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To honour the 100 years anniversary of the first publication about African swine fever (ASF) a webinar with a particular focus on disease control in the smallholder sector was organized. This article is based on the webinar, summarizing the early history of ASF research, reflecting on the current global disease situation and bringing forward some suggestions that could contribute towards achieving control of ASF. The first description of ASF by R. Eustace Montgomery in 1921 laid the foundations for what we know about the disease today. Subsequent research confirmed its association with warthogs and soft ticks of the Ornithodoros moubata complex. During the latter half of the 21st century, exponential growth of pig production in Africa has led to a change in the ASF-epidemiology pattern. It is now dominated by a cycle involving domestic pigs and pork with virus spread driven by people. In 2007, a global ASF epidemic started, reaching large parts of Europe, Asia and the Americas. In Europe, this epidemic has primarily affected wild boar. In Asia, wild boar, smallholders and industrialized pig farms have been affected with impact on local, national and international pig value chains. Globally and historically, domestic pigs in smallholder settings are most frequently affected and the main driver of ASF virus transmission. Awaiting a safe and efficacious vaccine, we need to continue focus on other measures, such as biosecurity, for controlling the disease. However, smallholders face specific challenges linked to poverty and other structural factors in implementing biosecurity measures that can prevent spread. Improving biosecurity in the smallholder sector thus remains an important tool for preventing and controlling ASF. In this regard, interdisciplinary research can help to find new ways to promote safe practices, facilitate understanding and embrace smallholders' perspectives, engage stakeholders and adjust prevention and control policies to improve implementation.

Following the first detection of

EFSA received a mandate from the European Commission to assess the effectiveness of some of the control measures against diseases included in the Category A list according to Regulation (EU) 2016/429 on transmissible animal diseases ('Animal Health Law’). This opinion belongs to a series of opinions where these control measures will be assessed, with this opinion covering the assessment of control measures for glanders. In this opinion, EFSA and the AHAW Panel of experts review the effectiveness of: (i) clinical and laboratory sampling procedures, (ii) monitoring period and (iii) the minimum radius of the protection and surveillance zone, and the minimum length of time the measures should be applied in these zones. The general methodology used for this series of opinions has been published elsewhere. Considering the epidemiology and distribution of glanders, it was foreseen that three different situations could lead to a suspicion of the disease. Sampling procedures were defined for each of the three different suspicion types, which can also be applied in most of the other scenarios assessed. The monitoring period (6 months) was assessed as effective in all scenarios. The AHAW Panel of experts considered the minimum radius and duration of the existing protection and surveillance zone, set at the establishment level, effective. Recommendations provided for each of the scenarios assessed aim to support the European Commission in the drafting of further pieces of legislation, as well as for plausible ad hoc requests in relation to glanders.

African swine fever vaccines could pose risk of causing disease and spreading the virus further

Small ruminants support the livelihoods of millions of poor pastoralist and sedentary households around the world. While pastoralists are generally not amongst the poorest in terms of assets, they are frequently marginalised in terms of their access to political power, health and education. This study was undertaken among pastoralist households keeping small ruminants in four regions of the country of Georgia. Small ruminants are an important cultural, social and economic asset in Georgia and are mainly managed in a transhumant pastoralist system. Georgia suffered its first, and so far only outbreak of peste des petits ruminants (PPR) in 2016. This qualitative interview study was designed to acquire contextual understanding of local small ruminant husbandry and the livelihood situations of the participating pastoralists, and to detect historical, unreported PPR outbreaks. Focus group discussions comprising participatory epidemiology tools and other forms of interviews were used to explore small ruminant management, disease spectrum and management, and animal health priorities. The participants had experienced a wide variety of animal health constraints, with intestinal worms, braxy, piroplasmosis, pasture-related problems, predators and lameness emerging as priorities. No historic, unreported PPR outbreak was detected in this study, and PPR was not a priority for participants. Instead, the day-to-day reality of animal health for the pastoralists was characterised by co-infections of mainly endemic pathogens, and problems related to other challenges such as access to land, feed and genetic resources. The rationale behind the participants’ prioritisation of animal health problems was supported by the need to pay extra attention to animals in order to avoid risk factors, keep animals healthy and minimise the negative impact of diseases or management problems; the various epidemiological and clinical parameters of the prioritised diseases; the economic impact of the specific problems and the zoonotic potential of diseases and predation. Even within regions, and within seemingly socially and culturally homogenous groups, there were important local differences in the problems faced by pastoralists that affect their livestock management. This study underlines the importance of a contextualised understanding of the local disease panorama and complexities in the livelihood situations of rural people when designing actions to improve animal health in general or, more specifically, passive surveillance as well as prevention or control measures. Finally, it is concluded that to achieve such an understanding, there is a need for participatory, scoping-style studies that specifically acknowledge diversity and power relations.

