Enhanced passive surveillance for early detection of African and classical swine fevers.

African swine fever (ASF) and classical swine fever (CSF) are transboundary animal diseases (TADs) of pigs. Much effort and resources are regularly put into preventing these diseases' introduction in free areas. Passive surveillance activities bring the highest chances for the early detection of TAD incursions because they are routinely and widely conducted at farms, and because these activities focus on the time between introduction and when the first sample is sent for diagnostic testing. The authors proposed the implementation of an enhanced passive surveillance (EPS) protocol based on collecting data through participatory surveillance actions using an objective and adaptable scoring system to aid the early detection of ASF or CSF at the farm level. The protocol was applied in two commercial pig farms for ten weeks in the Dominican Republic, which is a CSF- and ASF-infected country. This study was a proof of concept, based on the EPS protocol to aid detection of substantial variations in the risk score triggering testing. One of the followed farms had score variation, which triggered testing of the animals, although the test results were negative. The study enables assessment of some of the weaknesses associated with passive surveillance and provides lessons applicable to the problem. Results demonstrate the potential for overcoming some issues preventing the broad application of EPS protocols and suggest that standardised approaches may contribute to the early detection of CSF and ASF introductions.

La peste porcine africaine (PPA) et la peste porcine classique (PPC) sont des maladies animales transfrontalières touchant les porcs. De nombreux efforts et ressources sont régulièrement alloués pour prévenir l'introduction de ces maladies dans des zones indemnes. Les activités de surveillance passive offrent les meilleures perspectives de détection précoce des incursions de maladies animales transfrontalières parce qu'elles sont menées de manière systématique et exhaustive dans les élevages, et parce qu'elles se concentrent sur la période entre l'introduction de la maladie et le moment où le premier échantillon est envoyé au laboratoire pour analyse. Les auteurs proposent la mise en oeuvre d'un protocole de surveillance passive renforcée fondé sur la collecte de données via des actions de surveillance participative utilisant un système de notation objectif et adaptable, en vue d'une détection précoce de la PPA et de la PPC dans les élevages. Ce protocole a été appliqué en République dominicaine, pays infecté par la PPA et la PPC, dans deux élevages porcins commerciaux pendant dix semaines. Cette étude était destinée à valider le principe de la méthode et se fondait sur le protocole de surveillance passive renforcée pour mieux détecter les variations substantielles de la note de risque qui conduisent à tester les animaux. L'un des élevages suivis a présenté une variation de cette note, ce qui a conduit à tester les animaux mais les tests se sont révélés négatifs. L'étude permet d'évaluer certaines des faiblesses associées à la surveillance passive et apporte des enseignements applicables à ce problème. Les résultats illustrent le potentiel de l'approche à surmonter certaines des problématiques empêchant l'application extensive des protocoles de surveillance passive renforcée. Ils suggèrent également que des approches normalisées pourraient contribuer à la détection précoce des cas d'introduction de la PPC et de la PPA.

La peste porcina africana (PPA) y la peste porcina clásica (PPC) son enfermedades animales transfronterizas que afectan al cerdo. Periódicamente se dedican grandes esfuerzos y cuantiosos recursos a evitar que estas enfermedades penetren en zonas que están exentas de ellas. Las actividades de vigilancia pasiva son las más eficaces para detectar con prontitud toda incursión de enfermedades animales transfronterizas, no solo por la regularidad y amplitud con que se llevan a cabo en las explotaciones, sino también porque inciden específicamente en el intervalo entre la penetración de una enfermedad y el momento en que se envía la primera muestra para que sea sometida a pruebas de diagnóstico. Los autores propusieron que se aplicara un protocolo de vigilancia pasiva reforzada que reposaba en la obtención de datos mediante actividades de vigilancia participativa, empleando para ello un sistema objetivo y adaptable de puntuación que ayudaba a detectar con prontitud la presencia en las explotaciones de PPA o PPC. Dicho protocolo fue aplicado a lo largo de diez semanas en dos explotaciones porcinas industriales de la República Dominicana, país en el que ambas infecciones están presentes. El estudio, que sirvió para poner a prueba la idea, pasaba por la aplicación del protocolo de vigilancia pasiva reforzada para ayudar a detectar variaciones sustanciales de la puntuación del nivel de riesgo que activa la realización de pruebas. En una de las explotaciones estudiadas se produjo una variación de la puntuación, cosa que activó la realización de pruebas en los animales, aunque estas arrojaron resultado negativo. El estudio aquí descrito permite evaluar algunos de los puntos débiles de la vigilancia pasiva y extraer enseñanzas aplicables al problema. Los resultados demuestran que es posible salvar algunas de las dificultades que impiden la aplicación generalizada de protocolos de vigilancia pasiva reforzada y dejan pensar que quizá el uso de planteamientos normalizados pueda ayudar a detectar con prontitud los casos de penetración de PPC o PPA.

