Source: OHIO STATE UNIVERSITY submitted to NRP
THE MOLECULAR PHYSIOLOGY OF KIDNEY FUNCTION IN MOSQUITOES: IDENTIFICATION OF MOLECULAR TARGETS AND DISCOVERY OF INSECTICIDES FOR MOSQUITO CONTROL
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
0229723
Grant No.
(N/A)
Cumulative Award Amt.
(N/A)
Proposal No.
(N/A)
Multistate No.
(N/A)
Project Start Date
Jun 1, 2012
Project End Date
Sep 30, 2017
Grant Year
(N/A)
Program Code
[(N/A)]- (N/A)
Recipient Organization
OHIO STATE UNIVERSITY
1680 MADISON AVENUE
WOOSTER,OH 44691
Performing Department
Entomology
Non Technical Summary
Mosquitoes are vectors of numerous pathogens that present a significant challenge to the health and well-being of humans and animals. Unfortunately, the present arsenal of insecticides used for mosquito control is rapidly losing its effectiveness, because insecticide resistance is evolving in mosquitoes. Thus, there is an urgent need to develop new insecticides with novel mechanisms of action to control the spread of mosquito-borne diseases. In response to this need, the present Hatch proposal aims to facilitate the development of a new generation of insecticides that will kill mosquitoes by attacking their analog of a kidney--i.e., the Malpighian tubules. The objectives of the proposal are to: 1) identify genes that play key roles in the renal functions of adult female mosquitoes (the blood-feeding life stage); and 2) discover chemical compounds that kill mosquitoes by impairing their renal functions. If the objectives are met, then the proposed Hatch studies have potential to catalyze the development of a new generation of insecticides for controlling mosquitoes.
Animal Health Component
(N/A)
Research Effort Categories
Basic
(N/A)
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3053110102050%
3113110118025%
7213110118025%
Knowledge Area
311 - Animal Diseases; 305 - Animal Physiological Processes; 721 - Insects and Other Pests Affecting Humans;

Subject Of Investigation
3110 - Insects;

Field Of Science
1180 - Pharmacology; 1020 - Physiology;
Goals / Objectives
The goal of the first objective is to identify genes expressed in Malpighian (renal) tubules that contribute to the enhanced excretory functions of blood-fed mosquitoes. In particular, genes that encode ion transporters will be focused on, given the known importance of these proteins in Malpighian tubule function and the expertise of the principal investigator. The objectives will be accomplished by completing two specific aims: 1) identification of the ion-transporter genes that are expressed differentially in Malpighian tubules after female mosquitoes ingest a blood meal, and 2) validation of the identified genes in Malpighian tubule function during a blood meal. The goal of the second objective is to discover small-molecule insecticides that kill mosquitoes by impairing Malpighian tubule function. This objective will be accomplished by a collaborative multi-institutional team (led by the principal investigator at the Ohio State University). This team will complete two specific aims: 1) discover and optimize small molecule antagonists of specific mosquito ion transporters identified in Objective 1, and 2) assess the effects of the small-molecule antagonists on isolated Malpighian tubules and adult female mosquitoes.
Project Methods
Objective 1 Specific Aim 1.1. This aim will be accomplished using next-generation sequencing technology (RNA-seq) at the Molecular and Cellular Imaging Center (MCIC) of the OARDC. In brief, gene expression in Malpighian tubules will be assessed at specific time points (i.e., 6, 24, and 48 hrs) after female mosquitoes ingest a blood meal from a membrane-feeding device. Working with experts at the MCIC, cDNA libraries of Malpighian tubules at the aforementioned time points will be generated and sequenced. The data will be assembled in silico to establish a blood-feeding transcriptome for Malpighian tubules and subsequently analyzed to identify genes that exhibit dramatic changes in expression during the processing of a blood meal. Such genes will be noted as genes of interest. Specific Aim 1.2. This aim will be accomplished using a functional genetics approach. In particular, double-stranded RNA (dsRNA) against a gene of interest will be injected into the hemolymph of female mosquitoes using a microinjection workstation in the laboratory of the principal investigator. Injection of dsRNA should initiate an RNAi pathway that will knock down expression of the particular gene of interest and its encoded protein. Once knock down of a gene is verified, the mosquitoes will be fed a blood meal via a membrane-feeding device. The mosquitoes will be monitored for 5 days after the ingestion of blood to assess for mortality and desirable sub-lethal effects (e.g., decreased egg production, decreased digestion efficiency). Objective 2 Specific Aim 2.1. The goal of this aim is to discover and optimize small-molecule antagonists of ion transporters that are identified in Objective 1. This aim will be accomplished in collaboration with the Dr. Jerod S. Denton of Vanderbilt University. In brief, a cDNA that corresponds to a gene of interest will be cloned by the principal investigator. The cDNA will then be sent to Dr. Denton for heterologous expression in a cell line. Once expressed, Dr. Denton will develop a fluorescence-based assay to measure function of the encoded protein and screen a library of small molecules for antagonists of the protein. Discovered antagonists will be designated for downstream medicinal chemistry efforts (performed by Vanderbilt) to improve their potency and specificity for the mosquito protein. Once optimized, the antagonist will be assessed for its potential as an insecticide as described below. Specific Aim 2.2. The goal of this aim is to confirm that an optimized small-molecule antagonist (see previous aim) elicits the desired lethal effects on adult female mosquitoes. That is, the antagonist will be injected into the hemolymph of adult female mosquitoes using a microinjection workstation in the laboratory of the principal investigator at the OARDC. The survival of the injected mosquitoes will be monitored for 24 hrs. If no mortality is observed, then mosquitoes will be fed blood and monitored for 5 days as described above in Objective 1.2. Parallel experiments will also be conducted on mosquitoes fed a sucrose diet to determine if the antagonists have distinct effects on sugar-fed mosquitoes.

