besnoiti cause a chronic and debilitating condition called bovine besnoitiosis that has emerged in Europe during the last two decades, mainly due to limitations in its control associated to the absence of vaccines and therapeutical tools. Although the exact transmission pathway of B. besnoiti is currently unknown, it is assumed that the parasite might have an indirect life cycle with a carnivore as definitive host. Current lack of studies in wildlife might underestimate the importance of free-living species in the epidemiology of B. besnoiti. Thus, the aim of the present study is to assess the presence of Besnoitia spp. in free-ranging mesocarnivores in Spain. DNA was searched by PCR on faeces collected from wild carnivores as a first approach to determine which species could be considered as potential candidates for definitive hosts in further research. For this purpose, a total of 352 faecal samples from 12 free-living wild carnivore species belonging to the Canidae, Felidae, Herpestidae, Mustelidae, Procyonidae, and Viverridae families were collected in seven Spanish regions. PCR testing showed that Besnoitia spp. DNA was present in four faecal samples from red foxes collected in western Spain, an area with the greatest density of extensively reared cattle and associated to high incidence of bovine besnoitiosis in the country. To date, this is the first report of a Besnoitia besnoiti-like sequence (99.57% homology) from carnivore faeces in a worldwide context. Red foxes might contribute to the epidemiology of B. besnoiti, although further studies, mostly based on bioassay, would be needed to elucidate the accuracy and extent of these interesting findings.

African swine fever (ASF) is one of the most severe infectious diseases of pigs. In this study, a LAMP assay coupled with the CRISPR Cas12a system was established in one tube for the detection of the ASFV p72 gene. The single-strand DNA-fluorophore-quencher (ssDNA-FQ) reporters and CRISPR-derived RNA (crRNAs) were screened and selected for the CRISPR detection system. In combination with LAMP amplification assay, the detection limit for the LAMP-CRISPR assay can reach 7 copies/μl of p72 gene per reaction. Furthermore, this method displays no cross-reactivity with other porcine DNA or RNA viruses. The performance of the LAMP-CRISPR assay was compared with real-time qPCR tests for clinical samples, a good consistency between the LAMP-CRISPR assay and real-time qPCR was observed. In the current study, a LAMP coupled with the CRISPR detection method was developed. The method shed a light on the convenient, portable, low cost, highly sensitive and specific detection of ASFV, demonstrating a great application potential for monitoring on-site ASFV in the field.

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, correct waste disposal and swill feeding.

This study evaluates through modeling the possible individual and combined effect of three populational parameters of pathogens (reproduction rate; rate of novelty emergence; and propagule size) on the colonization of new host species – putatively the most fundamental process leading to the emergence of new infectious diseases. The results are analyzed under the theoretical framework of the Stockholm Paradigm using IBM simulations to better understand the evolutionary dynamics of the pathogen population and the possible role of Ecological Fitting. The simulations suggest that all three parameters positively influence the success of colonization of new hosts by a novel parasite population but contrary to the prevailing belief, the rate of novelty emergence (e.g. mutations) is the least important factor. Maximization of all parameters result in a synergetic facilitation of the colonization and emulates the expected scenario for pathogenic microorganisms. The simulations also provide theoretical support for the retention of the capacity of fast-evolving lineages to retro-colonize their previous host species/lineage by ecological fitting. Capacity is, thus, much larger than we can anticipate. Hence, the results support the empirical observations that opportunity of encounter (i.e. the breakdown in mechanisms for ecological isolation) is an fundamental determinant to the emergence of new associations - in special of Emergent Infectious Diseases - and the dynamics of host exploration, as observed in SARS-CoV-2. Insights on the dynamics of Emergent Infectious Diseases derived from the simulations and from the Stockholm Paradigm are discussed.