Estimation of foot-and-mouth disease windborne transmission risk from USA beef feedlots.

Windborne spread of foot-and-mouth disease (FMD) requires specific epidemiological and meteorological conditions, thus modeling the risk of windborne spread involves integrating epidemiological and meteorological models. The objective of this study was to investigate the potential risk of windborne spread of FMD from an infected US feedlot using an integrated modeling approach, and to identify factors that determine this risk. To address this objective, we integrated a within-herd epidemiological model and an advanced atmospheric dispersion model, and calculated infection risk dependent on exposed herd size. A previously developed epidemiological model was used to simulate the spread of FMD through a typical U.S. feedlot, while the National Oceanic and Atmospheric Administration's (NOAA) HYSPLIT atmospheric dispersion model, which has been validated for FMD modeling, was used to model virus dispersion. Infection risk for exposed herds was calculated as a binomial probability accounting for dose and exposed herd size. We modeled risk of windborne spread from a typical 4000 head feedlot in the U.S. state of Iowa (IA), and a typical 48,000 head feedlot in the U.S. state of Kansas (KS) during winter and summer seasons. The risk of windborne spread of FMD varied based on weather/season conditions, estimated average viral shedding rate per head, size of infected herd, and size of exposed herd. In the baseline Kansas scenario (KS/103/W), the median of the maximum daily risk of infecting a 1000-head exposed herd ranged from 58.16 % at 1 km to 0.78 % at 10 km (Table 4). In the baseline Iowa scenario (IA/103/W), the median of the maximum daily risk of infecting a 1000-head exposed herd ranged from 21.78 % at 1 km to 0.05 % at 10 km (Table 4). The minimum control area recommended by the United States Department of Agriculture (USDA) in an FMD outbreak is 10 km from the infected premise. Our results indicate that significant risk of windborne spread may extend beyond 10 km in certain situations. This is particularly a concern in areas where there are large feedlots in relatively close proximity, such as in southwestern Kansas. Our model may be useful as a research tool in the absence of an outbreak and may help direct surveillance and response efforts in the event of an outbreak.

Foot-and-mouth disease virus transmission dynamics and persistence in a herd of vaccinated dairy cattle in India.

Foot-and-mouth disease (FMD) is an important transboundary disease with substantial economic impacts. Although between-herd transmission of the disease has been well studied, studies focusing on within-herd transmission using farm-level outbreak data are rare. The aim of this study was to estimate parameters associated with within-herd transmission, host physiological factors and FMD virus (FMDV) persistence using data collected from an outbreak that occurred at a large, organized dairy farm in India. Of 1,836 regularly vaccinated, adult dairy cattle, 222 had clinical signs of FMD over a 39-day period. Assuming homogenous mixing, a frequency-dependent compartmental model of disease transmission was built. The transmission coefficient and basic reproductive number were estimated to be between 16.2-18.4 and 67-88, respectively. Non-pregnant animals were more likely to manifest clinical signs of FMD as compared to pregnant cattle. Based on oropharyngeal fluid (probang) sampling and FMDV-specific RT-PCR, four of 36 longitudinally sampled animals (14%) were persistently infected carriers 10.5 months post-outbreak. There was no statistical difference between subclinical and clinically infected animals in the duration of the carrier state. However, prevalence of NSP-ELISA antibodies differed significantly between subclinical and clinically infected animals 12 months after the outbreak with 83% seroprevalence amongst clinically infected cattle compared to 69% of subclinical animals. This study further elucidates within-herd FMD transmission dynamics during the acute-phase and characterizes duration of FMDV persistence and seroprevalence of FMD under natural conditions in an endemic setting.

Genetic diversity and comparison of diagnostic tests for characterization of foot-and-mouth disease virus strains from Pakistan 2008-2012.