Progress 06/01/12 to 09/30/17

Outputs
Target Audience:Target audiences include basic scientists, medical and veterinary professionals, and vector-management specialists who are faced with the challenge of controlling disease-spreading mosquitoes that are resistant to traditional insecticides. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project has providing training and professional development opportunities for the individuals listed below who are trainees of the principal investigator (Dr. Peter M. Piermarini): Postdoctoral trainees: Dr. Megha Kalsi Doctoral students: Dr. Travis L. Calkins (Graduated in May 2017), Dr. Liu Yang (Graduate in August 2017) Undergraduate students: Mr. Andrew DeLaat Visiting Scholars: Ms. Edna Alfaro Inocente Research assistants: Ms. Nuris Acosta How have the results been disseminated to communities of interest?Results of our work have been disseminated through peer-reviewed journals, invited seminars at other universities, and invited and submitted presentations at local, regional, national, and international conferences. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? This project is revealing novel molecular targets and chemicals to guide the development of a new generation of insecticides that target the 'kidneys' of mosquitoes for combatting insecticide resistant mosquitoes. The products of this research have the potential to slow the spread of mosquito-borne diseases, such as malaria, dengue fever, West Nile fever, and Zika, which collectively debilitate the health and well-being of hundreds of millions of people around the globe each year. This project is also providing new molecular insights into the contributions that the mosquito 'kidney' makes to the physiological processing of blood meals by mosquitoes, which has been an understudied area of mosquito biology. Thus, the products of this research are revealing new molecular and physiological mechanisms used by mosquitoes--and perhaps other blood-feeding insects-- that allow them to efficiently metabolize the large blood meals that they ingest. Efforts on the project have identified nearly 4,000 genes in the 'kidneys' (i.e., Malpighian tubules) of mosquitoes whose expression levels are significantly affected during the processing of blood meals. Functional pathway analyses of the differentially-expressed genes indicate that the Malpighian tubules undergo a functional transformation in which they reduce investment in pathways associated with urine production and increase investment in pathways associated with the detoxification and excretion of metabolites. Genes associated with the latter pathways are now being considered as targets for insecticides that may limit the number of blood meals mosquitoes can ingest during their lifetime, thereby slowing the transmission of mosquito-borne pathogens. Furthermore, in a collaborative effort with Vanderbilt University and Cornell University, this project is discovering chemicals that kill mosquitoes by disrupting Malpighian tubule function. We have discovered chemicals that interfere with the function of potassium channels in mosquito Malpighian tubules; the chemicals compromise the abilities of mosquitoes to excrete urine and fly. We have now identified similar chemicals that show a high potency for perturbing the potassium channels of mosquitoes, but not those of humans. Moreover, these chemicals show no toxic effects on honey bees. These chemicals are being further developed and will provide the basis for the development of novel, safe insecticides.

Publications

  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Li Y, Piermarini PM, Esquivel CJ, Drumm HE, Schilkey FD, Hansen IA (2017) RNA-Seq Comparison of Larval and Adult Malpighian Tubules of the Yellow Fever Mosquito Aedes aegypti Reveals Life Stage-Specific Changes in Renal Function. Front Physiol 8: 283.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Yang L, Piermarini PM (2016) Molecular expression of aquaporin mRNAs in the northern house mosquito, Culex pipiens. J Insect Physiol 96: 35-44.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Swale DR, Engers DW, Bollinger SR, Gross A, Inocente EA, Days E, Kanga F, Johnson RM, Yang L, Bloomquist JR, Hopkins CR, Piermarini PM, Denton JS (2016) An insecticide resistance-breaking mosquitocide targeting inward rectifier potassium channels in vectors of Zika virus and malaria. Sci Rep 6: 36954.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Kolosov D*, Piermarini PM, Kelly SP, ODonnell MJ. Cell-specific ion transport mechanisms in the Malpighian tubules of the larval cabbage looper Trichoplusia ni. Poster presentation, 26th Annual Biochemistry and Comparative Physiology Conference, Rice Lake, Canada. January, 27th-29th 2017.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Rakotondraibe LH, Alfaro E, Piermarini PM, Drew M, Lee N (2017) Bioactive secondary metabolites of three species of Madagascan endemic genus Cinnasmosma (Canellaceae) Annual meeting of the American Society of Pharmacognosy (ASP). Portland, Oregon: July 29th-August 2nd 2017
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Yang L, Gardiner MM and Piermarini PM (2017) The impact of urban greenspaces management on the mosquito community. NCB-ESA 2017, Indianapolis, IN. Symposia: Medical Entomology in a Changing World. Invited Talk.
  • Type: Other Status: Published Year Published: 2017 Citation: Piermarini PM (2017) Mosquito Molecular Physiology in a Time of Zika: targeting mosquito renal function for vector control. University of Sao Paulo, Brazil, Department of Parasitology Invited Seminar, January
  • Type: Other Status: Published Year Published: 2017 Citation: Piermarini PM (2017) Mosquito Molecular Physiology in a Time of Zika: targeting mosquito renal function for vector control. University of Cincinnati, Department of Biology Invited Seminar, February
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Piermarini PM, DeLaat A, Calkins TL (2017) Gap junctions in the crop of the yellow fever mosquito Aedes aegypti. Insect Biotech Conference Niagara on the Lake, ON, June
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Piermarini PM (2017) Discovery of natural and synthetic small molecule mosquitocides with novel mechanisms of action. 15th International Dengue, Zika, and Chikungunya Course, Pedro Kouri Institute of Tropical Medicine, Havana, Cuba, August.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Yang Z, Statler B-M, Calkins TL, Alfaro E, Esquivel CJ, Rouhier MF, Denton JS, Piermarini PM (2017) Dynamic expression of genes encoding subunits of inward rectifier potassium (Kir) channels in the yellow fever mosquito Aedes aegypti. Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology 204: 35-44.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Yang L, Denlinger DL, Piermarini PM (2017) The diapause program impacts renal excretion and molecular expression of aquaporins in the northern house mosquito, Culex pipiens. Journal of Insect Physiology 98: 141-148.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Piermarini PM, Esquivel CJ, Denton JS (2017) Malpighian Tubules as Novel Targets for Mosquito Control. Int J Environ Res Public Health 14.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Piermarini PM, Akuma DC, Crow JC, Jamil TL, Kerkhoff WG, Viel KCMF, Gillen CM (2017) Differential expression of putative sodium-dependent cation-chloride cotransporters in Aedes aegypti. Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology 214: 40-49.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Oca-Aguilar ACM, Mikery-Pacheco O, Castillo A, Rebollar-Tellez EA, Piermarini PM, Ibanez-Bernal S (2017) Morphology variation of Lutzomyia cruciata eggs (Diptera: Psychodidae: Phlebotominae) in southern Mexico. Zootaxa 4258: 477-489.
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Montes de Oca-Aguilar AC, Rebollar-Tellez EA, Piermarini PM, Ibanez-Bernal S (2017) Descriptions of the Immature Stages of Lutzomyia (Tricholateralis) cruciata (Coquillett) (Diptera: Psychodidae, Phlebotominae). Neotrop Entomol 46: 66-85.


