Progress 08/01/09 to 07/31/14
Outputs
Target Audience: Fellow scientists, policy makers, veterinarians, food-safety agencies, animal industry, pest control agencies, farmers and ranchers, funding agencies, dog owners, preventive medicine, public health organizations. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? Zurek: I have trained 2 Ph.D students and 1 MS, 1 postdoctoral research associate, and 2 undrgraduate students. I have taught Animal health entomology with enrolment of 70 students. Ortigao: I have taught ENTOM930 -Molecular Biology Lab (2 credits) - module "Working with RNA" and lectured in ENTOM 830 Molecular Entomology (Fall 2014). I have also served as a graduate student advisor for 2 students (Matt Heerman - PhD Entomology at KSU 2015, Maricela Robles-Murguia - MS Entomology at KSU Dec 2013 (left KSU on March 2014). I have served as a co-major advisor - 4 students (Cesar Raimundo Lima Costa Júnior - PhD in Genetics, Federal University of Pernambuco - UFPE 2015; Juliana Malta - PhD, Federal University of Vicosa (UFV) 2016; Tales Pascini - PhD, UFV 2017; Glenda Sena - MS, Federal University of Amazonas (UFAM-INPA) 2014. I have been on the advisory committee for 4 students (Kenner Morais - PhD, UFV 2014; Tales Pascini - MS, UFV 2014; Mustapha Soumalia - MS Entomology at KSU 2014; Donghun (Andy) Kim - PhD Entomology at KSU 2014). How have the results been disseminated to communities of interest? The MoU between K-State and the National Research Institute of the Amazon (INPA) set up in 2013 was finally signed into agreement in 2014, benefiting faculty and students engaged in studies of the Amazon Region in Brazil. I have represented the Entomology Department during visit to the Federal University of Vicosa in July-August 2014 and also provided guidance on sand fly rearing techniques to Genesis Lab, Wellington, CO. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
We hypothesized that as the microbial community of animal manure changes over time, it plays an important role in stable fly fitness. Two-choice bioassays were conducted using two week old horse manure (control) and aging horse manure (fresh to 5 week old) to evaluate the effect of manure age on stable fly oviposition. Our data showed that fresh feces did not stimulate oviposition and that the attractiveness increased as manure aged but started to decline after 3 weeks. Bioassays assessing the effect of manure age at the time of oviposition on larval development demonstrated that 1 to 3 week old manure supported larval development significantly better than fresh, 4, and 5 week old manure. In addition, adult fitness (body size) was significantly higher in flies from 1 and 2 week old manure comparing to that of all other treatments. It is likely that stable flies use bacteria-derived cues emitted from fresh manure to avoid competition with other insects and 4 ≥ week old manure is depleted of nutrients and microorganisms necessary for oviposition and larval development. Analysis of the bacterial community of aging horse manure by 454-pyrosequencing of 16S rDNA revealed a great reduction in bacterial diversity and richness from fresh to 1-5 week old manure and a major shift from strict anaerobes in fresh manure to facultative anaerobes and strict aerobes in aged manure. Overall, the microbial community of 2 and 3 week old horse manure with its dominant bacterial taxa Rhizobium, Devosia, and Brevudiomonas stimulated stable fly oviposition the most and provided a suitable habitat for larval development. These bacteria represent the candidates for studies focused on better understanding of stable fly - microbial interactions. We also used several measures to test the hypothesis that house flies, Musca domestica (L.), acquire and disseminate antibiotic resistant and potentially virulent E. faecalis from wastewater treatment facilities (WWTF) to the surrounding urban environment. House flies and sludge from four WWTF (1-4) as well as house flies from three urban sites close to WWTF-1 were collected and cultured for enterococci. Enterococci were identified, quantified, screened for antibiotic resistance and virulence traits, and assessed for clonality. Of the eleven antibiotics tested, E. faecalis was most commonly resistant to tetracycline, doxycycline, streptomycin, gentamicin and erythromycin and these traits were intraspecies horizontally transferrable by in vitro conjugation. Profiles of E. faecalis (prevalence, antibiotic resistance, and virulence traits) from each of WWTF sludge and associated house flies were similar, indicating that flies successfully acquired these bacteria from this substrate. The greatest number of E. faecalis with antibiotic resistance and virulence factors (i.e., gelatinase, cytolysin, enterococcus surface protein, and aggregation substance) originated from WWTF-1 that processed meat waste from a nearby commercial meat processing plant, suggesting an agricultural rather than human clinical source of these isolates. E. faecalis from house flies collected from three sites 0.7 - 1.5 km away from WWTF-1 were also similar in their antibiotic resistance profiles; however, antibiotic resistance was significantly less frequent. Clonal diversity assessment using pulsed-field gel electrophoresis revealed the same clones of E. faecalis from sludge and house flies from WWTF-1 but not from the three urban sites close to WWTF-1. This study demonstrates that house flies acquire antibiotic resistant enterococci from WWTF and potentially disseminate them to the surrounding environment Sand fly colonies have been established. We developed infection models for studies of sand fly-Leishmania interactions and produced antisera used in immunological and molecular approaches. RNAi platform for studies on reverse genetics has been developed. We also characterized the role of PM-associated proteins and Leishmnaia development and developed paratransgenic platform. Several aspects of innate immunity in sand flies that can be used for insect control have been characterized.The collaboration with investigators at the University of New Mexico led to new findings regarding how bacteria colonizes the sand fly gut and induce differential immune responses. We have continued to explore the development of a paratransgenic platform to control sand flies, and aspects of this approach were detailed on a book chapter published. We have submitted grant applications to Gates Foundation (pre-proposal) and NIH ICIDR (international collaboration of infectious disease research). I also have obtained a very competitive grant from the Brazilian Government under the program "Sciences without Borders" allowing the continuation of some of my international program in Brazil, at UFV in Vicosa working with Dr G Martins on studies focused on physiology of mosquitoes and sand flies (see publications). We continued our work with investigators from the National Research Institute of the Amazon (INPA) and published results of one study on the response of human patients to Leishmania infection bringing new insights into the immunity responses to cutaneous leishmaniasis. I also continued to host international researchers in my lab, and in 2014 there were five students and investigators who visited my lab. My international program is relevant to the K-State 2025 goals. From the studies I have maintained, several peer reviewed publications were generated in 2013; we are currently working on finalizing an additional four publications also related to those studies (Ortigao).
