In Silico Characterization of Intracellular Localization Signals and Structural Features of Mosquito Densovirus (MDV) Viral Proteins.

As entomopathogenic viruses, mosquito densoviruses (MDVs) are widely studied for their potential as biocontrol agents and molecular laboratory tools for mosquito manipulation. The nucleus of the mosquito cell is the site for MDV genome replication and capsid assembly, however the nuclear localization signals (NLSs) and nuclear export signals (NES) for MDV proteins have not yet been identified. We carried out an in silico analysis to identify putative NLSs and NESs in the viral proteins of densoviruses that infect diverse mosquito genera (Aedes, Anopheles, and Culex) and identified putative phosphorylation and glycosylation sites on these proteins. These analyses lead to a more comprehensive understanding of how MDVs are transported into and out of the nucleus and lay the foundation for the potential use of densoviruses in mosquito control and basic research.

Community Perception Regarding Mosquito-Borne Diseases in Some Selected Areas of Ganjam District of Odisha State, India.

Mosquito-borne diseases are a major public health concern. Because of absence of effective therapy and vaccination to treat and prevent such vector-borne diseases, these initial cases have triggered strong concerns within medical communities, turning a bad dream into reality. To date, reducing the mosquito vector population remains the main strategy to control the transmission of mosquito borne diseases. In this connection, community-based perception recognized as an essential tool for long-term vector control management. In this study, a community perception survey was carried out in six selected study areas of Ganjam district of Odisha. A total of 3257 inhabitants from randomly chosen 600 no. of houses from 06 selected study areas were interviewed using a pre-tested structured questionnaire about various mosquito vector-borne diseases. As an outcome, we found that only 36% of people have basic knowledge and idea about vector-borne diseases, out of which only 14% know Anopheles as a causative agent whereas Aedes and Culex were very least concern. Regarding the source of knowledge, electronic media was the principal source of information, while the involvements of health personnel were least mentioned. Results of the current study indicates the necessity to plan health education program to bring important perceptions regarding vector-borne diseases at school and college level because the infrequent source of knowledge is the main cause of an increase in the disease incidence.

SNP barcoding based on decision tree algorithm: A new tool for identification of mosquito species with special reference to Anopheles.

Molecular taxonomy based identification of species in the form of DNA barcodes are extensively used in evolutionary systematics. Almost all the DNA barcodes contain detailed information of the barcoding gene along with uninformative sequences of a particular species. Therefore, a technique is highly essential to remove or to reduce the number of uninformative sequences and ought to create species-specific barcodes for differentiation. The actual variation in genetic sequences, called single nucleotide polymorphism (SNP) genotyping, can be utilized to develop a new tool for rapid, reliable, and high-throughput assay to distinguish the known species. SNPs act as important hereditary markers for uncovering the evolutionary history and normal genetic polymorphisms. Keeping in mind, we propose a decision tree-based barcoding (DTB) algorithm for generating SNP barcodes from the DNA barcoding sequence of several evolutionarily related species to accurately identify a single species. To address this issue, we analyzed mitochondrial COI gene sequences of 64 species of Anopheles mosquitoes. After alignment and truncating, 32 SNPs were discovered in COI gene sequences of Anopheles mosquitoes and then computed to set up the decision rule for constructing the decision tree. The decision tree based barcoding algorithm generates 126 nodes and 32 loci for discriminating 64 Anopheles mosquito species. Finally, we concluded that the DTB method is useful and effective for generating sequence tags for Anopheles mosquito species identification.

Morphological and genetic analyses of the first record of longrakered trevally, Ulua mentalis (Perciformes: Carangidae) and of the pinjalo snapper, Pinjalo pinjalo (Perciformes: Lutjanidae) in the Odisha coast, Bay of Bengal.

Identification of fish species have so far been carried out mostly by classical morpho-taxonomy. In the present study, however, an attempt has been taken to identify two species of fishes Ulua mentalis and Pinjalo pinjalo of order Perciformes which happens to be the first record in Odisha coast Bay of Bengal, India during the year 2015, using DNA barcoding technique for reconfirmation over conventional morpho-taxonomy. During recent past, study of molecular-taxonomical profile of mitochondrial DNA in general and Cytochrome Oxidase subunit I (COI) gene in particular has gained enormous importance for accurate identification of species. In the present study, the partial COI sequence of Ulua mentalis and Pinjalo pinjalo were generated. Analysis using the COI gene produced phylogenetic trees in concurrence with other multi gene studies and we came across the identical phylogenetic relationship considering Neighbor-Joining and Maximum Likelihood tree. Moreover, these molecular data set further testified in Bayesian framework to reevaluate the exact taxonomic groupings within the family. Surprisingly, Ulua mentalis and Pinjalo pinjalo seems to be closely related to their sister taxa.

Factors influencing infection and transmission of Anopheles gambiae densovirus (AgDNV) in mosquitoes.

