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In this dissertation, we examine how advective and diffusive flows are created in the insect respiratory system, using a combination of direct biological studies and computational fluid dynamics simulations. The insect respiratory system differs significantly from the vertebrate respiratory system. While mammals use oxygen-carrying molecules such as hemoglobin, insects do not, favoring the direct delivery of oxygen to the tissues. An insect must balance advective flow with diffusive flux in order to sustain the appropriate oxygen concentrations at the tissue level. To better understand flow creation mechanisms, we studied the Madagascar hissing cockroach. In Chapter One, we used x-ray imaging to identify how tracheal tube compression, spiracular valving, and abdominal pumping coordinate to produce unidirectional flow during active respiration. In Chapter Two, we altered the environmental conditions by exposing the animals to various levels of hypoxia and hyperoxia, then examined how they changed their respiratory behaviors. In Chapter Three, we used our previous findings to construct a simulated insect respiratory system to parametrically study the effects of network geometry and valve timing on the creation of unidirectional advective flow and diffusive flux. These results can be used to inform future studies of the insect respiratory system, as well as act as the basis for bio-inspired microfluidic devices. / Doctor of Philosophy / The insect respiratory system works through the direct delivery of oxygen to the tissues. This occurs via a complex network of pumps, tubes, valves, and other structures that facilitate flow. These mechanisms allow insects to survive and prosper under a wide range of environmental and physiological conditions. While these structures have been studied extensively in a wide range of insect species, there are still many aspects of the respiratory system that remain unexplored. Here, we use the Madagascar hissing cockroach to examine how both bulk flow and diffusion are created in some types of insect respiratory systems. First, in Chapter One, we studied the animal under normal environmental conditions in order to determine how abdominal pumping, tracheal tube collapse, and spiracular valving are coordinated. Then, in Chapter Two, we exposed the animals to a range of oxygen concentrations to identify how the animals respond to varying environmental conditions. Finally, in Chapter Three, we constructed a simulated insect respiratory system to parametrically study the effects of network geometry and valve timing on the creation of advective and diffusive flow. By combining these three studies, we were able to improve our understanding of flow creation in the insect respiratory system.

The efficacy of two different autonomous Ultraviolet-C (UV-C) units on two spotted spidermite, Tetranychus urticae Koch (Acari: Tetranychidae), and greenhouse whitefly, Trialeurodes vaporariorum. (Hemiptera: Aleyrodidae). was tested in four commercial strawberry fields in California. In 2020, California strawberry growers used 12 million pounds of active ingredients (AIs) to keep pests under economic thresholds. While pesticides are an important part of an integrated pest management program, their use poses threats to the environment and human health. To protect agricultural workers, the public and the environment, regulators in California have consistently limited or removed the use of certain pesticide AIs. For strawberry growers in the state, this trend signifies that reliance on pesticides may not be a sustainable approach. Resistant pest populations further intensify the need for more sustainable pest control measures. Tetranychus urticae populations in California strawberry production have documented resistance to at least three AIs; abamectin, bifenazate, and hexythiazox. Trialeurodes vaporariorum populations in California strawberry production have documented resistance to five AIs; imidacloprid, thiamethoxam, dinotefuran, acetamiprid, and chlorpyrifos. To address this issue, stakeholders must invest in research and development of new non-chemical control methods. One of these methods is the use of autonomous UV-C for pest control. These systems emit UV-C radiation, which affects the cellular and genetic structure of organisms, leading to detrimental effects. One unit, by Tric Robotics, emits an avg of 250 J m−2 of UV-C irradiation and was tested in three different strawberry fields. Another unit by Saga Robotics emits an avg of 200 J m−2 was tested on a single field different from the first three. In each field, a plot was treated with UV-C twice weekly at night. Untreated controls were not possible with this study, so UV-C has been tested against each grower’s standard (GS) treatment. Twenty leaflets from the middle and 20 bottom tier of the strawberry plant canopy were collected weekly to assess the presence of T. urticae and T. vaporariorum. Samples were compared using Kruskal-Wallis rank sum tests with Bonferroni adjusted Dunn tests in the R statistical environment. The data showed that UV-C may be an effective tool for the control of certain T. vaporariorum stages in strawberry. Follow-up studies with untreated controls are needed to validate the efficacy of UV-C on T. urticae and further validate the positive results observed on T. vaporariorum.

