Search

Foot-and-mouth disease (FMD) is a severe and extremely contagious viral disease of cloven-hoofed domestic and wild animals, which leads to serious economic losses to the livestock industry globally. FMD is caused by the FMD virus (FMDV), a positive-strand RNA virus that belongs to the genus Aphthovirus, within the family Picornaviridae. Early detection and characterization of FMDV strains are key factors to control new outbreaks and prevent the spread of the disease. Here, we describe a direct RNA sequencing method using Oxford Nanopore Technology (ONT) Flongle flow cells on MinION Mk1C (or GridION) to characterize FMDV. This is a rapid, low cost, and easily deployed point of care (POC) method for a near real-time characterization of FMDV in endemic areas or outbreak investigation sites.

African swine fever (ASF) is an acute, highly hemorrhagic viral disease in domestic pigs and wild boars. The disease is caused by African swine fever virus, a double stranded DNA virus of the Asfarviridae family. ASF can be classified into 25 different genotypes, based on a 478 bp fragment corresponding to the C-terminal sequence of the B646L gene, which is highly conserved among strains and encodes the major capsid protein p72. The C-terminal end of p72 has been used as a PCR target for quick diagnosis of ASF, and its characterization remains the first approach for epidemiological tracking and identification of the origin of ASF in outbreak investigations. Recently, a new classification of ASF, based on the complete sequence of p72, reduced the 25 genotypes into only six genotypes; therefore, it is necessary to have the capability to sequence the full-length B646L gene (p72) in a rapid manner for quick genotype characterization. Here, we evaluate the use of an amplicon approach targeting the whole B646L gene, coupled with nanopore sequencing in a multiplex format using Flongle flow cells, as an easy, low cost, and rapid method for the characterization and genotyping of ASF in real-time.

Abstract Background African swine fever virus (ASFV) infections in Africa cause hemorrhagic disease in domestic pigs and is maintained by a sylvatic cycle in warthogs. It is endemic in Uganda, leading to significant economic losses. Previous studies performed in rural areas and in Kampala had differing diagnostic results. The purpose of this study was to provide a robust spatial, temporal, and diagnostic summary of pigs slaughtered in the greater Kampala metropolitan area over the course of one year. This study characterized 1208 to 1323 serum, blood, and tissue samples collected from pigs at six abattoirs in the greater Kampala metropolitan area of Uganda monthly from May 2021 through June 2022. Validated and standardized serologic and molecular diagnostics were used. Results Only 0.15% of pigs had detectable antibodies against ASFV, suggesting low survival rates or pre-clinical diagnosis. Yet, 59.5% of pigs were positive for ASFV DNA. Blood had the lowest detection rate (15.3%) while tonsil and lymph nodes had the highest (38% and 37.5%, respectively), spleen samples (31.5%) were in between. Agreement between sample types was fair to moderate overall. A significant seasonality of ASFV infections emerged with infections found predominately in the dry seasons. Spatial assessments revealed that the greater Kampala metropolitan area abattoirs have a catchment area that overlaps with Uganda’s most pig dense regions. Conclusions Pigs at greater Kampala metropolitan area abattoirs can be sentinels for acute disease throughout the pig dense region of Uganda, particularly in the dry seasons. The high prevalence detected suggests that pigs are sold in response to local reports of ASFV infections (panic sales). Serological surveillance is not useful, as very few pigs seroconverted in this study prior to slaughter. In contrast, tissue samples of pigs can be used to detect disease using qPCR methods.

Crimean-Congo hemorrhagic fever (CCHF) and Rift Valley fever (RVF) are among the list of emerging zoonotic diseases that require special attention and priority. RVF is one of the six priority diseases selected by the Senegalese government. Repeated epidemic episodes and sporadic cases of CCHF and RVF in Senegal motivated this study, involving a national cross-sectional serological survey to assess the distribution of the two diseases in this country throughout the small ruminant population. A total of 2127 sera from small ruminants (goat and sheep) were collected in all regions of Senegal. The overall seroprevalence of CCHF and RVF was 14.1% (IC 95%: 12.5–15.5) and 4.4% (95% CI: 3.5–5.3), respectively. The regions of Saint-Louis (38.4%; 95% CI: 30.4–46.2), Kolda (28.3%; 95% CI: 20.9–35.7), Tambacounda (22.2%; 95% CI: 15.8–28.6) and Kédougou (20.9%; 95% CI: 14.4–27.4) were the most affected areas. The risk factors identified during this study show that the age, species and sex of the animals are key factors in determining exposure to these two viruses. This study confirms the active circulation of CCHF in Senegal and provides important and consistent data that can be used to improve the surveillance strategy of a two-in-one health approach to zoonoses.

