First description of a lesion in the upper digestive mucosa associated with a novel gammaherpesvirus in a striped dolphin (Stenella coeruleoalba) stranded in the Western Mediterranean Sea.

BACKGROUND: A wide variety of lesions have been associated with herpesvirus in cetaceans. However, descriptions of herpesvirus infections in the digestive system of cetaceans are scarce. CASE REPORT: A young female striped dolphin stranded in the Valencian Community (Spain) on the 6th August 2021. The animal showed external macroscopic lesions suggestive of an aggressive interaction with bottlenose dolphins (rake marks in the epidermis). Internally, the main findings included congestion of the central nervous system and multiple, well-defined, whitish, irregularly shaped, proliferative lesions on the oropharyngeal and laryngopharyngeal mucosa. Histopathology revealed lymphoplasmacytic and histiocytic meningoencephalitis, consistent with neuro brucellosis. The oropharyngeal and laryngopharyngeal plaques were comprised histologically of focally extensive epithelial hyperplasia. As part of the health surveillance program tissue samples were tested for cetacean morbillivirus using a real-time reverse transcription-PCR, for Brucella spp. using a real-time PCR, and for herpesvirus using a conventional nested PCR. All samples were negative for cetacean morbillivirus; molecular positivity for Brucella spp. was obtained in pharyngeal tonsils and cerebrospinal fluid; herpesvirus was detected in a proliferative lesion in the upper digestive mucosa. Phylogenetic analysis showed that the herpesvirus sequence was included in the Gammaherpesvirinae subfamily. This novel sequence showed the greatest identity with other Herpesvirus sequences detected in skin, pharyngeal and genital lesions in five different species. CONCLUSIONS: To the best of the authors' knowledge, this is the first report of a proliferative lesion in the upper digestive mucosa associated with gammaherpesvirus posititvity in a striped dolphin (Stenella coeruleoalba).

Alpha- and gammaherpesviruses in stranded striped dolphins (Stenella coeruleoalba) from Spain: first molecular detection of gammaherpesvirus infection in central nervous system of odontocetes.

BACKGROUND: Herpesvirus infections in cetaceans have always been attributed to the Alphaherpesvirinae and Gammaherpesvirinae subfamilies. To date, gammaherpesviruses have not been reported in the central nervous system of odontocetes. CASE PRESENTATION: A mass stranding of 14 striped dolphins (Stenella coeruleoalba) occurred in Cantabria (Spain) on 18th May 2019. Tissue samples were collected and tested for herpesvirus using nested polymerase chain reaction (PCR), and for cetacean morbillivirus using reverse transcription-PCR. Cetacean morbillivirus was not detected in any of the animals, while gammaherpesvirus was detected in nine male and one female dolphins. Three of these males were coinfected by alphaherpesviruses. Alphaherpesvirus sequences were detected in the cerebrum, spinal cord and tracheobronchial lymph node, while gammaherpesvirus sequences were detected in the cerebrum, cerebellum, spinal cord, pharyngeal tonsils, mesenteric lymph node, tracheobronchial lymph node, lung, skin and penile mucosa. Macroscopic and histopathological post-mortem examinations did not unveil the potential cause of the mass stranding event or any evidence of severe infectious disease in the dolphins. The only observed lesions that may be associated with herpesvirus were three cases of balanitis and one penile papilloma. CONCLUSIONS: To the authors' knowledge, this is the first report of gammaherpesvirus infection in the central nervous system of odontocete cetaceans. This raises new questions for future studies about how gammaherpesviruses reach the central nervous system and how infection manifests clinically.

Immune related genes as markers for monitoring health status of honey bee colonies.

BACKGROUND: Honey bee population decline threatens the beekeeping sector, agriculture and global biodiversity. Early detection of colony mortality may facilitate rapid interventions to contain and prevent mortality spread. Among others, deformed wing virus (DWV) is capable of inducing colony losses, especially when combined with Varroa destructor mite. Since the bee immune system plays a crucial role in ensuring that bees are able to face these pathogens, we explored whether expression of immune genes could serve as biomarkers of colony health. RESULTS: Herein, we describe a preliminary immunological marker composed of two immune genes (relish and defensin), which provide insight on honey bee antiviral defense mechanism. Of the tested genes, relish expression correlated with the presence of DWV-Varroa complex, while decreased defensin expression correlated with poor resistance to this complex. CONCLUSIONS: The monitoring of these genes may help us to better understand the complex physiology of honey bees's immune system and to develop new approaches for managing the health impacts of DWV infection and varroa infestation in the field.