We report the laboratory analysis of 125 clinical samples from suspected cases of foot-and-mouth disease (FMD) in cattle and Asian buffalo collected in Pakistan between 2008 and 2012. Of these samples, 89 were found to contain viral RNA by rRT-PCR, of which 88 were also found to contain infectious FMD virus (FMDV) by virus isolation (VI), with strong correlation between these tests (κ = 0.96). Samples that were VI-positive were serotyped by antigen detection ELISA (Ag-ELISA) and VP1 sequence acquisition and analysis. Sequence data identified FMDV serotypes A (n = 13), O (n = 36) and Asia-1 (n = 41), including three samples from which both serotypes Asia-1 and O were detected. Serotype A viruses were classified within three different Iran-05 sublineages: HER-10, FAR-11 and ESF-10. All serotype Asia-1 were within Group VII (Sindh-08 lineage), in a genetic clade that differs from viruses isolated prior to 2010. All serotypes O were classified as PanAsia-2 within two different sublineages: ANT-10 and BAL-09. Using VP1 sequencing as the gold standard for serotype determination, the overall sensitivity of Ag-ELISA to correctly determine serotype was 74%, and serotype-specific sensitivity was 8% for serotype A, 88% for Asia-1 and 89% for O. Serotype-specific specificity was 100% for serotype A, 93% for Asia-1 and 94% for O. Interestingly, 12 of 13 serotype A viruses were not detected by Ag-ELISA. This study confirms earlier accounts of regional genetic diversity of FMDV in Pakistan and highlights the importance of continued validation of diagnostic tests for rapidly evolving pathogens such as FMDV.

Quantitative characteristics of the foot-and-mouth disease carrier state under natural conditions in India.

The goal of this study was to characterize the properties and duration of the foot-and-mouth disease (FMD) carrier state and associated serological responses subsequent to vaccination and naturally occurring infection at two farms in northern India. Despite previous vaccination of cattle in these herds, clinical signs of FMD occurred in October 2013 within a subset of animals at the farms containing juvenile-yearling heifers and steers (Farm A) and adult dairy cattle (Farm B). Subsequent to the outbreak, FMD virus (FMDV) asymptomatic carriers were identified in both herds by seroreactivity to FMDV non-structural proteins and detection of FMDV genomic RNA in oropharyngeal fluid. Carriers' seroreactivity and FMDV genome detection status were subsequently monitored monthly for 23 months. The mean extinction time of the carrier state was 13.1 ± 0.2 months, with extinction having occurred significantly faster amongst adult dairy cattle at Farm B compared to younger animals at Farm A. The rate of decrease in the proportion of carrier animals was calculated to be 0.07 per month. Seroprevalence against FMDV non-structural proteins decreased over the course of the study period, but was found to increase transiently following repeated vaccinations. These data provide novel insights into viral and host factors associated with the FMDV carrier state under natural conditions. The findings reported herein may be relevant to field veterinarians and governmental regulatory entities engaged in FMD response and control measures.

Efficacy of a high potency O1 Manisa monovalent vaccine against heterologous challenge with foot-and-mouth disease virus of O/SEA/Mya-98 lineage in sheep.

Potency tests for commercial oil-adjuvanted foot-and-mouth disease (FMD) vaccines are usually carried out in cattle, using a full dose (2 ml) of vaccine and homologous virus challenge. However, in sheep the recommended vaccine dose is half of the cattle dose (1 ml) and most vaccines have not been potency tested for this species, especially with heterologous viruses. To determine the efficacy of a high potency (>6PD50) FMD virus (FMDV) O1Manisa vaccine in sheep, we carried out a study using a heterologous FMDV (FMDV O/SKR/2010 - Mya-98 strain) challenge. Groups of seven animals each were vaccinated with 2×, 1×, 1/2× or 1/4× dose (2 ml, 1 ml, 0.5 ml or 0.25 ml respectively) and challenged at 7 days post vaccination (dpv). Only 3 of the 7 sheep in the group vaccinated with 2 ml were protected. With 2 additional groups, receiving double or single doses and challenged at 14 dpv, 4 of 7 sheep were protected in each group. None of the sheep had measurable neutralising antibodies against the vaccine or challenge virus at 7 dpv. However, all vaccinated animals challenged at 14 dpv had a homologous neutralising response against FMDV O1 Manisa on the day of challenge and all but one animal also had a heterologous response to FMDV O/SKR/2010. Infectious FMDV and viral RNA could be found in nasal swabs between 1 and 6 days post challenge (dpc) in most vaccinated sheep, but those vaccinated with higher doses or challenged at 14 dpv showed significant decreases in the level of FMDV detection. Intermittent virus shedding was noticed between 1 and 35 dpc in all vaccinated groups, but persistent infection could be demonstrated only in 4 sheep (20%). This study showed that at the recommended dose, a high potency (>6 PD50) FMDV O1Manisa vaccine does not protect sheep against a heterologous challenge at 7 dpv. However, partial protection was observed when a double dose was used at 7 dpv or when double or single dose vaccinated sheep were challenged at 14 dpv.

