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Thesis (M.S.)--University of Florida, 2003. / Title from title page of source document. Includes vita. Includes bibliographical references.

Call number: LD2668 .T4 1979 A36 / Master of Science

Although it is sympatric with tsetse flies, Stomoxys calcitrans is not a biological vector of trypanosomes. It is known that haemolymph (HL) and midgut reservoir zone (RH) lectins regulate parasitic infections in some dipteran insects. Agglutinins (lectins) were detected in HL and RH from unfed stable flies (maximum titre 2-6). Increased haemagglutination activity resulted post-feeding (maximum titre 2- 16 - 2- 18). Optimum titres varied according to agglutinogen type and mammalian blood source. Rabbit erythrocytes produced the highest haemagglutination titres followed by human group B, human group 0, horse, human group A, human group AB and sheep. Stomoxys haemagglutination activity was found to be 1.5 - 2.5 times stronger than that of Glossina. Whole blood-fed flies produced the highest titre (2-18), compared to glucose-fed insects, against rabbit erythrocytes. Anti-Trypanosoma brucei brucei titres ranged from 2-6 - 2-7 in both tissues. Similar results were obtained with Leishmania hertigi and Crithidia!asciculata. Purification of the samples was performed in order to draw conclusions with confidence regarding the physico-chemical properties of the agglutinins (lectins) and in order to determine the molecular weight of the agglutinins. Protein contents ofHL and RH samples of flies aged < 12 hours to 3 days were determined. They were 25 - 28 mg/ml and 6.4 mg/ml respectively. Protein contents increased with age reaching 32 mg/ml for HL and 7.2 mg/ml for RH at day 14 post-emergence (p.e.). The contents then started to decrease reaching 22 mg/ml for HL and 5.6 mg/ml for RIi at day 28 p.e. Purified lectins constitute 4.3% of the total protein contents in RH samples (having molecular weights of 26,302 Da, 16,218 Da and 14,028 Da) and, approximately twice, 9.47% of the total protein contents, in HL samples (having similar molecular weights of 28,300 Da, 16,218 Da and 14,600 Da). HL and RH anti-parasite and anti-erythrocyte agglutinins (lectins) were basic glycoproteins in nature, calcium ion dependent for activity, heat labile, freeze-thaw sensitive and required slightly acid to alkaline pH conditions for optimum agglutination. Lectins were specific for galactosyl and glucosyl moieties. In vivo sugar inhibition of RH lectin activity resulted in three-fold increased S. calcitrans mortalities post- T.b. brucei infection, compared to the controls, suggesting a lectin parasite-killing function. However, sugar inhibition of lectins did not lead to transformation of trypanosomes to procyclic forms or to infection of the fly.

Doctor of Philosophy / Department of Entomology / Alberto B. Broce / Ludek Zurek / Stable fly, Stomoxys calcitrans (L.), and house fly, Musca domestica L., are two major pests affecting both confined and pastured livestock in the United States. It costs livestock producers millions of dollars annually to reduce populations of these two pests. Control of stable flies and house flies based on chemical insecticides is only marginally effective and unsustainable in the long term due to the development of insecticide resistance. This has created a demand for alternative methods which are environmentally friendly and cost effective for the management of these pests. Information on stable fly and house fly oviposition behavior and the aggregation and segregation of their immatures may help in an integrated pest management control program for these pests. This research identified specific bacterial species from the surface of stable fly eggs which are suspected of releasing chemical cues used to induce gravid females to oviposit at sites where eggs have been deposited and inhibit additional deposition of eggs in the same habitat when it is already colonized. My research also showed that stable fly and house fly larvae tend to be aggregated in distribution, even in apparently homogenous habitats, and to be spatially segregated from each other. Finally, I evaluated the vector competence of stable flies for an emerging food-borne pathogen, Enterobacter sakazakii, showing that this fly species is potentially a good vector for this pathogen.

Doctor of Philosophy / Department of Entomology / Ludek Zurek / Stable flies (Stomoxys calcitrans L.) are blood-feeding insects with a great negative impact on livestock resulting in annual losses around $2 billion in the USA alone. One of the main reasons for such an impact is because stable fly management is very difficult. Stable fly larvae develop primarily in animal manure and live bacteria were shown to be essential for stable fly development. I hypothesized that the microbial community of horse manure changes over time and plays an important role in stable fly fitness. Two-choice bioassays were conducted using 2 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. The results showed that fresh manure did not stimulate 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.