African swine fever virus (ASFV) is the causative agent of African swine fever (ASF), a highly infectious and lethal disease of domesticated swine. Outbreaks of ASF have been mostly restricted to the continent of Africa. The outbreaks that have occurred outside of Africa were controlled by extensive depopulation of the domesticated pig population. However, in 2007, an outbreak occurred in the country of Georgia, where ASFV infected wild pigs and quickly spread across eastern Europe. Since the reintroduction of ASF into Europe, variants of the current pandemic strain, ASFV Georgia 2007/01 (ASFV-G), which is classified as Genotype 2 based on p72 sequencing, have been reported in countries within western Europe, Asia, and the island of Hispaniola. Additionally, isolates collected in 2020 confirmed the presence of variants of ASFV-G in Nigeria. Recently, we reported similar variants of ASFV-G collected from domestic pigs suspected of dying of ASF in Ghana in 2022. Here, we retroactively report, based on full-length sequencing, that similar variants were present in Ghana in 2021. The SNP analysis revealed derivatives of ASFV with distinct genetic markers. Furthermore, we identified three full-length ASFV genomes as Genotype 1, indicating that there were two genotypes circulating in proximity during the 2021 ASF outbreaks in Ghana.

Obtaining a complete good-quality sequence and annotation for the long double-stranded DNA genome of the African swine fever virus (ASFV) from next-generation sequencing (NGS) technology has proven difficult, despite the increasing availability of reference genome sequences and the increasing affordability of NGS. A gap analysis conducted by the global African swine fever research alliance (GARA) partners identified that a standardized, automatic pipeline for NGS analysis was urgently needed, particularly for new outbreak strains. Whilst there are several diagnostic and research labs worldwide that collect isolates of the ASFV from outbreaks, many do not have the capability to analyze, annotate, and format NGS data from outbreaks for submission to NCBI, and some publicly available ASFV genomes have missing or incorrect annotations. We developed an automated, standardized pipeline for the analysis of NGS reads that directly provides users with assemblies and annotations formatted for their submission to NCBI. This pipeline is freely available on GitHub and has been tested through the GARA partners by examining two previously sequenced ASFV genomes; this study also aimed to assess the accuracy and limitations of two strategies present within the pipeline: reference-based (Illumina reads) and de novo assembly (Illumina and Nanopore reads) strategies.

Rift Valley fever (RVF) in ungulates and humans is caused by a mosquito-borne RVF phlebovirus (RVFV). Live attenuated vaccines are used in livestock (sheep and cattle) to control RVF in endemic regions during outbreaks. The ability of two or more different RVFV strains to reassort when co-infecting a host cell is a significant veterinary and public health concern due to the potential emergence of newly reassorted viruses, since reassortment of RVFVs has been documented in nature and in experimental infection studies. Due to the very limited information regarding the frequency and dynamics of RVFV reassortment, we evaluated the efficiency of RVFV reassortment in sheep, a natural host for this zoonotic pathogen. Co-infection experiments were performed, first in vitro in sheep-derived cells, and subsequently in vivo in sheep. Two RVFV co-infection groups were evaluated: group I consisted of co-infection with two wild-type (WT) RVFV strains, Kenya 128B-15 (Ken06) and Saudi Arabia SA01-1322 (SA01), while group II consisted of co-infection with the live attenuated virus (LAV) vaccine strain MP-12 and a WT strain, Ken06. In the in vitro experiments, the virus supernatants were collected 24 h post-infection. In the in vivo experiments, clinical signs were monitored, and blood and tissues were collected at various time points up to nine days post-challenge for analyses. Cell culture supernatants and samples from sheep were processed, and plaque-isolated viruses were genotyped to determine reassortment frequency. Our results show that RVFV reassortment is more efficient in co-infected sheep-derived cells compared to co-infected sheep. In vitro, the reassortment frequencies reached 37.9% for the group I co-infected cells and 25.4% for the group II co-infected cells. In contrast, we detected just 1.7% reassortant viruses from group I sheep co-infected with the two WT strains, while no reassortants were detected from group II sheep co-infected with the WT and LAV strains. The results indicate that RVFV reassortment occurs at a lower frequency in vivo in sheep when compared to in vitro conditions in sheep-derived cells. Further studies are needed to better understand the implications of RVFV reassortment in relation to virulence and transmission dynamics in the host and the vector. The knowledge learned from these studies on reassortment is important for understanding the dynamics of RVFV evolution.