African swine fever virus transmission cycles in Central Europe: Evaluation of wild boar-soft tick contacts through detection of antibodies against Ornithodoros erraticus saliva antigen.

BACKGROUND: African swine fever (ASF) is one of the most complex viral diseases affecting both domestic and wild pigs. It is caused by ASF virus (ASFV), the only DNA virus which can be efficiently transmitted by an arthropod vector, soft ticks of the genus Ornithodoros. These ticks can be part of ASFV-transmission cycles, and in Europe, O. erraticus was shown to be responsible for long-term maintenance of ASFV in Spain and Portugal. In 2014, the disease has been reintroduced into the European Union, affecting domestic pigs and, importantly, also the Eurasian wild boar population. In a first attempt to assess the risk of a tick-wild boar transmission cycle in Central Europe that would further complicate eradication of the disease, over 700 pre-existing serum samples from wild boar hunted in four representative German Federal States were investigated for the presence of antibodies directed against salivary antigen of Ornithodoros erraticus ticks using an indirect ELISA format. RESULTS: Out of these samples, 16 reacted with moderate to high optical densities that could be indicative of tick bites in sampled wild boar. However, these samples did not show a spatial clustering (they were collected from distant geographical regions) and were of bad quality (hemolysis/impurities). Furthermore, all positive samples came from areas with suboptimal climate for soft ticks. For this reason, false positive reactions are likely. CONCLUSION: In conclusion, the study did not provide stringent evidence for soft tick-wild boar contact in the investigated German Federal States and thus, a relevant involvement in the epidemiology of ASF in German wild boar is unlikely. This fact would facilitate the eradication of ASF in the area, although other complex relations (wild boar biology and interactions with domestic pigs) need to be considered.

Evidence of shared bovine viral diarrhea infections between red deer and extensively raised cattle in south-central Spain.

BACKGROUND: Bovine viral diarrhea virus (BVDV) is a pestivirus that affects cattle production worldwide and that can infect other ungulates such as cervids and even wild boar (Sus scrofa). It is believed that domestic livestock can become infected through contact with wild animals, though it is known that infection can spread among wild animals in the absence of contact with livestock. Little is known about the sharing of BVDV infection between wild and domestic animals in the same habitat, which is important for designing eradication campaigns and preventing outbreaks, especially on hunting estates with high animal densities. RESULTS: We assessed the sharing of BVDV infections among hunted red deer, wild boar and cattle in south-central Spain. Sampled red deer (Cervus elaphus; n = 267) and wild boar (n = 52) were located on 19 hunting estates, and cattle (n = 180) were located on 18 nearby farms. We used ELISA kits for the serological screening, Taqman RT-PCR assay for the virus determination, and subsequent phylogenetic analysis for 17 RT-PCR positive sample amplicons. Fifty-two red deer (19.5%) and 82 cattle (45.6%) samples tested positive by ELISA. A high apparent prevalence (22.47%) was obtained for red deer, while only five cattle farms tested positive by RT-PCR. Conversely, no wild boar tested positive by both ELISA or RT-PCR. Eleven red deer (4.1%) tested positive by both ELISA and RT-PCR; these animals may have been sampled during the last phase of viremia, or they may represent previously exposed individuals infected by a different BVDV strain. The amplicons shared 92.7-100% identity and fell within the BVDV subgroup 1b, although nine of these (from four red deer and five cattle pools) formed a separate branch. This suggests that there might be a common BVDV infecting both cattle and red deer. Higher red deer abundance was significantly associated with greater risk that extensively raised cattle would test positive for BVDV by ELISA. CONCLUSIONS: Our findings suggest that BVDV is circulating between cattle and red deer populations in proximity, but further work is required to determine whether they share the same strain(s). These results suggest the potential of BVDV to serve as a surveillance marker in these shared habitats. High seroprevalence of BVDV in red deer from our study area suggests that although BVDV infection is common, animals usually survive the infection. Further research is needed to verify and investigate the role of red deer as a BVDV reservoir.