Publications
- Type:
Journal Articles
Status:
Under Review
Year Published:
2014
Citation:
Schaut RG, Robles-Murguia M, Juelsgaard R, Esch KJ, Bartholomay LC, Ramalho-Ortigao M, Petersen CA (2014)
Vertically-transmitted Leishmania infantum from North American dogs maintains ability for vector-borne transmission in
Lutzomyia spp. Emerging Infectious Diseases, In review.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Robles-Murguia M, Bloedow N, Murray, L, Ramalho-Ortigao, M (2014) Effect of mouse antisera targeting the Phlebotomus
papatasi midgut chitinase PpChit1 on sand fly physiology and fitness. Mem Inst Oswaldo Cruz, 109:1-6 DOI:
10.1590/0074-0276140382
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Espir TT, Figueira LP, Naiff MF, Costa A, Ramalho-Ortigao M, Malheiro A, Franco A (2014) The role of inflammatory, antiinflammatory
and regulatory cytokines in patients infected with cutaneous leishmaniasis in Amazonas State, Brazil. J
Immunol Res, Volume 2014, Article ID 481750 http://dx.doi.org/10.1155/2014/481750
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Geraci NS, Mukbel RM, Kemp MT, Wadsworth MN, Lesho E, Stayback, GM Champion MM, Bernard MA, Abo-Shehada
M, Coutinho-Abreu IV, Ramalho-Ortig�o M, Hanafi, HA, Fawaz EY, El-Hossary SS, Wortmann G, Hoel DF, McDowell MA.
(2014) Profiling of human acquired immunity against the salivary proteins of Phlebotomus papatasi reveals clusters of
differential immunoreactivity. Am J Trop Med Hyg. 90(5):923-38. doi: 10.4269/ajtmh.13-0130PMID:24615125
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Assis WA, Malta J, Pimenta PF, Ramalho-Ortig�o JM, Martins GF (2014) The characterization of the fat bodies and
oenocytes in the adult females of the sand fly vectors Lutzomyia longipalpis and Phlebotomus papatasi. Arthropod Struct
Dev. May 23. pii: S1467-8039(14)00042-5. doi: 10.1016/j.asd.2014.05.002.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Ghosh A., M. Akhtar, C. Holderman, and L. Zurek* (2014). Significance and survival of enterococci during the house fly
development. Journal of Medical Entomology 51: 63-67.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Doud C.W., H.M. Scott, and L. Zurek* (2014). Role of house flies in the ecology of Enterococcus faecalis from wastewater
treatment facilities. Microbial Ecology 67:380-391
|
Progress 01/01/13 to 09/30/13
Outputs
Target Audience: Fellow scientists, policy makers, veterinarians, food-safety agencies, animal industry, pest control agencies, farmers and ranchers, funding agencies, dog owners, preventive medicine, public health officials, members of the ASTMH, CDC, vector biology students and scientists, individuals living in endemic areas of transmission of leishmaniasis. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? Sand fly Genome Steering Committee BEI Resources Review Committee for Sand Fly Research Member of One Health Kansas to provide educational opportunities for K-12 students, the general public, undergraduate and graduate students, and professionals in areas of my expertise Set up new Memorandum of Understanding (MoU) between K-State and the National Research Institute of the Amazon (INPA). This new MoU will benefit faculty and students engaged in studies focused on biology of the Amazon Region in Brazil. In 2013 I began new collaborative research and student mentoring at INPA. Represented the Entomology Department during visit to the Federal University of Vicosa in August 2013. Established new connections with University of Sao Paulo – ESALQ Provided expert comments for The Scientists News and Opinions column by Sabrina Richards on the ability of GM modified insects to stop disease (http://www.the-scientist.com/?articles.view/articleNo/34005/title/Will-GM-Insects-Help-Stop-Disease-/) Provided guidance on sand fly rearing techniques to Genesis Lab, Wellington, CO Provided expertise on sand fly biology to Dr Rita Mukhopadhyay at Florida International University. How have the results been disseminated to communities of interest? PI, graduate students, undergraduate students, and postodoctoral reserach associates in of both laboratories have given a total 27 presentations at local, national and international scientirfic meetings. What do you plan to do during the next reporting period to accomplish the goals? We will continue investigating the role of muscoid flies in the ecology of 8 serotypes of Shiga-toxigenic Escherichia coli (STEC) by conducting laboratory bioassays as well as field studies in cattle productions (feedlots and dairies). (Zurek) We plan to submit a manuscript on the microbial ecology of stable flies (role of bacteria in oviposition, larval development, and fitness of stable flies developing in horse manure). (Zurek) I plan to continue the collaborations I have established in the last fewyearsthat will permit the work outlined to be completed successfully. I alsoplan to secure funding from various agencies, such as NIH, NSF, and others. I also plan the hiring of additionalPhD students whowill also add to the program. (Ortigao)
Impacts
What was accomplished under these goals?
