Phylogeographic analysis of foot-and-mouth disease virus serotype O dispersal and associated drivers in East Africa.

The continued endemicity of foot and mouth disease virus (FMDV) in East Africa has significant implications for livestock production and poverty reduction, yet its complex epidemiology in endemic settings remains poorly understood. Identifying FMDV dispersal routes and drivers of transmission is key to improved control strategies. Environmental heterogeneity and anthropogenic drivers (e.g., demand for animal products) can impact viral spread by influencing host movements. Here, we utilized FMDV serotype O VP1 genetic sequences and corresponding spatiotemporal data in order to (i) infer the recent dispersal history, and (II) investigate the impact of external factors (cattle density, human population density, proximity to livestock markets, and drought) on dispersal velocity, location, and direction of FMDV serotype O in East Africa. We identified statistical evidence of long-distance transmission events, and we found that FMDV serotype O tends to remain circulating in areas of high cattle density, high human population density, and in close proximity to livestock markets. The latter two findings highlight the influence of anthropogenic factors on FMDV serotype O spread in this region. These findings contribute to the understanding of FMDV epidemiology in East Africa and can help guide improved control measures.

Evidence of subclinical foot-and-mouth disease virus infection in young calves born from clinically recovered cow under natural condition.

Foot-and-mouth disease (FMD) is a highly contagious and economically important, transboundary viral disease of cloven-hoofed animals. It is known that an asymptomatic, persistent FMD virus (FMDV) infection may occur subsequent to acute or subclinical FMDV infection in adult ruminants. However, virus persistence in young calves has not been studied. In the current investigation, FMDV infection parameters were examined for calves born to FMD-clinically recovered cows (CRC), asymptomatic cows from infected herds (ASC) and cows from with no history of FMD (NHF). The study was conducted in natural condition after FMD outbreaks in two dairy herds in India. No calves described herein had any clinical signs of FMD. Six out of 12 calves born to CRC had detectable FMDV RNA in oesophageal-pharyngeal fluid consistent with asymptomatic FMDV infection. Three of the 12 calves of CRC group had seroreactivity against FMDV non-structural proteins. One calf had detectable FMDV RNA at two consecutive samplings at 2 months apart. However, infectious FMDV was not isolated from any calf in the study. None of the calves in the ASC or NHF groups had any evidence of FMDV infection. Overall, these data are consistent with earlier report on calves having been infected in utero. Further investigation of FMDV persistence in calves under controlled conditions may lead to greater understanding of the viral pathogenesis.

Constitutively Active IRF7/IRF3 Fusion Protein Completely Protects Swine against Foot-and-Mouth Disease.

UNLABELLED: Foot-and-mouth disease (FMD) remains one of the most devastating livestock diseases around the world. Several serotype-specific vaccine formulations exist, but they require about 5 to 7 days to induce protective immunity. Our previous studies have shown that a constitutively active fusion protein of porcine interferon (IFN) regulatory factors (IRF) 7 and 3 [IRF7/3(5D)] strongly induced type I IFN and antiviral genes in vitro and prevented mortality in an FMD mouse model when delivered with a replication-defective adenoviral vector [Ad5-poIRF7/3(5D)]. Here, we demonstrate that pigs treated with 10(8), 10(9), or 10(10) PFU of Ad5-poIRF7/3(5D) 24 h before FMDV challenge were fully protected from FMD clinical signs and did not develop viremia, virus shedding or antibodies against FMDV nonstructural proteins. Pigs treated with Ad5-poIRF7/3(5D) had higher levels of IFN and antiviral activity in serum, and upregulated expression of several IFN-stimulated genes in peripheral blood mononuclear cells, compared to pigs treated with Ad5-Blue vector control. Importantly, treatment of porcine cultured cells with Ad5-poIRF7/3(5D) inhibited the replication of all 7 FMDV serotypes. In vitro experiments using cultured embryonic fibroblasts derived from IFN receptor knockout mice suggested that the antiviral response induced by Ad5-poIRF7/3(5D) was dependent on type I and III IFN pathways; however, experiments with mice demonstrated that a functional type I IFN pathway mediates Ad5-poIRF7/3(5D) protection conferred in vivo Our studies demonstrate that inoculation with Ad5-poIRF7/3(5D) completely protects swine against FMD by inducing a strong type I IFN response and highlights its potential application to rapidly and effectively prevent FMDV replication and dissemination. IMPORTANCE: Foot-and-mouth disease virus (FMDV) causes a fast-spreading disease that affects farm animals, with economically and socially devastating consequences. Our study shows that inoculation with a constitutively active transcription factor, namely, a fusion protein of porcine interferon (IFN) regulatory factors (IRF) 7 and 3 delivered by an adenovirus vector [Ad5-poIRF7/3(5D)], is a new effective treatment to prevent FMD in swine. Animals pretreated with Ad5-poIRF7/3(5D) 1 day before being exposed to FMDV were completely protected from viral replication and clinical disease. It is noteworthy that the doses of Ad5-poIRF7/3(5D) required for protection are lower than those previously reported for similar approaches using Ad5 vectors delivering type I, II, or III IFN, suggesting that this novel strategy would be economically appealing to counteract FMD. Our results also indicate that a dynamic interplay among different components of pigs' innate immune defenses allows potent antiviral effects after Ad5-poIF7/3(5D) administration.