An Integrative Analysis of Foot-and-Mouth Disease Virus Carriers in Vietnam Achieved Through Targeted Surveillance and Molecular Epidemiology.

Foot-and-mouth disease (FMD) is a major constraint to transboundary trade in animal products, yet much of its natural ecology and epidemiology in endemic regions is still poorly understood. To address this gap, a multidisciplinary, molecular and conventional epidemiological approach was applied to an investigation of endemic FMD in Vietnam. Within the study space, it was found that 22.3% of sampled ruminants had previously been infected with FMD virus (FMDV), of which 10.8% were persistent, asymptomatic carriers (2.4% of the total population). Descriptive data collected from targeted surveillance and a farm questionnaire showed a significantly lower prevalence of FMDV infection for dairy farms. In contrast, farms of intermediate size and/or history of infection in 2010 were at increased risk of FMD exposure. At the individual animal level, buffalo had the highest exposure risk (over cattle), and there was spatial heterogeneity in exposure risk at the commune level. Conversely, carrier prevalence was higher for beef cattle, suggesting lower susceptibility of buffalo to persistent FMDV infection. To characterize virus strains currently circulating in Vietnam, partial FMDV genomic (VP1) sequences from carrier animals collected between 2012 and 2013 (N = 27) and from FMDV outbreaks between 2009 and 2013 (N = 79) were compared by phylogenetic analysis. Sequence analysis suggested that within the study period, there were two apparent novel introductions of serotype A viruses and that the dominant lineage of serotype O in Vietnam shifted from SEA/Mya-98 to ME-SA/PanAsia. FMDV strains shared close ancestors with FMDV from other South-East Asian countries indicating substantial transboundary movement of the predominant circulating strains. Close genetic relationships were observed between carrier and outbreak viruses, which may suggest that asymptomatic carriers of FMDV contribute to regional disease persistence. Multiple viral sequences obtained from carrier cattle over a 1-year period had considerable within-animal genetic variation, indicating within-host virus evolution.

Early Detection of Foot-And-Mouth Disease Virus from Infected Cattle Using A Dry Filter Air Sampling System.

Foot-and-mouth disease (FMD) is a highly contagious livestock disease of high economic impact. Early detection of FMD virus (FMDV) is fundamental for rapid outbreak control. Air sampling collection has been demonstrated as a useful technique for detection of FMDV RNA in infected animals, related to the aerogenous nature of the virus. In the current study, air from rooms housing individual (n = 17) or two groups (n = 4) of cattle experimentally infected with FDMV A24 Cruzeiro of different virulence levels was sampled to assess the feasibility of applying air sampling as a non-invasive, screening tool to identify sources of FMDV infection. Detection of FMDV RNA in air was compared with first detection of clinical signs and FMDV RNA levels in serum and oral fluid. FMDV RNA was detected in room air samples 1-3 days prior (seven animals) or on the same day (four animals) as the appearance of clinical signs in 11 of 12 individually housed cattle. Only in one case clinical signs preceded detection in air samples by one day. Overall, viral RNA in oral fluid or serum preceded detection in air samples by 1-2 days. Six individually housed animals inoculated with attenuated strains did not show clinical signs, but virus was detected in air in one of these cases 3 days prior to first detection in oral fluid. In groups of four cattle housed together, air detection always preceded appearance of clinical signs by 1-2 days and coincided more often with viral shedding in oral fluid than virus in blood. These data confirm that air sampling is an effective non-invasive screening method for detecting FMDV infection in confined to enclosed spaces (e.g. auction barns, milking parlours). This technology could be a useful tool as part of a surveillance strategy during FMD prevention, control or eradication efforts.