Fluorescent microbead-based immunoassay for anti-Erysipelothrix rhusiopathiae antibody detection in cetaceans.

A fluorescent microbead-based immunoassay (FMIA) for detection of anti-Erysipelothrix rhusiopathiae antibodies in pigs was adapted for use in cetaceans. The FMIA was validated and adjusted using serum samples from 10 vaccinated captive bottlenose dolphins Tursiops truncatus collected between 1 and 13 mo after immunization. The technique was then used to analyze specimens from 15 free-ranging cetaceans stranded alive on the Valencian Mediterranean coast between 2006 and 2014: 11 striped dolphins Stenella coeruleoalba, 3 Risso's dolphins Grampus griseus and 1 bottlenose dolphin Tursiops truncatus. One of these wild animals was confirmed to have died from E. rhusiopathiae septicemia, but no anti-E. rhusiopathiae antibodies were detected in its serum, pericardial fluid or milk samples. Another free-ranging individual, which lacked any signs or lesions that might be indicative of E. rhusiopathiae infection, showed high fluorescence intensity similar to that measured in captive dolphins at 6-13 mo after vaccination. These results suggest that this animal underwent an E. rhusiopathiae infection several months before stranding. The findings in the present study suggest that FMIA can be useful for detecting anti-E. rhusiopathiae antibodies in cetaceans, and its application to free-ranging animals is particularly interesting because of the great value of these specimens. Furthermore, the FMIA can be multiplexed to allow the determination of up to 100 analytes per sample in a single well, thereby reducing the cost, time and sample volume needed.

First molecular determination of herpesvirus from two mysticete species stranded in the Mediterranean Sea.

BACKGROUND: Herpesvirus can infect a wide range of animal species: mammals, birds, reptiles, fish, amphibians and bivalves. In marine mammals, several alpha- and gammaherpesvirus have been identified in some cetaceans and pinnipeds species. To date, however, this virus has not been detected in any member of the Balaenoptera genus. CASE PRESENTATION: Herpesvirus was determined by molecular methods in tissue samples from a male fin whale juvenile (Balaenoptera physalus) and a female common minke whale calf (Balaenoptera acutorostrata) stranded on the Mediterranean coast of the Region of Valencia (Spain). Samples of skin and penile mucosa from the fin whale and samples of skin, muscle and central nervous system tissue from the common minke whale tested positive for herpesvirus based on sequences of the DNA polymerase gene. Sequences from fin whale were identical and belonged to the Alphaherpesvirinae subfamily. Only members of the Gammaherpesvirinae subfamily were amplified from the common minke whale, and sequences from the muscle and central nervous system were identical. Sequences in GenBank most closely related to these novel sequences were viruses isolated from other cetacean species, consistent with previous observations that herpesviruses show similar phylogenetic branching as their hosts. CONCLUSIONS: To our knowledge, this is the first molecular determination of herpesvirus in the Balaenoptera genus. It shows that herpesvirus should be included in virological evaluation of these animals.

Novel adenovirus detected in captive bottlenose dolphins (Tursiops truncatus) suffering from self-limiting gastroenteritis.

BACKGROUND: Adenoviruses are common pathogens in vertebrates, including humans. In marine mammals, adenovirus has been associated with fatal hepatitis in sea lions. However, only in rare cases have adenoviruses been detected in cetaceans, where no clear correlation was found between presence of the virus and disease status. CASE PRESENTATION: A novel adenovirus was identified in four captive bottlenose dolphins with self-limiting gastroenteritis. Viral detection and identification were achieved by: PCR-amplification from fecal samples; sequencing of partial adenovirus polymerase (pol) and hexon genes; producing the virus in HeLa cells, with PCR and immunofluorescence detection, and with sequencing of the amplified pol and hexon gene fragments. A causative role of this adenovirus for gastroenteritis was suggested by: 1) we failed to identify other potential etiological agents; 2) the exclusive detection of this novel adenovirus and of seropositivity for canine adenoviruses 1 and 2 in the four sick dolphins, but not in 10 healthy individuals of the same captive population; and 3) the virus disappeared from feces after clinical signs receded. The partial sequences of the amplified fragments of the pol and hexon genes were closest to those of adenoviruses identified in sea lions with fatal adenoviral hepatitis, and to a Genbank-deposited sequence obtained from a harbour porpoise. CONCLUSION: These data suggest that adenovirus can cause self-limiting gastroenteritis in dolphins. This adenoviral infection can be detected by serology and by PCR detection in fecal material. Lack of signs of hepatitis in sick dolphins may reflect restricted tissue tropism or virulence of this adenovirus compared to those of the adenovirus identified in sea lions. Gene sequence-based phylogenetic analysis supports a common origin of adenoviruses that affect sea mammals. Our findings suggest the need for vigilance against adenoviruses in captive and wild dolphin populations.