The Foot-and-Mouth Disease Carrier State Divergence in Cattle.

UNLABELLED: The pathogenesis of persistent foot-and-mouth disease virus (FMDV) infection was investigated in 46 cattle that were either naive or had been vaccinated using a recombinant, adenovirus-vectored vaccine 2 weeks before challenge. The prevalence of FMDV persistence was similar in both groups (62% in vaccinated cattle, 67% in nonvaccinated cattle), despite vaccinated cattle having been protected from clinical disease. Analysis of antemortem infection dynamics demonstrated that the subclinical divergence between FMDV carriers and animals that cleared the infection had occurred by 10 days postinfection (dpi) in vaccinated cattle and by 21 dpi in nonvaccinated animals. The anatomic distribution of virus in subclinically infected, vaccinated cattle was restricted to the pharynx throughout both the early and the persistent phases of infection. In nonvaccinated cattle, systemically disseminated virus was cleared from peripheral sites by 10 dpi, while virus selectively persisted within the nasopharynx of a subset of animals. The quantities of viral RNA shed in oropharyngeal fluid during FMDV persistence were similar in vaccinated and nonvaccinated cattle. FMDV structural and nonstructural proteins were localized to follicle-associated epithelium of the dorsal soft palate and dorsal nasopharynx in persistently infected cattle. Host transcriptome analysis of tissue samples processed by laser capture microdissection indicated suppression of antiviral host factors (interferon regulatory factor 7, CXCL10 [gamma interferon-inducible protein 10], gamma interferon, and lambda interferon) in association with persistent FMDV. In contrast, during the transitional phase of infection, the level of expression of IFN-λ mRNA was higher in follicle-associated epithelium of animals that had cleared the infection. This work provides novel insights into the intricate mechanisms of FMDV persistence and contributes to further understanding of this critical aspect of FMDV pathogenesis. IMPORTANCE: The existence of a prolonged, asymptomatic carrier state is a political impediment for control and potential eradication of foot-and-mouth disease (FMD). When FMD outbreaks occur, they are often extinguished by massive depopulation of livestock due to the fear that some animals may have undiagnosed subclinical infection, despite uncertainty over the biological relevance of FMD virus (FMDV) persistence. The work described here elucidates aspects of the FMDV carrier state in cattle which may facilitate identification and/or abrogation of asymptomatic FMDV infection. The divergence between animals that clear infection and those that develop persistent infection was demonstrated to occur earlier than previously established. The host antiviral response in tissues maintaining persistent FMDV was downregulated, whereas upregulation of IFN-λ mRNA was found in the epithelium of cattle that had recently cleared the infection. This suggests that the clearing of FMDV infection is associated with an enhanced mucosal antiviral response, whereas FMDV persistence is associated with suppression of the host antiviral response.

