We are pleased to introduce the collection Frontiers in Tropical Diseases – Highlights in Vector Biology. This collection will welcome and showcase a selection of high impact articles authored by leaders in the field. The work presented here highlights the broad diversity of research performed across the Vector Biology section, and aims to put a spotlight on the main areas of interest. This collection aims to further support Frontiers’ strong community by shining a spotlight on our authors' highly impactful research.

University of Buea

Jigjiga University

Ain Shams University

Uncover a broad range of topics in the field of Vector Biology through this impactful collection.

. Vector-borne diseases are a global threat, and this research topic focuses on the development of drugs, vaccines, and alternative vector control strategies to combat them. <ul> <li>The Zika virus is a vector-borne flavivirus that has been detected in 87 countries worldwide and can lead to complications such as microcephaly in newborns and Guillain-Barré syndrome (GBS) in adults.</li> <li>Anopheles darlingi is a major vector of malaria in South and Central America, and insecticides have led to the selection of resistance alleles in malaria vectors.</li> <li>Live microbe vaccines are effective in preventing infectious diseases, and a flying vaccinator is a conceptually engineered hematophagous insect that could potentially deliver genetically attenuated virus vaccines via mosquito saliva.</li> <li>Ticks infesting cattle in the western region of Djurdjura in Algeria have an affinity for the inguinal region and necks.</li> <li>A rapid, simple, and versatile method for detecting Zika virus-derived DNA from mosquitoes has been developed, and this may provide a promising field-surveillance method for arbovirus circulation via vector mosquitoes.</li> <li>13 species of sandflies were identified in Chad, including the vector of human cutaneous leishmaniasis, Phlebotomus duboscqi.</li> <li>31 differentially expressed genes in the midgut tissue of mosquitoes infected with Dengue-2 virus have been identified, which could help develop novel dengue control strategies.</li> </ul>

Frontiers in Tropical Diseases

Vector Biology

Highlights in Vector Biology

The Zika virus (ZIKV) is a vector-borne flavivirus that has been detected in 87 countries worldwide. Outbreaks of ZIKV infection have been reported from various places around the world and the disease has been declared a public health emergency of international concern. ZIKV has two modes of transmission: vector and non-vector. The ability of ZIKV to vertically transmit in its competent vectors, such as Aedes aegypti and Aedes albopictus, helps it to cope with adverse conditions, and this could be the reason for the major outbreaks that occur from time to time. ZIKV outbreaks are a global threat and, therefore, there is a need for safe and effective drugs and vaccines to fight the virus. In more than 80% of cases, ZIKV infection is asymptomatic and leads to complications, such as microcephaly in newborns and Guillain–Barré syndrome (GBS) in adults. Drugs such as sofosbuvir, chloroquine, and suramin have been found to be effective against ZIKV infections, but further evaluation of their safety in pregnant women is needed. Although temoporfin can be given to pregnant women, it needs to be tested further for side effects. Many vaccine types based on protein, vector, DNA, and mRNA have been formulated. Some vaccines, such as mRNA-1325 and VRC-ZKADNA090-00-VP, have reached Phase II clinical trials. Some new techniques should be used for formulating and testing the efficacy of vaccines. Although there have been no recent outbreaks of ZIKV infection, several studies have shown continuous circulation of ZIKV in mosquito vectors, and there is a risk of re-emergence of ZIKV in the near future. Therefore, vaccines and drugs for ZIKV should be tested further, and safe and effective therapeutic techniques should be licensed for use during outbreaks.

The Zika virus (ZIKV) is a vector-borne flavivirus that has been detected in 87 countries worldwide. Outbreaks of ZIKV infection have been reported from various places around the world and the disease has been declared a public health emergency of international concern. ZIKV has two modes of transmission: vector and non-vector. The ability of ZIKV to vertically transmit in its competent vectors, such as Aedes aegypti and Aedes albopictus, helps it to cope with adverse conditions, and this could be the reason for the major outbreaks that occur from time to time. ZIKV outbreaks are a global threat and, therefore, there is a need for safe and effective drugs and vaccines to fight the virus. In more than 80% of cases, ZIKV infection is asymptomatic and leads to complications, such as microcephaly in newborns and Guillain–Barré syndrome (GBS) in adults. Drugs such as sofosbuvir, chloroquine, and suramin have been found to be effective against ZIKV infections, but further evaluation of their safety in pregnant women is needed. Although temoporfin can be given to pregnant women, it needs to be tested further for side effects. Many vaccine types based on protein, vector, DNA, and mRNA have been formulated. Some vaccines, such as mRNA-1325 and VRC-ZKADNA090-00-VP, have reached Phase II clinical trials. Some new techniques should be used for formulating and testing the efficacy of vaccines. Although there have been no recent outbreaks of ZIKV infection, several studies have shown continuous circulation of ZIKV in mosquito vectors, and there is a risk of re-emergence of ZIKV in the near future. Therefore, vaccines and drugs for ZIKV should be tested further, and safe and effective therapeutic techniques should be licensed for use during outbreaks.