First molecular detection and characterization of herpesvirus and poxvirus in a Pacific walrus (Odobenus rosmarus divergens).

BACKGROUND: Herpesvirus and poxvirus can infect a wide range of species: herpesvirus genetic material has been detected and amplified in five species of the superfamily Pinnipedia; poxvirus genetic material, in eight species of Pinnipedia. To date, however, genetic material of these viruses has not been detected in walrus (Odobenus rosmarus), another marine mammal of the Pinnipedia clade, even though anti-herpesvirus antibodies have been detected in these animals. CASE PRESENTATION: In February 2013, a 9-year-old healthy captive female Pacific walrus died unexpectedly at L'Oceanografic (Valencia, Spain). Herpesvirus was detected in pharyngeal tonsil tissue by PCR. Phylogenetic analysis revealed that the virus belongs to the subfamily Gammaherpesvirinae. Poxvirus was also detected by PCR in skin, pre-scapular and tracheobronchial lymph nodes and tonsils. Gross lesions were not detected in any tissue, but histopathological analyses of pharyngeal tonsils and lymph nodes revealed remarkable lymphoid depletion and lymphocytolysis. Similar histopathological lesions have been previously described in bovine calves infected with an alphaherpesvirus, and in northern elephant seals infected with a gammaherpesvirus that is closely related to the herpesvirus found in this case. Intracytoplasmic eosinophilic inclusion bodies, consistent with poxviral infection, were also observed in the epithelium of the tonsilar mucosa. CONCLUSION: To our knowledge, this is the first molecular identification of herpesvirus and poxvirus in a walrus. Neither virus was likely to have contributed directly to the death of our animal.

Modular framework to assess the risk of African swine fever virus entry into the European Union.

BACKGROUND: The recent occurrence and spread of African swine fever (ASF) in Eastern Europe is perceived as a serious risk for the pig industry in the European Union (EU). In order to estimate the potential risk of ASF virus (ASFV) entering the EU, several pathways of introduction were previously assessed separately. The present work aimed to integrate five of these assessments (legal imports of pigs, legal imports of products, illegal imports of products, fomites associated with transport and wild boar movements) into a modular tool that facilitates the visualization and comprehension of the relative risk of ASFV introduction into the EU by each analyzed pathway. RESULTS: The framework's results indicate that 48% of EU countries are at relatively high risk (risk score 4 or 5 out of 5) for ASFV entry for at least one analyzed pathway. Four of these countries obtained the maximum risk score for one pathway: Bulgaria for legally imported products during the high risk period (HRP); Finland for wild boar; Slovenia and Sweden for legally imported pigs during the HRP. Distribution of risk considerably differed from one pathway to another; for some pathways, the risk was concentrated in a few countries (e.g., transport fomites), whereas other pathways incurred a high risk for 4 or 5 countries (legal pigs, illegal imports and wild boar). CONCLUSIONS: The modular framework, developed to estimate the risk of ASFV entry into the EU, is available in a public domain, and is a transparent, easy-to-interpret tool that can be updated and adapted if required. The model's results determine the EU countries at higher risk for each ASFV introduction route, and provide a useful basis to develop a global coordinated program to improve ASFV prevention in the EU.

Geospatial analysis for strategic wildlife disease surveillance: African swine fever in South Korea (2019-2021).