The results showed that fresh manure did not stimulate stable fly oviposition and that the attractiveness increased as manure aged but started to decline after 3 weeks. Stable fly eggs artificially placed on 1, 2, and 3 week old manure resulted in significantly higher survival and heavier adults comparing to those developing in fresh, 4, and 5 week old manure. Analysis of the bacterial community of aging horse manure by 454-pyrosequencing of 16S rDNA revealed a major shift from strict anaerobes (e.g. Clostridium, Eubacterium, Prevotella, Bacteroidales) in fresh manure to facultative anaerobes and strict aerobes (e.g. Rhizobium, Devosia, Brevundimonas, Sphingopyxis, Comamonas, Pseudomonas) in 1-5 week old manure. Identified volatile compounds emitted from 2 and 3 week old horse manure included phenol, indole, p-cresol, and m-cresol. However, none of them stimulated stable fly oviposition in two-choice assays. In conclusion, the microbial community of 2 and 3 week old horse manure stimulates stable fly oviposition and provides a suitable habitat for stable fly development. Manure at this stage should be the main target for disrupting the stable fly life cycle. Volatile compounds acting as oviposition stimulants/attractants and their specific bacterial origin remain to be determined. Better understanding of stable fly microbial ecology is critical for development of novel management strategies based on alteration of the microbial community of stable fly habitat to generate a substrate that is non-conducive to fly oviposition and/or larval development. (Zurek) House flies and sludge from four WWTF (1-4) as well as house flies from three urban sites close to WWTF-1 were collected and cultured for enterococci. Enterococci were identified, quantified, screened for antibiotic resistance and virulence traits, and assessed for clonality. Of the eleven antibiotics tested, E. faecalis was most commonly resistant to tetracycline, doxycycline, streptomycin, gentamicin and erythromycin and these traits were intra-species horizontally transferrable by in vitro conjugation. Profiles of E. faecalis (prevalence, antibiotic resistance, and virulence traits) from each of WWTF sludge and associated house flies were similar, indicating that flies successfully acquired these bacteria from this substrate. The greatest number of E. faecalis with antibiotic resistance and virulence factors (i.e., gelatinase, cytolysin, enterococcus surface protein, and aggregation substance) originated from WWTF-1 that processed meat waste from a nearby commercial meat processing plant, suggesting an agricultural rather than human clinical source of these isolates. E. faecalis from house flies collected from three sites 0.7 - 1.5 km away from WWTF-1 were also similar in their antibiotic resistance profiles; however, antibiotic resistance was significantly less frequent. Clonal diversity assessment using pulsed-field gel electrophoresis revealed the same clones of E. faecalis from sludge and house flies from WWTF-1 but not from the three urban sites close to WWTF-1. This study demonstrates that house flies acquire antibiotic resistant enterococci from WWTF and potentially disseminate them to the surrounding environment. (Zurek) The collaboration with Ravi Durvasula and Ivy Horwitz (University of New Mexico) led to new research focused on innate immunity in sand fly larvae, and new findings regarding how the sand fly larvae responds to infection with different bacterial species. We have also continued to explore the possibility of developing a paratransgenesis platform to control sand flies by using a novel fluorescent-labeled single chain antibody to target docking of the parasites to the sand fly midgut. Data regarding these studies were presented at WorldLeish 5 in Brazil, and the ASTMH meeting in Washington DC. We are currently working on a grant application to NIH within the ICIDR (International Collaboration of Infection Diseases Research) and on several manuscripts. A new collaboration was started with Dr. Rita Mukhopadhyay (Florida International University) involving RNAi knock-down using the sand fly model to determine how aquaporins may interfere with host-seeking behavior (a new NIH proposal is being submitted in 2014 to support these studies and my role is that of collaborator). Regarding my international collaborations: at UFV, Vicosa (work with Dr G Martins), our studies focused on many aspects of vector biology and physiology of mosquitoes and sand flies (see publications), and led to a grant submitted and approved by the Brazilian funding agency CNPq for CY2014-2016; a new collaborative program was established with investigators from the National Research Institute of the Amazon (INPA) following a visit by INPA investigators to my lab in Fall of 2012 with a subsequent visit I made to INPA between April and May 2013; at FIOCRUZ, in Rio de Janeiro, we continued to work on sand fly and Leishmania interaction. My international program is relevant to the K-State 2025 goals. From the studies have maintained, several peer reviewed publications were generated in 2013; we are currently working on finalizing an additional four publications also related to those studies. (Ortigao)
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Scott J.G., C.A. Leichter, F.D.Rinkevihc, S.A Harris, C. Su, L.C. Aberegg, R. Moon, C.J. Geden, A.C. Gerry, D.B. Taylor, R.L. Byford, W. Watson, G. Johnson, and L. Zurek (2013). Insecticide resistance in house flies from the United States: Resistance levels and frequency of pyrethroid resistance alleles. Pesticide Biochemistry and Physiology 107:377-384.