Infection Dynamics of Foot-and-Mouth Disease Virus in Cattle Following Intranasopharyngeal Inoculation or Contact Exposure.

For the purpose of developing an improved experimental model for studies of foot-and-mouth disease virus (FMDV) infection in cattle, three different experimental systems based on natural or simulated natural virus exposure were compared under standardized experimental conditions. Ante-mortem infection dynamics were characterized in cattle exposed to FMDV through a novel, simulated natural intranasopharyngeal (INP) inoculation system or through standardized and controlled systems of within- or between-species direct contact exposure (cattle-to-cattle or pig-to-cattle). All three systems were efficient in causing synchronous, generalized foot-and-mouth disease in cattle exposed to one of three different strains of FMDV representing serotypes O, A and Asia1. There was more within-group variation in the timing of clinical infection following natural and simulated natural virus exposure systems when compared with the conventionally used system of needle inoculation (intraepithelial lingual inoculation). However, the three optimized exposure systems described herein have the advantage of closely simulating field conditions by utilizing natural routes of primary infection, thereby facilitating engagement of mucosal host defence mechanisms. Overall, it is concluded that INP inoculation and standardized systems of direct contact exposure provide effective alternatives to conventional (needle) inoculation systems for studies in which it is desirable to simulate the natural biology of FMDV infection.

Serotype Diversity of Foot-and-Mouth-Disease Virus in Livestock without History of Vaccination in the Far North Region of Cameroon.

Little information is available about the natural cycle of foot-and-mouth disease (FMD) in the absence of control measures such as vaccination. Cameroon presents a unique opportunity for epidemiological studies because FMD vaccination is not practiced. We carried out a prospective study including serological, antigenic and genetic aspects of FMD virus (FMDV) infections among different livestock production systems in the Far North of Cameroon to gain insight into the natural ecology of the virus. We found serological evidence of FMDV infection in over 75% of the animals sampled with no significant differences of prevalence observed among the sampled groups (i.e. market, sedentary, transboundary trade and mobile). We also found antibodies reactive to five of the seven FMDV serotypes (A, O, SAT1, SAT2 and SAT3) among the animals sampled. Finally, we were able to genetically characterize viruses obtained from clinical and subclinical FMD infections in Cameroon. Serotype O viruses grouped into two topotypes (West and East Africa). SAT2 viruses grouped with viruses from Central and Northern Africa, notably within the sublineage causing the large epidemic in Northern Africa in 2012, suggesting a common origin for these viruses. This research will guide future interventions for the control of FMD such as improved diagnostics, guidance for vaccine formulation and epidemiological understanding in support of the progressive control of FMD in Cameroon.

Detection of Foot-and-mouth Disease Virus RNA and Capsid Protein in Lymphoid Tissues of Convalescent Pigs Does Not Indicate Existence of a Carrier State.

A systematic study was performed to investigate the potential of pigs to establish and maintain persistent foot-and-mouth disease virus (FMDV) infection. Infectious virus could not be recovered from sera, oral, nasal or oropharyngeal fluids obtained after resolution of clinical infection with any of five FMDV strains within serotypes A, O and Asia-1. Furthermore, there was no isolation of live virus from tissue samples harvested at 28-100 days post-infection from convalescent pigs recovered from clinical or subclinical FMD. Despite lack of detection of infectious FMDV, there was a high prevalence of FMDV RNA detection in lymph nodes draining lesion sites harvested at 35 days post-infection, with the most frequent detection recorded in popliteal lymph nodes (positive detection in 88% of samples obtained from non-vaccinated pigs). Likewise, at 35 dpi, FMDV capsid antigen was localized within follicles of draining lymph nodes, but without concurrent detection of FMDV non-structural protein. There was a marked decline in the detection of FMDV RNA and antigen in tissue samples by 60 dpi, and no antigen or viral RNA could be detected in samples obtained at 100 dpi. The data presented herein provide the most extensive investigation of FMDV persistence in pigs. The overall conclusion is that domestic pigs are unlikely to be competent long-term carriers of infectious FMDV; however, transient persistence of FMDV protein and RNA in lymphoid tissues is common following clinical or subclinical infection.

Infection dynamics of foot-and-mouth disease virus in pigs using two novel simulated-natural inoculation methods.