Since the confirmation of African swine fever (ASF) in South Korea in 2019, its spread, predominantly in wild boars, has been a significant concern. A key factor in this situation is the lack of identification of risk factors by surveillance bias. The unique orography, characterized by high mountains, complicates search efforts, leading to overlooked or delayed case detection and posing risks to the swine industry. Additionally, shared rivers with neighboring country present a continual threat of virus entry. This study employs geospatial analysis and statistical methods to 1) identify areas at high risk of ASF occurrence but possibly under-surveilled, and 2) indicate strategic surveillance points for monitoring the risk of ASF virus entry through water bodies and basin influences. Pearson's rho test indicated that elevation (rho = -0.908, p-value < 0.001) and distance from roads (rho = -0.979, p-value < 0.001) may have a significant impact on limiting surveillance activities. A map of potential under-surveilled areas was created considering these results and was validated by a chi-square goodness-of-fit test (X-square = 208.03, df = 1, p-value < 0.001). The strong negative correlation (rho = -0.997, p-value <0.001) between ASF-positive wild boars and distance from water sources emphasizes that areas surrounding rivers are one of the priority areas for monitoring. The subsequent hydrological analyses provided important points for monitoring the risk of virus entry via water from the neighboring country. This research aims to facilitate early detection and prevent further spread of ASF.

Landscape connectivity for predicting the spread of ASF in the European wild boar population.

African swine fever (ASF) is an infectious and highly fatal disease affecting wild and domestic swine, which is unstoppably spreading worldwide. In Europe, wild boars are one of the main drivers of spread, transmission, and maintenance of the disease. Landscape connectivity studies are the main discipline to analyze wild-species dispersal networks, and it can be an essential tool to predict dispersal-wild boar movement routes and probabilities and therefore the associated potential ASF spread through the suitable habitat. We aimed to integrate wild boar habitat connectivity predictions with their occurrence, population abundance, and ASF notifications to calculate the impact (i.e., the capacity of a landscape feature to favor ASF spread) and the risk (i.e., the likelihood of a habitat patch becoming infected) of wild boar infection across Europe. Furthermore, we tested the accuracy of the risk of infection by comparing the results with the temporal distribution of ASF cases. Our findings identified the areas with the highest impact and risk factors within Europe's central and Eastern regions where ASF is currently distributed. Additionally, the impact factor was 31 times higher on habitat patches that were infected vs non-infected, proving the utility of the proposed approach and the key role of wild boar movements in ASF-spread. All data and resulting maps are openly accessible and usable.

Application of machine learning with large-scale data for an effective vaccination against classical swine fever for wild boar in Japan.

Classical swine fever has been spreading across the country since its re-emergence in Japan in 2018. Gifu Prefecture has been working diligently to control the disease through the oral vaccine dissemination targeting wild boars. Although vaccines were sprayed at 14,000 locations between 2019 and 2020, vaccine ingestion by wild boars was only confirmed at 30% of the locations. Here, we predicted the vaccine ingestion rate at each point by Random Forest modeling based on vaccine dissemination data and created prediction surfaces for the probability of vaccine ingestion by wild boar using spatial interpolation techniques. Consequently, the distance from the vaccination point to the water source was the most important variable, followed by elevation, season, road density, and slope. The area under the curve, model accuracy, sensitivity, and specificity for model evaluation were 0.760, 0.678, 0.661, and 0.685, respectively. Areas with high probability of wild boar vaccination were predicted in northern, eastern, and western part of Gifu. Leave-One-Out Cross Validation results showed that Kriging approach was more accurate than the Inverse distance weighting method. We emphasize that effective vaccination strategies based on epidemiological data are essential for disease control and that our proposed tool is also applicable for other wildlife diseases.

Unusual striped dolphin mass mortality episode related to cetacean morbillivirus in the Spanish Mediterranean sea.