Malaria occurs in a wide variety of environments. This tends to complicate the development of control strategies. In addition, vectorial capacity (the probability that a given vector will acquire, incubate, and transmit a disease organism) has been shown to vary greatly among both individuals and populations of vector species. Variability is influenced by both intrinsic and environmental factors. Understanding the effect of the environment on the vectorial capacity of mosquitoes would aid in the development and implementation of new control strategies. Body size and adult nutritional status of adult mosquitoes are two major factors thought to influence vectorial capacity. Both are influenced by changes in the environment. Various pesticides used in the control of mosquito populations also have been reported to influence acquisition and transmission of malaria parasites. Studies reported here focus on the impact of these extrinsic factors on the ability of the mosquito to acquire, incubate and transmit malaria parasites. Four hypotheses were tested in these experiments. These were: (1) large mosquitoes transmit malaria to more hosts during multiple feeding than small mosquitoes, (2) sugar taken before an infected blood meal and blood taken after an infected blood meal will increase acquisition and transmission of malaria, (3) mosquitoes exposed to sublethal doses of Bti will be less likely to acquire and transmit malaria than unexposed mosquitoes, and (4) developing oocysts promote the survivorship of the vector by inducing delays in subsequent egg-laying and blood-feeding. Two different models, an Anopheles stephensi - Plasmodium berghei - mouse model and an Aedes aegypti - Plasmodium gallinaceum - chicken model, were used to test these hypotheses. Both models were used to test the first hypothesis. This hypothesis was found to be true for Aedes aegypti. The hypothesis was also found to be true for Anopheles stephensi if the mosquitoes fed daily, but not true if they fed in quick succession. The Aedes aegypti model was used to test the second hypothesis. This hypothesis was not found to be true. The Anopheles stephensi model was used to test the third hypothesis. This hypothesis was not found to be true. The fourth hypothesis was tested using the Aedes aegypti model. The work presented here does not support the hypothesis. However, these experiments were preliminary, so more work is needed before any conclusions can be drawn.

Embryo growth of oviparous animals depends upon utilization of nutritive proteins, primarily the glycoprotein vitellin (Vt). These proteins are usually extraovarian in origin and accumulate in the oocyte through receptor-mediated endocytosis. This event is well characterized for both insects and vertebrates. In contrast, the mechanisms of yolk protein utilization are not understood. In this study, the German cockroach Blattella germanica was used as a model insect system to explore the components that initiate and regulate Vt processing during early embryo development. In B. germanica, Vt processing is initiated at days 3-4 postoviposition at 30$\sp\circ$C. The yolk of freshly oviposited eggs assayed in vitro for protease was devoid of activity but protease specific activity increased dramatically during embryo development. This activity correlated temporally with Vt processing in vivo suggesting that the protease is necessary for Vt processing. The protease was purified from yolk at day 6 postoviposition by gel filtration and affinity chromatography and classified as a cysteine protease. Its molecular weight, estimated by SDS-PAGE and immunoblotting, was approximately 29 kDa. Its pH optimum was 5.5, within the pH range of yolk granules. The purified protease degraded Vt in vitro yielding peptides of the same apparent molecular weights as Vt processed in vivo. Acidification of day 0 yolk in vitro induced protease activity suggesting that a latent protease is present in eggs early in embryo development. The latent protease activity was purified from yolk at day 0 postoviposition by successive use of gel filtration, Mono Q FPLC, and hydrophobic interaction chromatography. The purified latent protease had a molecular weight of approximately 40 kDa and could be activated in vitro into a cysteine protease of 29 kDa. The conversion depended on acidification and was blocked by the cysteine protease inhibitor E-64, suggesting the activation is autocatalytic. Kinetic studies showed that activation occurred by intermolecular catalysis. The pH-activity and inhibitor sensitivity profile of the in vitro-activated protease matched those of the protease suggesting that the latent protease is the proenzyme of the protease. Active site derivatization of the 40 kDa proprotease revealed that its conversion to the 29 kDa protease in vivo occurs as Vt processing begins, suggesting that Vt processing is regulated through the protease. The proenzyme activation and pH optimum data of the purified protease emphasize that yolk granule acidification is an important cellular factor for the regulation of Vt processing by B. germanica in vivo. A murine polyclonal antibody against purified proprotease was made monospecific by affinity column chromatography. Using this antibody, the proprotease was detected in fat bodies and ovaries of vitellogenic females by immunoblotting. This result is consistent with the hypothesis that the protease precursor is synthesized extraovarily, probably in the fat body.