Progress 10/01/15 to 09/30/16

Outputs
Target Audience:Target audiences include basic scientists, medical and veterinary professionals, and vector-management specialists who are faced with the challenge of controlling disease-spreading mosquitoes that are resistant to traditional insecticides. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project has providing training and professional development opportunities for the individuals listed below who are trainees of the principal investigator (Dr. Peter M. Piermarini): Doctoral students: Ms. Liu Yang, Mr. Travis L. Calkins Masters students: Mr. Carlos Esquivel (Graduated in May 2016) Undergraduate students: Ms. Jasmine Cruz High school students: Ms. Margot Shaya Visiting Scholars: Dr. Zhongxia Yang, Ms. Edna Alfaro Inocente Research assistants: Ms. Nuris Acosta How have the results been disseminated to communities of interest?Results of our work have been disseminated through peer-reviewed journals, invited seminars at other universities, and invited and submitted presentations at local, regional, national, and international conferences. What do you plan to do during the next reporting period to accomplish the goals?We plan to continue our investigations determining the genes that are expressed in the Malpighian tubules of mosquitoes and the respective roles they play in the processing of blood meals. We predict that inhibiting the expression or function of these genes will make a blood meal lethal to mosquitoes. We also plan to continue our development of new insecticides that target potassium channels in mosquitoes without effects on the potassium channels of humans and beneficial insects. Such chemicals may provide prototypes for the development of novel, safe insecticides.

Impacts
What was accomplished under these goals? This project is revealing novel molecular targets and chemicals to guide the development of a new generation of insecticides that target the 'kidneys' of mosquitoes for combatting insecticide resistant mosquitoes. The products of this research have the potential to slow the spread of mosquito-borne diseases, such as malaria, dengue fever, West Nile fever, and Zika, which collectively debilitate the health and well-being of hundreds of millions of people around the globe each year. This project is also providing new molecular insights into the contributions that the mosquito 'kidney' makes to the physiological processing of blood meals by mosquitoes, which has been an understudied area of mosquito biology. Thus, the products of this research are revealing new molecular and physiological mechanisms used by mosquitoes--and perhaps other blood-feeding insects--that allow them to efficiently metabolize the large blood meals that they ingest. Efforts on the project have identified nearly 4,000 genes in the 'kidneys' (i.e., Malpighian tubules) of mosquitoes whose expression levels are significantly affected during the processing of blood meals. Functional pathway analyses of the differentially-expressed genes indicate that the Malpighian tubules undergo a functional transformation in which they reduce investment in pathways associated with urine production and increase investment in pathways associated with the detoxification and excretion of metabolites. Genes associated with the latter pathways are now being considered as targets for insecticides that may limit the number of blood meals mosquitoes can ingest during their lifetime, thereby slowing the transmission of mosquito-borne pathogens. Furthermore, in a collaborative effort with Vanderbilt University and Cornell University, this project is discovering chemicals that kill mosquitoes by disrupting Malpighian tubule function. We have discovered chemicals that interfere with the function of potassium channels in mosquito Malpighian tubules; the chemicals compromise the abilities of mosquitoes to excrete urine and fly. We have now identified similar chemicals that show a high potency for perturbing the potassium channels of mosquitoes, but not those of humans. Moreover, these chemicals show no toxic effects on honey bees. These chemicals are being further developed and will provide the basis for the development of novel, safe insecticides.

Publications

  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Esquivel CJ, Cassone BJ, Piermarini PM (2016) A de novo transcriptome of the Malpighian tubules in non-blood-fed and blood-fed Asian tiger mosquitoes Aedes albopictus: insights into diuresis, detoxification, and blood meal processing. PeerJ 4: e1784.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Montes de Oca-Aguilar AC, Rebollar-Tellez EA, Piermarini PM, Ibanez-Bernal S (2016) Descriptions of the Immature Stages of Lutzomyia (Tricholateralis) cruciata (Coquillett) (Diptera: Psychodidae, Phlebotominae). Neotrop Entomol.
  • Type: Book Chapters Status: Published Year Published: 2016 Citation: Piermarini PM (2016) Renal Excretory Processes in Mosquitoes. In: Raikhel AS, editor. Advances in Insect Physiology. Oxford: Academic Press. pp. 393-433.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Piermarini PM, Cassone BJ, Esquivel CJ Molecular, biochemical, and physiological evidence for a functional transition in the 'kidneys' of adult female mosquitoes during the processing of blood meals. Presented at the Annual Conference of Entomological Society of America in the PBT Section symposium entitled Water and Ion Homeostasis-Role of Aquaporins and Other Channel Proteins, Minneapolis, MN, USA (2015).
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Yang L, Denlinger DL, Piermarini PM Effects of diapause on the excretory physiology and expression of aquaporins in the West Nile vector Culex pipiens Presented at the Annual Conference of the American Society for Tropical Medicine and Hygiene in the symposium entitled The next generation of medical entomologists, Philadelphia, PA, USA (2015).
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Piermarini PM, Cassone BJ, Esquivel CJ The Malpighian tubule transcriptome of the Asian tiger mosquito Aedes albopictus: new insights into blood meal processing by mosquito kidneys. Presented at the Annual Conference of the American Society for Tropical Medicine and Hygiene, Philadelphia, PA, USA (2015).
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Piermarini PM, Beyenbach KW, Denton JS Exploiting the kidneys of mosquitoes for the development of novel insecticides Presented at the Annual Conference of the American Society for Tropical Medicine and Hygiene, Philadelphia, PA, USA (2015).
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Piermarini PM "New molecular and physiological targets for mosquitocide development" Presented at the Annual Conference of the North Central Branch of the Entomological Society of America, Cleveland, OH, USA (2016)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Piermarini PM "Mosquito molecular physiology in a time of Zika" Presented at the Insect Biotech Conference, St. Catharines, Ontario, Canada (2016)
  • Type: Other Status: Other Year Published: 2016 Citation: Piermarini PM " Mosquito molecular physiology in a time of Zika" Presented in New Mexico State University's Department of Biology Seminar Series. Las Cruces, NM, USA (2016)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Piermarini PM, Beyenbach KW, Denton JS "The molecular physiology of inward rectifier potassium (Kir) channels in mosquitoes" Presented at the International Congress of Entomology, Orlando, FL, USA (2016)
  • Type: Other Status: Other Year Published: 2016 Citation: Piermarini PM, Beyenbach KW, Denton JS Exploiting the kidneys of mosquitoes for the development of novel insecticides Presented at the International Congress of Entomology, Orlando, FL, USA (2016)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: 2016 Calkins, T.L., and Piermarini, P.M., 2016. Gap junctions and the mosquito blood meal. International Congress of Entomology and Entomological Society of America meeting. Orlando, Florida, August 27, 2016.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: 2016 Calkins, T.L., and Piermarini, P.M., 2016. A role of gap junctions in mosquito blood meal processing. Entomological Society of America Annual North Central Branch Meeting, Cleveland, OH. June 6, 2015 (Oral Presentation)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Calkins, T.L., and Piermarini, P.M., 2016. Pharmacological and genetic evidence for gap junctions as potential new insecticide targets in the yellow fever mosquito, Aedes aegypti. Ohio Agriculture Research and Development Center Annual Research Conference, Wooster, OH. April 21, 2016.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Calkins, T.L. and Piermarini, P.M. 2016 Gap junctions as potential new insecticide targets in the yellow fever mosquito, Aedes aegypti. American Chemical Society, Annual meeting, Philadelphia, Pennsylvania, August, 22, 2016 (Oral Presentation).
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Liu Yang and Peter M. Piermarini. Diapause suppresses the diuretic capacity and molecular expression of aquaporins in the West Nile vector mosquito, Culex pipiens. Presented at the Annual Conference of the American Society for Tropical Medicine and Hygiene, Philadelphia, PA, USA (2015).
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Liu Yang and Peter M. Piermarini. The molecular expression of aquaporin genes in the northern house mosquito, Culex pipiens. Ohio Agriculture Research and Development Center Annual Research Conference, Wooster, OH. April 2016.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Liu Yang and Peter M. Piermarini. Molecular expression of aquaporin mRNAs in the northern house mosquito, Culex pipiens. Presented at the International Congress of Entomology. Orlando, Florida, 2016.