To monitor a state of disease freedom and to ensure a timely detection of new introductions of disease, sur-veillance programmes need be evaluated prior to implementation. We present a strategy to evaluate surveillance of Mycobacterium avium subsp. paratuberculosis (MAP) using simulated testing of bulk milk in an infectious dis-ease spread model. MAP is a globally distributed, chronic infectious disease with substantial animal health impact. Designing surveillance for this disease poses specific challenges because methods for surveillance evaluation have focused on estimating surveillance system sensitivity and probability of freedom from disease and do not account for spread of disease or complex and changing population structure over long periods. The aims of the study were to 1. define a model that describes the spread of MAP within and between Swedish herds; 2. define a method for simulation of imperfect diagnostic testing in this framework; 3. to compare surveillance strategies to support surveillance design choices. The results illustrate how this approach can be used to identify differences between the probability of detecting disease in the population based on choices of the number of herds sampled and the use of risk-based or random selection of these herds. The approach was also used to assess surveillance to detect introduction of disease and to detect a very low prevalence endemic state. The use of bulk milk sampling was determined to be an effective method to detect MAP in the population with as few as 500 herds tested per year if the herd-level prevalence was 0.2 %. However, detection of point introductions in the population was unlikely in the 13-year simulation period even if as many as 2000 herds were tested per year. Interestingly, the use of a risk-based selection strategy was found to be a disadvantage to detect MAP given the modelled disease dynamics.&nbsp

Rift Valley fever (RVF) is a vector-borne disease transmitted by a broad spectrum of mosquito species, especially Aedes and Culex genus, to animals (domestic and wild ruminants and camels) and humans. Rift Valley fever is endemic in sub-Saharan Africa and in the Arabian Peninsula, with periodic epidemics characterised by 5–15 years of inter-epizootic periods. In the last two decades, RVF was notified in new African regions (e.g. Sahel), RVF epidemics occurred more frequently and low-level enzootic virus circulation has been demonstrated in livestock in various areas. Recent outbreaks in a French overseas department and some seropositive cases detected in Turkey, Tunisia and Libya raised the attention of the EU for a possible incursion into neighbouring countries. The movement of live animals is the most important pathway for RVF spread from the African endemic areas to North Africa and the Middle East. The movement of infected animals and infected vectors when shipped by flights, containers or road transport is considered as other plausible pathways of introduction into Europe. The overall risk of introduction of RVF into EU through the movement of infected animals is very low in all the EU regions and in all MSs (less than one epidemic every 500 years), given the strict EU animal import policy. The same level of risk of introduction in all the EU regions was estimated also considering the movement of infected vectors, with the highest level for Belgium, Greece, Malta, the Netherlands (one epidemic every 228–700 years), mainly linked to the number of connections by air and sea transports with African RVF infected countries. Although the EU territory does not seem to be directly exposed to an imminent risk of RVFV introduction, the risk of further spread into countries neighbouring the EU and the risks of possible introduction of infected vectors, suggest that EU authorities need to strengthen their surveillance and response capacities, as well as the collaboration with North African and Middle Eastern countries. info:eu-repo/semantics/publishedVersion

International audience; African swine fever is a devastating disease that can result in death in almost all infected pigs. The continuing spread of African swine fever from Africa to Europe and recently to the high-pig production countries of China and others in Southeast Asia threatens global pork production and food security. The African swine fever virus is an unusual complex DNA virus and is not related to other viruses. This has presented challenges for vaccine development, and currently none is available. The virus is extremely well adapted to replicate in its hosts in the sylvatic cycle in East and South Africa. Its spread to other regions, with different wildlife hosts, climatic conditions, and pig production systems, has revealed unexpected epidemiological scenarios and different challenges for control. Here we review the epidemiology of African swine fever in these different scenarios and methods used for control. We also discuss progress toward vaccine development and research priorities to better understand this complex disease and improve control.