The life history and behavioral ecology of Myiopharus doryphorae (Riley) and Myiopharus aberrans (Townsend), important parasitoids of the Colorado potato beetle (CPB) Leptinotarsa decemlineata (Say), were investigated through a series of field and laboratory studies. The recruitment-recruitment method of determining percentage parasitism was compared with traditional methods in assessing population dynamics of the CPB and Myiopharus. Over the three-year period of the study, percentage parasitism calculated from traditional foliage sampling showed an erratic pattern. Revised estimates employing the recruitment method revealed a consistent 30-50% mortality of CPB larvae due to parasitism even when the CPB prepupal population density reached 80 per square meter per generation, showing that Myiopharus spp. can cause high levels of mortality to CPB larvae at higher host densities than has been reported in most previous field studies. Field sampling demonstrated that M. doryphorae and M. aberrans overwinter as first-instar larvae within adult diapausing CPB and complete their development the following spring after the emergence of the parasitized beetles. Growth-chamber studies were conducted to quantify development of summer-generation M. doryphorae at different life stages. During the first four days after being larviposited, these M. doryphorae remain as first-instar larvae but grow an average of 0.45 $\pm$ 0.03 mm prior to the prepupal stage of their hosts, in which the parasitoids complete development. During this latter period, development rates of M. doryphorae were found to track closely those of the CPB itself when modeled as a nonlinear function of temperature assuming cessation of growth outside the approximate range of 4-34$\sp\circ$C. The model appears to require additional adjustment at temperatures below 10$\sp\circ$C. Laboratory studies showed that M. doryphorae do not discriminate between CPB larvae fed sublethal doses of B. thuringiensis and larvae not fed with B. thuringiensis. M. doryphorae appear more sensitive to CPB larval movement than to the presence or absence of B. thuringiensis. Through field studies a series of behaviors was identified and their frequency and duration were quantified for the two Myiopharus spp., which appeared not to vary their allocation of time in response to each other's presence in the same field. A significant difference was found between the frequencies of larviposition by the two Myiopharus species across ranges of temperature and time of day. Behavioral studies led to the discovery that larvipositing M. doryphorae and M. aberrans females discriminate between parasitized and non-parasitized host larvae, rejecting the former on contact and failing to larviposit in them when other potential hosts are available. This discrimination breaks down to some extent late in the growing season when CPB larvae of appropriate stages are rare; breakdown of host discrimination is accompanied, however, by the defense of recently parasitized hosts by females of both Myiopharus species, and is followed by the switch of larvipositing M. aberrans from larval hosts to adult CPB which are more common at this time of year.

Thesis (PhD)--Stellenbosch University, 2004. / ENGLISH ABSTRACT: In surveys carried out on poultry farms in the Western Cape Province, South Africa, the larvae of the flies Musca domestica L., Fannia canicularis (L.) and Leptocera sp. were found to be numerous. The majority of M domestica larvae and the mite Proctolaelaps sp. were recovered from manure with temperatures between 20 and 26°C while the larvae of F. canicularis and Leptocera sp. were mostly found between 14 and 22°C. Predominant predatory arthropods in the manure were the histerid Carcinops pumilio (Erichson), the staphylinid Philonthus sordidus (Gravenhorst) and the mite Macrocheles muscaedomesticae (Scopoli). These had a manure temperature tolerance of between 12 and 31°C. Macrocheles muscaedomesticae and P. sordidus preferred fresh manure, whilst C pumilio preferred aged manure. Therefore these two predators could complement one another in integrated pest management programmes. From a study of the vertical cross-sectional profile of the manure cone it was clear that the larvae of the prey species M domestica and F. canicularis and predator P. sordidus distinctly occupy the crest of the manure cone. The succession of arthropods in accumulating poultry manure and the effect of manure height on their numbers was studied. The prey species M domestica and F. canicularis were the first colonizers followed by the predators P. sordidus and M muscaedomesticae. Carcinops pumilio was a late colonizer. The numbers of M domestica and F. canicularis larvae were negatively correlated with the increase in manure height whilst the numbers