Search

The reservoir for zoonotic o'nyong-nyong virus (ONNV) has remained unknown since this virus was first recognized in Uganda in 1959. Building on existing evidence for mosquito blood-feeding on various frugivorous bat species in Uganda, and seroprevalence for arboviruses among bats in Uganda, we sought to assess if serum samples collected from bats in Uganda demonstrated evidence of exposure to ONNV or the closely related zoonotic chikungunya virus (CHIKV). In total, 652 serum samples collected from six bat species were tested by plaque reduction neutralization test (PRNT) for neutralizing antibodies against ONNV and CHIKV. Forty out of 303 (13.2%) Egyptian rousettes from Maramagambo Forest and 1/13 (8%) little free-tailed bats from Banga Nakiwogo, Entebbe contained neutralizing antibodies against ONNV. In addition, 2/303 (0.7%) of these Egyptian rousettes contained neutralizing antibodies to CHIKV, and 8/303 (2.6%) contained neutralizing antibodies that were nonspecifically reactive to alphaviruses. These data support the interepidemic circulation of ONNV and CHIKV in Uganda, although Egyptian rousette bats are unlikely to serve as reservoirs for these viruses given the inconsistent occurrence of antibody-positive bats.

Rift Valley fever virus (RVFV) is a mosquito-transmitted virus with proven ability to emerge into naïve geographic areas. Limited field evidence suggests that RVFV is transmitted vertically from parent mosquito to offspring, but until now this mechanism has not been confirmed in the laboratory. Furthermore, this transmission mechanism has allowed for the prediction of RVFV epizootics based on rainfall patterns collected from satellite information. However, in spite of the relevance to the initiation of epizootic events, laboratory confirmation of vertical transmission has remained an elusive research aim for thirty-five years. Herein we present preliminary evidence of the vertical transmission of RVFV by Culex tarsalis mosquitoes after oral exposure to RVFV. Progeny from three successive gonotrophic cycles were reared to adults, with infectious RVFV confirmed in each developmental stage. Virus was detected in ovarian tissues of parental mosquitoes 7 days after imbibing an infectious bloodmeal. Infection was confirmed in progeny as early as the first gonotrophic cycle, with infection rates ranging from 2.0-10.0%. Virus titers among progeny were low, which may indicate a host mechanism suppressing replication., Competing Interests: The authors have declared that no competing interests exist.

Bunyaviruses (Negarnaviricota: Bunyavirales) are a large and diverse group of viruses that include important human, veterinary, and plant pathogens. The rapid characterization of known and new emerging pathogens depends on the availability of comprehensive reference sequence databases that can be used to match unknowns, infer evolutionary relationships and pathogenic potential, and make response decisions in an evidence-based manner. In this study, we determined the coding-complete genome sequences of 99 bunyaviruses in the Centers for Disease Control and Prevention's Arbovirus Reference Collection, focusing on orthonairoviruses (family Nairoviridae), orthobunyaviruses (Peribunyaviridae), and phleboviruses (Phenuiviridae) that either completely or partially lacked genome sequences. These viruses had been collected over 66 years from 27 countries from vertebrates and arthropods representing 37 genera. Many of the viruses had been characterized serologically and through experimental infection of animals but were isolated in the pre-sequencing era. We took advantage of our unusually large sample size to systematically evaluate genomic characteristics of these viruses, including reassortment, and co-infection. We corroborated our findings using several independent molecular and virologic approaches, including Sanger sequencing of 197 genome segments, and plaque isolation of viruses from putative co-infected virus stocks. This study contributes to the described genetic diversity of bunyaviruses and will enhance the capacity to characterize emerging human pathogenic bunyaviruses., Competing Interests: The authors have declared that no competing interests exist.