Pathogenesis of virulent and attenuated foot-and-mouth disease virus in cattle.

BACKGROUND: Understanding the mechanisms of attenuation and virulence of foot-and-mouth disease virus (FMDV) in the natural host species is critical for development of next-generation countermeasures such as live-attenuated vaccines. Functional genomics analyses of FMDV have identified few virulence factors of which the leader proteinase (Lpro) is the most thoroughly investigated. Previous work from our laboratory has characterized host factors in cattle inoculated with virulent FMDV and attenuated mutant strains with transposon insertions within Lpro. METHODS: In the current study, the characteristics defining virulence of FMDV in cattle were further investigated by comparing the pathogenesis of a mutant, attenuated strain (FMDV-Mut) to the parental, virulent virus from which the mutant was derived (FMDV-WT). The only difference between the two viruses was an insertion mutation in the inter-AUG region of the leader proteinase of FMDV-Mut. All cattle were infected by simulated-natural, aerosol inoculation. RESULTS: Both viruses were demonstrated to establish primary infection in the nasopharyngeal mucosa with subsequent dissemination to the lungs. Immunomicroscopic localization of FMDV antigens indicated that both viruses infected superficial epithelial cells of the nasopharynx and lungs. The critical differences between the two viruses were a more rapid establishment of infection by FMDV-WT and quantitatively greater virus loads in secretions and infected tissues compared to FMDV-Mut. The slower replicating FMDV-Mut established a subclinical infection that was limited to respiratory epithelial sites, whereas the faster replication of FMDV-WT facilitated establishment of viremia, systemic dissemination of infection, and clinical disease. CONCLUSION: The mutant FMDV was capable of achieving all the same early pathogenesis landmarks as FMDV-WT, but was unable to establish systemic infection. The precise mechanism of attenuation remains undetermined; but current data suggests that the impaired replication of the mutant is more responsible for attenuation than differences in host immunological factors. These results complement previous studies by providing data of high-granularity describing tissue-specific tropism of FMDV and by demonstrating microscopic localization of virulent and attenuated clones of the same field-strain FMDV.

Phylodynamics of foot-and-mouth disease virus O/PanAsia in Vietnam 2010-2014.

Foot-and-mouth disease virus (FMDV) is endemic in Vietnam, a country that plays an important role in livestock trade within Southeast Asia. The large populations of FMDV-susceptible species in Vietnam are important components of food production and of the national livelihood. In this study, we investigated the phylogeny of FMDV O/PanAsia in Vietnam, reconstructing the virus' ancestral host species (pig, cattle or buffalo), clinical stage (subclinical carrier or clinically affected) and geographical location. Phylogenetic divergence time estimation and character state reconstruction analyses suggest that movement of viruses between species differ. While inferred transmissions from cattle to buffalo and pigs and from pigs to cattle are well supported, transmission from buffalo to other species, and from pigs to buffalo may be less frequent. Geographical movements of FMDV O/PanAsia virus appears to occur in all directions within the country, with the South Central Coast and the Northeast regions playing a more important role in FMDV O/PanAsia spread. Genetic selection of variants with changes at specific sites within FMDV VP1 coding region was different depending on host groups analyzed. The overall ratio of non-synonymous to synonymous nucleotide changes was greater in pigs compared to cattle and buffalo, whereas a higher number of individual amino acid sites under positive selection were detected in persistently infected, subclinical animals compared to viruses collected from clinically diseased animals. These results provide novel insights to understand FMDV evolution and its association with viral spread within endemic countries. These findings may support animal health organizations in their endeavor to design animal disease control strategies in response to outbreaks.

Systemic immune response and virus persistence after foot-and-mouth disease virus infection of naïve cattle and cattle vaccinated with a homologous adenovirus-vectored vaccine.