African swine fever virus (ASFV) is the causative agent of a severe and highly contagious viral disease affecting domestic and wild swine. The current ASFV pandemic strain has a high mortality rate, severely impacting pig production and, for countries suffering outbreaks, preventing the export of their pig products for international trade. Early detection and diagnosis of ASFV is necessary to control new outbreaks before the disease spreads rapidly. One of the rate-limiting steps to identify ASFV by next-generation sequencing platforms is library preparation. Here, we investigated the capability of the Oxford Nanopore Technologies’ VolTRAX platform for automated DNA library preparation with downstream sequencing on Nanopore sequencing platforms as a proof-of-concept study to rapidly identify the strain of ASFV. Within minutes, DNA libraries prepared using VolTRAX generated near-full genome sequences of ASFV. Thus, our data highlight the use of the VolTRAX as a platform for automated library preparation, coupled with sequencing on the MinION Mk1C for field sequencing or GridION within a laboratory setting. These results suggest a proof-of-concept study that VolTRAX is an effective tool for library preparation that can be used for the rapid and real-time detection of ASFV.

African swine fever (ASF) is a lethal disease of domestic pigs that has been causing outbreaks for over a century in Africa ever since its first discovery in 1921. Since 1957, there have been sporadic outbreaks outside of Africa; however, no outbreak has been as devastating and as far-reaching as the current pandemic that originated from a 2007 outbreak in the Republic of Georgia. Derivatives with a high degree of similarity to the progenitor strain, ASFV-Georgia/2007, have been sequenced from various countries in Europe and Asia. However, the current strains circulating in Africa are largely unknown, and 24 different genotypes have been implicated in different outbreaks. In this study, ASF isolates were collected from samples from swine suspected of dying from ASF on farms in Ghana in early 2022. While previous studies determined that the circulating strains in Ghana were p72 Genotype I, we demonstrate here that the strains circulating in 2022 were derivatives of the p72 Genotype II pandemic strain. Therefore, this study demonstrates for the first time the emergence of Genotype II ASFV in Ghana.

Blood samples were collected from pigs at six abattoirs in the Kampala, Uganda metropolitan area from May 2021 through June 2022, and tested for African swine fever virus. Thirty-one samples with cycle threshold values < 26 from pigs with different geographic origins, clinical and pathologic signs, and Ornithodoros moubata exposure underwent whole genome sequencing. The p72 gene was used to genotype the isolates, and all were found to be genotype IX; whole genome sequences to previous genotype IX isolates confirmed their similarity. Six of the isolates had enough coverage to evaluate single nucleotide polymorphisms (SNPs). Five of the isolates differed from historic regional isolates, but had similar SNPs to one another, and the sixth isolate also differed from historic regional isolates, but also differed from the other five isolates, even though they are all genotype IX. Whole genome sequencing data provide additional detail on viral evolution that can be useful for molecular epidemiology, and understanding the impact of changes in genes to disease phenotypes, and may be needed for vaccine targeting should a commercial vaccine become available. More sequencing of African swine fever virus isolates is needed in Uganda to understand how and when the virus is changing.