Understanding vertebrate-vector interactions is vitally important for understanding the transmission dynamics of arthropod-vectored pathogens and depends on the ability to accurately identify the vertebrate source of blood-engorged arthropods in field collections using molecular methods. A decade ago, molecular techniques being applied to arthropod blood meal identification were thoroughly reviewed, but there have been significant advancements in the techniques and technologies available since that time. This review highlights the available diagnostic markers in mitochondrial and nuclear DNA and discusses their benefits and shortcomings for use in molecular identification assays. Advances in real-time PCR, high resolution melting analysis, digital PCR, next generation sequencing, microsphere assays, mass spectrometry, and stable isotope analysis each offer novel approaches and advantages to bloodmeal analysis that have gained traction in the field. New, field-forward technologies and platforms have also come into use that offer promising solutions for point-of-care and remote field deployment for rapid bloodmeal source identification. Some of the lessons learned over the last decade, particularly in the fields of DNA barcoding and sequence analysis, are discussed. Though many advancements have been made, technical challenges remain concerning the prevention of sample degradation both by the arthropod before the sample has been obtained and during storage. This review provides a roadmap and guide for those considering modern techniques for arthropod bloodmeal identification and reviews how advances in molecular technology over the past decade have been applied in this unique biomedical context.

Accurate species-level identification of the source of arthropod bloodmeals is important for deciphering blood feeding patterns of field-collected specimens. Cytochrome c oxidase I (COI) mitochondrial gene sequencing has been used for this purpose; however, species resolution can be difficult to obtain from certain vertebrate genera, including Odocoileus. Sanger sequencing of mitochondrial genes was employed to identify the bloodmeal source of wild-caught mosquitoes trapped in Greeley, Colorado. Initial sequencing of the COI gene of mitochondrial DNA in bloodmeals was inadequate for species-level resolution of bloodmeals from deer in the genus Odocoileus, with current databases returning low fidelity matches to multiple genera. The use of the hypervariable D loop of the control region provided species-level identification of white-tailed deer (Order: Artiodactyla, Family: Cervidae, Odocoileus virginianus); however, taxonomic identification was successful only to genus for mule (O. hemionus hemionus) and black-tailed deer (O. hemionus columbianus). We advocate the use of multiple loci for bloodmeal analysis and the buildout of available databases to include multiple mitochondrial reference genes for reliable host species identification., (© The Author(s) 2020. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The yellow fever mosquito, Aedes aegypti, serves as the primary vector for epidemic transmission of yellow fever, dengue, Zika (ZIKV), and chikungunya viruses to humans. Control of Ae. aegypti is currently limited to insecticide applications and larval habitat management; however, to combat growing challenges with insecticide resistance, novel genetic approaches for vector population reduction or transmission interruption are being aggressively pursued. The objectives of this study were to assess the ability of the Ae. aegypti antiviral exogenous-small interfering RNA (exo-siRNA) response to inhibit ZIKV infection and transmission, and to identify the optimal RNA interference (RNAi) target region in the ZIKV genome. We accomplished these objectives by in vitro transcription of five long double-stranded RNAs (dsRNAs) from the genome region spanning the NS2B-NS3-NS4A genes, which were the most highly conserved among ZIKV RNA sequences representing both East and West African and Asian-American clades, and evaluation of the ability of these dsRNAs to trigger an effective antiviral exo-siRNA response after intrathoracic injection into Ae. aegypti. In a pilot study, five ZIKV dsRNAs were tested by intrathoracic inoculation of 250 ng dsRNA into groups of approximately 5-day-old mosquitoes. Three days post-inoculation, mosquitoes were provided an infectious blood-meal containing ZIKV strain PRVABC59 (Puerto Rico), MR766 (Uganda), or 41525 (Senegal). On days 7 and 14 post-infection individual whole mosquito bodies were assessed for ZIKV infectious titer by plaque assays. Based on the results of this initial assessment, three dsRNAs were selected for further evaluation of viral loads of matched body and saliva expectorants using a standardized infectious dose of 1 × 10 7  PFU/mL of each ZIKV strain. Fourteen days post-exposure to ZIKV, paired saliva and carcass samples were harvested from individual mosquitoes and assessed for ZIKV RNA load by qRT-PCR. Injection of each of the three dsRNAs resulted in significant inhibition of replication of all three strains of ZIKV in mosquito bodies and saliva. This study lays critical groundwork for pursuing ZIKV transmission-blocking strategies that exploit the Ae. aegypti exo-siRNA response for arbovirus suppression in natural populations., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Rift Valley fever virus (RVFV) poses a major threat of introduction to several continents, including North America. Such an introduction could cause significant losses to the livestock industry, in addition to substantial human morbidity and mortality. Because of the opportunistic blood host selection of Culex tarsalis mosquitoes, we hypothesized that this species could be an important bridge vector of RVFV near feedlots in the event of an introduction. We investigated the mosquito community composition at livestock feedlots and surrounding natural and residential areas to determine differences in mosquito relative abundance and blood feeding patterns attributed to cattle feeding operations. DNA extracted from abdomens of blood-fed mosquitoes were sequenced to determine host identity. Multivariate regression analyses revealed differences between mosquito community assemblages at feedlots and non-feedlot sites (p < 0.05), with this effect driven largely by differential abundances of Aedes vexans (p adj  < 0.05). Mosquito diversity was lower on feedlots than surrounding areas for three out of four feedlots. Culex tarsalis was abundant at both feedlots and nearby sites. Diverse vertebrate blood meals were detected in Cx. tarsalis at non-feedlot sites, with a shift towards feeding on cattle at feedlots. These data support a potential for Cx. tarsalis to serve as a bridge vector of RVFV between livestock and humans in Colorado., (© 2019 Blackwell Verlag GmbH.)