Zhu J.J., B.J. Wienhold, J. Wehrle, D. Davis, H. Chen, D. Taylor, K. Friesen and L. Zurek (2013)Efficacy and longevity of newly developed catnip oil microcapsules against stable fly oviposition and larval growth. Medical and Veterinary Entomology DOI: 10.1111/mve.12029.
Doud C.W., H. M. Scott and L. Zurek (2013). Role of house flies in the ecology of Enterococcus faecalis from wastewater treatment facilities. Microbial Ecology DOI 10.1007/s 00248-013-0337-6.
Le�dido AC, Ramalho-Ortig�o M, Martins GF (2013) The ultrastructure of the Aedes aegypti heart. Arthropod Struct Dev. Nov;42(6):539-50. doi: 10.1016/j.asd.2013.09.005. Epub 2013 Oct 1. PMID: 4095854
Sigle LT, Ramalho-Ortig�o M. (2013) Kazal-type serine proteinase inhibitors in the midgut of Phlebotomus papatasi. Mem Inst Oswaldo Cruz. Sep;108(6):671-8. doi: 10.1590/0074-0276108062013001. PMID: 24037187
Pascini TV Ramalho-Ortig�o M, Martins GF. (2013) The Fine Structure of the Spermatheca in Anopheles aquasalis (Diptera: Culicidae). Ann Ent Soc of America 106(6):857-867. DOI: http://dx.doi.org/10.1603/AN13079
Coutinho-Abreu IV, Sharma NK, Robles-Murguia M, Ramalho-Ortigao M (2013) Characterization of Phlebotomus papatasi peritrophins, and the role of PpPer1 in Leishmania major survival in its natural vector. PLoS Neglected Tropical Diseases, PLoS Negl Trop Dis; 7(3):e2132. doi: 10.1371/journal.pntd.0002132. Epub 2013 Mar 14. PMID: 23516661
Abrudan J, Ramalho-Ortigao M, Lawyer P, Kamhawi S, Rowton ED, Lehane MJ, Bates PA, Valenzuela JG, Tomlinson C, Appelbaum E, Moeller D, Thiesing B, Dillon R, Clifton S, Lobo NF, Wilson RK, Collins FH, McDowell MA. (2013) The characterization of the Phlebotomus papatasi transcriptome. Insect Biochemistry and Molecular Biology, Insect Mol Biol. Apr;22(2):211-32. doi: 10.1111/imb.12015. Epub 2013 Feb 7.
|
Progress 01/01/12 to 12/31/12
Outputs
OUTPUTS: Our data have been presented at international, national, and local scientific meetings. This includes the Annual Meeting of the Society for Vector Ecology, St. Augustine, FL, Sept. 27-29, 2012, presented by C.W. Doud with a title: Role of house flies in the ecology of Enterococcus faecalis from wastewater treatment facilities; the North Central Branch Meeting of ESA, June 3-6, 2013, presented by T. Albuquerque with the title: Effect of microbial community of horse manure on stable fly (Stomoxys calcitrans) fitness; the KSU Research Forum, March 7, 2012, presented by T. Albuquerque with the title: Microbial ecology of stable flies: Effect of bacterial community of aging horse manure on stable fly oviposition and larval development; the 60th Annual ESA Meeting, Knoxville, TN, Nov. 11-14,2012, presented by L.Zurek with the title: Significance of the midgut microbial community in the vector competence of Phlebotomus duboscqi for Leishmania major; and the Annual Meeting of the Missouri Valley Branch of the American Society of Microbiology, April 12-14, 2012 in Manhattan, KS presented by Dr. A. Ghosh with the title: Pre-harvest food safety: Role of houseflies in the ecology of Escherichia coli O157:H7 in a cattle feedlot. Activities: Research proposals submitted; NIH RO1 "P. papatasi Molecules: Gene Function and TBV Candidates" ($1.2 million); NIH R21 No cost extension (NCE); NIH IRIDA "Salivary gland protein profiles of P. papatasi ($718,000); DARPA (with Tona Melgarejo, PI) proposal "Hyena Immune Defense Strategies ($2 million); Deployed Warfare Fighter Program (pre-proposal) "Hyena Defense Strategies to Combat Vector-Borne Pathogens" ($750,000); Summer undergrad student training; mentored grad students and postdocs; co-advised MS and PhD students in Brazil; faculty selection committee at UFV; collaborations in Brazil; MoU with Entomology Dept at Vicosa; MoU with Dept of Genetics, UFPE; Hosted investigators Brazil's INPA. Invited Talks: International: XVIII International Congress Trop Med and Malaria,Sept 2012,Rio de Janeiro Universidade Federal de Vicosa National: 2012 ESA meeting MUVE ESA 2012 meeting, poster "The role of midgut immunity and homeostasis in Leishmania survival, development, and transmission by the sand fly L longipalpis".Presenter Matt Heerman UTMB,Galveston, TX Biological Sciences, Pitt State University, KS CMG, College of Vet Sciences, K-State Services:Committee assignments:National Member of Sand fly Genome Steering Committee Ad Hoc reviewer for NIH Vector Biology Session Regional:Member of One Health Student committee Pitts State University Task force on university infrastructure MoU between K-State and UFV and UFPE College:College