ZIKV: Epidemiology, infection mechanism and current therapeutics

Anopheles darlingi is a major vector of both Plasmodium falciparum and Plasmodium vivax in South and Central America including French Guiana. However, the vector competence and physiology of this mosquito species have been scarcely studied due to difficulties in rearing it in the laboratory. Here, we report the successful establishment of a robust colony, from a mosquito collection in French Guiana. We describe our mosquito colonization procedure with relevant information on environmental conditions, mating ability, larval development, and survival, recorded over the first six critical generations. Experimental infection showed that our An. darlingi colony has a moderate permissiveness to in vitro produced gametocytes of the P. falciparum NF54 strain originating from Africa. This colony, which has reached its 21st generation, will allow further characterization of An. darlingi life-history traits and of Plasmodium–mosquito interactions with South American malaria parasites.

Anopheles darlingi is a major vector of both Plasmodium falciparum and Plasmodium vivax in South and Central America including French Guiana. However, the vector competence and physiology of this mosquito species have been scarcely studied due to difficulties in rearing it in the laboratory. Here, we report the successful establishment of a robust colony, from a mosquito collection in French Guiana. We describe our mosquito colonization procedure with relevant information on environmental conditions, mating ability, larval development, and survival, recorded over the first six critical generations. Experimental infection showed that our An. darlingi colony has a moderate permissiveness to in vitro produced gametocytes of the P. falciparum NF54 strain originating from Africa. This colony, which has reached its 21st generation, will allow further characterization of An. darlingi life-history traits and of Plasmodium–mosquito interactions with South American malaria parasites.

Establishment of a colony of Anopheles darlingi from French Guiana for vector competence studies on malaria transmission

Extensive use of insecticides has led to the selection of resistance alleles in malaria vectors threatening the control programs. Even if mosquitoes are not killed directly in the contact of insecticide-treated bed nets, their capacity to transmit malaria parasite could be decreased because of the consequences on their life-history traits after repeated exposure. The current work investigated the effects of organochlorine, carbamate, organophosphate, and pyrethroid insecticide exposure on the reproductive success in Anopheles gambiae s.s. Two Anopheles gambiae strains, AcerKis, KisKdr, were used. According to WHO recommendations, female mosquitoes of these resistant strains were exposed to discriminant doses of DDT, chlorpyriphos-methyl, bendiocarb, and permethrin insecticides. Surviving mosquitoes were then fed and allowed to lay eggs. Fecundity was assessed by examining the number of eggs per mosquito, the number of larvae per egg batch and larval hatching rates were used to evaluate the fertility. The data showed that AcerKis females surviving chlorpyriphos-methyl exposure significantly laid few eggs. No significant difference in the hatching rate was noticed in AcerKis females exposed to bendiocarb compared to their control. No significant effect on the fecundity and fertility was observed in KisKdr females exposed to permethrin. Our finding showed that organophosphate insecticides represented here by chlorpyriphos-methyl could hamper egg-laying in insensitive acetylcholinesterase An. gambiae female mosquitoes. This knowledge could help design alternative vector control strategies targeting fecundity and fertility in resistant malaria vectors.

Extensive use of insecticides has led to the selection of resistance alleles in malaria vectors threatening the control programs. Even if mosquitoes are not killed directly in the contact of insecticide-treated bed nets, their capacity to transmit malaria parasite could be decreased because of the consequences on their life-history traits after repeated exposure. The current work investigated the effects of organochlorine, carbamate, organophosphate, and pyrethroid insecticide exposure on the reproductive success in Anopheles gambiae s.s. Two Anopheles gambiae strains, AcerKis, KisKdr, were used. According to WHO recommendations, female mosquitoes of these resistant strains were exposed to discriminant doses of DDT, chlorpyriphos-methyl, bendiocarb, and permethrin insecticides. Surviving mosquitoes were then fed and allowed to lay eggs. Fecundity was assessed by examining the number of eggs per mosquito, the number of larvae per egg batch and larval hatching rates were used to evaluate the fertility. The data showed that AcerKis females surviving chlorpyriphos-methyl exposure significantly laid few eggs. No significant difference in the hatching rate was noticed in AcerKis females exposed to bendiocarb compared to their control. No significant effect on the fecundity and fertility was observed in KisKdr females exposed to permethrin. Our finding showed that organophosphate insecticides represented here by chlorpyriphos-methyl could hamper egg-laying in insensitive acetylcholinesterase An. gambiae female mosquitoes. This knowledge could help design alternative vector control strategies targeting fecundity and fertility in resistant malaria vectors.