Crimean–Congo hemorrhagic fever virus (CCHFV) is a widely distributed tickborne zoonotic agent that infects a variety of host species. There is a lack of information on the true geographic distribution of the prevalence and risk of CCHFV in West Africa. A countrywide cross-sectional study involving 1413 extensively managed indigenous small ruminants and cattle at livestock sales markets and in village herds, respectively, was carried out in The Gambia. In sheep, an overall anti-CCHFV antibody prevalence of 18.9% (95% CI: 15.5–22.8%), goats 9.0% (95% CI: 6.7–11.7%), and cattle 59.9% (95% CI: 54.9–64.7%) was detected. Significant variation (p < 0.05) in the prevalence of anti-CCHFV antibodies at sites in the five administrative regions (sheep: 4.8–25.9%; goats: 1.8–17.1%) and three agroecological zones (sheep: 8.9–32.9%; goats: 4.1–18.0%) was also observed. Comparatively, higher anti-CCHFV antibody prevalence was detected in cattle (33.3–84.0%) compared to small ruminants (1.8–8.1%). This study represents the first countrywide investigation of the seroprevalence of CCHFV in The Gambia, and the results suggest potential circulation and endemicity of the virus in the country. These data provide critical information vital to the development of informed policies for the surveillance, diagnosis, and control of CCFHV infection in The Gambia and the region.

Arboviruses are important public health threats in many regions of the world. Nigeria has experienced outbreaks of arboviruses over the past decades, leading to concerns of widespread endemicity, which are frequently misdiagnosed. This study aimed to determine the seroprevalence of dengue virus (DENV) (a flavivirus) and chikungunya virus (CHIKV) (an alphavirus) infections in three major population centers of Nigeria. A convenience sample of 701 sera was collected from both healthy and febrile participants between August 2010 and March 2018. Sera were tested for prior exposure to CHIKV virus and DENV using indirect IgG ELISA. Results showed that 54.1% (379/701) of participants were seropositive for anti-DENV antibodies, 41.3% (290/701) were seropositive for anti-CHIKV antibodies, and 20.1% (141/701) had previous exposure to both. The seropositivity for prior CHIKV exposure and prior exposure to DENV and CHIKV was significantly associated with age (CHIKV: OR = 2.7 (95% CI: 1.7–4.3); DENV and CHIKV: OR = 2.2 (95% CI: 1.2–4.0) for adults compared to participants under 18 years old). Overall, the high seropositivity across all age groups suggests that arboviral infections are prevalent in Nigeria and indicates that surveillance and further epidemiological studies are required to determine the true burden of these infections and the spectrum of diseases associated with these exposures.

African swine fever virus (ASFV) is the etiological agent of African swine fever (ASF), a disease of domestic and wild swine that has spread throughout a large geographical area including Central Europe, East and Southeast Asia, and Southern Africa. Typically, the clinical presentation of the disease in affected swine heavily depends on the virulence of the ASFV strain. Very recently, ASFV was detected in the Dominican Republic (DR) and Haiti, constituting the first diagnosis of ASFV in more than 40 years in the Western hemisphere. In this report, the clinical presentation of the disease in domestic pigs inoculated with an ASFV field strain isolated from samples collected in the DR (ASFV-DR21) was observed. Two groups of domestic pigs were inoculated either intramuscularly (IM) or oronasally (ON) with ASFV-DR21 (104 hemadsorbing dose-50% (HAD50)). A group of naïve pigs (designated as the contact group) was co-housed with the ASFV-DR21 IM-inoculated animals to evaluate ASFV transmission and disease manifestation. Animals inoculated IM with ASFV-DR21 developed an acute disease leading to humane euthanasia at approximately day 7 post-inoculation (pi). Interestingly, animals inoculated via the ON route with ASFV-DR21 developed a heterogeneous pattern of disease kinetics. One animal developed an acute form of the disease and was euthanized on day 7 pi, another animal experienced a protracted presentation of the disease with euthanasia by day 16 pi, and the remaining two animals presented a milder form of the disease, surviving through the 28-day observational period. The contact animals also presented with a heterogenous presentation of the disease. Three of the animals presented protracted but severe forms of the disease being euthanized at days 14, 15 and 21 pi. The other two animals presented with a milder form of the disease, surviving the entire observational period. In general, virus titers in the blood of animals in all study groups closely followed the clinical presentation of the disease, both in length and extent. Importantly, all animals presenting with a prolonged form of the disease, as well as those surviving throughout the observational period, developed a strong ASFV-specific antibody response. These results suggest that ASFV-DR21, unless inoculated parenterally, produces a spectrum of clinical disease, with some animals experiencing an acute fatal form while others presented with a mild transient disease accompanied by the induction of a strong antibody response. At the time of publication, this is the first report characterizing the virulent phenotype of an ASFV field strain isolated from samples collected in the DR during the 2021 outbreak and provides information that may be used in developing epidemiological management measures to control ASF on the island of Hispaniola.