Progress 10/01/14 to 09/30/15

Outputs
Target Audience:Target audiences include basic scientists, medical and veterinary professionals, and vector-management specialists who are faced with the challenge of controlling disease-spreading mosquitoes that are resistant to traditional insecticides. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project has providing training and professional development opportunities for the individuals listed below who are trainees of the principal investigator (Dr. Peter M. Piermarini): Doctoral students: Ms. Liu Yang, Mr. Travis L. Calkins Masters students: Mr. Carlos Esquivel Undergraduate students: Ms. Elizabeth Statler Visiting Scholars: Dr. Zhongxia Yang, Ms. Edna Alfaro Inocente Research assistant: Ms. Nuris Acosta How have the results been disseminated to communities of interest?Results of our work have been disseminated through peer-reviewed journals, invited seminars at other universities, and invited and submitted presentations at local, regional, national, and international meetings. What do you plan to do during the next reporting period to accomplish the goals?We plan to continue our investigations into the genes that are expressed in the Malpighian tubules of mosquitoes and determine which roles they play in the processing of blood meals. We will interfere with the expression of these genes using RNA interference and gene editing approaches (e.g., CRISPR/cas9). We predict that perturbing the expression of these genes will make a blood meal lethal to mosquitoes. In addition, we plan to continue our development of new insecticides that target potassium channels in mosquitoes without effects on the potassium channels of humans and beneficial insects. Such chemicals may provide prototypes for the development of novel, safe insecticides.

Impacts
What was accomplished under these goals? This project is revealing novel molecular targets and chemicals to guide the development of a new generation of insecticides that target the 'kidneys' of mosquitoes for combatting insecticide resistant mosquitoes. The products of this research have the potential to slow the spread of mosquito-borne diseases, such as malaria, dengue fever, and West Nile fever, which collectively debilitate the health and well-being of hundreds of millions of people around the globe each year. This project is also providing new molecular insights into the contributions that the mosquito 'kidney' makes to the physiological processing of blood meals by mosquitoes, which has been an understudied area of mosquito biology. Thus, the products of this research are revealing new physiological mechanisms used by mosquitoes--and perhaps other blood-feeding insects--that allow them to efficiently metabolize the large blood meals that they ingest. Efforts on the project have identified over 3,000 genes in the 'kidneys' (i.e., Malpighian tubules) of mosquitoes whose expression levels are significantly affected during the processing of blood meals. These genes are now being prioritized as targets for insecticides that may limit the number of blood meals mosquitoes can ingest during their lifetime, thereby slowing the transmission of mosquito-borne pathogens. Furthermore, in a collaborative effort with Vanderbilt University and Cornell University, this project is discovering chemicals that kill mosquitoes by disrupting Malpighian tubule function. We have discovered chemicals that interfere with the function of potassium channels in mosquito Malpighian tubules; the chemicals compromise the abilities of mosquitoes to excrete urine and fly. We have now identified similar chemicals that show a high potency for perturbing the potassium channels of mosquitoes, but not those of humans. Moreover, these chemicals show no toxic effects on honey bees. These chemicals are being further developed and will provide the basis for the development of novel, safe insecticides.