African swine fever (ASF) is a devastating disease of pigs. Without a vaccine, early detection and rapid diagnosis of ASF is a crucial step towards effective disease control. In many countries where ASF is endemic, laboratory infrastructure including sampling and sample shipment is inadequate, and a rapid laboratory confirmation would require that the diagnosis is performed at regional laboratories close to the pig farms of concern, or even at the farm-side. This study intended to evaluate measures including sample preparation methods, a dried-down assay, and a portable, battery-powered real-time PCR instrument, to improve molecular diagnosis under field conditions. A simple dilution of blood samples, either in Phosphate-buffered saline or a commercial buffer, worked similarly to beads-based nucleic acid extraction using a magnet as the core equipment; the latter method did work as well for those samples with low viral load or high Ct values. The real-time PCR assay using a Universal ProbeLibrary (UPL) probe tolerated suspected inhibitory substances present in the prepared samples better, whereas the dried-down assay had a higher diagnostic sensitivity. Additionally, an inhibition control assay proved to be helpful in avoiding false negative results when interpreting negative results of samples that might be of low quality or with inadequate reduction in inhibitory substances. When tested with synthetic DNA standards, the portable instrument performed at a level approaching stationary thermocyclers. In summary, the developments of suitable sample preparation methods, robust and thermal-stable real-time PCR assays with inhibition control, and battery-powered portable thermocyclers with middle-throughput offer one way forward to provide rapid, reliable molecular diagnosis under challenging field conditions.

The African swine fever epizootic in central and eastern European Union member states has a newly identified component involving virus transmission by wild boar and virus survival in the environment. Insights led to an update of the 3 accepted African swine fever transmission models to include a fourth cycle: wild boar-habitat.

Background In Africa, intensified pig production is frequently accompanied by increased occurrence of African swine fever (ASF) outbreaks, leading to high case fatality rates and socio-economic impact for the farmers. ASF control relies on prevention of disease transmission and control of outbreaks. The aim of this study was to increase the understanding on how the knowledge of ASF epidemiology and control can be transferred into successfully implemented biosecurity interventions on farm and community level. Structured interviews with 200 randomly selected, pig-keeping households in northern Uganda were undertaken three times. Perceptions related to general biosecurity and hypothetical control interventions and attitudes towards pig farming were investigated by measuring the agreement to statements using a Likert scale. Results Respondents generally conveyed positivism towards pig farming, biosecurity, and the potential of biosecurity for preventing ASF outbreaks. These positive attitudes, as well as the will to invest in biosecurity, were reduced in households that had experienced ASF outbreaks. Among the control interventions change of boots before entering the pig stable was highly accepted and seasonal adaptation of pig rearing times accepted on medium level. Statements on preventive sales of healthy pigs in connection with outbreaks and on buying pork products from slaughter operations receiving ASF-contact pigs received low acceptance, increasing, however, for households that had experienced ASF outbreaks. Consumption of pork from ASF infected pigs was generally not accepted, medium level of agreement was expressed for statements on the zoonotic potential of ASF and for neutralizing ASF by cooking. Conclusions To gain in-depth understanding of the complexity of people’s behaviour, reasoning and decision-making processes, deeper involvement of the social sciences and a qualitative research approach might be used for further studies. Communicating information regarding the ASF not being zoonotic, and how the virus is neutralized will be important for increasing acceptance and enhancing implementation for the hypothetical control interventions preventive sales, safe slaughter, and consumption of processed and safe pork. Likewise, participatory development to adopt any control interventions to the local context on community level will be necessary for successful implementation. Electronic supplementary material The online version of this article (10.1186/s12917-019-2005-7) contains supplementary material, which is available to authorized users.