of Ag Diversity Committee Entomology Head Search Committee Department: Member (Chair)of Planning Committee Member of Seminar Committee Manuscript reviews Service to professional and scientific organizations Scientific Meeting Co-organizer of ENTOMOL5, Recife, Brazil Committees:Voting member of American Committee on Medical Entomology (ACME) Voting member of the ESA-MUVE Selection committee for faculty at UFV Sand Fly Genome Steering Committee Editor or editorial boards Guest editor J Trop Medicine PARTICIPANTS: Graduate students: Thais Albuquerque , Carl Doud, Postdoctoral fellows: Anuradha Ghosh, Kamila Koci Collaborators: Jerry Zhu and D. Taylor (USDA, Lincoln, NE), Marcelo Ortigao (KSU Entomology) TARGET AUDIENCES: Pest control operators, medical and veterinary entomologists, farmers and ranchers, microbial ecologists, general public, military. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Species diversity and seasonal abundance of muscoid flies (Diptera: Muscidae) developing in biosolid cake (dewatered biosolids) stored at a wastewater treatment facility in northeastern Kansas were evaluated. Emergence traps were deployed for 22 weeks. In total, 11,349 muscoid flies were collected emerging from the biosolid cake. Stable flies (Stomoxys calcitrans (L.) and house flies (Musca domestica (L.), represented 80 and 18% of the muscoid flies, respectively. An estimated 550 stable flies and 220 house flies per square-meter of surface area developed in the biosolid cake annually producing 450,000 stable flies and 175,000 house flies. Stable fly emergence was seasonally bimodal with a primary peak in mid-July and a secondary peak in late August. House fly emergence peaked with the first stable fly emergence peak and then declined gradually for the remainder of the year. House flies tended to emerge from the biosolid cake sooner after its deposition than did stable flies. In addition, house fly emergence was concentrated around midsummer whereas stable fly emergence began earlier in the spring and continued later into the fall. Biosolid age and temperature were the most important parameters affecting emergence for house flies and stable flies, whereas precipitation was not important for either species. This study highlights the importance of biosolid cake as a larval developmental habitat for stable flies and house flies. Objective 3 Milestone experiments and data have been obtained and published. Among critical milestones - Working and maintaining sand fly colonies. - Maintenance of sand flies and Leishmania cell lines in vitro - Infection of sand flies - Antisera production - RNAi-based silencing and effects of Leishmania development - Expression analyses of sand fly midgut and salivary gland genes - Expression and purification of recombinant proteins Objective 4 Our studies led to report of sand flies in Kansas for the first time and to new research towards development of novel treatment against canine visceral leishmaniasis.
Publications
- Doud C.W., D.B. Taylor and L. Zurek (2012). Dewatered sewage biosolids provide a productive larval habitat for stable flies and house flies (Diptera: Muscidae). Journal of Medical Entomology 49:286-292.
- Peterkova-Koci K., M.Robles-Murguia, M. Ramalho-Ortigao, L. Zurek (2012). Significance of bacteria in oviposition and larval development of the sand fly Lutzomyia longipalpis. Parasites & Vectors 5:145.
- Abrudan J, Ramalho-Ortigao M, Lawyer P, Kamhawi S, Rowton ED, Lehane MJ, Bates PA, Valenzuela JG, Tomlinson C, Appelbaum E, Moeller D, Thiesing B, Dillon R, Clifton S, Lobo NF, Wilson RK, Collins FH, McDowell MA (2012). The characterization of the Phlebotomus papatasi transcriptome. Insect Biochemistry and Molecular Biology, in print.
- Coutinho-Abreu IV, Sharma NK, Robles-Murguia M, Ramalho-Ortigao M (2012) Characterization of Phlebotomus papatasi peritrophins, and the role of PpPer1 in Leishmania major survival in its natural vector. PLoS Neg Trop Dis
- Martins GF, Ramalho-Ortigao JM (2012). Oenocytes in insects. Invertebrate Survival Journal 9: 139-152
- Weng J-L, Young SL, Gordon DM, Claborn D, Petersen C, Ramalho-Ortigao M (2012) First report of Phlebotomine sand flies (Diptera: Psychodidae) in Kansas and Missouri, and a PCR method to distinguish Lutzomyia shannoni from Lutzomyia vexator. Journal of Medical Entomology 49(6): 1460-1465.
- Peterkova-Koci K, Robles-Murguia M, Ramalho-Ortigao M, Zurek L (2012) Significance of bacteria in oviposition and larval development of the sand fly Lutzomyia longipalpis. Parasites and Vectors 24, 5:145.