Western equine encephalitis (WEE) was once prevalent and routinely isolated from mosquitoes in Colorado; however, isolations of Western equine encephalitis virus (WEEV) have not been reported from mosquito pools since the early 1990s. The objective of the present study was to test pools of Culex tarsalis (Coquillett) mosquitoes sampled from Weld County, CO, in 2016 for evidence of WEEV infection. Over 7,000 mosquitoes were tested, but none were positive for WEEV RNA. These data indicate that WEEV either was not circulating enzootically in Northern Colorado, was very rare, and would require much more extensive mosquito sampling to detect, or was heterogeneously distributed spatially and temporally and happened to not be present in the area sampled during 2016. Even though the reported incidence of WEE remains null, screening for WEEV viral RNA in mosquito vectors offers forewarning toward the detection and prevention of future outbreaks., (© The Author(s) 2018. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Introduction: A number of arboviruses have previously been isolated from naturally-infected East African bats, however the role of bats in arbovirus maintenance is poorly understood. The aim of this study was to investigate the exposure history of Ugandan bats to a panel of arboviruses. Materials and methods: Insectivorous and fruit bats were captured from multiple locations throughout Uganda during 2009 and 2011-2013. All serum samples were tested for neutralizing antibodies against West Nile virus (WNV), yellow fever virus (YFV), dengue 2 virus (DENV-2), Zika virus (ZIKV), Babanki virus (BBKV), and Rift Valley fever virus (RVFV) by plaque reduction neutralization test (PRNT). Sera from up to 626 bats were screened for antibodies against each virus. Results and Discussion:  Key findings include the presence of neutralizing antibodies against RVFV in 5/52 (9.6%) of little epauletted fruit bats ( Epomophorus labiatus ) captured from Kawuku and 3/54 (5.6%) Egyptian rousette bats from Kasokero cave. Antibodies reactive to flaviviruses were widespread across bat taxa and sampling locations. Conclusion: The data presented demonstrate the widespread exposure of bats in Uganda to arboviruses, and highlight particular virus-bat associations that warrant further investigation., Competing Interests: No potential conflict of interest was reported by the authors.

A large number of arthropod-borne viruses are endemic to East Africa. As a part of the process of undertaking a systematic characterization of the mosquito fauna of Uganda, we examined mosquitoes collected from 2008 through early 2012 for known and novel viruses. In all, 8,288 mosquito pools containing 157,554 mosquitoes were tested. Twenty-nine isolations of 11 different viruses were made from mosquitoes of nine distinct species and from pools identified only to genus Culex. Identified viruses were from family Togaviridae, alphaviruses Sindbis and Babanki viruses; family Rhabdoviridae, hapaviruses Mossuril and Kamese viruses; family Flaviviridae, flaviviruses West Nile and Usutu viruses; family Phenuiviridae, phlebovirus Arumowot virus; and family Peribunyaviridae, orthobunyaviruses Witwatersrand, Pongola, and Germiston viruses. In addition, a novel orthobunyavirus, provisionally named Mburo virus, was isolated from Coquillettidia metallica (Theobald). This is the first report of Babanki, Arumowot, and Mossuril virus isolation from Uganda., (Published by Oxford University Press on behalf of Entomological Society of America 2017. This work is written by US Government employees and is in the public domain in the US.)