Publications

  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Hine RM, Rouhier MF, Park ST, Qi Z, Piermarini PM, Beyenbach KW (2014) The excretion of NaCl and KCl loads in mosquitoes: 1. Control data. Am J Physiol Regul Integr Comp Physiol 307: R837-R849.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Rouhier MF, Hine RM, Park ST, Raphemot R, Denton JS, Piermarini PM, Beyenbach KW (2014) The excretion of NaCl and KCl loads in mosquitoes: 2. Effects of the small molecule Kir channel modulator VU573 and its inactive analog VU342. Am J Physiol Regul Integr Comp Physiol 307: R850-R861.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Beyenbach KW, Yu Y, Piermarini PM, Denton J (2015) Targeting renal epithelial channels for the control of insect vectors. Tissue Barriers 3: e1081861.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Calkins TL, Piermarini PM (2015) Pharmacological and genetic evidence for gap junctions as potential new insecticide targets in the yellow fever mosquito, Aedes aegypti. PLoS ONE 10: e0137084.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Calkins TL, Woods-Acevedo MA, Hildebrandt O, Piermarini PM (2015) The molecular and immunochemical expression of innexins in the yellow fever mosquito, Aedes aegypti: Insights into putative life stage- and tissue-specific functions of gap junctions. Comp Biochem Physiol B Biochem Mol Biol 183C: 11-21.
  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Piermarini PM, Dunemann SM, Rouhier MF, Calkins TL, Raphemot R, Denton JS, Hine RM, Beyenbach KW (2015) Localization and role of inward rectifier K channels in Malpighian tubules of the yellow fever mosquito Aedes aegypti. Insect Biochem Mol Biol Epub ahead of print.
  • Type: Book Chapters Status: Published Year Published: 2015 Citation: Piermarini P, Gillen C (2015) Non-traditional Models: The Molecular Physiology of Sodium and Water Transport in Mosquito Malpighian Tubules. In: Hyndman KA, Pannabecker TL, editors. Sodium and Water Homeostasis: Springer New York. pp. 255-278.
  • Type: Other Status: Other Year Published: 2014 Citation: Piermarini PM "Eliciting renal failure in mosquitoes." Presented at College of Wooster Department of Biology Seminar Series. Wooster, Ohio, United States. (2014)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Piermarini PM, Denton JS, Beyenbach KW "Exploiting the Renal Homeostatic Mechanisms of Mosquitoes for Novel Vector Control." Presented at 2014 APS Intersociety Meeting: Comparative Approaches to Grand Challenges in Physiology. San Diego, California, United States. (2014)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Piermarini PM A brief history of Klaus W. Beyenbachs contributions to the field of renal epithelial physiology. Presented at the Society for Experimental Biology Annual Meeting. Prague, Czech Republic. (2015).
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Alfaro EA, Yang L, Acosta, N, Piermarini PM "Evaluation of small molecules as insecticides for the control of mosquitoes (Aedes aegypti) and aphids (Aphis glycines)." Presented at 9th Annual International Scholar Research Exposition. Columbus, Ohio, United States. (2014)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Alfaro EA, Yang L, Acosta, N, Piermarini PM "Evaluation of small molecules as insecticides for the control of mosquitoes (Aedes aegypti) and aphids (Aphis glycines)." Presented at 2015 OARDC Annual Research Conference: Can We Protect Water Quality?: The Importance of Watersheds. Columbus, Ohio, United States. (2015)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Calkins TL, Piermarini PM "Toxicological effects of gap junction inhibitors on the yellow fever mosquito, Aedes aegypti." Presented at 2015 OARDC Annual Research Conference: Can We Protect Water Quality?: The Importance of Watersheds. Columbus, Ohio, United States. (2015)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Esquivel CJ, Piermarini PM "Physiological and biochemical assays confirm a functional transition in the Malpighian tubules after a blood meal in the invasive Asian tiger mosquito Aedes albopictus." Presented at 2015 OARDC Annual Research Conference: Can We Protect Water Quality?: The Importance of Watersheds. Columbus, Ohio, United States. (2015)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Calkins TL, Piermarini PM "Toxicological effects of gap junction inhibitors on the yellow fever mosquito, Aedes aegypti." Presented at the North Central Branch of the Entomological Society of America Annual Meeting. Manhattan, Kansas, United States. (2015)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Calkins TL, Piermarini PM "The molecular physiology of gap junctions (innexins) in the mosquito Aedes aegypti: integrating pharmacology and functional genetics." Presented at Ohio Valley Entomological Association Annual Meeting. Columbus, Ohio, United States. (2014)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Esquivel CJ, Cassone BJ, Piermarini PM "Transcriptomic evidence of a functional transition of the Malpighian tubules after a blood meal in the Asian tiger mosquito Aedes albopictus." Presented at Ohio Valley Entomological Association Annual Meeting. Columbus, Ohio, United States. (2014)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Esquivel CJ, Cassone BJ, Piermarini PM "Transcriptomic evidence of a functional transition of the Malpighian tubules after a blood meal in the Asian tiger mosquito Aedes albopictus." Presented at Entomological Society of America Annual Meeting. Portland, Oregon, United States. (2014)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Yang L, Piermarini PM "Effects of diapause on the excretory physiology of the northern house mosquito, Culex pipiens." Presented at Entomological Society of America Annual Meeting. Portland, Oregon, United States. (2014)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2015 Citation: Calkins TL, Piermarini PM Elucidating the function of gap junctions in the yellow fever mosquito, Aedes aegypti, utilizing toxicological and molecular approaches. Presented at the Society for Experimental Biology Annual Meeting. Prague, Czech Republic. (2015)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Calkins TL, Piermarini PM "The molecular physiology of gap junctions (innexins) in the mosquito Aedes aegypti: integrating pharmacology and functional genetics." Presented at Entomological Society of America Annual Meeting. Portland, Oregon, United States. (2014)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Yang L, Piermarini PM "Effects of diapause on the excretory physiology of the northern house mosquito, Culex pipiens." Presented at the Ohio Valley Entomological Association Annual Meeting. Columbus, Ohio, United States. (2014)


Progress 10/01/13 to 09/30/14

Outputs
Target Audience: Target audiences include basic scientists, medical and veterinary professionals, and vector-management specialists who are faced with the challenge of controlling disease-spreading mosquitoes that are resistant to traditional insecticides. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? The project has providing training and professional development opportunities for the individuals listed below who are trainees of the principal investigator (Dr. Peter M. Piermarini): Postdoctoral researcher: Dr. Matthew F. Rouhier Doctoral student: Ms. Liu Yang Masters students: Mr. Travis L. Calkins; Mr. Carlos Esquivel Undergraduate students: Ms. Sheila Amoko, Mr. Mikal A. Woods, Ms. Jacque Garcia Visiting Scholars: Ms. Rosa Witty, Ms. Edna Alfaro Inocente Research assistant: Ms. Nuris Acosta How have the results been disseminated to communities of interest? Results of our work have been disseminated through peer-reviewed journals, invited seminars at other universities, and invited and submitted presentations at local, regional, national, and international meetings. What do you plan to do during the next reporting period to accomplish the goals? We plan to continue our investigations into which genes expressed in the Malpighian tubules of mosquitoes play key roles in the processing of blood meals by interfering with the expression of those genes whose expression is more abundant after a blood meal using RNA interference. We predict that perturbing the expression of these genes will make a blood meal lethal to mosquitoes. In addition, we plan to continue our development of new insecticides that target potassium channels in mosquitoes. With collaborators at Vanderbilt and Cornell Universities, we are in search of chemicals that will show a high potency for perturbing the potassium channels of mosquitoes, but not those of humans and other animals. Such chemicals may provide the basis for the development of novel, safe insecticides.