- Pascini TV, Ramalho-Ortigao M, Martins (2012) GF Morphological and morphometrical assessment of spermathecae of Aedes aegypti females. Mem Institute Oswaldo Cruz 107(6):705-12
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Progress 01/01/11 to 12/31/11
Outputs
OUTPUTS: Our data have been presented at international, national and local scientific meetings. This includes 55th Annual Meeting of Livestock Insect Workers Conference and 56th AAVP (American Association of Veterinary Parasitologists), St. Louis, MO, July 16-19, presented by R.L. Moon with a title: Winter debris-cleanup deadlines based on a ground-truthed degree-day model; Ecological Genomics Meeting, Kansas City, MO., Nov. 4-6, 2011. Presented by K. Koci K, Microbial ecology of Lutzomyia longipalpis; COBRE (Center of Biomedical Research Excellence), Epithelial Biology, Meeting, KSU, Manhattan. October 13, presented by L. Zurek with a title: Progress on the microbial ecology and immunity of sand flies; and International Symposium on Phlebotomine Sand Flies, Kusadasi, Turkey, April 25-30, 2011 presented by L. Zurek with a title: Microbial ecology and epithelial immunity of Lutzomyia longipalpis and Phlebotomus papatasi. PARTICIPANTS: Graduate students: Thais Albuquerque , Carl Doud, Postdoctoral fellows: Anuradha Ghosh, Kamila Koci Collaborators: Roger Moon (collaborator, University of Minnesota), Jerry Zhu (USDA, Lincoln, NE), Marcelo Ortigao (KSU Entomology) TARGET AUDIENCES: Pest control operators, medical and veterinary entomologists, farmers and ranchers, microbial ecologists, general public, military. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Stable flies are perennial pests of confined and grazing cattle and horses throughout the world. In northern temperate latitudes, attack rates increase from zero in winter to peaks in June-September, depending on location and weather. Accumulating evidence indicates the spring increase is from larvae that develop in urine- and manure-soiled animal bedding and winter feeding debris. If this hypothesis is true, then spring cleanups could help minimize summer stable fly populations, provided debris is destroyed in a timely manner, before adults emerge. Laboratory studies demonstrate development time from egg to adult depends on incubation temperature, and mathematical models can be used to predict field development times from matching field temperature records. We conducted a field study to measure field development times, by planting cohorts of stable fly eggs in field containers of artificial medium. Observed times from egg to 50% emergence ranged from 2 to 8 weeks, depending on starting date and geographic location. In turn, the field times and matching min-max NOAA weather records were used to derive a best-fit degree-day model to predict development times for hypothetical cohorts starting on an arbitrary date at an arbitrary location. Model parameters were a base temperature of 3.7 degrees C, a ceiling temperature of 32 degrees D, and a cumulative 369 degree-days. We then used the model and the phenology modeling package NAPPFAST to predict and map emergence dates for hypothetical cohorts of flies originating as eggs on 1 January, and mapped emergence dates using historical weather data. Maps will be presented to summarize first emergence dates using national weather data from individual years and historical averages. Maps of weekly isopleths can be used to help producers schedule spring breeding site cleanup operations.
Publications
- Ahmad A, A. Ghosh, C. Schal, and L. Zurek (2011). Insects in confined swine operations carry a large antibiotic resistant and potentially virulent enterococcal community. BMC Microbiology 11:23.
- Doud C.W. and L. Zurek (2012) Enterococcus faecalis OG1RF:pMV158 survives and proliferates in the house fly digestive tract. Journal of Medical Entomology 9:15-155.
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Progress 01/01/10 to 12/31/10
Outputs
OUTPUTS: (ZUREK) Our data have been presented at international, national and local scientific meetings and have been incorporated into extension presentations given across Kansas and surrounding states. Albuquerque, T.A. and L. Zurek. 2010. Microbial community of aging horse manure affects stable fly (Stomoxys calcitrans) oviposition and larval development. Annual Research-Extension Update of the Department of Entomology, Manhattan, KS. (22-23 November, 2010) Albuquerque, T.A. and L. Zurek 2010. Microbial Community Structure of Aging Horse Manure and Stable Fly Oviposition Behavior. Annual Meeting of Livestock Insect Workers Conference, Knoxville, TN. (27-30 June 2010) Albuquerque, T.A. and L. Zurek 2010. Effect of Bacterial Community Structure of Aging Horse Manure on the Stable Fly Oviposition Behavior. Missouri Valley American Society for Microbiology (ASM) Branch Annual Meeting, Manhattan, KS. (09-10 April, 2010) Zurek L. 2010. House fly-microbial interactions from public health perspective. Annual Meeting of the Society for Vector Ecology. Raleigh, North Carolina (Sept. 25-30). Zurek L. 2010. Microbial ecology of the sand fly, Lutzomyia longipalpis. Dept. of Entomology, North Carolina State University Sept. 27. (ORTIGAO)Activities: 2010 was a significant year in terms goals achieved. My research program has been recognized nationally and internationally. Grant proposals were funded, including R21, a CRDF, and two ARRAs. FIRCA and PUF grants were submitted. Travel grants (4) funded by AGC and VP Research. Key experiments outlined in my grants were completed and results published. A new graduate course was offered on vector-borne diseases, and I continued mentoring PhD and MS students. I coordinated training of summer students, mentored undergraduates and postdocs, co-advised MS students from Pitt State from UFPE (Brazil). A provisional patent application was issued by KSURF (Disclosure No. 10-18) for PpChit1 as TBV. Events: (International) Lectured on "Molecular aspects of sand fly interaction with Leishmania", Brazil, September 13th-17th. Talk "RNA interference in sand flies" (coordinator and session chair), IV Workshop on Molecular Biology