In mid-2015, Salvador, Brazil, reported an outbreak of Guillain-Barré syndrome (GBS), coinciding with the introduction and spread of Zika virus (ZIKV). We found that GBS incidence during April-July 2015 among those ≥12 years of age was 5.6 cases/100,000 population/year and increased markedly with increasing age to 14.7 among those ≥60 years of age. We conducted interviews with 41 case-patients and 85 neighborhood controls and found no differences in demographics or exposures prior to GBS-symptom onset. A higher proportion of case-patients (83%) compared to controls (21%) reported an antecedent illness (OR 18.1, CI 6.9-47.5), most commonly characterized by rash, headache, fever, and myalgias, within a median of 8 days prior to GBS onset. Our investigation confirmed an outbreak of GBS, particularly in older adults, that was strongly associated with Zika-like illness and geo-temporally associated with ZIKV transmission, suggesting that ZIKV may result in severe neurologic complications.

Understanding the ability of the chikungunya virus (CHIKV) to be transmitted by Aedes vectors in the Americas is critical for assessing epidemiological risk. One element that must be considered is the minimum infectious dose of virus that can lead to transmission following the extrinsic incubation period. This study aimed to determine the minimum infection rate for the two Aedes species studied. The results revealed that doses as low as 3.9 log10 plaque-forming units per mL (pfu/mL) of an Asian genotype CHIKV strain can lead to transmission by Ae. albopictus, and doses of at least 5.3 log10 pfu/mL from the same strain are needed for transmission from Ae. aegypti. These low infecting doses suggest that infected individuals may be infectious for almost the entire period of their viremia, and therefore, to prevent further cases, measures should be taken to prevent them from getting bitten by mosquitoes during this period.

Dengue virus (DENV) and West Nile virus (WNV) are important reemerging arboviruses that are under-recognized in many parts of Africa due to lack of surveillance. As a part of a study on flavivirus, alphavirus, and parasite exposure in coastal Kenya, we measured neutralizing antibody against DENV and, to evaluate assay specificity, WNV in serum samples that tested positive for serum anti-DENV IgG by enzyme-linked immunosorbent assay. Of 830 anti-DENV IgG-positive samples that were tested for neutralizing activity, 488 (58.8%) neutralized DENV and 94 (11.3%) neutralized WNV. Of children ≤ 10 years of age, 23% and 17% had serum neutralizing antibody to DENV and WNV, respectively, indicating that DENV and WNV transmission has occurred in this region within the past decade. The results suggest that ongoing DENV and WNV transmission continues on the coast of Kenya and supports a need for routine arboviral surveillance in the area to detect and respond to future outbreaks., (© The American Society of Tropical Medicine and Hygiene.)

Highlands J virus (HJV) is an alphavirus closely related to western equine encephalitis virus (WEEV) and eastern equine encephalitis virus (EEEV). HJV is an avian pathogen with the potential for disruption of poultry operations, but is not known to cause human or equine disease. HJV has only been identified in the eastern United States and is thought to have a transmission cycle similar to that of EEEV involving Culiseta melanura mosquitoes and birds. However, HJV is more genetically similar to WEEV and it remains unclear if it may be transmitted by Culex species mosquitoes like WEEV. Seven strains of HJV were characterized to assess this potential. Phylogenetic analysis of whole genome sequences revealed four distinct HJV lineages (lineages 1-4), and vector competence studies in Cx. tarsalis with four of the HJV strains from different lineages yielded two distinct infection patterns. Lineage 1 strains had low infection rates, while lineages 2 and 4 had significantly higher infection rates similar to those previously published for WEEV. The average mosquito body viral titer was highest at 8 dpi (6.60-7.26 log10 pfu equivalents/body), and head titers at all time points ranged between 6.01 and 6.80 log10 pfu equivalents/head. Nearly 45% of mosquitoes infected with strain AB-80-9 were able to transmit virus in saliva with an average titer of 5.02 log10 pfu equivalents/saliva. A single amino acid difference between high and low infectivity phenotypes was identified at genome position 8605, in the E2 gene. A nonpolar glycine was present in the low infectivity lineage 1 strains, while an acidic glutamic acid was present in the higher infectivity lineage 2 and 4 strains. This study demonstrates HJV transmission by Cx. tarsalis mosquitoes and clearly identifies the potential for transmission in the western United States. Two infection phenotypes were exhibited, indicating the need for further studies to understand Culex species transmission patterns.