African swine fever (ASF) was first described in 1921 as a highly fatal and contagious disease which caused severe outbreaks among settlers’ pigs in British East Africa. Since then the disease has expanded its geographical distribution and is currently present in large parts of Africa, Europe and Asia and considered a global threat. Although ASF is typically associated with very high case fatality rates, a certain proportion of infected animals will recover from the infection and survive. Early on it was speculated that such survivors may act as carriers of the virus, and the importance of such carries for disease persistence and spread has since then almost become an established truth. However, the scientific basis for such a role of carriers may be questioned. With this in mind, the objective of this study was to review the available literature in a systematic way and to evaluate the available scientific evidence. The selection of publications for the review was based on a database search, followed by a stepwise screening process in order to exclude duplicates and non-relevant publications based on pre-defined exclusion criteria. By this process the number of publications finally included was reduced from the 3664 hits identified in the initial database search to 39 publications, from which data was then extracted and analysed. Based on this it was clear that a definition of an ASF virus carrier is lacking, and that in general any survivor or seropositive animal has been referred to as carrier. It was also clear that evidence of any significant role of such a carrier is absent. Two types of “survivors” could be defined: 1) pigs that do not die but develop a persistent infection, characterised by periodic viraemia and often but not always accompanied by some signs of subacute to chronic disease, and 2) pigs which clear the infection independently of virulence of the virus, and which are not persistently infected and will not present with prolonged virus excretion. There is no evidence that suggests that any of these categories of survivors can be considered as “healthy” carriers, i.e. pigs that show no sign of disease but can transmit the virus to in-contact pigs. However, localized virus persistence in lymphoid tissues may occur to some extent in any of the categories of survivors, which in theory may cause infection after oral uptake. To what extent this is relevant in reality, however, can be questioned given the virus dose generally needed for oral infection.

In 2007 African swine fever (ASF) arrived at a Black Sea harbour in Georgia and in 2014 the infection reached the European Union (EU), where it still expands its territory. ASF is a fatal viral disease affecting domestic pigs and wild boar of all ages with clinical presentations ranging from per-acute to chronic disease, including apparently asymptomatic courses. Until the detection of the first case inside the EU, infections in the current epidemic were mainly seen among pig farms with generally low biosecurity, and with incidental spill over to the wild boar population. In the EU, however, the infection survived locally in the wild boar population independently from outbreaks in domestic pigs, with a steady and low prevalence. Apart from the wild boar population and the habitat, the current epidemic recognizes humans as the main responsible for both long distance transmission and virus introduction in the domestic pig farms. This underlines the importance to include social science when planning ASF-prevention, −control, or -eradication measures. Based on experiences, knowledge and data gained from the current epidemic this review highlights some recent developments in the epidemiological understanding of ASF, especially concerning the role of wild boar and their habitats in ASF epidemiology. In this regard, the qualities of three epidemiological traits: contagiousity, tenacity, and case fatality rate, and their impact on ASF persistence and transmission are especially discussed.

Due to the current unavailability of vaccines or treatments for African swine fever (ASF), which is caused by African swine fever virus (ASFV), rapid and reliable detection of the virus is essential for timely implementation of emergency control measures and differentiation of ASF from other swine diseases with similar clinical presentations. Here, an improved PCR assay was developed and evaluated for sensitive and universal detection of ASFV. Primers specific for ASFV were designed based on the highly conserved region of the vp72 gene sequences of all ASFV strains available in GenBank, and the PCR assay was established and compared with two OIE-validated PCR tests. The analytic detection limit of the PCR assay was 60 DNA copies per reaction. No amplification signal was observed for several other porcine viruses. The novel PCR assay was more sensitive than two OIE-validated PCR assays when testing 14 strains of ASFV representing four genotypes (I, V, VIII and IX) from diverse geographical areas. A total of 62 clinical swine blood samples collected from Uganda were examined by the novel PCR, giving a high agreement (59/62) with a superior sensitive universal probe library-based real-time PCR. Eight out of 62 samples tested positive, and three samples with higher Ct values (39.15, 38.39 and 37.41) in the real-time PCR were negative for ASFV in the novel PCR. In contrast, one (with a Ct value of 29.75 by the real-time PCR) and two (with Ct values of 29.75 and 33.12) ASFV-positive samples were not identified by the two OIE-validated PCR assays, respectively. Taken together, these data show that the novel PCR assay is specific, sensitive, and applicable for molecular diagnosis and surveillance of ASF.