of Disease Vectors, Recife, Brazil. Talk "Ecological genetics of sand fly salivary gland genes" (Grad student) (National) Biology Department, Pitt State Univ., "Introduction to vector biology", November 16th; At the 59th ASTMH meeting, Atlanta, GA, November 3rd-7th, presented talk "RNAi silencing of P. papatasi midgut molecules and effects n Le. major development"; At the 59th ASTMH; Poster "Dissecting the molecular architecture of sand fly peritrophic matrix, a barrier for Leishmania development" Coutinho-Abreu IV et al; At the 4th AGC Symposium, June 10th -13th, Kansas City, MO; Poster "Reverse Genetics in Sand flies: Targeting Midgut Molecules Affects Leishmania Development" Coutinho-Abreu et al. Services: Meeting organization Co-organized the ENTOMOL4 - Workshop on Molecular Biology of Vectors, Brazil; Session co-Chair, 59th ASTMH, Atlanta, November 3rd-7th Committee assignments: National Sand Fly Genome Steering Committee; Sand Fly Genome Consortium; American Committee on Medial Entomology PARTICIPANTS: Maricela Robles-Murguia (Research Assistant); Alicia Sy (Research Assistant); Nathan Elliott (Undergraduate Student); Emma Hayes (Undergraduate Student) TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Zurek: We have investigated the changes in aging (fresh to five weeks old, on weekly basis) horse manure from the perspectives of its attractiveness for stable fly oviposition, its suitability for stable fly larval development, and its changes in terms of bacterial community structure and resulting emissions of volatile compounds that likely play a role in stable fly oviposition. Our results show that two weeks old manure is the most attractive for stable fly oviposition and fresh and 4-5 weeks old manure do not stimulate the oviposition. Surprisingly, fresh manure does support stable fly larval development although into a lesser degree that that of 2 weeks old manure. Four to five weeks old manure is not the suitable habitat for larval development. Great changes were detected in the bacterial community structure. Fresh horse manure contains primarily strictly anaerobic bacteria (clostridia, Bacteroides) that almost disappear in one week old manure. One to five week old manure is primarily colonized by aerobic and facultative anaerobic bacteria. Analysis of volatile compounds is currently in progress. This study shows that management strategies to control stable flies need to focus of 2-3 weeks old manure and fresh and >3 weeks old manure does not contain stable fly larvae. Our study shows a promising starting point for exploiting stable fly-bacterial associations for development of novel approaches for management based on better understanding of stable fly microbial ecology. Objective 3 Milestone experiments and data have been obtained and published. Among critical milestones - Working sand fly colonies. - Maintenance of Leishmania lines in vitro - Infection of sand flies - Antisera production - RNAi-based silencing and effects of Leishmania development - Expression analyses of sand fly midgut and salivary gland genes - Expression and purification of recombinant proteins Objective 4 New R21 (NIH) was funded to assess the role of US-native sand flies in transmission of CVL to Foxhounds. New RA was hired and sand fly trapping indicate expansion of L. shannoni. A complete study is schedule for 2011-2012. Preliminary data on exposure of Foxhounds to sand fly is encouraging.
Publications
- Martins GF, Serrao JE, Ramalho-Ortigao M, Pimenta PFP. 2010. Histochemical and ultrastructural studies of the mosquito Aedes aegypti fat body: effect of aging and diet type. Accepted for publication in Microscopy Research and Techniques.
- Coutinho-Abreu IV, Wadsworth M, Stayback G, Ramalho-Ortigao M, McDowell MA (2010) Differential expression of salivary gland genes in the female sand fly Phlebotomus papatasi. J Med Entomol 47(6):1146-1155.
- Coutinho-Abreu IV, Sharma NK, Robles-Murguia M, Ramalho-Ortigao M (2010) Targeting the midgut secreted PpChit1 reduces Leishmania major development in its natural vector, the sand fly Phlebotomus papatasi . PLoS Neglected Tropical Diseases 4(11): e901 doi:10.1371/journal.pntd.0000901
- Martins GF, Guedes BAM, Silva LM, Serrao JE, Fortes-Dias CL, Ramalho-Ortigao JM, Pimenta PFP. 2010. Isolation, primary culture and morphological characterization of oenocytes from Aedes aegypti pupae. Cell and Tissue Research, submitted and accepted for publication.
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Progress 01/01/09 to 12/31/09
Outputs
OUTPUTS: Our data have been presented at international, national and local scientific meetings and have been incorporated into extension presentations given across Kansas and surrounding states. Zurek, L. 2009. Role of house flies in dissemination of E. coli O157. Annual Meeting of the Multi-state project S1030, Baton Rouge, LA, Jan. 9-11. Albuquerque, T., A. Broce, and L. Zurek. 2009. Effect of age and microbial community structure of horse manure on the stable fly, Stomoxys calcitrans (L.), oviposition behavior Annual Meeting of LIWC, French Lick, IN, June 21-24. Zhu, J.J., D.R. Berkebile, and L. Zurek. 2009 Chemical Ecology of Stable Fly and its Future Practical Applications in Control. 