Organophosphate Insecticide Exposure Impacts Reproductive Success in Insensitive Acetylcholinesterase Anopheles gambiae Mosquitoes

Live microbe vaccines are designed to elicit strong cellular and antibody responses without developing the symptoms of the disease, and these are effective in preventing infectious diseases. A flying vaccinator (also known as a flying syringe) is a conceptual, genetically engineered hematophagous insect that is used to deliver vaccines such as an antigen from a parasite produced in mosquito saliva; bites from such insects may elicit antibody production by immunizing the host with an antigen through blood-feeding. In addition to a simple vaccine antigen, a flying vaccinator may potentially load a live attenuated microbe with an appropriate mechanism for sustaining its constitutive proliferation in the insect. In this study, a recombinant vesicular stomatitis virus (VSV) lacking the glycoprotein gene (VSV-G) was used to produce replication-restricted VSV (rrVSV) containing GFP. Transgenic Anopheles stephensi mosquitoes, in which the salivary glands expressed a VSV-G gene driven by an aapp salivary gland-specific promoter, were generated and injected intraperitoneally with rrVSV. The injected rrVSV entered the cells of the salivary gland and stimulated endogenous production of progeny rrVSV particles, as seen in rrVSV-infected Drosophila melanogaster expressing VSV-G. These data suggested the possibility of developing a valuable tool for delivering genetically attenuated virus vaccines via mosquito saliva, although efficient replication-restricted virus production is required.

Live microbe vaccines are designed to elicit strong cellular and antibody responses without developing the symptoms of the disease, and these are effective in preventing infectious diseases. A flying vaccinator (also known as a flying syringe) is a conceptual, genetically engineered hematophagous insect that is used to deliver vaccines such as an antigen from a parasite produced in mosquito saliva; bites from such insects may elicit antibody production by immunizing the host with an antigen through blood-feeding. In addition to a simple vaccine antigen, a flying vaccinator may potentially load a live attenuated microbe with an appropriate mechanism for sustaining its constitutive proliferation in the insect. In this study, a recombinant vesicular stomatitis virus (VSV) lacking the glycoprotein gene (VSV-G) was used to produce replication-restricted VSV (rrVSV) containing GFP. Transgenic Anopheles stephensi mosquitoes, in which the salivary glands expressed a VSV-G gene driven by an aapp salivary gland-specific promoter, were generated and injected intraperitoneally with rrVSV. The injected rrVSV entered the cells of the salivary gland and stimulated endogenous production of progeny rrVSV particles, as seen in rrVSV-infected Drosophila melanogaster expressing VSV-G. These data suggested the possibility of developing a valuable tool for delivering genetically attenuated virus vaccines via mosquito saliva, although efficient replication-restricted virus production is required.

Generation of Transgenic Mosquitoes Harboring a Replication-Restricted Virus

The present study aimed to determine the phenology and predilection sites of ticks infesting cattle in the western region of Djurdjura (North Algeria) from November 2018 and October 2020. Nineteen cattle farms located in thirteen localities within four provinces were visited monthly for two years. Among the 289 examined cattle, 64.36 ± 2.81% (n=189) were infested by ticks. Of the 10,243 collected ticks, the most abundant tick species was Rhipicephalus bursa (31.57 ± 0.46%), followed by R. annulatus (31.26 ± 0.45%), Hyalomma marginatum (10.67 ± 0.30%), H. lusitanicum (7.02 ± 0.25%), H. excavatum (5.52±%0.22), H. scupense (4.27 ± 0.19%) and H. impeltatum (3.32 ± 0.17%). Ticks of the Hyalomma species were present throughout the year but in a limited number during the winter. H. scupense and H. impletatum showed similar activity from March to October and peaked in April and July, respectively (P<0.001). H. lusitanicum and H. excavatum were active from April to November and both peaked in September and October respectively. Rhipicephalus bursa was collected from April to August and R. annulatus from July to October. R. turanicus was active between April and June. Hyalomma genus had an affinity of attachment for the inguinal region (P<0.001). All parasitic stages of R. annulatus had an affinity for the necks (P<0.001). Nymphs of R. bursa prefer the neck (P<0.001), the adults attach to the perennial and inguinal regions (P<0.001). R. turanicus attach to the ears (P<0.001). As far as we know, this is the first study on the phenology and predilection sites of cattle ticks in the west region of Djurdjura in Algeria.