This study examined the influence of informal milk delivery chains on the risk of human exposure to Brucella spp. through milk consumption in two regions of Uganda (Gulu and Soroti Districts). The work involved describing milk delivery chains, investigating brucellosis awareness amongst milk deliverers and determining the presence of Brucella spp. antibodies in cattle milk on delivery to primary collection points (boiling points and dairies). Milk samples (n = 331) were collected from deliverers at primary collection points and from street vendors at point of sale and analysed using indirect enzyme-linked immunosorbent assay (I-ELISA). A written questionnaire was used to collect data from deliverers (n = 279) on their milk delivery chains and their brucellosis awareness. The most common delivery points in Gulu District were small dairies and in Soroti District boiling points. The presence of Brucella spp. antibodies in milk samples was higher in Soroti (40 %) than in Gulu (11 %) (P

Equine Influenza Virus (EIV) is a major cause of respiratory disease in horses and the virus constantly undergoes antigenic drift. Here we characterize and describe the HA1 and the NA genes of H3N8 within samples obtained from outbreaks in Sweden during November-December 2011. Both clade 1 and clade 2 viruses of the Florida sublineage were identified. The index case of clade 2 was transported to Sweden from Spain through the Netherlands, whereas the clade 1 had its origin from a Swedish stud farm. The clade 1 virus was efficiently spread between training yards by unvaccinated young horses, but vaccinated horses were also presented with clinical signs of respiratory disease. No virus of the Eurasian lineage was isolated during this outbreak. Clade 1 has previously been described in outbreaks in numerous of other countries, but this is the first time it has been detected in Sweden. The results from this study shows the importance of including both clade 1 and clade 2 of the Florida sublineage in equine influenza vaccines, supporting the ESP and OIE recommendations.

Control of African swine fever (ASF) in countries in Eastern, Central and Southern Africa (ECSA) is particularly complex owing to the presence of all three known epidemiological cycles of maintenance of the virus, namely an ancient sylvatic cycle involving the natural hosts and vectors of the disease as well as domestic cycles with and without involvement of natural vectors. While the situation is well documented in some of the countries, for others very little information is available. In spite of the unfavourable ASF situation, the pig population in the sub-region has grown exponentially in recent decades and is likely to continue to grow in response to rapid urban growth resulting in increasing demand for animal protein by populations that are no longer engaged in livestock production. Better management of ASF will be essential to permit the pig sector to reach its full potential as a supplier of high quality protein and a source of income to improve livelihoods and create wealth. No vaccine is currently available and it is likely that, in the near future, the sub-region will continue to rely on the implementation of preventive measures, based on the epidemiology of the disease, to avoid both the devastating losses that outbreaks can cause and the risk the sub-region poses to other parts of Africa and the world. The current situation in the ECSA sub-region is reviewed and gaps in knowledge are identified in order to support ongoing strategy development for managing ASF in endemic areas.

Uganda is a low-income country with the largest pig population in East Africa. Pig keeping has a large potential, commercially and as a tool for poverty reduction, but African swine fever (ASF) is a major hurdle for development of the sector. The objective of this study was to evaluate knowledge, attitudes and practices related to ASF in the smallholder pig production value chain in northern Uganda. The study included three separate series of participatory rural appraisals (PRA), comprising purposively selected farmers and other actors in the pig production value chain. In the PRAs, various participatory epidemiology tools were used. A total of 49 PRAs and 574 participants, representing 64 different villages, were included. The results indicate that participants were well aware of the clinical signs of ASF, routes for disease spread and measures for disease control. However, awareness of the control measures did not guarantee their implementation. A majority of middlemen and butchers acknowledged having sold live pigs, carcasses or pork they believed infected with ASF. Outbreaks of ASF had a strong negative impact on participants' socio-economic status with loss of revenue and reversal into more severe poverty. In conclusion, lack of knowledge is not what is driving the continuous circulation of ASF virus in this setting. To control ASF and reduce its impact, initiatives that stimulate changes in management are needed. Because the behaviour of all actors in the value chain is largely influenced by the deep rural poverty in the region, this needs to be combined with efforts to reduce rural poverty.