African swine fever (ASF) is a viral disease that affects members of the Suidae family. The notifiable disease is considered a major threat to the pig industry, animal health, and food security worldwide. According to the European Food Safety Authority, ASF virus (ASFV) survival and transmission in feed and feed materials is a major research gap. Against this background, the objective of this study was to determine the survival of ASFV on re-contaminated spray dried porcine plasma (SDPP) when stored at two different temperatures. To this means, commercial SDPP granules were contaminated with high titers of ASFV in a worst-case re-contamination scenario. Three samples per time point and temperature condition were subjected to blind passaging on macrophage cultures and subsequent haemadsorption test to determine residual infectivity. In addition, viral genome was detected by real-time PCR. The results indicate that heavily re-contaminated SDPP stored at 4°C remains infectious for at least five weeks. In contrast, contaminated SDPP stored at room temperature displayed a distinct ASFV titer reduction after one week and complete inactivation after two weeks. In conclusion, the residual risk of ASFV transmission through re-contaminated SDPP is low, if SDPP is stored at room temperature for a period of at least two weeks before feeding.

Introduction: Despite eradication and control measures applied across Europe, bovine tuberculosis (bTB) remains a constant threat. In Belgium, after several years of bTB disease freedom status, routine movement testing, as currently practiced, revealed itself inadequate to detect some sporadic breakdown herds. The aim of this study was to strike the balance between cost and effectiveness of different surveillance system components to identify sustainable alternatives for early detection and substantiation of freedom of bTB while maintaining acceptance of these amongst the different animal health stakeholders. Methods: Stochastic iteration model was built to simulate, first, the expected current surveillance system performance in terms of sensitivity and specificity of detection. These results were then descriptively compared to observed field results. Secondly, the cost and effectiveness of simulated alternative surveillance components were quantified. To measure impact of key assumptions (i.e. regarding diagnostic tests and true prevalence), sensitivity analysis was performed. Results: Discrepancies between the predicted and observed performance of bTB surveillance in Belgium were observed. Secondly, simulated alternatives revealed that targeted IFN-γ as well serological testing with Antibody ELISA towards risk herds would enable increasing the overall cost and effectiveness of the Belgian bTB surveillance system. Sensitivity analysis showed that results remained constant despite modification of some key assumptions. Discussion: Performance of current bTB surveillance system performance in Belgium was questionable. This exercise highlighted that not only sensitivity, but specificity is a key driver for surveillance performance. The quantitative and participative conceptual framework revealed itself a useful tool to allow evidence-based decision making regarding future tuberculosis surveillance in Belgium, as required by the international standards.

Bovine tuberculosis (bTB) challenges intensive dairy production in Ethiopia and implementation of the test and slaughter control strategy is not economically acceptable in the country. Vaccination of cattle with Bacillus Calmette-Guerin (BCG) could be an important adjunct to control, which would require a diagnostic test to differentiate Mycobacterium bovis (M. bovis)-infected and BCG-vaccinated animals (DIVA role). This study describes evaluation of a DIVA skin test (DST) that is based on a cocktail (DSTc) or fusion (DSTf) of specific (ESAT-6, CFP-10 and Rv3615c) M. bovis proteins in Zebu-Holstein crossbred cattle in Ethiopia. The study animals used were 74 calves (35 BCG-vaccinated and 39 unvaccinated) aged less than three weeks at the start and 68 known bTB positive cows. Six weeks after vaccination, the 74 calves were tested with DSTc and the single intradermal cervical comparative tuberculin (SICCT) test. The cows were tested with DSTc and SICCT test. Reactions to DSTc were not observed in BCG-vaccinated and unvaccinated calves while SICCT test reactions were detected in vaccinated calves. DSTc reactions were detected in 95.6% of the cows and single intradermal tuberculin (SIT) positive reactions were found in 98.2% (95% confidence interval, CI, 92.1–100%). The sensitivity of DSTc was 95.6% (95% CI, 87.6–99.1%), and significantly (P<0.001) higher than the sensitivity (75%, 95% CI, 63.0-84.7%) of the SICCT test at 4mm cutoff. DSTf and DSTc reactions were correlated (r = 0.75; 95% CI =0.53–0.88). In conclusion, DSTc could differentiate M. bovis-infected from BCG-vaccinated cattle in Ethiopia. DST had higher sensitivity than the SICCT test. Hence, DSTc could be used as a diagnostic tool for bTB if BCG vaccination is implemented for the control of bTB in Ethiopia and other countries.