The present study aimed to determine the phenology and predilection sites of ticks infesting cattle in the western region of Djurdjura (North Algeria) from November 2018 and October 2020. Nineteen cattle farms located in thirteen localities within four provinces were visited monthly for two years. Among the 289 examined cattle, 64.36 ± 2.81% (n=189) were infested by ticks. Of the 10,243 collected ticks, the most abundant tick species was Rhipicephalus bursa (31.57 ± 0.46%), followed by R. annulatus (31.26 ± 0.45%), Hyalomma marginatum (10.67 ± 0.30%), H. lusitanicum (7.02 ± 0.25%), H. excavatum (5.52±%0.22), H. scupense (4.27 ± 0.19%) and H. impeltatum (3.32 ± 0.17%). Ticks of the Hyalomma species were present throughout the year but in a limited number during the winter. H. scupense and H. impletatum showed similar activity from March to October and peaked in April and July, respectively (P&lt;0.001). H. lusitanicum and H. excavatum were active from April to November and both peaked in September and October respectively. Rhipicephalus bursa was collected from April to August and R. annulatus from July to October. R. turanicus was active between April and June. Hyalomma genus had an affinity of attachment for the inguinal region (P&lt;0.001). All parasitic stages of R. annulatus had an affinity for the necks (P&lt;0.001). Nymphs of R.&nbsp;bursa prefer the neck (P&lt;0.001), the adults attach to the perennial and inguinal regions (P&lt;0.001). R. turanicus attach to the ears (P&lt;0.001). As far as we know, this is the first&nbsp;study on the phenology and predilection sites of cattle ticks in the west region of Djurdjura in Algeria.

Seasonal Dynamics and Predilection Sites of Ticks (Acari: Ixodidae) Feeding on Cows in the Western Parts of the Djurdjura, Algeria

Detection of infectious viruses in mosquitoes is one of the prerequisite measures to monitor the prevalence of vector-borne viral diseases. Determining which mosquitoes are currently infected with arboviruses such as Zika, dengue, and chikungunya virus is not yet practical in endemic areas due to multiple causes including the difficulty of dealing with the virus’ unstable RNA. In this study, instead of handling viral RNA, virus-derived DNA (vDNA) was introduced as a target template for nucleic acid amplification. In combination with loop-mediated isothermal amplification (LAMP), we examined a LAMP-based vDNA detection assay (vDNA-LAMP) targeting Zika virus (ZIKV). The vDNA-LAMP reaction amplifying part of the NS3 region of ZIKV successfully detected its vDNA from crude DNA purified from artificially infected cultured cells and Aedes mosquitoes. This rapid, simple, and versatile method may provide a promising field-surveillance method for arbovirus circulation via vector mosquitoes.

Detection of infectious viruses in mosquitoes is one of the prerequisite measures to monitor the prevalence of vector-borne viral diseases. Determining which mosquitoes are currently infected with arboviruses such as Zika, dengue, and chikungunya virus is not yet practical in endemic areas due to multiple causes including the difficulty of dealing with the virus’ unstable RNA. In this study, instead of handling viral RNA, virus-derived DNA (vDNA) was introduced as a target template for nucleic acid amplification. In combination with loop-mediated isothermal amplification (LAMP), we examined a LAMP-based vDNA detection assay (vDNA-LAMP) targeting Zika virus (ZIKV). The vDNA-LAMP reaction amplifying part of the NS3 region of ZIKV successfully detected its vDNA from crude DNA purified from artificially infected cultured cells and Aedes mosquitoes. This rapid, simple, and versatile method may provide a promising field-surveillance method for arbovirus circulation via vector mosquitoes.

