Mosquito and Arbovirus Activity

Mosquitoes are public health concerns as vectors of arthropod-borne viruses (arboviruses) and/or as nuisances to humans, so surveillance efforts are important to determine areas and times that may pose a risk. The Horticulture Research Station (HRS), Ames, Iowa, consistently yields mosquitoes that are positive for West Nile virus (WNV). Objectives were to continue to assess human risk by monitoring both mosquitoes and sentinel chickens, which serve as vertebrate hosts for arbovirus

In Vivo Titration and Development of a Challenge Model for White Spot Syndrome Virus (WSSV) in Pacific White Shrimp (Litopeneus vannamei)

A challenge model was developed using a controlled bioassay system for estimation of lethal infective doses (LD 50) of White Spot Syndrome Virus (WSSV), as a model system to be used in further WSSV studies

Landscape, demographic, entomological, and climatic associations with human disease incidence of West Nile virus in the state of Iowa, USA

<p>Abstract</p> <p>Background</p> <p>West Nile virus (WNV) emerged as a threat to public and veterinary health in the Midwest United States in 2001 and continues to cause significant morbidity and mortality annually. To investigate biotic and abiotic factors associated with disease incidence, cases of reported human disease caused by West Nile virus (WNV) in the state of Iowa were aggregated by census block groups in Iowa for the years 2002–2006. Spatially explicit data on landscape, demographic, and climatic conditions were collated and analyzed by census block groups. Statistical tests of differences between means and distributions of landscape, demographic, and climatic variables for census block groups with and without WNV disease incidence were carried out. Entomological data from Iowa were considered at the state level to add context to the potential ecological events taking place.</p> <p>Results</p> <p>Numerous statistically significant differences were shown in the means and distributions of various landscape and demographic variables for census block groups with and without WNV disease incidence. Census block groups with WNV disease incidence had significantly lower population densities than those without. Landscape variables showing differences included stream density, road density, land cover compositions, presence of irrigation, and presence of animal feeding operations. Statistically significant differences in the annual means of precipitations, dew point, and minimum temperature for both the year of WNV disease incidence and the prior year, were detected in at least one year of the analysis for each parameter. However, the differences were not consistent between years.</p> <p>Conclusion</p> <p>The analysis of human WNV disease incidence by census block groups in Iowa demonstrated unique landscape, demographic, and climatic associations. Our results indicate that multiple ecological WNV transmission dynamics are most likely taking place in Iowa. In 2003 and 2006, drier conditions were associated with WNV disease incidence. In a significant novel finding, rural agricultural settings were shown to be strongly associated with human WNV disease incidence in Iowa.</p

Biodistribution of degradable polyanhydride particles in Aedes aegypti tissues

Insecticide resistance poses a significant threat to the control of arthropods that transmit disease agents. Nanoparticle carriers offer exciting opportunities to expand the armamentarium of insecticides available for public health and other pests. Most chemical insecticides are delivered by contact or feeding, and from there must penetrate various biological membranes to reach target organs and kill the pest organism. Nanoparticles have been shown to improve bioactive compound navigation of such barriers in vertebrates, but have not been well-explored in arthropods. In this study, we explored the potential of polyanhydride micro- and nanoparticles (250 nm– 3 μm), labeled with rhodamine B to associate with and/or transit across insect biological barriers, including the cuticle, epithelium, midgut and ovaries, in female Ae. aeygpti mosquitoes. Mosquitoes were exposed using conditions to mimic surface contact with a residual spray or paint, topical exposure to mimic contact with aerosolized insecticide, or per os in a sugar meal. In surface contact experiments, microparticles were sometimes observed in association with the exterior of the insect cuticle. Nanoparticles were more uniformly distributed across exterior tissues and present at higher concentrations. Furthermore, by surface contact, topical exposure, or per os, particles were detected in internal organs. In every experiment, amphiphilic polyanhydride nanoparticles associated with internal tissues to a higher degree than hydrophobic nanoparticles. In vitro, nanoparticles associated with Aedes aegypti Aag2 cells within two hours of exposure, and particles were evident in the cytoplasm. Further studies demonstrated that particle uptake is dependent on caveolae-mediated endocytosis. The propensity of these nanoparticles to cross biological barriers including the cuticle, to localize in target tissue sites of interest, and to reach the cytoplasm of cells, provides great promise for targeted delivery of insecticidal candidates that cannot otherwise reach these cellular and subcellular locations