African swine fever (ASF) is a highly contagious transboundary viral hemorrhagic disease of domestic and wild pigs presenting a significant threat to the global swine industry. Following its introduction in Caucasus, Georgia, in 2007, the genome of the genotype II of African swine fever virus (ASFV) strain Georgia-07 and its derivatives accumulated significant mutations, resulting in the emergence of genetic variants within short epidemiological timescales as it spreads and infects different hosts in diverse ecosystems, causing outbreaks in Europe, South Asia, South East Asia and the Caribbean. This suggests that ASFV, with a comparatively large and complex DNA genome, is susceptible to genetic mutations including deletions and that although the virus is environmentally stable, it is genetically unstable. This has implications for the development of vaccines and diagnostic tests for disease detection and surveillance. Analysis of the ASFV genome revealed recombination hotspots, which in double-stranded DNA (dsDNA) viruses represent key drivers of genetic diversity. The ability of pox virus, a dsDNA virus with a genome complexity similar to ASFV, regaining virulence following the deletion of a virulence gene via gene amplification, coupled with the recent emergence and spread of live-attenuated ASFV vaccine strains causing disease and death in pigs in China, raise legitimate concerns around the use of live-attenuated ASFV vaccines in non-endemic regions to control the potential introduction. Further research into the risk of using live-attenuated ASFV in non-endemic regions is highly needed.

Rift Valley fever virus (RVFV) is a mosquito-borne zoonotic pathogen that causes periodic outbreaks of abortion in ruminant species and hemorrhagic disease in humans in sub-Saharan Africa. These outbreaks have a significant impact on veterinary and public health. Its introduction to the Arabian Peninsula in 2003 raised concerns of further spread of this transboundary pathogen to non-endemic areas. These concerns are supported by the presence of competent vectors in many non-endemic countries. There is no licensed RVF vaccine available for humans and only a conditionally licensed veterinary vaccine available in the United States. Currently employed modified live attenuated virus vaccines in endemic countries lack the ability for differentiating infected from vaccinated animals (DIVA). Previously, the efficacy of a recombinant subunit vaccine based on the RVFV Gn and Gc glycoproteins, derived from the 1977 human RVFV isolate ZH548, was demonstrated in sheep. In the current study, cattle were vaccinated subcutaneously with the Gn only, or Gn and Gc combined, with either one or two doses of the vaccine and then subjected to heterologous virus challenge with the virulent Kenya-128B-15 RVFV strain, isolated from Aedes mosquitoes in 2006. The elicited immune responses by some vaccine formulations (one or two vaccinations) conferred complete protection from RVF within 35 days after the first vaccination. Vaccines given 35 days prior to RVFV challenge prevented viremia, fever and RVFV-associated histopathological lesions. This study indicates that a recombinant RVFV glycoprotein-based subunit vaccine platform is able to prevent and control RVFV infections in target animals.

Rift Valley fever virus, a zoonotic arbovirus, poses major health threats to livestock and humans if introduced into the United States. White-tailed deer, which are abundant throughout the country, might be sentinel animals for arboviruses. We determined the susceptibility of these deer to this virus and provide evidence for a potentially major epidemiologic role.