LAMP Detection of Virus-Derived DNA of Zika Virus in Vector Mosquito

Leishmaniasis transmitted by sandflies is an important vector-borne disease. In Chad, information on sandflies is outdated, and so this study was designed to update the sandfly fauna. Sandflies were collected in five health districts representing two geoclimatic zones using sticky traps and pyrethrum sprays in indoor and peridomestic habitats between September 2019 and December 2020. All collected sandfly specimens were identified based on species-level morphological characters. A total of 2,015 specimens belonging to 13 species of sandflies (one Phlebotomus and twelve Sergentomyia) were collected and identified. The vector of human cutaneous leishmaniasis, Phlebotomus duboscqi, represents 0.65% of all collected sandflies and is the only representative of the Phlebotomus genus to be collected predominantly inside human dwellings. Phlebotomus orientalis, the vector of visceral leishmaniasis previously collected in Chad in 1976 was not found in this study. Sergentomyia clydei, Sergentomyia schwetzi, Sergentomyia antennata and Sergentomyia africana were the most abundant species collected with 44.71%; 22.73%; 16.03% and 11.17%, respectively. Sergentomyia schwetzi and Sergentomyia dubia, the two species involved in the transmission of canine leishmaniasis, were found in five and four study sites respectively. According to our results, the sandfly fauna of the two geoclimatic zones of Chad is comprised of 13 species. Our data showed that, unlike P. orientalis which was not found, P. duboscqi is present in four of the five sites surveyed. Therefore, these areas are at risk and remain potential foci of cutaneous leishmaniasis. However, the need for further studies such as vector species detection, their seasonal fluctuations and their vector competence.

Leishmaniasis transmitted by sandflies is an important vector-borne disease. In Chad, information on sandflies is outdated, and so this study was designed to update the sandfly fauna. Sandflies were collected in five health districts representing two geoclimatic zones using sticky traps and pyrethrum sprays in indoor and peridomestic habitats between September 2019 and December 2020. All collected sandfly specimens were identified based on species-level morphological characters. A total of 2,015 specimens belonging to 13 species of sandflies (one Phlebotomus and twelve Sergentomyia) were collected and identified. The vector of human cutaneous leishmaniasis, Phlebotomus duboscqi, represents 0.65% of all collected sandflies and is the only representative of the Phlebotomus genus to be collected predominantly inside human dwellings. Phlebotomus orientalis, the vector of visceral leishmaniasis previously collected in Chad in 1976 was not found in this study. Sergentomyia clydei, Sergentomyia schwetzi, Sergentomyia antennata and Sergentomyia africana were the most abundant species collected with 44.71%; 22.73%; 16.03% and 11.17%, respectively. Sergentomyia schwetzi and Sergentomyia dubia, the two species involved in the transmission of canine leishmaniasis, were found in five and four study sites respectively. According to our results, the sandfly fauna of the two geoclimatic zones of Chad is comprised of 13 species. Our data showed that, unlike P. orientalis which was not found, P. duboscqi is present in four of the five sites surveyed. Therefore, these areas are at risk and remain potential foci of cutaneous leishmaniasis. However, the need for further studies such as vector species detection, their seasonal fluctuations and their vector competence.

Distribution and Diversity of Sand Fly Species (Diptera: Psychodidae, Phlebotominae) in Two Geoclimatic Zones of Chad

Understanding the mosquito antiviral response could reveal target pathways or genes of interest that could form the basis of new disease control applications. However, there is a paucity of data in the current literature in understanding antiviral response during the replication period. To illuminate the gene expression patterns in the replication stage, we collected gene expression data at 2.5 days after Dengue-2 virus (DENV-2) infection. We sequenced the whole transcriptome of the midgut tissue and compared gene expression levels between the control and virus-infected group. We identified 31 differentially expressed genes. Based on their function, we identified that those genes fell into two major functional categories - (1) nucleic acid/protein process and (2) immunity/oxidative stress response. Our study has identified candidate genes that can be followed up for gene overexpression/inhibition experiments to examine if the perturbed gene interaction may impact the mosquito’s immune response against DENV. This is an important step to understanding how mosquitoes eliminate the virus and provides an important foundation for further research in developing novel dengue control strategies.

Understanding the mosquito antiviral response could reveal target pathways or genes of interest that could form the basis of new disease control applications. However, there is a paucity of data in the current literature in understanding antiviral response during the replication period. To illuminate the gene expression patterns in the replication stage, we collected gene expression data at 2.5 days after Dengue-2 virus (DENV-2) infection. We sequenced the whole transcriptome of the midgut tissue and compared gene expression levels between the control and virus-infected group. We identified 31 differentially expressed genes. Based on their function, we identified that those genes fell into two major functional categories - (1) nucleic acid/protein process and (2) immunity/oxidative stress response. Our study has identified candidate genes that can be followed up for gene overexpression/inhibition experiments to examine if the perturbed gene interaction may impact the mosquito&#x2019;s immune response against DENV. This is an important step to understanding how mosquitoes eliminate the virus and provides an important foundation for further research in developing novel dengue control strategies.