Anopheles gambiae densovirus (AgDNV) is a potential microbial agent for paratransgenesis and gene transduction in An. gambiae, the major vector of human malaria in sub-Saharan Africa. Understanding the interaction between AgDNV and An. gambiae is critical for using AgDNV in a basic and applied manner for Anopheles gene manipulation. Here, we tested the effects of mosquito age, sex, blood feeding status, and potential for horizontal transmission using an enhanced green fluorescent protein (EGFP) reporter AgDNV system. Neither mosquito age at infection nor feeding regime affected viral titers. Female mosquitoes were more permissive to viral infection than males. Despite low viral titers, infected males were able to venereally transmit virus to females during mating, where the virus was localized with the transferred sperm in the spermathecae. These findings will be useful for designing AgDNV-based strategies to manipulate Anopheles gambiae.

In vitro and in vivo host range of Anopheles gambiae densovirus (AgDNV).

AgDNV is a powerful gene transduction tool and potential biological control agent for Anopheles mosquitoes. Using a GFP reporter virus system, we investigated AgDNV host range specificity in four arthropod cell lines (derived from An. gambiae, Aedes albopictus and Drosophila melanogaster) and six mosquito species from 3 genera (An. gambiae, An. arabiensis, An. stephensi, Ae. albopictus, Ae. aegypti and Culex tarsalis). In vitro, efficient viral invasion, replication and GFP expression was only observed in MOS55 An. gambiae cells. In vivo, high levels of GFP were observed in An. gambiae mosquitoes. Intermediate levels of GFP were observed in the closely related species An. arabiensis. Low levels of GFP were observed in An. stephensi, Ae. albopictus, Ae. aegypti and Cx. tarsalis. These results suggest that AgDNV is a specific gene transduction tool for members of the An. gambiae species complex, and could be potentially developed into a biocontrol agent with minimal off-target effects.

Cycloxygenase-2 (COX-2)--a potential target for screening of small molecules as radiation countermeasure agents: an in silico study.

Cyclooxygenase-2 (COX-2) is well established for its role in inflammation, cancer and has also been reported to play a significant role in radiation induced inflammation and bystander effect. It has already been reported to have a role in protection against radiation induced damage, suggesting it to be an important target for identifying novel radiation countermeasure agents. Present study aims at identifying novel small molecules from pharmacopeia using COX-2 as target in silico. Systematic search of the molecules that are reported to exhibit radiation protection revealed that around 30% (40 in 130) of them have a role in inflammation and a small percentage of these molecules (20%; 8 in 40) are reported to act as non-steroidal anti-inflammatory drugs (NSAIDS). Docking studies further clarified that antiinflammatory compounds exhibited higher binding energy (BE). Out of 15 top hits, 14 molecules are reported to have anti-inflammatory property, suggesting the significant role of COX-2 in radiation protection. Further, Johns Hopkins Clinical Compound Library (JHCCL), a collection of small molecule clinical compounds, was screened virtually for COX-2 inhibition by docking approach. Docking of around 1400 small molecules against COX-2, leads to identification of a number of previously unreported molecules, which are likely to act as radioprotectors.

Silica nanoparticle: a potential new insecticide for mosquito vector control.

Presently, there is a need for increased efforts to develop newer and effective methods to control mosquito vectors as the existing chemical and biological methods are not as effective as in earlier period owing to different technical and operational reasons. The use of nanomaterial products in various sectors of science including health increased during the last decade. We tested three types of nanosilica, namely lipophilic, hydrophilic and hydrophobic, to assess their larvicidal, pupicidal and growth inhibitor properties and also their influence on oviposition behaviour (attraction/deterrence) of mosquito species that transmit human diseases, namely malaria (Anopheles), yellow fever, chickungunya and dengue (Aedes), lymphatic filariasis and encephalitis (Culex and Aedes). Application of hydrophobic nanosilica at 112.5 ppm was found effective against mosquito species tested. The larvicidal effect of hydrophobic nanosilica on mosquito species tested was in the order of Anopheles stephensi > Aedes aegypti > Culex quinquefasciatus, and the pupicidal effect was in the order of A. stephensi > C. quinquefasciatus > Ae. aegypti. Results of combined treatment of hydrophobic nanosilica with temephos in larvicidal test indicated independent toxic action without any additive effect. This is probably the first report that demonstrated that nanoparticles particularly nanosilica could be used in mosquito vector control.

Chlorfenapyr: a new insecticide with novel mode of action can control pyrethroid resistant malaria vectors.