Sunguru virus (SUNV), a novel virus belonging to the highly diverse Rhabdoviridae family, was isolated from a domestic chicken in the district of Arua, Uganda, in 2011. This is the first documented isolation of a rhabdovirus from a chicken. SUNV is related to, but distinct from, Boteke virus and other members of the unclassified Sandjimba group. The genome is 11056 nt in length and contains the five core rhabdovirus genes plus an additional C gene (within the ORF of a phosphoprotein gene) and a small hydrophobic protein (between the matrix and glycoprotein genes). Inoculation of vertebrate cells with SUNV resulted in significant viral growth, with a peak titre of 7.8 log10 p.f.u. ml(-1) observed in baby hamster kidney (BHK) cells. Little to no growth was observed in invertebrate cells and in live mosquitoes, with Anopheles gambiae mosquitoes having a 47.4% infection rate in the body but no dissemination of the virus to the salivary glands; this suggests that this novel virus is not arthropod borne as some other members of the family Rhabdoviridae.

Vector-borne and zoonotic pathogens have comprised a significant proportion of the emerging infectious diseases in humans in recent decades. The role of many wildlife species as reservoirs for arthropod-borne viral pathogens is poorly understood. We investigated the exposure history of various African wildlife species from the Congo basin to mosquito-borne flaviviruses and alphaviruses by testing archived serum samples. Sera from 24 African forest buffalo (Syncerus caffer nanus), 34 African elephants (Loxodonta africana), 40 duikers (Cephalophus and Philantomba spp.), 25 mandrills (Mandrillus sphinx), 32 mountain gorillas (Gorilla beringei beringei), five Grauer's gorillas (Gorilla beringei graueri), two L'Hoest's monkeys (Cercopithecus lhoesti), two golden monkeys (Cercopithecus kandti), and three chimpanzees (Pan troglodytes) sampled between 1991 and 2009 were tested for antibodies against chikungunya virus (CHIKV), o'nyong-nyong virus (ONNV), West Nile virus (WNV), dengue 2 virus (DENV-2), and yellow fever virus (YFV) by plaque reduction neutralization test. Specific neutralizing antibodies against ONNV were found in African forest buffalo in the Democratic Republic of the Congo (DRC) and Gabon, duikers in the DRC, and mandrills in Gabon, providing novel evidence of enzootic circulation of ONNV in these countries. African forest buffalo in the DRC and Gabon also demonstrated evidence of exposure to CHIKV, WNV, and DENV-2, while mandrills in Gabon were antibody positive for CHIKV, DENV-2, WNV, and YFV. All of the elephants tested had a strong neutralizing antibody response to WNV. We also document results from a survey of gorillas for arboviruses, of which 4/32 (13%) had antibody to an alphavirus or flavivirus. Overall, our results demonstrate a high prevalence of neutralizing antibodies against multiple arboviruses in wildlife in equatorial Africa.