In order to characterize foot-and-mouth disease virus (FMDV) infection dynamics in pigs, two simulated-natural inoculation systems were developed and evaluated. Intra-oropharyngeal (IOP) and intra-nasopharyngeal (INP) inoculation both enabled precise control of dose and timing of inoculation while simulating field exposure conditions. There were substantial differences between outcomes of infections by the two routes. IOP inoculation resulted in consistent and synchronous infection, whereas INP inoculation at similar doses resulted in delayed, or completely absent infection. All pigs that developed clinical infection had detectable levels of FMDV RNA in their oropharynx directly following inoculation. Furthermore, FMDV antigens were localized to the oropharyngeal tonsils suggesting a role in early infection. The utility of IOP inoculation was further demonstrated in a vaccine-challenge experiment. Thus, the novel system of IOP inoculation described herein, offers a valid alternative to traditionally used systems for FMDV inoculation of pigs, applicable for experimental studies of FMDV pathogenesis and vaccinology.

Interaction of foot-and-mouth disease virus nonstructural protein 3A with host protein DCTN3 is important for viral virulence in cattle.

UNLABELLED: Nonstructural protein 3A of foot-and-mouth disease virus (FMDV) is a partially conserved protein of 153 amino acids in most FMDVs examined to date. The role of 3A in virus growth and virulence within the natural host is not well understood. Using a yeast two-hybrid approach, we identified cellular protein DCTN3 as a specific host binding partner for 3A. DCTN3 is a subunit of the dynactin complex, a cofactor for dynein, a motor protein. The dynactin-dynein duplex has been implicated in several subcellular functions involving intracellular organelle transport. The 3A-DCTN3 interaction identified by the yeast two-hybrid approach was further confirmed in mammalian cells. Overexpression of DCTN3 or proteins known to disrupt dynein, p150/Glued and 50/dynamitin, resulted in decreased FMDV replication in infected cells. We mapped the critical amino acid residues in the 3A protein that mediate the protein interaction with DCTN3 by mutational analysis and, based on that information, we developed a mutant harboring the same mutations in O1 Campos FMDV (O1C3A-PLDGv). Although O1C3A-PLDGv FMDV and its parental virus (O1Cv) grew equally well in LFBK-αvß6, O1C3A-PLDGv virus exhibited a decreased ability to replicate in primary bovine cell cultures. Importantly, O1C3A-PLDGv virus exhibited a delayed disease in cattle compared to the virulent parental O1Campus (O1Cv). Virus isolated from lesions of animals inoculated with O1C3A-PLDGv virus contained amino acid substitutions in the area of 3A mediating binding to DCTN3. Importantly, 3A protein harboring similar amino acid substitutions regained interaction with DCTN3, supporting the hypothesis that DCTN3 interaction likely contributes to virulence in cattle. IMPORTANCE: The objective of this study was to understand the possible role of a FMD virus protein 3A, in causing disease in cattle. We have found that the cellular protein, DCTN3, is a specific binding partner for 3A. It was shown that manipulation of DCTN3 has a profound effect in virus replication. We developed a FMDV mutant virus that could not bind DCTN3. This mutant virus exhibited a delayed disease in cattle compared to the parental strain highlighting the role of the 3A-DCTN3 interaction in virulence in cattle. Interestingly, virus isolated from lesions of animals inoculated with mutant virus contained mutations in the area of 3A that allowed binding to DCTN3. This highlights the importance of the 3A-DCTN3 interaction in FMD virus virulence and provides possible mechanisms of virus attenuation for the development of improved FMD vaccines.

Early adaptive immune responses in the respiratory tract of foot-and-mouth disease virus-infected cattle.

Foot-and-mouth disease (FMD) is a highly contagious viral disease which affects both domestic and wild biungulate species. This acute disease, caused by the FMD virus (FMDV), usually includes an active replication phase in the respiratory tract for up to 72 h postinfection, followed by hematogenous dissemination and vesicular lesions at oral and foot epithelia. The role of the early local adaptive immunity of the host in the outcome of the infection is not well understood. Here we report the kinetics of appearance of FMDV-specific antibody-secreting cells (ASC) in lymphoid organs along the respiratory tract and the spleen in cattle infected by aerosol exposure. While no responses were observed for up to 3 days postinfection (dpi), all animals developed FMDV-ASC in all the lymphoid organs studied at 4 dpi. Tracheobronchial lymph nodes were the most reactive organs at this time, and IgM was the predominant isotype, followed by IgG1. Numbers of FMDV-ASC were further augmented at 5 and 6 dpi, with an increasing prevalence in upper respiratory organs. Systemic antibody responses were slightly delayed compared with the local reaction. Also, IgM was the dominant isotype in serum at 5 dpi, coinciding with a sharp decrease of viral RNA detection in peripheral blood. These results indicate that following aerogenous administration, cattle develop a rapid and vigorous genuine local antibody response throughout the respiratory tract. Time course and isotype profiles indicate the presence of an efficient T cell-independent antibody response which drives the IgM-mediated virus clearance in cattle infected by FMDV aerosol exposure.