BACKGROUND: In order to investigate host factors associated with the establishment of persistent foot-and-mouth disease virus (FMDV) infection, the systemic response to vaccination and challenge was studied in 47 steers. Eighteen steers that had received a recombinant FMDV A vaccine 2 weeks earlier and 29 non-vaccinated steers were challenged by intra-nasopharyngeal deposition of FMDV A24. For up to 35 days after challenge, host factors including complete blood counts with T lymphocyte subsets, type I/III interferon (IFN) activity, neutralizing and total FMDV-specific antibody titers in serum, as well as antibody-secreting cells (in 6 non-vaccinated animals) were characterized in the context of viral infection dynamics. RESULTS: Vaccination generally induced a strong antibody response. There was a transient peak of FMDV-specific serum IgM in non-vaccinated animals after challenge, while IgM levels in vaccinated animals did not increase further. Both groups had a lasting increase of specific IgG and neutralizing antibody after challenge. Substantial systemic IFN activity in non-vaccinated animals coincided with viremia, and no IFN or viremia was detected in vaccinated animals. After challenge, circulating lymphocytes decreased in non-vaccinated animals, coincident with viremia, IFN activity, and clinical disease, whereas lymphocyte and monocyte counts in vaccinated animals were unaffected by vaccination but transiently increased after challenge. The CD4(+)/CD8(+) T cell ratio in non-vaccinated animals increased during acute infection, driven by an absolute decrease of CD8(+) cells. CONCLUSIONS: The incidence of FMDV persistence was 61.5 % in non-vaccinated and 54.5 % in vaccinated animals. Overall, the systemic factors examined were not associated with the FMDV carrier/non-carrier divergence; however, significant differences were identified between responses of non-vaccinated and vaccinated cattle.

Cell culture adaptation mutations in foot-and-mouth disease virus serotype A capsid proteins: implications for receptor interactions.

In this study we describe the adaptive changes fixed on the capsid of several foot-and-mouth disease virus serotype A strains during propagation in cell monolayers. Viruses passaged extensively in three cell lines (BHK-21, LFBK and IB-RS-2) consistently gained positively charged amino acids in the putative heparin-sulfate-binding pocket (VP2 ßE-ßF loop, VP1 C-terminus and VP3 ß-B knob) surrounding the fivefold symmetry axis (VP1 ßF-ßG loop) and at other discrete sites on the capsid (VP3 ßG-ßH loop, VP1 C-terminus, VP2 ßC strand and VP1 ßG-ßH loop). A lysine insertion in the VP1 ßF-ßG loop of two of the BHK-21-adapted viruses supports the biological advantage of positively charged residues acquired in cell culture. The charge transitions occurred irrespective of cell line, suggesting their possible role in ionic interaction with ubiquitous negatively charged cell-surface molecules such as glycosaminoglycans (GAG). This was supported by the ability of the cell-culture-adapted variants to replicate in the integrin-deficient, GAG-positive CHO-K1 cells and their superior fitness in competition assays compared with the lower passage viruses with WT genotypes. Substitutions fixed in the VP1 ßG-ßH loop (-3, -2 and +2 'RGD' positions) or in the structural element known to be juxtaposed against that loop (VP1 ßB-ßC loop) suggest their possible role in modulating the efficiency and specificity of interaction of the 'RGD' motif with αv-integrin receptors. The nature and location of the substitutions described in this study could be applied in the rapid cell culture adaptation of viral strains for vaccine production.

Expression of porcine fusion protein IRF7/3(5D) efficiently controls foot-and-mouth disease virus replication.

