A compendium of molecules involved in vector-pathogen interactions pertaining to malaria

Malaria is a vector-borne disease causing extensive morbidity, debility and mortality. Development of resistance to drugs among parasites and to conventional insecticides among vector-mosquitoes necessitates innovative measures to combat this disease. Identification of molecules involved in the maintenance of complex developmental cycles of the parasites within the vector and the host can provide attractive targets to intervene in the disease transmission. In the last decade, several efforts have been made in identifying such molecules involved in mosquito-parasite interactions and, subsequently, validating their role in the development of parasites within the vector. In this study, a list of mosquito proteins, which facilitate or inhibit the development of malaria parasites in the midgut, haemolymph and salivary glands of mosquitoes, is compiled. A total of 94 molecules have been reported and validated for their role in the development of malaria parasites inside the vector. This compendium of molecules will serve as a centralized resource to biomedical researchers investigating vector-pathogen interactions and malaria transmission. © 2013 Sreenivasamurthy et al.; licensee BioMed Central Ltd

Integrating transcriptomic and proteomic data for accurate assembly and annotation of genomes

© 2017 Wong et al.; Published by Cold Spring Harbor Laboratory Press. Complementing genome sequence with deep transcriptome and proteome data could enable more accurate assembly and annotation of newly sequenced genomes. Here, we provide a proof-of-concept of an integrated approach for analysis of the genome and proteome of Anopheles stephensi, which is one of the most important vectors of the malaria parasite. To achieve broad coverage of genes, we carried out transcriptome sequencing and deep proteome profiling of multiple anatomically distinct sites. Based on transcriptomic data alone, we identified and corrected 535 events of incomplete genome assembly involving 1196 scaffolds and 868 protein-coding gene models. This proteogenomic approach enabled us to add 365 genes that were missed during genome annotation and identify 917 gene correction events through discovery of 151 novel exons, 297 protein extensions, 231 exon extensions, 192 novel protein start sites, 19 novel translational frames, 28 events of joining of exons, and 76 events of joining of adjacent genes as a single gene. Incorporation of proteomic evidence allowed us to change the designation of more than 87 predicted noncoding RNAs to conventional mRNAs coded by protein-coding genes. Importantly, extension of the newly corrected genome assemblies and gene models to 15 other newly assembled Anopheline genomes led to the discovery of a large number of apparent discrepancies in assembly and annotation of these genomes. Our data provide a framework for how future genome sequencing efforts should incorporate transcriptomic and proteomic analysis in combination with simultaneous manual curation to achieve near complete assembly and accurate annotation of genomes

Dataset on fat body proteome of Anopheles stephensi Liston

Fat body from Anopheles stephensi female mosquitoes were dissected and processed for proteomic analysis. Both SDS-PAGE and basic Reverse Phase Liquid Chromatography-based fractionation strategies were used to achieve a broad coverage of protein identification. The fractionated peptides were then analyzed on a high-resolution mass spectrometer. Searching the raw data against the protein database of An. stephensi resulted in identification of 4535 proteins, which is, to our knowledge, the largest catalog of fat body proteome in any mosquito vector species reported so far. Bioinformatics analysis on these fat body proteins suggested the enrichment of biological processes including carbon and lipid metabolism, amino acid metabolism, signal peptide processing and oxidation-reduction. In addition, using proteogenomic approaches, 43 novel proteins were identified, which were not listed in the annotated gene annotations of An. stephensi. The data used in the analysis are related to the article ‘Integrating transcriptomic and proteomic data for accurate assembly and annotation of genomes’ (Prasad et al., 2017)

Proteome data of Anopheles stephensi hemolymph using high resolution mass spectrometry

The article provides insights into the protein expression in Anopheles stephensi hemolymph. We carried out data acquisition using a high-resolution LTQ-Orbitrap Velos mass spectrometer to identify the hemolymph proteins of An. stephensi. Experimentally derived mass spectrometry data was analyzed using Proteome Discoverer 2.1 software using two different search algorithms SEQUEST and MASCOT. A total of 1091 proteins were identified from the hemolymph. The identified proteins were categorized for their role in biological processes and molecular functions. The interactions between these proteins were predicted using STRING online tool. Relation can be drawn between the data provided in this study to the already published article “Integrating transcriptomics and proteomics data for accurate assembly and annotation of genomes” (Prasad et al., 2017) [1]