La mouche charbonneuse, Stomoxys calcitrans, est une mouche hématophage commune et cosmopolite. Elle est le vecteur de nombreux agents pathogènes dont Besnoitia besnoiti, une coccidie parasite des bovins à l’origine de la formation de kystes cutanés. La compétence vectorielle de S. calcitrans pour B. besnoiti reste cependant mal connue. Dans ce travail, la persistance des formes bradyzoites et tachyzoites de B. besnoiti a été évaluée sur les pièces buccales et dans le tractus digestif des mouches. Dans ces conditions expérimentales, B. besnoiti persiste deux jours au maximum chez les stomoxes. La destruction rapide du parasite chez ce vecteur suggère que la transmission vectorielle mécanique ne peut avoir lieu que très peu de temps après un repas contaminant sur un hôte bovin infecté. De plus, nous avons étudié la capacité des stomoxes à transmettre les formes bradyzoites ou tachyzoites à des lapins, utilisés ici comme animaux modèles de besnoitiose. Des repas interrompus de stomoxes ont été réalisés sur des bovins en phase chronique de besnoitiose (source de formes bradyzoites dans leur peau) ou bien sur du sang contaminé avec des tachyzoites de culture. Le repas a ensuite été complété sur des lapins. Les résultats de cette étude ont montré que 300 stomoxes sont suffisants pour transmettre des bradyzoites virulents d’un bovin à un lapin qui a montré des signes de besnoitiose aigue et une séroconversion. Dans la troisième partie de ce travail, des méthodes alternatives aux insecticides ont été testées afin de proposer des stratégies de contrôle efficaces et pérennes contre ces vecteurs. L’attractivité pour les stomoxes de sept types d’écrans bleus, présentant entre eux de légères variations de réflectance autour de 460 nm, a été étudiée, de même que les facteurs influençant l’efficacité des pièges (hauteur par rapport au sol, orientation est ou ouest de l’écran, proximité de différents hôtes ou d’un tas de fumier). Les résultats ont montré une très grande attractivité, de très hauts niveaux de capture et une grande sélectivité de tous les écrans testés. Un total de 164692 stomoxes a été capturé au cours de l’étude (soit 24 jours de piégeage avec 4 écrans par jour et 8 heures par jour). La moitié inférieure des écrans a capturé la majorité des stomoxes (70%), de même, le nombre de stomoxes capturés par la partie de l’écran orientée à l’Est est plus important que sur la face orientée à l’Ouest. Ces éléments soulignent l’intérêt de cetype de dispositifs dans la lutte contre les stomoxes. / The Stable fly, Stomoxys calcitrans, is a common and worldwide-distributed blood-feeding fly. It is the vector of numerous pathogens including Besnoitia besnoiti, a cyst-forming coccidian parasite of cattle. Vector competency of S. calcitrans for B. besnoiti remains poorly understood. We investigated here the persistence of bradyzoites and tachyzoites forms of B. besnoiti on mouthparts and in digestive tract of S. calcitrans. In experimental conditions, B. besnoiti proved to persist until two days in stable flies. The fast destruction of parasite within this vector suggests that mechanical transmission could occur a short time only after a contaminated blood meal. Moreover, we investigated the competence of stable flies to transmit immediately B besnoiti bradyzoites and tachyzoites to susceptible rabbits after an interrupted blood meal on chronically infected cows (source of bradyzoites) or on a tachyzoite-contaminated blood meal. The results show that 300 S. calcitrans only were able to transmit virulent bradyzoites to rabbits which suffered from clinical signs of acute besnoitiosis and seroconverted. In the third part of this study, we examined sustainable methods as alternative to insecticides but efficient to control this pest. Seven different types of blue screens, with slight differences in their reflectance around 460 nm, were tested in order to evaluate their attractiveness for stable flies and to determine the factors associated with the efficiency of trapping (height of screens, east versus west side of blue screen, proximity of different hosts and manure). The results showed high attractiveness, high number of captures and very high selectivity of all tested screens, with a total number of 164692 stable flies during the study (24 days of capture, 4 blue screens per day, 8 hours per day). The results showed that the lower part of the blue screens caught the majority of stable flies, whereas the east orientation was the most attractive which caught up to 60 % of the total number captured of stable flies. Taking together, these results are highlighting the interest of these new devices in the future control of stable flies.

Master of Science / Entomology / Ludek Zurek / House flies (Musca domestica L.) and stable flies (Stomoxys calcitrans L.) are insects of medical and veterinary importance. House flies are recognized as mechanical vectors of human foodborne pathogens and stable files are known for their painful bites resulting in reduction of body weight gain and milk production in cattle. The larval development of both fly species takes place in decaying organic materials (primarily animal manure), resulting in large fly populations in confined cattle environments. Shiga toxin-producing Escherichia coli (STEC) are a major foodborne pathogen. Cattle are the asymptomatic reservoir of STEC with bacteria being released to the environment via their feces. STEC O157 is the main serogroup causing human illness. However, infections with non-O157 STEC are increasing: more than 70% of non-O157 infections are caused by six serogroups of non-O157, referred as "Big six" (O26, O45, O103, O111, O121, and O145). In addition, there was a large 2011 outbreak in Europe caused by STEC O104. The objectives of my thesis were: 1) To assess the prevalence of seven serogroups of non-O157 STEC (O26, O45, O103, O104, O111, O121, and O145) (STEC-7) in house flies and stable flies collected from confined cattle environments; 2) To investigate the vector competence of house flies for non-O157 STEC-7. A total of 463 house flies from feedlots and dairies from six states, and 180 stable flies collected from a feedlot in Nebraska were processed for the isolation and identification of STEC-7 using a culture-based approach followed by PCR for the confirmation of serogroups, and virulence genes. A total of 34.3% of house flies and 1.1% of stable flies tested positive for at least one serogroup of E. coli of interest, and 1.5% of house flies harbored STEC with the Shiga-toxin gene (stx1). No STEC were detected in stable flies. Vector competence bioassays for non-O157 STEC revealed that house flies can carry non-O157 STEC for at least six days with the exception STEC O145. Overall, the findings of this research demonstrate that house flies, but not stable flies, likely play an important role in the ecology and transmission of non-O157 STEC in confined cattle environments.