UNLABELLED: Several studies have demonstrated that the delivery of type I, II, or III interferons (IFNs) by inoculation of a replication-defective human adenovirus 5 (Ad5) vector expressing IFNs can effectively control foot-and-mouth disease (FMD) in cattle and swine during experimental infections. However, relatively high doses are required to achieve protection. In this study, we identified the functional properties of a porcine fusion protein, poIRF7/3(5D), as a biotherapeutic and enhancer of IFN activity against FMD virus (FMDV). We showed that poIRF7/3(5D) is a potent inducer of type I IFNs, including alpha IFN (IFN-α), IFN-ß, and IFN-ω but not type III IFN (interleukin-28B), without inducing cytotoxicity. Expression of poIRF7/3(5D) significantly and steadily reduced FMDV titers by up to 6 log10 units in swine and bovine cell lines. Treatment with an IFN receptor inhibitor (B18R) combined with an anti-IFN-α antibody neutralized the antiviral activity in the supernatants of cells transduced with an Ad5 vector expressing poIRF7/3(5D) [Ad5-poIRF7/3(5D)]. However, several transcripts with known antiviral function, including type I IFNs, were still highly upregulated (range of increase, 8-fold to over 500-fold) by poIRF7/3(5D) in the presence of B18R. Furthermore, the sera of mice treated with Ad5-poIRF7/3(5D) showed antiviral activity that was associated with the induction of high levels of IFN-α and resulted in complete protection against FMDV challenge at 6, 24, or 48 h posttreatment. This study highlights for the first time the antiviral potential of Ad5-poIRF7/3(5D) in vitro and in vivo against FMDV. IMPORTANCE: FMD remains one of the most devastating diseases that affect livestock worldwide. Effective vaccine formulations are available but are serotype specific and require approximately 7 days before they are able to elicit protective immunity. We have shown that vector-delivered IFN is an option to protect animals against many FMDV serotypes as soon as 24 h and for about 4 days postadministration. Here we demonstrate that delivery of a constitutively active transcription factor that induces the production of endogenous IFNs and potentially other antiviral genes is a viable strategy to protect against FMD.

Interrogating Genomes and Geography to Unravel Multiyear Vesicular Stomatitis Epizootics.

We conducted an integrative analysis to elucidate the spatial epidemiological patterns of the Vesicular Stomatitis New Jersey virus (VSNJV) during the 2014-15 epizootic cycle in the United States (US). Using georeferenced VSNJV genomics data, confirmed vesicular stomatitis (VS) disease cases from surveillance, and a suite of environmental factors, our study assessed environmental and phylogenetic similarity to compare VS cases reported in 2014 and 2015. Despite uncertainties from incomplete virus sampling and cross-scale spatial processes, patterns suggested multiple independent re-invasion events concurrent with potential viral overwintering between sequential seasons. Our findings pointed to a geographically defined southern virus pool at the US-Mexico interface as the source of VSNJV invasions and overwintering sites. Phylodynamic analysis demonstrated an increase in virus diversity before a rise in case numbers and a pronounced reduction in virus diversity during the winter season, indicative of a genetic bottleneck and a significant narrowing of virus variation between the summer outbreak seasons. Environment-vector interactions underscored the central role of meta-population dynamics in driving disease spread. These insights emphasize the necessity for location- and time-specific management practices, including rapid response, movement restrictions, vector control, and other targeted interventions.

Community composition of black flies during and after the 2020 vesicular stomatitis virus outbreak in Southern New Mexico, USA.

BACKGROUND: Vesicular stomatitis virus (VSV), a vector-borne pathogen of livestock, emerges periodically in the western US. In New Mexico (NM), US, most cases occur close to the Rio Grande River, implicating black flies (Simulium spp.) as a possible vector. In 2020, VS cases were reported in NM from April to May, although total black fly abundance remained high until September. We investigated the hypothesis that transience of local VSV transmission results from transient abundance of key, competent black fly species. Additionally, we investigated whether irrigation canals in southern NM support a different community of black flies than the main river. Lastly, to gain insight into the source of local black flies, in 2023 we collected black fly larvae prior to the release of water into the Rio Grande River channel. METHODS: We randomly sub-sampled adult black flies collected along the Rio Grande during and after the 2020 VSV outbreak. We also collected black fly adults along the river in 2021 and 2022 and at southern NM farms and irrigation canals in 2022. Black fly larvae were collected from dams in the area in 2023. All collections were counted, and individual specimens were subjected to molecular barcoding for species identification. RESULTS: DNA barcoding of adult black flies detected four species in 2020: Simulium meridionale (N = 158), S. mediovittatum (N = 83), S. robynae (N = 26) and S. griseum/notatum (N = 1). Simulium robynae was only detected during the VSV outbreak period, S. meridionale showed higher relative abundance, but lower absolute abundance, during the outbreak than post-outbreak period, and S. mediovittatum was rare during the outbreak period but predominated later in the summer. In 2022, relative abundance of black fly species did not differ significantly between the Rio Grande sites and farm and irrigation canals. Intriguingly, 63 larval black flies comprised 56% Simulium vittatum, 43% S. argus and 1% S. encisoi species that were either extremely rare or not detected in previous adult collections. CONCLUSIONS: Our results suggest that S. robynae and S. meridionale could be shaping patterns of VSV transmission in southern NM. Thus, field studies of the source of these species as well as vector competence studies are warranted.