Characterization of Newly Revealed Sequences in the Infectious Myonecrosis Virus Genome in \u3ci\u3eLitopenaeus vannamei\u3c/i\u3e

Infectious myonecrosis virus (IMNV) causes significant economic losses in farmed shrimp, where associated mortality in ponds can reach 70%. To explore host/pathogen interactions, a next-generation sequencing approach using lymphoid organ tissue from IMNV-infected Litopenaeus vannamei shrimp was conducted. Preliminary sequence assembly of just the virus showed that there were at least an additional 639 bp at the 5′ terminus and 23 nt at the 3′ terminus as compared with the original description of the IMNV genome (7561 nt). Northern blot and reverse transcription-PCR analysis confirmed the presence of novel sequence at both ends of the genome. Using 5′ RACE, an additional 4 nt were discovered; 3′ RACE confirmed the presence of 22 bp rather than 23 bp of sequence. Based on these data, the IMNV genome is 8226 bp in length. dsRNA was used to trigger RNA interference (RNAi) and suppress expression of the newly revealed genome sections at the 5′ end of the IMNV genome in IMNV-infected L. vannamei. An RNAi trigger targeting a 376 bp length of the 5′ UTR did not improve survival of infected shrimp. In contrast, an RNAi trigger targeting a 381 bp sequence in ORF1 improved survival to 82.2% as compared with 2.2% survival in positive control animals. These studies revealed the importance of the new genome sections to produce high-titre infection, and associated disease and mortality, in infected shrimp

Sequence-optimized and targeted double-stranded RNA as a therapeutic antiviral treatment against infectious myonecrosis virus in \u3ci\u3eLitopenaeus vannamei\u3c/i\u3e

Infectious myonecrosis virus (IMNV) is a significant and emerging pathogen that has a tremendous impact on the culture of the Pacific white shrimp Litopenaeus vannamei. IMNV first emerged in Brazil in 2002 and subsequently spread to Indonesia, causing large economic losses in both countries. No existing therapeutic treatments or effective interventions currently exist for IMNV. RNA interference (RNAi) is an effective technique for preventing viral disease in shrimp. Here, we describe the efficacy of a double-stranded RNA (dsRNA) applied as an antiviral therapeutic following virus challenge. The antiviral molecule is an optimized dsRNA construct that targets an IMNV sequence at the 5’ end of the genome and that showed outstanding antiviral protection previously when administered prior to infection. At least 50% survival is observed with a low dose of dsRNA administered 48 h post-infection with a lethal dose of IMNV; this degree of protection was not observed when dsRNA was administered 72 h post-infection. Additionally, administration of the dsRNA antiviral resulted in a significant reduction of the viral load in the muscle of shrimp that died from disease or survived until termination of the present study, as assessed by quantitative RT-PCR. These data indicate that this optimized RNAi antiviral molecule holds promise for use as an antiviral therapeutic against IMNV

RNA Nanovaccine Protects against White Spot Syndrome Virus in Shrimp

In the last 15 years, crustacean fisheries have experienced billions of dollars in economic losses, primarily due to viral diseases caused by such pathogens as white spot syndrome virus (WSSV) in the Pacific white shrimp Litopenaeus vannamei and Asian tiger shrimp Penaeus monodon. To date, no effective measures are available to prevent or control disease outbreaks in these animals, despite their economic importance. Recently, double-stranded RNA-based vaccines have been shown to provide specific and robust protection against WSSV infection in cultured shrimp. However, the limited stability of double-stranded RNA is the most significant hurdle for the field application of these vaccines with respect to delivery within an aquatic system. Polyanhydride nanoparticles have been successfully used for the encapsulation and release of vaccine antigens. We have developed a double-stranded RNA-based nanovaccine for use in shrimp disease control with emphasis on the Pacific white shrimp L. vannamei. Nanoparticles based on copolymers of sebacic acid, 1,6-bis(pcarboxyphenoxy) hexane, and 1,8-bis(p-carboxyphenoxy)-3,6-dioxaoctane exhibited excellent safety profiles, as measured by shrimp survival and histological evaluation. Furthermore, the nanoparticles localized to tissue target replication sites for WSSV and persisted through 28 days postadministration. Finally, the nanovaccine provided ~80% protection in a lethal WSSV challenge model. This study demonstrates the exciting potential of a safe, effective, and field-applicable RNA nanovaccine that can be rationally designed against infectious diseases affecting aquaculture