Impacts
What was accomplished under these goals? Mosquito-borne diseases (e.g., malaria, dengue fever, West Nile disease) are an emerging threat to the health and well-being of humans. Each year, such mosquito-borne diseases take an immense toll on global health. Hundreds of millions of people around the globe are infected with these diseases each year, which claim nearly one million lives. Because mosquitoes are rapidly evolving resistance to conventional insecticides, new insecticides are urgently needed to expand our chemical arsenal for controlling mosquitoes and slowing the spread of mosquito-borne diseases. This project is revealing novel molecular targets and chemicals to guide the development of a new generation of insecticides that target the 'kidneys' of mosquitoes for combatting insecticide resistant mosquitoes. The products of this research have the potential to slow the spread of mosquito-borne diseases, such as malaria, dengue fever, and West Nile fever, which collectively debilitate the health and well-being of hundreds of millions of people around the globe each year. This project is also providing new molecular insights into the contributions that the mosquito 'kidney' makes to the physiological processing of blood meals by mosquitoes, which has been an understudied area of mosquito biology. Thus, the products of this research are revealing new physiological mechanisms used by mosquitoes—and perhaps other blood-feeding insects—that allow them to efficiently metabolize the large blood meals that they ingest. Efforts on the project have identified nearly 2,000 genes in the 'kidneys' (i.e., Malpighian tubules) of mosquitoes whose expression levels are significantly affected during the processing of blood meals. These genes are now being prioritized as targets for insecticides that may limit the number of blood meals mosquitoes can ingest during their lifetime, thereby slowing the transmission of mosquito-borne pathogens. Furthermore, in a collaborative effort with Vanderbilt University and Cornell University, this project is discovering chemicals that kill mosquitoes by disrupting Malpighian tubule function. We have discovered chemicals that interfere with the function of potassium channels in mosquito Malpighian tubules; the chemicals compromise the abilities of mosquitoes to excrete urine and fly. With this important proof-of-concept, we have now identified similar chemicals that show a high potency for perturbing the potassium channels of mosquitoes, but not those of humans. These chemicals are being further developed and will provide the basis for the development of novel, safe insecticides

Publications

  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Raphemot R, Estevez-Lao TY, Rouhier MF, Piermarini PM, Denton JS, Hillyer JF, "Molecular and functional characterization of Anopheles gambiae inward rectifier potassium (Kir1) channels: A novel role in egg production". Insect Biochem Mol Biol. Vol. 51, 10-19. 2014.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Esquivel CJ, Cassone BJ, Piermarini PM, "Transcriptomic Evidence for a Dramatic Functional Transition of the Malpighian Tubules after a Blood Meal in the Asian Tiger Mosquito Aedes albopictus". PLoS Negl Trop Dis. Vol. 8, no. 6: e2929. 2014.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Rouhier MF, Raphemot R, Denton JS, Piermarini PM, "Pharmacological Validation of an Inward- Rectifier Potassium (Kir) Channel as an Insecticide Target in the Yellow Fever Mosquito Aedes aegypti". PLOS ONE. Vol. 9, no. 6: e100700. 2014.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Rouhier MF, Piermarini PM, "Identification of life-stage and tissue-specific splice variants of an inward rectifying potassium (Kir) channel in the yellow fever mosquito Aedes aegypti". Insect Biochem Mol Biol. Vol. 48, 91-99. 2014.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Zhang Q, Piermarini PM, Nachman RJ, Denlinger DL, "Molecular identification and expression analysis of a diapause hormone receptor in the corn earworm, Helicoverpa zea". Peptides. Vol. 53, 250-257. 2014.
  • Type: Journal Articles Status: Published Year Published: 2014 Citation: Piermarini PM, Calkins TL, "Evidence for intercellular communication in mosquito renal tubules: A putative role of gap junctions in coordinating and regulating the rapid diuretic effects of neuropeptides". Gen Comp Endocrinol Vol 203: 43-48. 2014
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Akuma DC, Piermarini PM, Gillen CM. "Tissue and developmental expression of three putative Na-coupled cation-chloride cotransporters in Aedes aegypti" In: Integr Comp Biol. Vol. 54. ( 2014 ): e235. ( Published ) http://www.sicb.org/meetings/2014/SICB2014AbstractBook.pdf
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Raphemot R, Days E, Estevez-Lao TY, Rouhier MF, Hilyer JF, Hopkins CR, Piermarini PM, Denton JS. "Development of pharmacological and genetic tools to explore the potential of inward rectifier potassium channels as novel insecticide targets" In: FASEB Journal. Vol. 28. ( 2014 ): 654.9. ( Published
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Raphemot R, Days E, Estevez-Lao TY, Swale D, Rouhier MF, Hilyer JF, Weaver CD, Hopkins CR, Beyenbach KW, Piermarini PM, Denton JS. "Reverse-genetic and pharmacologic strategies for probing mosquito inwad rectifier potassium channels as insecticide targets" In: Am J Trop Med Hyg. Vol. 89. ( 2013 ): 337. ( Published )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Piermarini PM, Presenter. "Excellence in Vector-Transmitted Diseases in Managed and Natural Ecosystems." Presented at PHPID Annual Member Meeting. Columbus, Ohio, United States. ( 2014 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Piermarini PM, Presenter. "Intercellular regulation of renal function in mosquitoes: Making connections with innexins." Presented at Entomological Society of America Annual Meeting. Austin, Texas, United States. ( 2013 ) http://www.entsoc.org/entomology2013
  • Type: Other Status: Other Year Published: 2014 Citation: Piermarini PM, Seminar Presenter. "The molecular mechanisms of 'kidney' function in mosquitoes: towards novel vector control strategies." Presented at Instituto de Biotechnologia y Ecologia Aplicada Research Seminar. Xalapa, Mexico. ( 2014 )Invited departmental seminar.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Calkins TL, Piermarini PM, Presenter. "Effects of gap junction inhibitors on the survival and diuretic capacity of adult female mosquitoes, Aedes aegypti." Presented at Entomological Society of America North Central Branch Annual Meeting. Des Moines, Iowa, United States. ( 2014 ) http://www.entsoc.org/PDF/NCB/2014NCBProgramBook.pdf [
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Piermarini PM, Presenter. "Eliciting renal failure in mosquitoes." Presented at Southern Salt, Water, and Kidney Club. Sarasota, Florida, United States. ( 2013 ) http://www.sswkc.org/Home/2013-program
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Calkins TL and Piermarini PM, Presenter. "Gap junctions in the yellow fever mosquito, Aedes aegypti." Presented at Ohio Valley Entomological Association Annual Meeting. Indianapolis, Indiana, United States. ( 2013 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Esquivel CJ, Cassone, BJ, and Piermarini PM, Presenter. "A blood meal elicits global, time-dependent changes in gene expression in the renal (Malpighian) tubules of the Asian tiger mosquito Aedes albopictus." Presented at Ohio Valley Entomological Association Annual Meeting. Indianapolis, Indiana, United States. ( 2013 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Yang L, Denlinger D, and Piermarini PM, Presenter. "Effects of diapause on the excretory physiology of the northern house mosquito, Culex pipiens." Presented at Ohio Valley Entomological Association Annual Meeting. Indianapolis, Indiana, United States. ( 2013 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2014 Citation: Esquivel CJ, Cassone BJ, Piermarni PM, Poster Presenter. "Transcriptomic evidence for dramatic functional transition of the Malpighian Tubules after a blood meal in the Asian tiger mosquito Aedes albopictus." Presented at 2014 OARDC Annual Research Conference: From Biology to Business -The Transformation Power of Big Data. Wooster, Ohio, United States. ( 2014 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Calkins TL and Piermarini PM, Poster Presenter. "Gap junctions in the yellow fever mosquito, Aedes aegypti." Presented at Entomological Society of America Annual Meeting. Austin, Texas, United States. ( 2013 ) http://www.entsoc.org/entomology2013
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Piermarini PM, Rouhier MF, Hine RM, Beyenbach KW, Raphemot R, Denton JS, Presenter. "Eliciting renal failure in mosquitoes." Presented at Entomological Society of America Annual Meeting. Austin, Texas, United States. ( 2013 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Price DP, Piermarini P, Drake LL, Rodriguez SD, Drumm H, Aguirre SE, Hansen IA, Poster Presenter. "RNA-seq comparison of larval and adult malpighian tubules of Aedes aegypt, the yellow fever mosquito." Presented at Entomological Society of America Annual Meeting. Austin, Texas, United States. ( 2013 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Rouhier MF, Piermarini PM, Poster Presenter. "Tissue and life stage specific alternative splicing of a Kir channel in mosquitoes." Presented at Entomological Society of America Annual Meeting. Austin, Texas, United States. ( 2013 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Swale DR, Raphemot R, Piermarini P, Denton JS, Presenter. "Development of insecticides targeting inward rectifying potassium channels in the malaria vector, Anopheles gambiae." Presented at Entomological Society of America Annual Meeting. Austin, Texas, United States. ( 2013 ) http://www.entsoc.org/entomology2013
  • Type: Journal Articles Status: Under Review Year Published: 2014 Citation: Calkins TL, Hildebrandt O, Acevedo M, Piermarini PM. "The molecular expression of genes encoding gap junctions (innexins) in the yellow fever mosquito, Aedes aegypti: insights into putative life stage- and tissue-specific functions of innexins in mosquitoes." submitted to: Comp Biochem Physiol B. 2014.