Modeling the 2014-2015 Vesicular Stomatitis Outbreak in the United States Using an SEIR-SEI Approach.

Vesicular stomatitis (VS) is a vector-borne livestock disease caused by the vesicular stomatitis New Jersey virus (VSNJV). This study presents the first application of an SEIR-SEI compartmental model to analyze VSNJV transmission dynamics. Focusing on the 2014-2015 outbreak in the United States, the model integrates vertebrate hosts and insect vector demographics while accounting for heterogeneous competency within the populations and observation bias in documented disease cases. Key epidemiological parameters were estimated using Bayesian inference and Markov chain Monte Carlo (MCMC) methods, including the force of infection, effective reproduction number (Rt), and incubation periods. The model revealed significant underreporting, with only 10-24% of infections documented, 23% of which presented with clinical symptoms. These findings underscore the importance of including competence and imperfect detection in disease models to depict outbreak dynamics and inform effective control strategies accurately. As a baseline model, this SEIR-SEI implementation is intended to serve as a foundation for future refinements and expansions to improve our understanding of VS dynamics. Enhanced surveillance and targeted interventions are recommended to manage future VS outbreaks.

A continuous bovine kidney cell line constitutively expressing bovine αvß6 integrin has increased susceptibility to foot-and-mouth disease virus.

Foot-and-mouth disease (FMD) is a worldwide problem limiting the trade of animals and their products from affected countries. The rapid isolation, serotyping, and vaccine matching of FMD virus from disease outbreaks is critical for enabling the implementation of effective vaccination programs and to stop the spread of infection during outbreaks. Some primary cells have been shown to be highly susceptible to most strains of FMD virus (FMDV) but are difficult and expensive to prepare and maintain. Since the αVß6 integrin is a principal receptor for FMDV, we transduced a bovine kidney cell line to stably express both the αV and ß6 bovine integrin subunits. This stable cell line (LFBK-αVß6) showed ß6 expression and enhanced susceptibility to FMDV infection for ≥ 100 cell passages. LFBK-αVß6 cells were highly sensitive for detecting all serotypes of FMDV from experimentally infected animals, including the porcinophilic FMDV strain O/TAW/97. In comparison to other cell types that are currently used for virus isolation, LFBK-αVß6 cells were more effective at detecting FMDV in clinical samples, supporting their use as a more sensitive tool for virus isolation.

A safe foot-and-mouth disease vaccine platform with two negative markers for differentiating infected from vaccinated animals.

Vaccination of domestic animals with chemically inactivated foot-and-mouth disease virus (FMDV) is widely practiced to control FMD. Currently, FMD vaccine manufacturing requires the growth of large volumes of virulent FMDV in biocontainment-level facilities. Here, two marker FMDV vaccine candidates (A(24)LL3D(YR) and A(24)LL3B(PVKV)3D(YR)) featuring the deletion of the leader coding region (L(pro)) and one of the 3B proteins were constructed and evaluated. These vaccine candidates also contain either one or two sets of mutations to create negative antigenic markers in the 3D polymerase (3D(pol)) and 3B nonstructural proteins. Two mutations in 3D(pol), H(27)Y and N(31)R, as well as RQKP(9-12)→PVKV substitutions, in 3B(2) abolish reactivity with monoclonal antibodies targeting the respective sequences in 3D(pol) and 3B. Infectious cDNA clones encoding the marker viruses also contain unique restriction endonuclease sites flanking the capsid-coding region that allow for easy derivation of custom designed vaccine candidates. In contrast to the parental A(24)WT virus, single A(24)LL3D(YR) and double A(24)LL3B(PVKV)3D(YR) mutant viruses were markedly attenuated upon inoculation of cattle using the natural aerosol or direct tongue inoculation. Likewise, pigs inoculated with live A(24)LL3D(YR) virus in the heel bulbs showed no clinical signs of disease, no fever, and no FMD transmission to in-contact animals. Immunization of cattle with chemically inactivated A(24)LL3D(YR) and A(24)LL3B(PVKV)3D(YR) vaccines provided 100% protection from challenge with parental wild-type virus. These attenuated, antigenically marked viruses provide a safe alternative to virulent strains for FMD vaccine manufacturing. In addition, a competitive enzyme-linked immunosorbent assay targeted to the negative markers provides a suitable companion test for differentiating infected from vaccinated animals.