Time-dependent biodistribution and transgene expression of a recombinant human adenovirus serotype 5-luciferase vector as a surrogate for rAd5-FMDV vaccines in cattle.

Replication-defective recombinant adenovirus 5 (rAd5) vectors carrying foot-and-mouth disease virus (FMDV) transgenes elicit a robust immune response to FMDV challenge in cattle; however mechanistic functions of vaccine function are incompletely understood. Recent efforts addressing critical interactions of rAd5 vectors with components of the bovine immune system have elucidated important aspects of induction of protective immunity against FMDV. In the current study, a rAd5-Luciferase (rAd5-Luc) surrogate vector was utilized for indirect assessment of rAd5-FMDV distribution during the first 48 hours post inoculation (hpi). To compare vector distribution dynamics and time-dependent transgene expression, bovine cells were inoculated in vitro with rAd5-FMDV and rAd5-Luc vectors. Superior transgene expression was detected in cells infected with rAd5-Luc compared to rAd5-FMDV. However, both vectors behaved remarkably similar in demonstrating elevated mRNA transcription at 24 and 48 hpi with peak occurrence of transgene expression at 48 hpi. Injection sites of cattle inoculated with rAd5-Luc contained mononuclear inflammatory infiltrates with hexon and transgene proteins associated with antigen-presenting cells. Luciferase activity, as well as microscopic detection of luciferase antigens, peaked at 24 hpi. Presence of viral mRNA also peaked at 24 hpi but unlike luciferase, remained strongly detected at 48 hpi. Cell-associated luciferase antigens were detected as early as 6 hpi at the cortical interfolicullar areas of local LN, indicating rapid trafficking of antigen-presenting cells to lymphoid tissues. This work provides mechanistic insights on rAd5-mediated immunity in cattle and will contribute to ongoing efforts to enhance rAd5-FMDV vaccine efficacy.

The pathogenesis of foot-and-mouth disease II: viral pathways in swine, small ruminants, and wildlife; myotropism, chronic syndromes, and molecular virus-host interactions.

Investigation into the pathogenesis of foot-and-mouth disease (FMD) has focused on the study of the disease in cattle with less emphasis on pigs, small ruminants and wildlife. 'Atypical' FMD-associated syndromes such as myocarditis, reproductive losses and chronic heat intolerance have also received little attention. Yet, all of these manifestations of FMD are reflections of distinct pathogenesis events. For example, naturally occurring porcinophilic strains and unique virus-host combinations that result in high-mortality outbreaks surely have their basis in molecular-, cellular- and tissue-level interactions between host and virus (i.e. pathogenesis). The goal of this review is to emphasize how the less commonly studied FMD syndromes and host species contribute to the overall understanding of pathogenesis and how extensive in vitro studies have contributed to our understanding of disease processes in live animals.

The pathogenesis of foot-and-mouth disease I: viral pathways in cattle.

In 1898, foot-and-mouth disease (FMD) earned a place in history as the first disease of animals shown to be caused by a virus. Yet, despite over a century of active investigation and elucidation of many aspects of FMD pathogenesis, critical knowledge about the virus-host interactions is still lacking. The aim of this review is to provide a comprehensive overview of FMD pathogenesis in cattle spanning from the earliest studies to recently acquired insights emphasizing works which describe animals infected by methodologies most closely resembling natural infection (predominantly aerosol or direct/indirect contact). The three basic phases of FMD pathogenesis in vivo will be dissected and characterized as: (i) pre-viraemia characterized by infection and replication at the primary replication site(s), (ii) sustained viraemia with generalization and vesiculation at secondary infection sites and (iii) post-viraemia/convalescence including resolution of clinical disease that may result in long-term persistent infection. Critical evaluation of the current status of understanding will be used to identify knowledge gaps to guide future research efforts.