Western equine encephalitis virus (WEEV) is a naturally occurring recombinant virus derived from ancestral Sindbis and Eastern equine encephalitis viruses. We previously showed that infection by WEEV isolates McMillan (McM) and IMP-181 (IMP) results in high (∼90-100%) and low (0%) mortality, respectively, in outbred CD-1 mice when virus is delivered by either subcutaneous or aerosol routes. However, relatively little is known about specific virulence determinants of WEEV. We previously observed that IMP infected Culex tarsalis mosquitoes at a high rate (app. 80%) following ingestion of an infected bloodmeal but these mosquitoes were infected by McM at a much lower rate (10%). To understand the viral role in these phenotypic differences, we characterized the pathogenic phenotypes of McM/IMP chimeras. Chimeras encoding the E2 of McM on an IMP backbone (or the reciprocal) had the most significant effect on infection phenotypes in mice or mosquitoes. Furthermore, exchanging the arginine, present on IMP E2 glycoprotein at position 214, for the glutamine present at the same position on McM, ablated mouse mortality. Curiously, the reciprocal exchange did not confer mouse virulence to the IMP virus. Mosquito infectivity was also determined and significantly, one of the important loci was the same as the mouse virulence determinant identified above. Replacing either IMP E2 amino acid 181 or 214 with the corresponding McM amino acid lowered mosquito infection rates to McM-like levels. As with the mouse neurovirulence, reciprocal exchange of amino acids did not confer mosquito infectivity. The identification of WEEV E2 amino acid 214 as necessary for both IMP mosquito infectivity and McM mouse virulence indicates that they are mutually exclusive phenotypes and suggests an explanation for the lack of human or equine WEE cases even in the presence of active transmission.

O'nyong nyong virus (ONNV) and Chikungunya virus (CHIKV) are two closely related alphaviruses with very different infection patterns in the mosquito, Anopheles gambiae. ONNV is the only alphavirus transmitted by anopheline mosquitoes, but specific molecular determinants of infection of this unique vector specificity remain unidentified. Fifteen distinct chimeric viruses were constructed to evaluate both structural and non-structural regions of the genome and infection patterns were determined through artificial infectious feeds in An. gambiae with each of these chimeras. Only one region, non-structural protein 3 (nsP3), was sufficient to up-regulate infection to rates similar to those seen with parental ONNV. When ONNV non-structural protein 3 (nsP3) replaced nsP3 from CHIKV virus in one of the chimeric viruses, infection rates in An. gambiae went from 0% to 63.5%. No other single gene or viral region addition was able to restore infection rates. Thus, we have shown that a non-structural genome element involved in viral replication is a major element involved in ONNV's unique vector specificity.

Emerging mosquito-borne alphavirus infections caused by chikungunya virus (CHIKV) or o'nyong-nyong virus (ONNV) are responsible for sporadic and sometimes explosive urban outbreaks. Currently, there is no licensed vaccine against either virus. We have developed a highly attenuated recombinant CHIKV candidate vaccine (CHIKV/IRES) that in preclinical studies was demonstrated to be safe, immunogenic and efficacious. In this study we investigated the potential of this vaccine to induce cross-protective immunity against the antigenically related ONNV. Our studies demonstrated that a single dose of CHIKV/IRES elicited a strong cross-neutralizing antibody response and conferred protection against ONNV challenge in the A129 mouse model. Moreover, CHIKV/IRES immune A129 dams transferred antibodies to their offspring that were protective, and passively transferred anti-CHIKV/IRES immune serum protected AG129 mice, independently of a functional IFN response. These findings highlight the potential of the CHIKV/IRES vaccine to protect humans against not only CHIKV but also against ONNV-induced disease., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Chikungunya virus (CHIKV) is the mosquito-borne alphavirus that is the etiologic agent of massive outbreaks of arthralgic febrile illness that recently affected millions of people in Africa and Asia. The only CHIKV vaccine that has been tested in humans, strain 181/clone 25, is a live-attenuated derivative of Southeast Asian human isolate strain AF15561. The vaccine was immunogenic in phase I and II clinical trials; however, it induced transient arthralgia in 8% of the vaccinees. There are five amino acid differences between the vaccine and its parent, as well as five synonymous mutations, none of which involves cis-acting genome regions known to be responsible for replication or packaging. To identify the determinants of attenuation, we therefore tested the five nonsynonymous mutations by cloning them individually or in different combinations into infectious clones derived from two wild-type (WT) CHIKV strains, La Reunion and AF15561. Levels of virulence were compared with those of the WT strains and the vaccine strain in two different murine models: infant CD1 and adult A129 mice. An attenuated phenotype indistinguishable from that of the 181/clone 25 vaccine strain was obtained by the simultaneous expression of two E2 glycoprotein substitutions, with intermediate levels of attenuation obtained with the single E2 mutations. The other three amino acid mutations, in nsP1, 6K, and E1, did not have a detectable effect on CHIKV virulence. These results indicate that the attenuation of strain 181/clone 25 is mediated by two point mutations, explaining the phenotypic instability observed in human vaccinees and also in our studies.