Exploring the Molecular Basis of Vesicular Stomatitis Virus Pathogenesis in Swine: Insights from Expression Profiling of Primary Macrophages Infected with M51R Mutant Virus.

Vesicular stomatitis virus (VSV) is an emergent virus affecting livestock in the US. Previously, using a recombinant VSV carrying the M51R mutation in the matrix protein (rNJ0612NME6-M51R), we evaluated the pathogenesis of this virus in pigs. Our results indicated that rNJ0612NME6-M51R represented an attenuated phenotype in in-vivo and in ex-vivo in pig macrophages, resembling certain clinical features observed in field VSV isolates. In order to gain more insight into the molecular basis leading to the attenuation of rNJ0612NME6-M51R in pigs, we conducted a microarray analysis to assess the gene expression profiles of primary porcine macrophages infected with rNJ0612NME6-M51R compared to its parental virus (rNJ0612NME6). Our results showed an overall higher gene expression in macrophages infected with rNJ0612NME6-M51R. Specifically, we observed that the pathways related with immune cytokine signaling and interferon (IFN)-related responses (including activation, signaling, induction, and antiviral mechanisms) were the ones comprising most of the relevant genes identified during this study. Collectively, the results presented herein highlight the relevance of type I interferon during the pathogenesis of VSV in pigs. The information generated from this study may represent a framework for future studies intended to understand the molecular bases of the pathogenesis of field strains in livestock.

Inferred Causal Mechanisms of Persistent FMDV Infection in Cattle from Differential Gene Expression in the Nasopharyngeal Mucosa.

Foot-and-mouth disease virus (FMDV) can persistently infect pharyngeal epithelia in ruminants but not in pigs. Our previous studies demonstrated that persistent FMDV infection in cattle was associated with under-expression of several chemokines that recruit immune cells. This report focuses on the analysis of differentially expressed genes (DEG) identified during the transitional phase of infection, defined as the period when animals diverge between becoming carriers or terminators. During this phase, Th17-stimulating cytokines (IL6 and IL23A) and Th17-recruiting chemokines (CCL14 and CCL20) were upregulated in animals that were still infected (transitional carriers) compared to those that had recently cleared infection (terminators), whereas chemokines recruiting neutrophils and CD8+ T effector cells (CCL3 and ELR+CXCLs) were downregulated. Upregulated Th17-specific receptor, CCR6, and Th17-associated genes, CD146, MIR155, and ThPOK, suggested increased Th17 cell activity in transitional carriers. However, a complex interplay of the Th17 regulatory axis was indicated by non-significant upregulation of IL17A and downregulation of IL17F, two hallmarks of TH17 activity. Other DEG suggested that transitional carriers had upregulated aryl hydrocarbon receptor (AHR), non-canonical NFκB signaling, and downregulated canonical NFκB signaling. The results described herein provide novel insights into the mechanisms of establishment of FMDV persistence. Additionally, the fact that ruminants, unlike pigs, produce a large amount of AHR ligands suggests a plausible explanation of why FMDV persists in ruminants, but not in pigs.

Genome of Bovine Viral Diarrhea Virus (BVDV) Contaminating a Continuous LFBK-αVß6 Cell Line.