26th Meeting of ISCE, University of Neuchatel, Switzerland 23-27 August 2009 Zurek, L. 2009. Microbial ecology of Muscoid flies. University of Florida, Gainesville, FL Oct. 22 (invited speaker). Doud, C. and Zurek, L. 2009. Role of house flies in dissemination of antibiotic resistant Enterococci from wastewater treatment plants. ESA Annual Meeting, Indianapolis, IN, Dec.13-16. Ortigao: Various experiments were completed according to outlined in my RO1 proposal (please see below for a list of the milestones achieved) and new data were presented at the following meetings: "Reverse genetics in sand flies: a path to assessing transmission blocking vaccine candidates", in Tunis, Tunisia. At the Society of Vector Ecology Meeting in Turkey "Phlebotomus papatasi salivary gland sequence variability and impact on defining vaccine candidates". At the 3rd Annual Arthropod Genomics Symposium, Kansas City, MO, I was one of the featured speakers and my presentation was "Sand flies functional genomics and beyond". At the ESA meeting in Indianapolis, IN. Talk entitled "Silencing midgut targets in sand flies: effects on physiology and Leishmania development". I was one of the speakers in the ESA section symposium "Novel Ideas in Sand Fly Research: Classification and Systematics, Epidemiology, and Molecular Interactions" which I organized. PARTICIPANTS: Graduate students: Thais Albuquerque, Carl Doud, Anuradha Ghosh, Iliano V. G Coutinho-Abreu, Leah Cox. Postdoctoral fellows: Anuradha Ghosh, Narinder Sharma. Collaborators: USDA-ARS, Lincoln, NE; Iowa State University; NIAID-NIH, Bethesda, MD; University of Notre Dame, Notre Dame, IN. TARGET AUDIENCES: Pest control operators, medical and veterinary entomologists, farmers and ranchers, restaurant management personnel, sanitarians, hygienists, microbial ecologists, clinicians, general public, military. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Zurek: Our results demonstrate stable fly/house fly - bacterial symbiosis; adult flies are capable of selecting an oviposition site based on microbially-derived stimuli that indicate the suitability of the substrate for larval development. The bacterial isolates that stimulate fly oviposition also support the larval development and in contrast, bacteria that do not stimulate or even repel flies from oviposition also do not support or even kill fly larvae. Our study shows a promising starting point for exploiting SF-bacterial associations for development of novel approaches for SF management. We have been able to show that muscoid flies have a close evolutionary relationship with microbes; the immature stages strictly depend on active microbial communities in the developmental habitat and females of these flies are capable of selecting of the new habitat for their offspring based on the cues emitted form the microbes. These studies provide a basis upon which oviposition behavior of muscoid flies and muscoid fly-bacterial association could be developed into novel approaches for integrated pest control. Our studies demonstrate that muscoid flies play an important role in the ecology of Escherichia coli O157:H7, Enterobacter sakazakii, and multi-drug resistant enterococci in environment and have to be seriously considered in designing of pre- and post-harvest food safety strategies. Ortigao: Establishment of working sand fly colonies. Culturing of Leishmania in vitro. Several antisera have been produced and tested by immunological and molecular approaches. We have produced dsRNA allowing us to perform RNAi experiments for several of the targets we are investigating. We have developed an efficient method for silencing sand fly transcripts and are the only laboratory in the country performing such studies. We have assessed the effect of RNAi silencing of midgut transcripts on sand flies. We have developed new protocols for enrichment of midgut proteins. We have cloned and are expressing several recombinant proteins from sand fly midgut. Assessed expression of several P. papatasi midgut transcripts via real time PCR analyses. Canine visceral leishmaniasis (CVL) is a serious disease infecting Foxhounds in the US. Three Foxhounds kennels in Kansas were affected by CVL, one near Pittsburg. Leishmania infantum the parasite causing CVL can be transmitted vertically but a sand fly route of transmission has not been ruled out. Lutzomyia shannoni sand flies were recently reported in locations throughout the Midwest, beyond their historic range. I started a collaboration with Dr David Gordon from Pittsburg State University to assess whether L. shannoni are present in Kansas. We plan to set up traps at different locations in the south east part of the state to determine whether L. shannoni has expanded into Kansas. A NIH R21 proposal submitted in July 2009 to fund the studies described above is currently pending.
Publications
- Talley J., A. Broce, and L. Zurek (2009). Characterization of the stable fly (Diptera: Muscidae) larval developmental habitat at round hay bale feeding sites. Journal of Medical Entomology 46: 1310-1319.
- Akhtar, M., H. Hirt, and L. Zurek (2009). Horizontal transfer of the tetracycline resistance gene tetM mediated by pCF10 among Enterococcus faecalis in the house fly alimentary canal. Microbial Ecology 59: 509-518.
- Macovei, L., A. Ghosh, V. Thomas, L. Hancock, S. Mahmood, and L. Zurek (2009). Enterococcus faecalis with the gelatinase phenotype regulated by the fsr-operon and with biofilm forming capacity are common in the agricultural environment. Environmental Microbiology. 11:154-1547.
- Ahmad, A. and L. Zurek (2009). Evaluation of metaflumizone granular bait for management of house flies. Medical and Veterinary Entomology 23: 167-169.
- Nagulapally, S.R., A. Ahmad, A. Henry, G.L. Marchin, L. Zurek, and A. Bhandari (2009). Occurrence of ciprofloxacin, trimethoprim-sulfamethoxazole, and vancomycin resistant bacteria in a municipal wastewater treatment plant. Water Environment Research 80: 82-90.
- Chakrabarti, S, S. Kambhampati, and L. Zurek (2010). Assessment of house fly dispersal between rural and urban habitats in Kansas, USA. Journal of Kansas Entomological Society (in press).
- Pitaluga AN, Beteille V, Lobo AR, Farias JRO, Davila AMR, Souza AA, Ramalho-Ortigao JM, Traub-Cseko YM (2009). EST sequencing of blood-fed and Leishmania-infected gut of Lutzomyia longipalpis, the principal visceral leishmaniasis vector in the Americas. Mol Genet Genomics 282(3):307-317.
- Coutinho-Abreu IV, Zhu K Y, Ramalho-Ortigao M (2009). Transgenesis and paratransgenesis to control insect-borne diseases: Current status and future challenges. Parasitol Int. Oct 9. [Epub ahead of print]
- Coutinho-Abreu IV, Ramalho-Ortigao M (2009). Transmission blocking vaccines to control insect-borne diseases. Mem Inst Oswaldo Cruz, in print.
- Ramalho-Ortigao M, Saraiva EM, Traub-Cseko YM (2009). Sand fly-Leishmania interactions: long relationships are not necessarily easy. The Open Parasitology Journal, in print
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