Epizootic hemorrhagic disease virus (EHDV) is an arthropod-transmitted RNA virus and the causative agent of epizootic hemorrhagic disease (EHD) in wild and domestic ruminants. In North America, white-tailed deer (WTD) experience the highest EHD-related morbidity and mortality, although clinical disease is reported in cattle during severe epizootics. No commercially licensed EHDV vaccine is available in North America. The objective of this study was to develop and evaluate a subunit vaccine candidate to control EHD in WTD. Recombinant VP2 (rVP2) outer capsid proteins of EHDV serotypes 2 (EHDV-2) and 6 (EHDV-6) were produced in a baculovirus-expression system. Mice and cattle vaccinated with EHDV-2 or EHDV-6 rVP2 produced homologous virus-neutralizing antibodies. In an immunogenicity/efficacy study, captive-bred WTD received 2 doses of EHDV-2 rVP2 or sham vaccine, then were challenged with wild-type EHDV-2 at 30 d post vaccination. None of the rVP2-vaccinated deer developed clinical disease, no viral RNA was detected in their blood or tissues (liver, lung, spleen, kidney), and no EHDV-induced lesions were observed. Sham-vaccinated deer developed clinical disease with viremia and typical EHD vascular lesions. Here, we demonstrate a rVP2 subunit vaccine that can provide protective immunity from EHDV infection and which may serve as an effective tool in preventing clinical EHD and reducing virus transmission.

Heartwater, or cowdriosis, is a tick-borne disease of domestic and wild ruminants that is endemic in the Caribbean and sub-Saharan Africa. The disease is caused by an intracellular pathogen, Ehrlichia ruminantium and may be fatal within days of the onset of clinical signs with mortality rates of up to 90% in susceptible hosts. Due to the presence of competent tick vectors in North America, there is substantial risk of introduction of heartwater with potentially devastating consequences to the domestic livestock industry. There is currently no reliable or safe vaccine for use globally. To develop a protective DIVA (differentiate infected from vaccinated animals) subunit vaccine for heartwater, we targeted the E. ruminantium immunodominant major antigenic protein1 (MAP1) with the hypothesis that MAP1 is a glycosylated protein and glycans contained in the antigenic protein are important epitope determinants. Using a eukaryotic recombinant baculovirus expression system, we expressed and characterized, for the first time, a glycoform profile of MAP1 of two Caribbean E. ruminantium isolates, Antigua and Gardel. We have shown that the 37-38 kDa protein corresponded to a glycosylated form of the MAP1 protein, whereas the 31-32 kDa molecular weight band represented the non-glycosylated form of the protein frequently reported in scientific literature. Three groups of sheep (n = 3-6) were vaccinated with increasing doses of a bivalent (Antigua and Gardel MAP1) rMAP1 vaccine cocktail formulation with montanide ISA25 as an adjuvant. The glycosylated recombinant subunit vaccine induced E. ruminantium-specific humoral and Th1 type T cell responses, which are critical for controlling intracellular pathogens, including E. ruminantium, in infected hosts. These results provide an important basis for development of a subunit vaccine as a novel strategy to protect susceptible livestock against heartwater in non-endemic and endemic areas.

First appearing in 2011 in Northern Europe, Schmallenberg virus (SBV), an Orthobunyavirus of the Simbu serogroup, is associated with clinical disease mainly in ruminants such as cattle, sheep and goats. The clinical signs are characterized by abortion and congenital deformities in newborns. The virus is transmitted by Culicoides midges of the Obsoletus complex. SBV infection induces a solid protective immunity that persists for at least 4 or 6 years in sheep and cattle, respectively. SBV infection can be diagnosed directly by real-time RT-qPCR and virus isolation or indirectly by serological assays. Three vaccines are commercially available in Europe. This article provides a comprehensive literature review on this emerging disease regarding pathogenesis, transmission, diagnosis, control and prevention. This review also highlights that although much has been learned since SBV’s first emergence, there are still areas that require further study to devise better mitigation strategies.

Rift Valley fever virus (RVFV) is a mosquito-borne zoonotic pathogen causing severe outbreaks in humans and livestock in sub-Saharan Africa and the Arabian Peninsula. Human infections are characterized by fever, sometimes leading to encephalitis, retinitis, hemorrhagic fever and occasionally death. There are currently no fully licensed vaccines or effective therapies for human use. Gene silencing mediated by double-stranded short interfering RNA (siRNA) is a sequence-specific, highly conserved mechanism in eukaryotes, which serves as an antiviral defence mechanism. Here, we demonstrate that siRNA duplexes directed against the RVFV nucleoprotein can effectively inhibit RVFV replication in human (MRC5 cells) and African green monkey cells (Vero E6 cells). Using these cells, we demonstrate that individual or complex siRNAs, targeting the RVFV nucleoprotein gene completely abrogate viral protein expression and prevent degradation of the host innate antiviral factor, protein kinase R (PKR). Importantly, pre-treatment of cells with the nucleoprotein-specific siRNAs markedly reduces the virus titer. The antiviral effect of the siRNAs was not attributable to interferon or the interferon response effector molecule, protein kinase R. Thus, the antiviral activity of RVFV nucleoprotein-specific siRNAs may provide novel therapeutic strategy against RVFV infections in animals and humans.