Universal primers for the amplification and sequence analysis pf actin-1 from diverse mosquito species

We report the development of universal primers for the reverse-transcription polymerase chain reaction (RT-PCR) amplification and nucleotide sequence analysis of actin cDNAs from taxonomically diverse mosquito species. Primers specific to conserved regions of the invertebrate actin-1 gene were designed after actin cDNA sequences of Anopheles gambiae, Bombyx mori, Drosophila melanogaster, and Caenorhabditis elegans. The efficacy of these primers was determined by RT-PCR with the use of total RNA from mosquitoes belonging to 30 species and 8 genera (Aedes, Anopheles, Culex, Deinocerites, Mansonia, Psorophora, Toxorhynchites, and Wyeomyia). The RT-PCR products were sequenced, and sequence data were used to design additional primers. One primer pair, denoted as Act-2F (5′-ATGGTCGGYATGGGNCAGAAGGACTC-3′) and Act-8R (5′-GATTCCATACCCAGGAAG-GADGG-3′), successfully amplified an RT-PCR product of the expected size (683-nt) in all mosquito spp. tested. We propose that this primer pair can be used as an internal control to test the quality of RNA from mosquitoes collected in vector surveillance studies. These primers can also be used in molecular experiments in which the detection, amplification or silencing of a ubiquitously expressed mosquito housekeeping gene is necessary. Sequence and phylogenetic data are also presented in this report

Diversity of Anaplasma and novel Bartonella species in Lipoptena fortisetosa collected from captive Eld’s deer in Thailand

Lipoptena insects are important ectoparasites of cervids and may affect humans that are incidentally bitten. The presence of zoonotic pathogen DNA, such as Anaplasma, and Bartonella, raises the importance of Lipoptena insects in veterinary and human medicine. Eld’s deer (Rucervus eldii thamin), an endangered wild ruminant in Thailand, are bred and raised in the open zoo. The semi-wild zoo environment suggests ectoparasite infestation and potential risk for mechanical transmission of pathogens to visitors, zoo workers, or other animals. However, epidemiology knowledge of pathogens related to endangered wild ruminants in Thailand is limited. This study aims to determine the prevalence and diversity of Anaplasma and Bartonella in the L. fortisetosa collected from captive Eld’s deer in Chon Buri, Thailand. Of the 91 Lipoptena DNA samples obtained, 42 (46.15%) and 25 (27.47%) were positive for Anaplasma and Bartonella by molecular detection, respectively. Further, 42 sequences of Anaplasma (4 nucleotide sequence types) showed 100% identity to those detected in other ruminants and blood-sucking ectoparasites. Twenty-five sequences of Bartonella (8 nucleotide sequence types) showed 97.35–99.11% identity to the novel Bartonella species from sika deer and keds in Japan. Phylogenetic trees revealed Anaplasma sequences were grouped with the clusters of A. bovis and other ruminant-related Anaplasma, while Bartonella sequences were clustered with the novel Bartonella species lineages C, D, and E, which originated from Japan. Interestingly, a new independent lineage of novel Bartonella species was found in obtained specimens. We report the first molecular detection of Anaplasma and Bartonella on L. fortisetosa, which could represent infectious status of captive Eld’s deer in the zoo. Wild animals act as reservoirs for many pathogens, thus preventive measures in surrounding areas should be considered to prevent pathogen infection among animals or potential zoonotic infection among humans