Here, we report the genome of bovine viral diarrhea virus 1 (BVDV-1) contaminating a continuous fetal bovine kidney cell line. The cell line (LFBK-αVß6) is used for the rapid isolation and serotyping of foot-and-mouth disease virus (FMDV). The sequence contains the full polyprotein-coding sequence and partial untranslated regions (UTRs).

Comparison of Endemic and Epidemic Vesicular Stomatitis Virus Lineages in Culicoides sonorensis Midges.

Vesicular stomatitis virus (VSV) primarily infects livestock and is transmitted by direct contact and vectored by Culicoides midges (Diptera: Ceratopogonidae). Endemic to Central and South America, specific VSV lineages spread northward out of endemic regions of Mexico and into the U.S. sporadically every five to ten years. In 2012, a monophyletic epidemic lineage 1.1 successfully spread northward into the U.S. In contrast, the closest endemic ancestor, lineage 1.2, remained circulating exclusively in endemic regions in Mexico. It is not clear what roles virus-animal interactions and/or virus-vector interactions play in the ability of specific viral lineages to escape endemic regions in Mexico and successfully cause outbreaks in the U.S., nor the genetic basis for such incursions. Whole-genome sequencing of epidemic VSV 1.1 and endemic VSV 1.2 revealed significant differences in just seven amino acids. Previous studies in swine showed that VSV 1.1 was more virulent than VSV 1.2. Here, we compared the efficiency of these two viral lineages to infect the vector Culicoides sonorensis (Wirth and Jones) and disseminate to salivary glands for subsequent transmission. Our results showed that midges orally infected with the epidemic VSV 1.1 lineage had significantly higher infection dissemination rates compared to those infected with the endemic VSV 1.2 lineage. Thus, in addition to affecting virus-animal interactions, as seen with higher virulence in pigs, small genetic changes may also affect virus-vector interactions, contributing to the ability of specific viral lineages to escape endemic regions via vector-borne transmission.

Ecological and Anthropogenic Spatial Gradients Shape Patterns of Dispersal of Foot-and-Mouth Disease Virus in Uganda.

Using georeferenced phylogenetic trees, phylogeography allows researchers to elucidate interactions between environmental heterogeneities and patterns of infectious disease spread. Concordant with the increasing availability of pathogen genetic sequence data, there is a growing need for tools to test epidemiological hypotheses in this field. In this study, we apply tools traditionally used in ecology to elucidate the epidemiology of foot-and-mouth disease virus (FMDV) in Uganda. We analyze FMDV serotype O genetic sequences and their corresponding spatiotemporal metadata from a cross-sectional study of cattle. We apply step selection function (SSF) models, typically used to study wildlife habitat selection, to viral phylogenies to show that FMDV is more likely to be found in areas of low rainfall. Next, we use a novel approach, a resource gradient function (RGF) model, to elucidate characteristics of viral source and sink areas. An RGF model applied to our data reveals that areas of high cattle density and areas near livestock markets may serve as sources of FMDV dissemination in Uganda, and areas of low rainfall serve as viral sinks that experience frequent reintroductions. Our results may help to inform risk-based FMDV control strategies in Uganda. More broadly, these tools advance the phylogenetic toolkit, as they may help to uncover patterns of spread of other organisms for which genetic sequences and corresponding spatiotemporal metadata exist.

Immunobiology of Crimean-Congo hemorrhagic fever.

Human infection with Crimean-Congo hemorrhagic fever virus (CCHFV), a tick-borne pathogen in the family Nairoviridae, can result in a spectrum of outcomes, ranging from asymptomatic infection through mild clinical signs to severe or fatal disease. Studies of CCHFV immunobiology have investigated the relationship between innate and adaptive immune responses with disease severity, attempting to elucidate factors associated with differential outcomes. In this article, we begin by highlighting unanswered questions, then review current efforts to answer them. We discuss in detail current clinical studies and research in laboratory animals on CCHF, including immune targets of infection and adaptive and innate immune responses. We summarize data about the role of the immune response in natural infections of animals and humans and experimental studies in vitro and in vivo and from evaluating immune-based therapies and vaccines, and present recommendations for future research.