Rift Valley Fever (RVF) is a mosquito-borne zoonotic disease that presents a substantial threat to human and public health. It is caused by Rift Valley fever phlebovirus (RVFV), which belongs to the genus Phlebovirus and the family Phenuiviridae within the order Bunyavirales. The wide distribution of competent vectors in non-endemic areas coupled with global climate change poses a significant threat of the transboundary spread of RVFV. In the last decade, an improved understanding of the molecular biology of RVFV has facilitated significant progress in the development of novel vaccines, including DIVA (differentiating infected from vaccinated animals) vaccines. Despite these advances, there is no fully licensed vaccine for veterinary or human use available in non-endemic countries, whereas in endemic countries, there is no clear policy or practice of routine/strategic livestock vaccinations as a preventive or mitigating strategy against potential RVF disease outbreaks. The purpose of this review was to provide an update on the status of RVF vaccine development and provide perspectives on the best strategies for disease control. Herein, we argue that the routine or strategic vaccination of livestock could be the best control approach for preventing the outbreak and spread of future disease.

Bacterial gene transcription is initiated by RNA polymerase containing a sigma factor. To understand gene regulation in Ehrlichia chaffeensis, an important tick-transmitted rickettsiae responsible for human monocytic ehrlichiosis, we initiated studies evaluating the transcriptional machinery of several genes of this organism. We mapped the transcription start sites of 10 genes and evaluated promoters of five genes (groE, dnaK, hup, p28-Omp14 and p28-Omp19 genes). We report here that the RNA polymerase binding elements of E. chaffeensis gene promoters are highly homologous for its only two transcription regulators, sigma 32 and sigma 70, and that gene expression is accomplished by either of the transcription regulators. RNA analysis revealed that although transcripts for both sigma 32 and sigma 70 are upregulated during the early replicative stage, their expression patterns remained similar for the entire replication cycle. We further present evidence demonstrating that the organism's -35 motifs are essential to transcription initiations. The data suggest that E. chaffeensis gene regulation has evolved to support the organism's growth, possibly to facilitate its intraphagosomal growth. Considering the limited availability of genetic tools, this study offers a novel alternative in defining gene regulation in E. chaffeensis and other related intracellular pathogens.

Recent outbreaks of Rift Valley fever in ruminant livestock, characterized by mass abortion and high mortality rates in neonates, have raised international interest in improving vaccine control strategies. Previously, we developed a reliable challenge model for sheep that improves the evaluation of existing and novel vaccines in sheep. This sheep model demonstrated differences in the pathogenesis of Rift Valley fever virus (RVFV) infection between two genetically-distinct wild-type strains of the virus, Saudi Arabia 2001 (SA01) and Kenya 2006 (Ken06). Here, we evaluated the pathogenicity of these two RVFV strains in mixed breed beef calves. There was a transient increase in rectal temperatures with both virus strains, but this clinical sign was less consistent than previously reported with sheep. Three of the five Ken06-infected animals had an early-onset viremia, one day post-infection (dpi), with viremia lasting at least three days. The same number of SA01-infected animals developed viremia at 2 dpi, but it only persisted through 3 dpi in one animal. The average virus titer for the SA01-infected calves was 1.6 logs less than for the Ken06-infected calves. Calves, inoculated with either strain, seroconverted by 5 dpi and showed time-dependent increases in their virus-neutralizing antibody titers. Consistent with the results obtained in the previous sheep study, elevated liver enzyme levels, more severe liver pathology and higher virus titers occurred with the Ken06 strain as compared to the SA01 strain. These results demonstrate the establishment of a virulent challenge model for vaccine evaluation in calves.