Animal production is important for the agricultural economy in low-income countries, but is threatened by infectious diseases. Serosurveys are conducted for different reasons such as disease detection, risk factor studies, disease monitoring and establishing disease-free status. Most reports on such serosurveys include some discussion about methodological constraints but still, by necessity, rely on serological results for case definition. This study uses a cross-sectional serosurvey for foot-and-mouth disease (FMD), Rift Valley fever (RVF) and contagious bovine pleuropneumonia (CBPP) in cattle in three districts in Western Uganda to illustrate the limitations of this approach, addressing the questions of what flaws can be expected in sampling and diagnostics and how these influence the results. The target was to collect blood samples from 60 cattle herds per district. To reflect the recent infection history of the herd, young animals (two to five years) were prioritised. The farmers were interviewed about management, cattle trade, cattle health and vaccination. Commercial ELISAs were used for serological analyses: for CBPP the IDEXX CBPP Mycoplasma mycoides subspecies mycoides antibody test kit, for RVF the ID Screen Rift Valley Fever competitive ELISA, and for FMD the PrioCHECK FMDV NS. Apparent prevalence, true prevalence and associations with herd characteristics were assessed. The sampling plans could not be entirely fulfilled, nor the number of tests run in the laboratory. There were reactors to all three diseases with an apparent prevalence of approximately 30 per cent for CBPP, 6 per cent for RVF and 7 per cent for FMD. Calculation of true prevalence based on test sensitivity and specificity resulted in a slightly higher prevalence figure for CBPP and lower figures for RVF and FMD. The study illustrates the importance of considering diagnostic test performance when interpreting results from serosurveys, and the challenge of representative sampling and laboratory work in low-income countries.

African swine fever (ASF) is one of the most important pig diseases, causing high case fatality rate and trade restrictions upon reported outbreaks. In Uganda, a low-income country with the largest pig population in East Africa, ASF is endemic. Animal disease impact is multidimensional and include social and economic impact along the value chain. In low-income settings, this impact keep people poor and push those that have managed to escape poverty back again. If the diseases can be controlled, their negative consequences can be mitigated. However, to successfully argue for investment in disease control, its cost-benefits need to be demonstrated. One part in the cost-benefit equations is disease impact quantification. The objective of this study was therefore to investigate the socio-economic impact of ASF outbreaks at household level in northern Uganda. In a longitudinal study, structured interviews with two hundred, randomly selected, pig-keeping households were undertaken three times with a six month interval. Questions related to family and pig herd demographics, pig trade and pig business. Associations between ASF outbreaks and economic and social impact variables were evaluated using linear regression models. The study showed that pigs were kept in extreme low-input-low-output farming systems involving only small monetary investments. Yearly incidence of ASF on household level was 19%. Increasing herd size was positively associated with higher economic output. The interaction between ASF outbreaks and the herd size showed that ASF outbreaks were negatively associated with economic output at the second interview occasion and with one out of two economic impact variables at the third interview occasion. No significant associations between the social impact variables included in the study and ASF outbreaks could be established. Trade and consumption of sick and dead pigs were coping strategies used to minimize losses of capital and animal protein. The results indicate that causality of social and economic impact of ASF outbreaks in smallholder systems is complex. Pigs are mostly kept as passive investments rather than active working capital, complicating economic analyses and further disqualifying disease control arguments based only on standard economic models.

In Uganda, a low-income country in east Africa, African swine fever (ASF) is endemic with yearly outbreaks. In the prevailing smallholder subsistence farming systems, farm biosecurity is largely non-existent. Outbreaks of ASF, particularly in smallholder farms, often go unreported, creating significant epidemiological knowledge gaps. The continuous circulation of ASF in smallholder settings also creates biosecurity challenges for larger farms. In this study, an on-going outbreak of ASF in an endemic area was investigated on farm level, including analyses of on-farm environmental virus contamination. The study was carried out on a medium-sized pig farm with 35 adult pigs and 103 piglets or growers at the onset of the outbreak. Within 3 months, all pigs had died or were slaughtered. The study included interviews with farm representatives as well as biological and environmental sampling. ASF was confirmed by the presence of ASF virus (ASFV) genomic material in biological (blood, serum) and environmental (soil, water, feed, manure) samples by real-time PCR. The ASFV-positive biological samples confirmed the clinical assessment and were consistent with known virus characteristics. Most environmental samples were found to be positive. Assessment of farm biosecurity, interviews, and the results from the biological and environmental samples revealed that breaches and non-compliance with biosecurity protocols most likely led to the introduction and within-farm spread of the virus. The information derived from this study provides valuable insight regarding the implementation of biosecurity measures, particularly in endemic areas.