Venezuelan equine encephalitis virus (VEEV) has been the causative agent for sporadic epidemics and equine epizootics throughout the Americas since the 1930s. In 1969, an outbreak of Venezuelan equine encephalitis (VEE) spread rapidly from Guatemala and through the Gulf Coast region of Mexico, reaching Texas in 1971. Since this outbreak, there have been very few studies to determine the northward extent of endemic VEEV in this region. This study reports the findings of serologic surveillance in the Gulf Coast region of Mexico from 2003-2010. Phylogenetic analysis was also performed on viral isolates from this region to determine whether there have been substantial genetic changes in VEEV since the 1960s. Based on the findings of this study, the Gulf Coast lineage of subtype IE VEEV continues to actively circulate in this region of Mexico and appears to be responsible for infection of humans and animals throughout this region, including the northern State of Tamaulipas, which borders Texas.

Chikungunya virus (CHIKV) is a reemerging mosquito-borne pathogen that has recently caused devastating urban epidemics of severe and sometimes chronic arthralgia. As with most other mosquito-borne viral diseases, control relies on reducing mosquito populations and their contact with people, which has been ineffective in most locations. Therefore, vaccines remain the best strategy to prevent most vector-borne diseases. Ideally, vaccines for diseases of resource-limited countries should combine low cost and single dose efficacy, yet induce rapid and long-lived immunity with negligible risk of serious adverse reactions. To develop such a vaccine to protect against chikungunya fever, we employed a rational attenuation mechanism that also prevents the infection of mosquito vectors. The internal ribosome entry site (IRES) from encephalomyocarditis virus replaced the subgenomic promoter in a cDNA CHIKV clone, thus altering the levels and host-specific mechanism of structural protein gene expression. Testing in both normal outbred and interferon response-defective mice indicated that the new vaccine candidate is highly attenuated, immunogenic and efficacious after a single dose. Furthermore, it is incapable of replicating in mosquito cells or infecting mosquitoes in vivo. This IRES-based attenuation platform technology may be useful for the predictable attenuation of any alphavirus.

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes explosive outbreaks of febrile illness associated with rash, and painful arthralgia. The CHIK vaccine strain 181/clone25 (181/25) developed by the United States Army Medical Research Institute of Infectious Diseases (USAMRIID) was shown to be well-tolerated and highly immunogenic in phase I and II clinical trials although it induced transient arthralgia in some healthy adult volunteers. In an attempt to better understand the host factors that are involved in the attenuating phenotype of CHIK 181/25 vaccine virus we conducted studies in interferon (IFN)-compromised mice and also evaluated its immunogenic potential and protective capacity. Infection of AG129 mice (defective in IFN-α/β and IFN-γ receptor signaling) with CHIK 181/25 resulted in rapid mortality within 3-4 days. In contrast, all infected A129 mice (defective in IFN-α/β receptor signaling) survived with temporary morbidity characterized by ruffled appearance and body weight loss. A129 heterozygote mice that retain partial IFN-α/β receptor signaling activity remained healthy. Infection of A129 mice with CHIK 181/25 induced significant levels of IFN-γ and IL-12 while the inflammatory cytokines, TNFα and IL-6 remained low. A single administration of the CHIK 181/25 vaccine provided both short-term and long-term protection (38 days and 247 days post-prime, respectively) against challenge with wt CHIKV-La Reunion (CHIKV-LR). This protection was at least partially mediated by antibodies since passively transferred immune serum protected both A129 and AG129 mice from wt CHIKV-LR and 181/25 virus challenge. Overall, these data highlight the importance of IFNs in controlling CHIK 181/25 vaccine and demonstrate the ability of this vaccine to elicit neutralizing antibody responses that confer short-and long-term protection against wt CHIKV-LR challenge., (Copyright © 2011 Elsevier Ltd. All rights reserved.)