BACKGROUND: Malaria vectors have acquired widespread resistance to many of the currently used insecticides, including synthetic pyrethroids. Hence, there is an urgent need to develop alternative insecticides for effective management of insecticide resistance in malaria vectors. In the present study, chlorfenapyr was evaluated against Anopheles culicifacies and Anopheles stephensi for its possible use in vector control. METHODS: Efficacy of chlorfenapyr against An. culicifacies and An. stephensi was assessed using adult bioassay tests. In the laboratory, determination of diagnostic dose, assessment of residual activity on different substrates, cross-resistance pattern with different insecticides and potentiation studies using piperonyl butoxide were undertaken by following standard procedures. Potential cross-resistance patterns were assessed on field populations of An. culicifacies. RESULTS: A dose of 5.0% chlorfenapyr was determined as the diagnostic concentration for assessing susceptibility applying the WHO tube test method in anopheline mosquitoes with 2 h exposure and 48 h holding period. The DDT-resistant/malathion-deltamethrin-susceptible strain of An. culicifacies species C showed higher LD50 and LD99 (0.67 and 2.39% respectively) values than the DDT-malathion-deltamethrin susceptible An. culicifacies species A (0.41 and 2.0% respectively) and An. stephensi strains (0.43 and 2.13% respectively) and there was no statistically significant difference in mortalities among the three mosquito species tested (p > 0.05). Residual activity of chlorfenapyr a.i. of 400 mg/m2 on five fabricated substrates, namely wood, mud, mud+lime, cement and cement + distemper was found to be effective up to 24 weeks against An. culicifacies and up to 34 weeks against An. stephensi. No cross-resistance to DDT, malathion, bendiocarb and deltamethrin was observed with chlorfenapyr in laboratory-reared strains of An. stephensi and field-caught An. culicifacies. Potentiation studies demonstrated the antagonistic effect of PBO. CONCLUSION: Laboratory studies with susceptible and resistant strains of An. culicifacies and An. stephensi, coupled with limited field studies with multiple insecticide-resistant An. culicifacies have shown that chlorfenapyr can be a suitable insecticide for malaria vector control, in multiple-insecticide-resistant mosquitoes especially in areas with pyrethroid resistant mosquitoes.

Malaria vector control: from past to future.

Malaria is one of the most common vector-borne diseases widespread in the tropical and subtropical regions. Despite considerable success of malaria control programs in the past, malaria still continues as a major public health problem in several countries. Vector control is an essential part for reducing malaria transmission and became less effective in recent years, due to many technical and administrative reasons, including poor or no adoption of alternative tools. Of the different strategies available for vector control, the most successful are indoor residual spraying and insecticide-treated nets (ITNs), including long-lasting ITNs and materials. Earlier DDT spray has shown spectacular success in decimating disease vectors but resulted in development of insecticide resistance, and to control the resistant mosquitoes, organophosphates, carbamates, and synthetic pyrethroids were introduced in indoor residual spraying with needed success but subsequently resulted in the development of widespread multiple insecticide resistance in vectors. Vector control in many countries still use insecticides in the absence of viable alternatives. Few developments for vector control, using ovitraps, space spray, biological control agents, etc., were encouraging when used in limited scale. Likewise, recent introduction of safer vector control agents, such as insect growth regulators, biocontrol agents, and natural plant products have yet to gain the needed scale of utility for vector control. Bacterial pesticides are promising and are effective in many countries. Environmental management has shown sufficient promise for vector control and disease management but still needs advocacy for inter-sectoral coordination and sometimes are very work-intensive. The more recent genetic manipulation and sterile insect techniques are under development and consideration for use in routine vector control and for these, standardized procedures and methods are available but need thorough understanding of biology, ethical considerations, and sufficiently trained manpower for implementation being technically intensive methods. All the methods mentioned in the review that are being implemented or proposed for implementation needs effective inter-sectoral coordination and community participation. The latest strategy is evolution-proof insecticides that include fungal biopesticides, Wolbachia, and Denso virus that essentially manipulate the life cycle of the mosquitoes were found effective but needs more research. However, for effective vector control, integrated vector management methods, involving use of combination of effective tools, is needed and is also suggested by Global Malaria Control Strategy. This review article raises issues associated with the present-day vector control strategies and state opportunities with a focus on ongoing research and recent advances to enable to sustain the gains achieved so far.

Development of larval thermotolerance and its impact on adult susceptibility to malathion insecticide and Plasmodium vivax infection in Anopheles stephensi.

The effect of the range of temperature on the thermal adaptation in Anopheles stephensi Liston 1901 was evaluated in the laboratory. Late third instar larvae of An. stephensi were exposed to variable temperatures viz. 37°C, 39°C, 41°C, 43°C and 45°C, and their lethal time to cause 50% mortality (LTM(50)) values were calculated. All larvae survived up to 39°C of exposure. However, at 45°C, they died within 30 min of exposure. Pre-exposure to variable temperatures and re-exposure to higher temperatures conferred adaptive thermotolerance. The larvae pre-adapted at 41°C that were re-exposed to 43°C, and larvae pre-adapted at 39°C that were re-exposed to 45°C, were found more thermotolerant than the thermally non-adapted larvae. Adaptive cross-tolerance to malathion was also induced by pre-exposing them to 37°C and 39°C. It suggests that temperature stress also play an important role in the development of adaptive cross-tolerance to other stress conditions. Similarly, the oocyst rate was relatively more in adults that emerged from larvae pre-adapted at 40°C as compared to adults that emerged from thermally non-adapted larvae and with lower oocyst load.