without antiviral) to 45% and 100% in presence of Acyclovir (10ml/ mL and 100 mg/mL, respectively). These results are correlated with the decreased of viral load measured in the presence of Acyclovir: 1010 genome copies/mLwith EHV-1 only,106 copies/mL with EHV-1 + Acyclovir 10 mg/mL and 105 copies/mLwith EHV-1 + Acyclovir 100 mg/ mL. In conclusion, the cellular impedance monitoring system is a new real-time method to monitor cell growth and viral proliferation in cell cultures. This technology is well adapted for high-throughput screening of antiviral molecules. In fact, the xCELLigence system enables continuous impedance measurement and quantification of cell adhesion, proliferation, cell death and detachment during viral infection.

African swine fever virus (ASFV) causes one of the most dreaded transboundary animal diseases (TADs) in Suidae. African swine fever (ASF) often causes high rates of morbidity and mortality, which can reach 100% in domestic swine. To date, serological diagnosis has the drawback of not being able to differentiate variants of this virus. Previous studies have identified the 22 genotypes based on sequence variation in the C-terminal region of the p72 gene, which has become the standard for categorizing ASFVs. This article describes a genotyping assay developed using a segment of PCR-amplified genomic DNA of approximately 450 bp, which encompasses the C-terminal end of the p72 gene. Complementary paired DNA probes of 15 or 17 bp in length, which are identical except for a single nucleotide polymorphism (SNP) in the central position, were designed to either individually or in combination differentiate between the 22 genotypes. The assay was developed using xMAP technology; probes were covalently linked to microspheres, hybridized to PCR product, labelled with a reporter and read in the Luminex 200 analyzer. Characterization of the sample was performed by comparing fluorescence of the paired SNP probes, that is, the probe with higher fluorescence in a complementary pair identified the SNP that a particular sample possessed. In the final assay, a total of 52 probes were employed, 24 SNP pairs and 4 for general detection. One or more samples from each of the 22 genotypes were tested. The assay was able to detect and distinguish all 22 genotypes. This novel assay provides a powerful novel tool for the simultaneous rapid diagnosis and genotypic differentiation of ASF.

Japanese Encephalitis Virus (JEV) is considered an important reproductive pathogen in pigs. Most studies of the reproductive impact of JEV have been conducted in areas where the disease occurs in seasonal epidemics. In this study, the associations between seropositivity for JEV, measured with an IgG ELISA, and the number of piglets born alive and stillborn were investigated in a tropical area endemic for JEV in Vietnam. Sixty percent of sows from four farms in the Mekong delta of Vietnam were seropositive to JEV and the Odds Ratio for a sow being infected was highest (6.4) in sows above 3.5 years (95% confidence interval 2.2–18.3). There was an association between increasing Optical Density (OD) values from the ELISA and the number of stillborn piglets in sows less than 1.5 years, but no effect of seropositivity could be shown when all sows were studied. OD values had an effect (p = 0.04) on the number of piglets born alive in the statistical analysis only when interacting with the effect of the breeds. An increase in mean OD value of the herd was correlated (p

To the Editor: The article by Decaro et al. (1) described an outbreak of severe respiratory disease in calves in Italy caused by an atypical bovine pestivirus. This report confirms our concern that this group of viruses is probably widespread (2) and present on >3 continents. Moreover, it demonstrates that atypical bovine pestiviruses are capable of causing disease in calves in the field and a clinical picture consistent with most bovine viral diarrhea virus (BVDV) infections that occur naturally or experimentally (3,4). Some key issues remain unknown. The origin of the bovine pestivirus and the route of introduction into the herd are unclear, although phylogenies demonstrated a close relationship between this virus strain from Italy and atypical bovine pestiviruses from Brazil. Batches of fetal bovine serum from Brazil have repeatedly been found to be contaminated with atypical bovine pestiviruses (2,3), and there is a risk for contamination of vaccines with these viruses. Animal trade and vaccines should be considered when conducting further investigations into this outbreak. The evolutionary relationship between the atypical and the recognized pestivirus species (1) needs to be clarified. Maximum-likelihood and Bayesian analyses of a concatenated dataset positioned atypical pestiviruses consistently in a clade sister to BVDV-1 and BVDV-2, and the larger clade was sister to pestivirus of giraffes (5). The same pattern was observed in complete genome phylogeny (1). The term atypical is not informative, and a new name has been proposed for these bovine pestiviruses (5). Outcomes of infections with these viruses are typical, but the viral antigens and phylogenies are unique.