Data from salivary gland proteome analysis of female Aedes aegypti Linn

Salivary gland proteins from female Aedes aegypti mosquito were extracted and analyzed on high-resolution mass spectrometry. Proteomic data was analysed using two search algorithms SEQUEST and Mascot, which results in acquisition of 83,836 spectra which were assigned to 5417 peptides belonging to 1208 proteins.These proteins were then assigned molecular functions and further analysis revealed biological processes they are involved in using Gene Ontology annotations. Several immunity related pathways were found to be enriched in salivary gland.The data of this study are also related to the research article “Mosquito-Borne Diseases and Omics: Salivary gland proteome of the female Aedes aegypti mosquito” (Dhawan et al., 2017) [1]. These data are deposited in ProteomeXchange in the public dataset PXD002468. In addition,a scientific interpretation of this dataset by Dhawan et al. [1] is available at http://dx.doi.org/10.1089/journal.omi.2016.0160. Keywords: Proteomics, Salivary gland, Immunogenic, Vectorborne Diseases, Aedes aegypti, Protein–protein interaction

Mapping Anopheles stephensi midgut proteome using high-resolution mass spectrometry

Anopheles stephensi Liston is one of the major vectors of malaria in urban areas of India. Midgut plays a central role in the vector life cycle and transmission of malaria. Because gene expression of An. stephensi midgut has not been investigated at protein level, an unbiased mass spectrometry-based proteomic analysis of midgut tissue was carried out. Midgut tissue proteins from female An. stephensi mosquitoes were extracted using 0.5% SDS and digested with trypsin using two complementary approaches, in-gel and in-solution digestion. Fractions were analysed on high-resolution mass spectrometer, which resulted in acquisition of 494,960 MS/MS spectra. The MS/MS spectra were searched against protein database comprising of known and predicted proteins reported in An. stephensi using Sequest and Mascot software. In all, 47,438 peptides were identified corresponding to 5,709 An. stephensi proteins. The identified proteins were functionally categorized based on their cellular localization, biological processes and molecular functions using Gene Ontology (GO) annotation. Several proteins identified in this data are known to mediate the interaction of the Plasmodium with vector midgut and also regulate parasite maturation inside the vector host. This study provides information about the protein composition in midgut tissue of female An. stephensi, which would be useful in understanding vector parasite interaction at molecular level and besides being useful in devising malaria transmission blocking strategies. The data of this study is related to the research article “Integrating transcriptomics and proteomics data for accurate assembly and annotation of genomes”

Proteome data of Anopheles stephensi ovary using high-resolution mass spectrometry

This article contains data on the proteins expressed in the ovaries of Anopheles stephensi, a major vector of malaria in India. Data acquisition was performed using a high-resolution Orbitrap-Velos mass spectrometer. The acquired MS/MS data was searched against An. stephensi protein database comprising of 11,789 sequences. Overall, 4407 proteins were identified, functional analysis was performed for the identified proteins and a protein-protein interaction map predicted. The data provided here is also related to a published article - “Integrating transcriptomics and proteomics data for accurate assembly and annotation of genomes” (Prasad et al., 2017) [1]

Mapping malaria vectors and insecticide resistance in a high-endemic district of Haryana, India: implications for vector control strategies

Abstract Background Achieving effective control and elimination of malaria in endemic regions necessitates a comprehensive understanding of local mosquito species responsible for malaria transmission and their susceptibility to insecticides. Methods The study was conducted in the highly malaria prone Ujina Primary Health Center of Nuh (Mewat) district of Haryana state of India. Monthly entomological surveys were carried out for adult mosquito collections via indoor resting collections, light trap collections, and pyrethrum spray collections. Larvae were also collected from different breeding sites prevalent in the region. Insecticide resistance bioassay, vector incrimination, blood meal analysis was done with the collected vector mosquitoes. Results A total of 34,974 adult Anopheles mosquitoes were caught during the survey period, out of which Anopheles subpictus was predominant (54.7%). Among vectors, Anopheles stephensi was predominant (15.5%) followed by Anopheles culicifacies (10.1%). The Human Blood Index (HBI) in the case of An. culicifacies and An. stephensi was 6.66 and 9.09, respectively. Vector incrimination results revealed Plasmodium vivax positivity rate of 1.6% for An. culicifacies. Both the vector species were found resistant to DDT, malathion and deltamethrin. Conclusion The emergence of insecticide resistance in both vector species, compromises the effectiveness of commonly used public health insecticides. Consequently, the implementation of robust insecticide resistance management strategies becomes imperative. To effectively tackle the malaria transmission, a significant shift in vector control strategies is warranted, with careful consideration and adaptation to address specific challenges encountered in malaria elimination efforts