BACKGROUND: In the last 20 years, Cetacean Morbillivirus (CeMV) has been responsible for many die-offs in marine mammals worldwide, as clearly exemplified by the two dolphin morbillivirus (DMV) epizootics of 1990-1992 and 2006-2008, which affected Mediterranean striped dolphins (Stenella coeruleoalba). Between March and April 2011, the number of strandings on the Valencian Community coast (E Spain) increased. CASE PRESENTATION: Necropsy and sample collection were performed in all stranded animals, with good state of conservation. Subsequently, histopathology, immunohistochemistry, conventional reverse transcription polymerase chain reaction (RT-PCR) and Universal Probe Library (UPL) RT-PCR assays were performed to identify Morbillivirus. Gross and microscopic findings compatible with CeMV were found in the majority of analyzed animals. Immunopositivity in the brain and UPL RT-PCR positivity in seven of the nine analyzed animals in at least two tissues confirmed CeMV systemic infection. Phylogenetic analysis, based on sequencing part of the phosphoprotein gene, showed that this isolate is a closely related dolphin morbillivirus (DMV) to that responsible for the 2006-2008 epizootics. CONCLUSION: The combination of gross and histopathologic findings compatible with DMV with immunopositivity and molecular detection of DMV suggests that this DMV strain could cause this die-off event.

Detection and assessment of electrocution in endangered raptors by infrared thermography.

BACKGROUND: Most European birds of prey find themselves in a poor state of conservation, with electrocution as one of the most frequent causes of unnatural death. Since early detection of electrocution is difficult, treatment is usually implemented late, which reduces its effectiveness. By considering that electrocution reduces tissue temperature, it may be detectable by thermography, which would allow a more rapid identification. Three individuals from three endangered raptor species [Spanish imperial eagle (Aquila adalberti), Lammergeier (Gypaetus barbatus) and Osprey (Pandion haliaetus)] were studied thermographically from the time they were admitted to a rehabilitation centre to the time their clinical cases were resolved. CASES PRESENTATION: The three raptors presented lesions lacking thermal bilateral symmetry and were consistent with electrocution of feet, wings and eyes, visible by thermography before than clinically; lesions were well-defined and showed a lower temperature than the surrounding tissue. Some lesions evolved thermally and clinically until the appearance of normal tissue recovered, while others evolved and became necrotic. A histopathological analysis of a damaged finger amputated off a Lammergeier, and the necropsy and histopathology examination of an osprey, confirmed the electrocution diagnosis. CONCLUSIONS: These results suggest that thermography is effective and useful for the objective and early detection and monitoring of electrocuted birds, and that it may prove especially useful for examining live animals that require no amputation or cannot be subjected to invasive histopathology.

Two cases of pseudohermaphroditism in loggerhead sea turtles Caretta caretta.

Two juvenile (curved carapace lengths: 28 and 30 cm) loggerhead sea turtles Caretta caretta with precocious male external characteristics were admitted to the ARCA del Mar rescue area at the Oceanogràfic Aquarium in Valencia, Spain, in 2009 and 2010. Routine internal laparoscopic examination and subsequent histopathology confirmed the presence of apparently healthy internal female gonads in both animals. Extensive tissue biopsy and hormone induction assays were consistent with female sex. To the best of our knowledge, this is the first report of pseudohermaphroditism in loggerhead sea turtles based on sexual external characteristics and internal laparoscopic examination. Our findings suggest that the practice of using external phenotypical characteristics as the basis for gender identification in sea turtles should be reevaluated. Future research should focus on detecting more animals with sexual defects and their possible effects on the sea turtle population.

What can we learn from the five-year African swine fever epidemic in Asia?

Today's global swine industry is exposed to the unprecedented threat of African swine fever (ASF). Asia, the site of the most recent epidemics, could serve as a huge viral reservoir for the rest of the world given the severity of the damage, the huge swine industry, and the high volume of trade with other countries around the world. As the majority of ASF notifications in Asia today originate from pig farms, the movement of live pigs and associated pork products are considered critical control points for disease management. Particularly, small-scale or backyard farms with low biosecurity levels are considered major risk factors. Meanwhile, wild boars account for most notified cases in some countries and regions, which makes the epidemiological scenario different from that in other Asian countries. As such, the current epidemic situation and higher risk factors differ widely between these countries. A variety of studies on ASF control have been conducted and many valuable insights have been obtained in Asia; nevertheless, the overall picture of the epidemic is still unclear. The purpose of this review is to provide an accurate picture of the epidemic situation across Asia, focusing on each subregion to comprehensively explain the disease outbreak. The knowledge gained from the ASF epidemics experienced in Asia over the past 5 years would be useful for disease control in areas that are already infected, such as Europe, as well as for non-affected areas to address preventive measures. To this end, the review includes two aspects: a descriptive analytical review based on publicly available databases showing overall epidemic trends, and an individualized review at the subregional level based on the available literature.