Progress 01/01/13 to 09/30/13

Outputs
Target Audience: Target audiences include basic scientists, medical and veterinary professionals, and vector-management specialists who are faced with the challenge of controlling disease-spreading mosquitoes that are resistant to traditional insecticides. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? The project has providing training and professional development opportunities for the individuals listed below who are trainees of the principal investigator (Dr. Peter M. Piermarini): Postdoctoral researcher: Dr. Matthew F. Rouhier Doctoral student: Ms. Liu Yang Masters students: Mr. Travis L. Calkins; Mr. Carlos Esquivel Undergraduate students: Ms. Sheila Amoko, Mr. Oliver Hildebrandt Visiting Scholars: Ms. Sonja Dunemann, Ms. Karolin Kramer, Mr. Carlos Esquivel Research assistant: Ms. Nuris Acosta How have the results been disseminated to communities of interest? Results of our work have been disseminated through peer-reviewed journals, invited seminars at other universities, and invited and submitted presentations at local, regional, national, and international meetings. What do you plan to do during the next reporting period to accomplish the goals? We plan to continue our investigations into which genes expressed in the Malpighian tubules of mosquitoes play key roles in the processing of blood meals by interfering with the expression of those genes whose expression is more abundant after a blood meal using RNA interference. We predict that perturbing the expression of these genes will make a blood meal lethal to mosquitoes. In addition, we plan to continue our development of new insecticides that target potassium channels in mosquitoes. With collaborators at Vanderbilt and Cornell Universities, we are in search of chemicals that will show a high potency for perturbing the potassium channels of mosquitoes, but not those of humans and other animals. Such chemicals may provide the basis for the development of novel, safe insecticides.

Impacts
What was accomplished under these goals? Mosquito-borne diseases (e.g., malaria, dengue fever, West Nile disease) are an emerging threat to the health and well-being of humans. Each year, such mosquito-borne diseases take an immense toll on global health. Hundreds of millions of people around the globe are infected with these diseases each year, which claim nearly one million lives. Because mosquitoes are rapidly evolving resistance to conventional insecticides, new insecticides are urgently needed to expand our chemical arsenal for controlling mosquitoes and slowing the spread of mosquito-borne diseases. This project is revealing novel molecular targets and chemicals to guide the development of a new generation of insecticides that target the ‘kidneys’ of mosquitoes for combatting insecticide resistant mosquitoes. The products of this research have the potential to slow the spread of mosquito-borne diseases, such as malaria, dengue fever, and West Nile fever, which collectively debilitate the health and well-being of hundreds of millions of people around the globe each year. This project is also providing new molecular insights into the contributions that the mosquito ‘kidney’ makes to the physiological processing of blood meals by mosquitoes, which has been an understudied area of mosquito biology. Thus, the products of this research are revealing new physiological mechanisms used by mosquitoes—and perhaps other blood-feeding insects—that allow them to efficiently metabolize the large blood meals that they ingest. Efforts on the project have identified nearly 2,000 genes in the ‘kidneys’ (i.e., Malpighian tubules) of mosquitoes whose expression levels are significantly affected during the processing of blood meals. These genes are now being prioritized as targets for insecticides that may limit the number of blood meals mosquitoes can ingest during their lifetime, thereby slowing the transmission of mosquito-borne pathogens. Furthermore, in a collaborative effort with Vanderbilt University and Cornell University, this project is discovering chemicals that kill mosquitoes by disrupting Malpighian tubule function. We have discovered a chemical that interferes with the function of potassium channels in mosquito Malpighian tubules; the chemical compromises the abilities of mosquitoes to excrete urine and fly. With this important proof-of-concept, the team is now in search of similar chemicals that will show a high potency for perturbing the potassium channels of mosquitoes, but not those of humans and other animals. These chemicals will provide the basis for the development of novel, safe insecticides.