The early pathogenesis of foot-and-mouth disease in cattle after aerosol inoculation. Identification of the nasopharynx as the primary site of infection.

To characterize the early events of foot-and-mouth disease virus (FMDV) infection in cattle subsequent to simulated natural exposure, 16 steers were aerosol inoculated with FMDV and euthanized at various times. Samples were collected from each steer antemortem (serum, nasal swabs, and oral swabs) and postmortem (up to 40 tissues per animal) and screened for FMDV by virus isolation and for FMDV RNA by real-time reverse transcription polymerase chain reaction. Tissues that tested positive for FMDV or viral RNA were examined by immunohistochemistry and multichannel immunofluorescence microscopy. In previremic steers, FMDV was most consistently localized to nasopharyngeal tissues, thereby indicating this region as the most important site of primary viral replication. The earliest site of microscopic localization of FMDV antigens was the lymphoid follicle-associated epithelium of the pharyngeal mucosa-associated lymphoid tissue of the nasopharynx at 6 hours postaerosolization. At early time points after aerosol inoculation, viral antigens colocalized with cytokeratin-positive pharyngeal epithelial cells; intraepithelial FMDV-negative, MHCII/CD11c-double-positive dendritic cells were present in close proximity to FMDV-positive cells. Onset of viremia coincided with marked increase of viral loads in pulmonary tissues and with substantial decrease of viral detection in nasopharyngeal tissues. These data indicate that subsequent to aerogenous exposure to FMDV, the temporally defined critical pathogenesis events involve (1) primary replication in epithelial cells of the pharyngeal mucosa-associated lymphoid tissue crypts and (2) subsequent widespread replication in pneumocytes in the lungs, which coincides with (3) the establishment of sustained viremia.

Agricultural diseases on the move early in the third millennium.

With few exceptions, the diseases that present the greatest risk to food animal production have been largely similar throughout the modern era of veterinary medicine. The current trend regarding the ever-increasing globalization of the trade of animals and animal products ensures that agricultural diseases will continue to follow legal and illegal trade patterns with increasing rapidity. Global climate changes have already had profound effects on the distribution of animal diseases, and it is an inevitable reality that continually evolving climatic parameters will further transform the ecology of numerous pathogens. In recent years, many agricultural diseases have given cause for concern regarding changes in distribution or severity. Foot-and-mouth disease, avian influenza, and African swine fever continue to cause serious problems. The expected announcement of the global eradication of rinderpest is one of the greatest successes of veterinary preventative medicine, yet the closely related disease peste des petits ruminants still spreads throughout the Middle East and Asia. The spread of novel strains of bluetongue virus across Europe is an ominous indicator that climate change is sure to influence trends in movement of agricultural diseases. Overall, veterinary practitioners and investigators are advised to not only maintain vigilance against the staple disease threats but to always be sufficiently broad-minded to expect the unexpected.

Hepatotoxicity associated with pyrrolizidine alkaloid (Crotalaria spp) ingestion in a horse on Easter Island.

Since 1984, a significant number of privately owned and feral horses on Easter Island have died of a syndrome consisting of progressive anorexia, weight loss, obtundation, and other central nervous system abnormalities. A single horse experiencing clinical signs of the reported syndrome was identified, examined and necropsied. Clinical signs included inappetence, emaciation, ataxia and icterus. Gross necropsy findings included hepatic enlargement and mottling, ascites and gastric impaction. Histopathological lesions included hepatic hemorrhage and necrosis, periportal megalocytosis, portal fibrosis, bile duct hyperplasia and multinucleate hepatocytes. Crotalaria grahamiana and C pallida, were identified in the pasture of the presenting horse, and found to be widespread on the island. Alkaloid fingerprinting identified grahamine, monocrotalin, and a grahamine analog in C grahamiana. A retrorsine analog and a senecionine analog were identified in C pallida. The highly characteristic lesions and the identification of pyrrolizidine alkaloids in the 2 plants strongly suggest that ingestion of 1 or both of the Crotalaria species led to chronic liver damage and hepatic encephalopathy in the presenting horse. Widespread distribution of C grahamiana on the island and reported temporal and seasonal trends in incidence among horses and cattle suggest that C grahamiana may be responsible for extensive morbidity and mortality among horses and cattle on Easter Island.