A useful tool for the safe diagnosis and control of the two main pandemics of the XXI century: COVID-19 and African Swine Fever disease.

The COVID-19 pandemic and the disease triggered by the African Swine Fever virus are currently two of the main problems regarding public and animal health, respectively. Although vaccination seems to be the ideal tool for controlling these diseases, it has several limitations. Therefore, early detection of the pathogen is critical in order to apply preventive and control measures. Real-time PCR is the main technique used for the detection of both viruses, which requires previous processing of the infectious material. If the potentially infected sample is inactivated at the time of sampling, the diagnosis will be accelerated, impacting positively on the diagnosis and control of the disease. Here, we evaluated the inactivation and preservation properties of a new surfactant liquid for non-invasive and environmental sampling of both viruses. Our results demonstrated that the surfactant liquid effectively inactivates SARS-CoV-2 and African Swine Fever virus in only five minutes, and allows for the preservation of the genetic material for long periods even at high temperatures such as 37°C. Hence, this methodology is a safe and useful tool for recovering SARS-CoV-2 and African Swine Fever virus RNA/DNA from different surfaces and skins, which has significant applied relevance in the surveillance of both diseases.

Quantitative risk assessment of African swine fever introduction into Spain by legal import of swine products.

African swine fever (ASF) is currently threatening the global swine industry. Its unstoppable global spread poses a serious risk to Spain, one of the world's leading producers. Over the past years, there has been an increased global burden of ASF not only in swine but also swine products. Unfortunately, many pigs are not diagnosed before slaughter and their products are used for human consumption. These ASF-contaminated products are only a source for new ASF outbreaks when they are consumed by domestic pigs or wild boar, which may happen either by swill feeding or landfill access. This study presents a quantitative stochastic risk assessment model for the introduction of ASF into Spain via the legal import of swine products, specifically pork and pork products. Entry assessment, exposure assessment, consequence assessment and risk estimation were carried out. The results suggest an annual probability of ASF introduction into Spain of 1.74 × 10-4, the highest risk being represented by Hungary, Portugal, and Poland. Monthly risk distribution is homogeneously distributed throughout the year. Illegal trade and pork product movement for own consumption (e.g., air and ship passenger luggage) have not been taken into account due to the lack of available, accredited data sources. This limitation may have influenced the model's outcomes and, the risk of introduction might be higher than that estimated. Nevertheless, the results presented herein would contribute to allocating resources to areas at higher risk, improving prevention and control strategies and, ultimately, would help reduce the risk of ASF introduction into Spain.

Epidemiological impacts of attenuated African swine fever virus circulating in wild boar populations.

African swine fever virus (ASFV) genotype II has been present in wild boar in the European Union since 2014. Control measures have reduced the incidence of the ASF, but highly virulent as well as attenuated ASFV strains continue to circulate. We present the intraherd epidemiological parameters of low and highly virulent ASFV in wild boar from experimental data, and for the first time, evaluate the impact of attenuated strain circulation through unique deterministic compartmental model simulations under various potential scenarios and hypotheses. Using an estimated PCR infectious threshold of TPCR = 36.4, we obtained several transmission parameters, like an Rx (experimental intraherd R0) value of 4.5. We also introduce two novel epidemiological parameters: infectious power and resistance power, which indicate the ability of animals to transmit the infection and the reduction in infectiousness after successive exposures to varying virulence strains, respectively. The presence of ASFV attenuated strains results in 4-17% of animals either remaining in a carrier state or becoming susceptible again when exposed to highly virulent ASFV for more than two years. The timing between exposures to viruses of different virulence also influences the percentage of animals that die or remain susceptible. The findings of this study can be utilized in epidemiological modelling and provide insight into important risk situations that should be considered for surveillance and future potential ASF vaccination strategies in wild boar.