Publications

  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Mamidala, P., Mittapelly, P., Jones, S. C., Piermarini, P. M., Mittapalli, O., "Molecular characterization of genes encoding inward rectifier potassium (Kir) channels in the bed bug (Cimex lectularius)". Comp Biochem Physiol B Biochem Mol Biol. Vol. 164, no. 4: 275-279. 2013.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Raphemot R, Rouhier MF, Hopkins CR, Gogliotti RD, Lovell KM, Hine RM Ghosalkar D, Longo A, Beyenbach KW, Denton JS, Piermarini PM, "Eliciting renal failure in mosquitoes with a small-molecule inhibitor of inward-rectifying potassium channels". PLoS ONE. Vol. 8, no. 5: e64905. 2013.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Piermarini, PM. "Cellular and molecular mechanisms of diuresis in mosquitoes." Presented at North American Society for Comparative Endocrinology. Queretaro, Mexico. (2013 )Invited international talk.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Piermarini, Peter M., Rouhier, Matthew F., Schepel, Matthew, Kosse, Christin, Beyenbach, Klaus W., "Cloning and functional characterization of inward-rectifying potassium (Kir) channels from Malpighian tubules of the mosquito Aedes aegypti". Insect Biochem Mol Biol. Vol. 43, no. 1: 75-90. 2013.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Miyauchi, Jeremy T., Piermarini, Peter M., Yang, Jason D., Gilligan, Diana M., Beyenbach, Klaus W., "Roles of PKC and phospho?adducin in transepithelial fluid secretion by Malpighian tubules of the yellow fever mosquito". Tissue Barriers. Vol. 1, no. 1: 7-6. 2013.
  • Type: Other Status: Other Year Published: 2013 Citation: Piermarini PM. "High-throughput discovery of chemicals that induce kidney failure in the malarial vector Anopheles gambiae." Presented at Innovative Vector Control Consortium ESAC-1 Meeting. Liverpool, United Kingdom. ( 2013 ) Presentation to the external scientific advisory committee-1 (ESAC-1) of the Innovative Vector Control Consortium.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Rouhier MF, Denton JS, Beyenbach KW, and Piermarini PM. "Molecular and functional characterization of inward-rectifying potassium (Kir) channels in the kidneys of mosquitoes: towards the development of new insecticides." Presented at Public Health Preparedness for Infectious Diseases (PHPID) program annual conference. Columubs, Ohio, United States. ( 2013 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Esquivel CJ, Cassone, BJ, and Piermarini PM. "A blood meal elicits global, time-dependent changes in gene expression in the renal (Malpighian) tubules of the Asian tiger mosquito Aedes albopictus." Presented at Public Health Preparedness for Infectious Diseases (PHPID) program annual conference. Columbus, Ohio, United States. ( 2013
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Yang L, Denlinger D, and Piermarini PM. "Effects of diapause on the excretory physiology of the northern house mosquito, Culex pipiens." Presented at North Central Branch of the Entomological Society of America Annual Meeting. Rapid City, South Dakota, United States. ( 2013 )
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Calkins TL and Piermarini PM. "Expression of transcripts encoding gap junctional proteins (innexins) in the yellow fever mosquito, Aedes aegypti." Presented at North Central Branch of the Entomological Society of America Annual Meeting. Rapid City, South Dakota, United States. (2013)
  • Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Yang L, Denlinger D, and Piermarini PM, Poster Presenter. "Effects of diapause on the excretory physiology of the northern house mosquito, Culex pipiens." Presented at the 6th International Congress of Society for Vector Ecology, La Quinta, CA. (2013)


Progress 01/01/12 to 12/31/12

Outputs
OUTPUTS: Results of our work have been disseminated through peer-reviewed journals, invited seminars at the Ohio State University and other universities, and invited and submitted presentations at local, regional, and national meetings. PARTICIPANTS: Members of the Piermarini laboratory that have contributed to the outputs include Dr. Matthew Rouhier (postdoctoral researcher), Ms. Nuris Acosta (part-time technician), Mr. Travis Calkins (full-time technician), Ms. Sonja Dunemann (visiting scholar from Germany), and Mr. Carlos Esquivel (visiting scholar from Guatemala). Additional participants include Dr. Jerod S. Denton, Mr. Rene Raphemot, and Dr. Corey Hopkins who are collaborators from Vanderbilt University, and Dr. Klaus W. Beyenbach and Ms. Rebecca Hine who are collaborators from Cornell University. TARGET AUDIENCES: Target audiences include basic scientists, medical and veterinary professionals, and vector-management specialists who are faced with the challenge of controlling disease-spreading mosquitoes that are resistant to traditional insecticides. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
This project focuses on developing a new generation of insecticides that will kill mosquitoes by attacking their analog of a kidney--i.e., the Malpighian tubules. This year we have commenced efforts to identify genes that play key roles in the renal functions of adult female mosquitoes (the blood-feeding life stage), and 2) discover chemical compounds that kill mosquitoes by impairing their renal functions. We are presently employing next-generation sequencing to identify genes that are expressed differentially in mosquito Malpighian tubules during the processing of a blood meal. Furthermore, using traditional RT-PCR we identified several genes of the innexin (gap junction) family that are expressed in Malpighian tubules; we are presently evaluating these genes as potential insecticidal targets. Our largest impact has been made in a collaborative project with researchers at Vanderbilt and Cornell Universities in which we have demonstrated proof-of-concept that disrupting Malpighian tubule function is a viable new approach for killing mosquitoes. In brief, we targeted a family of mosquito potassium channels that are expressed in Malpighian tubules. Using high-throughput drug discovery, we identified a small-molecule inhibitor of the mosquito potassium channels and showed that this molecule kills mosquitoes by disrupting their excretory physiology (i.e., it causes 'kidney' failure). We are presently screening for additional small-molecule inhibitors of potassium channels that are highly selective and potent for the mosquito channels vs. mammalian orthologs; once discovered we will attempt to develop these small molecules as new insecticides for combatting mosquitoes that are resistant to traditional control agents, such as pyrethroids and DDT.

Publications

  • Linser PJ, Neira Oviedo M, Hirata T, Seron TJ, Smith KE, Piermarini PM, and Romero MF (2012) Slc4-like anion transporters of the larval mosquito alimentary canal. J Insect Physiol 58: 551-562.
  • Blumenthal EM, and Piermarini PM (2012) Introduction to the special issue on molecular physiology of epithelial transport in insects - A tribute to William R. Harvey. J Insect Physiol 58: 427.