In this study, the complete genome sequence of a Newcastle disease virus (NDV) isolate collected from an outbreak in 1995 in chickens was fully characterized and compared with other NDV sequences. The genome was found to be 15,192 nucleotides long and to consist of six genes in the order 3'-NP-P-M-F-HN-L-5', similar to other avian paramyxoviruses type-I. However, a six-nucleotide insertion was observed in the 5' non-coding regions of the nucleoprotein (NP) gene, a feature that is unique to some NDV isolates. The isolate shows the amino acid sequence (112)RRQKRF(117) at the cleavage site of the F protein, which is identical to a known motif for virulent pathotypes of NDV. The phylogenetic analysis of the coding region of the F gene indicated that this isolate belongs to genotype VI, more specifically to genotype VId, along with isolates from the other European countries (Denmark, Switzerland and Austria). The same genotype caused outbreaks in the Middle East and Greece in the late 1960s, and in Hungary, in the early 1980s, suggesting a common source for these outbreaks.

The complete genome sequence of a Newcastle disease virus (NDV) isolated from a chicken in Sweden was determined and compared with other NDV sequences. The isolate was shown to belong to genotype VIIb, which arose in the Far East and spread around the world during the 1990s. It had a length of 15,192 bases and consisted of six genes in the order 3'-NP-P-M-F-HN-L-5'. The F protein cleavage site was 112-RRQRRF-117, corresponding to that of a virulent pathotype.

This study reports a novel method for the rapid detection and identification of the four recognized species in the pestivirus genus of the Flaviviridae family, i.e. classical swine fever virus (CSFV), border disease virus (BDV), bovine viral diarrhoea virus type 1 (BVDV1) and type 2 (BVDV2). The analysis of pestivirus PCR products was performed on microarrays by means of magnetic bead detection. The process utilizes an oligonucleotide array, onto which 5' biotinylated PCR products were hybridized, followed by visualization with streptavidin-coated magnetic particles by the naked eye, microscope or biochip reader. The assay was tested on a collection of pestiviruses that included all four species and allowed a specific and sensitive detection. Sensitivity was compared with other post-PCR detection methods, namely gel electrophoresis and suspension microarray. The results indicate that due to its high sensitivity, specificity and simple detection procedure, the magnetic bead assay provides a powerful tool for detection and identification of viral pathogens. Considering the simplicity of the assay, the protocols for hybridization and magnetic bead detection offer an emerging application for molecular diagnoses in virology that is amenable for use in a modestly equipped laboratory.

In this cross-sectional study, a stratified two-stage random sampling procedure was employed to select 221 dairy herds for bulk tank milk (BTM) sampling, and a subset of 55 dairy herds for individual blood sampling of a number of young animals (spot test), to predict presence or absence of current BVDV infection, and for data collection. The prediction was based on the high probability of seropositivity in groups of animals where PI animals are present because of the efficient spread of virus from PI animals to the surrounding group. BTM samples were collected in August 2003 (n = 192) and February 2004 (n = 195), and the 55 herds selected for spot testing and data collection were visited in December 2003. All samples were tested for presence of BVDV specific antibodies using a commercial indirect ELISA (SVANOVA Biotech AB, Uppsala, Sweden). The results demonstrated a very high level of exposure to BVDV in the region, and the proportion of herds with high antibody levels in the BTM was above 95% on both occasions. Despite this, almost two thirds of the herds had spot test results indicating absence of current infection, suggesting a high probability of self-clearance. A logistic regression model with the results from the spot tests as dependent variable was used to investigate possible herd and management factors associated with self-clearance, and suggested that this may occur regardless of herd size. Even though it is well established that the process of identification and elimination of PI animals is required within a systematic BVDV eradication programme, the present study strongly suggests that many herds may be cleared without intervention even in regions with high cattle density and high BVDV prevalence. Consequently, in any BVDV infected population (regardless of the herd-level BVDV seroprevalence), and at any given point of time, a large proportion of the herds will be free from infection due to self-clearance. Self-clearance is therefore a process that works in favour of any effort to control BVDV, which should be taken into account when planning and assessing the cost-effectiveness of a systematic control programme.