Susceptibility of wild and colonized Anopheles stephensi to Plasmodium vivax infection

Abstract Background As much as 80% of global Plasmodium vivax infections occur in South Asia and there is a shortage of direct studies on infectivity of P. vivax in Anopheles stephensi, the most common urban mosquito carrying human malaria. In this quest, the possible effects of laboratory colonization of mosquitoes on infectivity and development of P. vivax is of interest given that colonized mosquitoes can be genetically less divergent than the field population from which they originated. Methods Patient-derived P. vivax infected blood was fed to age-matched wild and colonized An. stephensi. Such a comparison requires coordinated availability of same-age wild and colonized mosquito populations. Here, P. vivax infection are studied in colonized An. stephensi in their 66th–86th generation and fresh field-caught An. stephensi. Wild mosquitoes were caught as larvae and pupae and allowed to develop into adult mosquitoes in the insectary. Parasite development to oocyst and sporozoite stages were assessed on days 7/8 and 12/13, respectively. Results While there were batch to batch variations in infectivity of individual patient-derived P. vivax samples, both wild and colonized An. stephensi were roughly equally susceptible to oocyst stage Plasmodium infection. At the level of sporozoite development, significantly more mosquitoes with sporozoite load of 4+ were seen in wild than in colonized populations. Conclusions Overall at the level of oocyst development, significant difference was found between the colonized and wild Anopheles stephensi in their susceptibility to P. vivax. For initial understanding of infections with local strains of P. vivax, colonized Anopheles stephensi will serve as a good model. For experiments, where high number of sporozoites are necessary, wild mosquitoes provide distinct advantage over the colonized vector populations. Understanding the molecular mechanism modulating this variability between these two populations will be prime area of focus in future studies

Dynamics of Plasmodium vivax sporogony in wild Anopheles stephensi in a malaria-endemic region of Western India

Abstract Background In global efforts to track mosquito infectivity and parasite elimination, controlled mosquito-feeding experiments can help in understanding the dynamics of parasite development in vectors. Anopheles stephensi is often accepted as the major urban malaria vector that transmits Plasmodium in Goa and elsewhere in South Asia. However, much needs to be learned about the interactions of Plasmodium vivax with An. stephensi. As a component of the US NIH International Center of Excellence for Malaria Research (ICEMR) for Malaria Evolution in South Asia (MESA), a series of membrane-feeding experiments with wild An. stephensi and P. vivax were carried out to better understand this vector-parasite interaction. Methods Wild An. stephensi larvae and pupae were collected from curing water in construction sites in the city of Ponda, Goa, India. The larvae and pupae were reared at the MESA ICEMR insectary within the National Institute of Malaria Research (NIMR) field unit in Goa until they emerged into adult mosquitoes. Blood for membrane-feeding experiments was obtained from malaria patients at the local Goa Medical College and Hospital who volunteered for the study. Parasites were counted by Miller reticule technique and correlation between gametocytaemia/parasitaemia and successful mosquito infection was studied. Results A weak but significant correlation was found between patient blood gametocytaemia/parasitaemia and mosquito oocyst load. No correlation was observed between gametocytaemia/parasitaemia and oocyst infection rates, and between gametocyte sex ratio and oocyst load. When it came to development of the parasite in the mosquito, a strong positive correlation was observed between oocyst midgut levels and sporozoite infection rates, and between oocyst levels and salivary gland sporozoite loads. Kinetic studies showed that sporozoites appeared in the salivary gland as early as day 7, post-infection. Conclusions This is the first study in India to carry out membrane-feeding experiments with wild An. stephensi and P. vivax. A wide range of mosquito infection loads and infection rates were observed, pointing to a strong interplay between parasite, vector and human factors. Most of the present observations are in agreement with feeding experiments conducted with P. vivax elsewhere in the world