De Novo Transcriptome Sequencing in Anopheles funestus Using Illumina RNA-Seq Technology

BACKGROUND: Anopheles funestus is one of the primary vectors of human malaria, which causes a million deaths each year in sub-Saharan Africa. Few scientific resources are available to facilitate studies of this mosquito species and relatively little is known about its basic biology and evolution, making development and implementation of novel disease control efforts more difficult. The An. funestus genome has not been sequenced, so in order to facilitate genome-scale experimental biology, we have sequenced the adult female transcriptome of An. funestus from a newly founded colony in Burkina Faso, West Africa, using the Illumina GAIIx next generation sequencing platform. METHODOLOGY/PRINCIPAL FINDINGS: We assembled short Illumina reads de novo using a novel approach involving iterative de novo assemblies and "target-based" contig clustering. We then selected a conservative set of 15,527 contigs through comparisons to four Dipteran transcriptomes as well as multiple functional and conserved protein domain databases. Comparison to the Anopheles gambiae immune system identified 339 contigs as putative immune genes, thus identifying a large portion of the immune system that can form the basis for subsequent studies of this important malaria vector. We identified 5,434 1:1 orthologues between An. funestus and An. gambiae and found that among these 1:1 orthologues, the protein sequence of those with putative immune function were significantly more diverged than the transcriptome as a whole. Short read alignments to the contig set revealed almost 367,000 genetic polymorphisms segregating in the An. funestus colony and demonstrated the utility of the assembled transcriptome for use in RNA-seq based measurements of gene expression. CONCLUSIONS/SIGNIFICANCE: We developed a pipeline that makes de novo transcriptome sequencing possible in virtually any organism at a very reasonable cost ($6,300 in sequencing costs in our case). We anticipate that our approach could be used to develop genomic resources in a diversity of systems for which full genome sequence is currently unavailable. Our An. funestus contig set and analytical results provide a valuable resource for future studies in this non-model, but epidemiologically critical, vector insect

Transmission-Blocking Vaccines: Focus on Anti-Vector Vaccines against Tick-Borne Diseases

Tick-borne diseases are a potential threat that account for significant morbidity and mortality in human population worldwide. Vaccines are not available to treat several of the tick-borne diseases. With the emergence and resurgence of several tick-borne diseases, emphasis on the development of transmission-blocking vaccines remains increasing. In this review, we provide a snap shot on some of the potential candidates for the development of anti-vector vaccines (a form of transmission-blocking vaccines) against wide range of hard and soft ticks that include Ixodes, Haemaphysalis, Dermacentor, Amblyomma, Rhipicephalus and Ornithodoros species

Ebola: translational science considerations

We are currently in the midst of the most aggressive and fulminating outbreak of Ebola-related disease, commonly referred to as “Ebola”, ever recorded. In less than a year, the Ebola virus (EBOV, Zaire ebolavirus species) has infected over 10,000 people, indiscriminately of gender or age, with a fatality rate of about 50%. Whereas at its onset this Ebola outbreak was limited to three countries in West Africa (Guinea, where it was first reported in late March 2014, Liberia, where it has been most rampant in its capital city, Monrovia and other metropolitan cities, and Sierra Leone), cases were later reported in Nigeria, Mali and Senegal, as well as in Western Europe (i.e., Madrid, Spain) and the US (i.e., Dallas, Texas; New York City) by late October 2014. World and US health agencies declared that the current Ebola virus disease (EVD) outbreak has a strong likelihood of growing exponentially across the world before an effective vaccine, treatment or cure can be developed, tested, validated and distributed widely. In the meantime, the spread of the disease may rapidly evolve from an epidemics to a full-blown pandemic. The scientific and healthcare communities actively research and define an emerging kaleidoscope of knowledge about critical translational research parameters, including the virology of EBOV, the molecular biomarkers of the pathological manifestations of EVD, putative central nervous system involvement in EVD, and the cellular immune surveillance to EBOV, patient-centered anthropological and societal parameters of EVD, as well as translational effectiveness about novel putative patient-targeted vaccine and pharmaceutical interventions, which hold strong promise, if not hope, to curb this and future Ebola outbreaks. This work reviews and discusses the principal known facts about EBOV and EVD, and certain among the most interesting ongoing or future avenues of research in the field, including vaccination programs for the wild animal vectors of the virus and the disease from global translational science perspective

Resting and feeding preferences of Anopheles stephensi in an urban setting, perennial for malaria

Background: The Indian city of Chennai is endemic for malaria and the known local malaria vector is Anopheles stephensi. Plasmodium vivax is the predominant malaria parasite species, though Plasmodium falciparum is present at low levels. The urban ecotype of malaria prevails in Chennai with perennial transmission despite vector surveillance by the Urban Malaria Scheme (UMS) of the National Vector Borne Disease Control Programme (NVBDCP). Understanding the feeding and resting preferences, together with the transmission potential of adult vectors in the area is essential in effective planning and execution of improved vector control measures. Methods: A yearlong survey was carried out in cattle sheds and human dwellings to check the resting, feeding preferences and transmission potential of An. stephensi. The gonotrophic status, age structure, resting and host seeking preferences were studied. The infection rate in An. stephensi and Anopheles subpictus were analysed by circumsporozoite ELISA (CS-ELISA). Results: Adult vectors were found more frequently and at higher densities in cattle sheds than human dwellings. The overall Human Blood Index (HBI) was 0.009 indicating the vectors to be strongly zoophilic. Among the vectors collected from human dwellings, 94.2% were from thatched structures and the remaining 5.8% from tiled and asbestos structures. 57.75% of the dissected vectors were nulliparous whereas, 35.83% were monoparous and the rest 6.42% biparous. Sporozoite positivity rate was 0.55% (4/720) and 1.92% (1/52) for An. stephensi collected from cattle sheds and human dwellings, respectively. One adult An. subpictus (1/155) was also found to be infected with P. falciparum. Conclusions: Control of the adult vector populations can be successful only by understanding the resting and feeding preferences. The present study indicates that adult vectors predominantly feed on cattle and cattle sheds are the preferred resting place, possibly due to easy availability of blood meal source and lack of any insecticide or repellent pressure. Hence targeting these resting sites with cost effective, socially acceptable intervention tools, together with effective larval source management to reduce vector breeding, could provide an improved integrated vector management strategy to help drive down malaria transmission and assist in India's plan to eliminate malaria by 2030

Existing Infection Facilitates Establishment and Density of Malaria Parasites in Their Mosquito Vector

Very little is known about how vector-borne pathogens interact within their vector and how this impacts transmission. Here we show that mosquitoes can accumulate mixed strain malaria infections after feeding on multiple hosts. We found that parasites have a greater chance of establishing and reach higher densities if another strain is already present in a mosquito. Mixed infections contained more parasites but these larger populations did not have a detectable impact on vector survival. Together these results suggest that mosquitoes taking multiple infective bites may disproportionally contribute to malaria transmission. This will increase rates of mixed infections in vertebrate hosts, with implications for the evolution of parasite virulence and the spread of drug-resistant strains. Moreover, control measures that reduce parasite prevalence in vertebrate hosts will reduce the likelihood of mosquitoes taking multiple infective feeds, and thus disproportionally reduce transmission. More generally, our study shows that the types of strain interactions detected in vertebrate hosts cannot necessarily be extrapolated to vectors