of the predators C pumilio and M muscaedomesticae were positively correlated with this. The total developmental time of C pumilio from egg to adult emergence at 30°C was 20.5 days. Two larval instars were recorded. The immature stages sustained about 50% mortality before adult emergence. Carcinops pumilio adults can live up to 130 days at 30°C and had a Type I survivorship curve. The intrinsic rate of natural increase (rm) was 0.064943 and net reproduction rate (Rs) was 20.191.The rate of oviposition was directly proportional to body size and weight. Increase in density had a dampening effect on fecundity and led to an increase in developmental time. Thus density will be a critical factor in any mass rearing programme for this predator of fly larvae. In addition, an increase in density stimulated the dispersal of C. pumilio. A crowding level of 50 C. pumilio adults per 200ml container resulted in the least dispersal which did not exceed 2.5% per day, compared with up to 24.0% at a crowding level of 400. The predation rate of flies by C. pumilio in the laboratory decreased with an increase in predator density, but increased with an increase in starvation. Starvation had no effect on predation rate of M muscaedomesticae. A technique exploiting Drosophila melanogaster Meig. as a source of prey was successfully developed for rearing C. pumilio, which could be of commercial value. Drosophila melanogaster appeared to be an ideal candidate as a source of prey for mass rearing C. pumilio due to its short developmental time of about ten days. It is also inexpensive and easy to breed, and has a very high biotic potential. Carcinops pumilio that were fed on artificial diet had a prolonged developmental time and increase in weight, and laid fewer eggs than those fed on natural diet. C. pumilio completed its development on the artificial diet and both the F, and F2 generations fed on an artificial diet were able to lay eggs. This could be the first step towards finding an artificial diet that would allow continuous rearing of C. pumilio and their availability at all times for utilization in the biological control of houseflies. / AFRIKAANSE OPSOMMING: In opnames wat op pluimveeplase in die Westelike Kaapprovinsie, Suid-Afrika uitgevoer is, is gevind dat die vliegsoorte Musca damestica L., Fannia canicularis (L.) en Leptacera sp. volop is. Die meeste van die M. damestica larwes en die myt Practalaelaps sp. is versamel in hoendermis met temperature tussen 20 en 26°C en die larwes van Ficanicularis en Leptacera tussen 14 en 22°C. Die volopste predatoriese arthropode in die mis was die histerid Carcinaps pumilia Erichson, die staphylinid Philanthus sardidus (Gravenhorst) en die myt Machrascheles muscaedamesticae (Scopoli). Hulle temperatuurtoleransie in die mis was tussen 12 en 31°C. Machrasceles muscaedamesticae en P. sardidus het vars mis verkies, terwyl C. pumilio voorkeur aan ouer mis gegee het. Gevolglik behoort hierdie twee predatore mekaar te komplementeer in programme van ge-integreerde plaagbestuur. In Studie van die vertikale deursnee-profiel van die miskeel het duidelik getoon dat die prooispesies M. damestica en en F canicularis en die predator P. sardidus in die kroon van die miskeel voorkom. Die opeenvolging van die arthropode in die ophopende hoendermis en die invloed van mishoogte op hulle getalle is bestudeer. Die prooispesies M damestica en F canicularis was die eerste koloniseerders, gevolg deur die predatore P. sardidus en M muscaedamesticae. Carcinaps pumilia was In laat koloniseerder. Die getalle van die larwes van M damestica en F. canicularis was negatief gekorreleer met die toename in mishoogte terwyl die getalle van die predartore C. pumilia en M. muscaedamesticae positief daarmee gekorreleer was. Die totale ontwikkelingstyd van C. pumilia van eier tot die volwassene se verskyning was 20.5 dae by 30°C. Twee larwale instars is gevind. Die onvolwasse stadia het 50% mortaliteit ondergaan voor die verskyning van die volwassenes. Die volwassenes van C. pumilia kan tot 130 dae lank by 30°C lewe en het In Tipe 1 oorlewingskurwe gehad. Die intrinsieke tempo van natuurlike toename (rm) was 0.064943 en die netto reproduksietempo (Rs) 20.191. Die tempo van eierlegging was in direkte verhouding tot die liggaamsgrootte en massa. Toename in digtheid het 'n onderdrukkende effek op vrugbaarheid gehad en tot 'n toename in ontwikkelingstyd gelei. Digtheid sal dus 'n kritieke faktor wees III emge program van massateling vir hierdie predator van vlieglarwes. Daarmee saam het 'n toename in digtheid die verspreiding van C. pumilio gestimuleer. By 'n digtheid van 50 C. pumilio volwassenes per houer het die verspreiding nie 2.5% oorskrei nie, in vergelyking met tot 24.0% by 'n dightheid van 400. Die predasietempo van vliee deur C. pumilio in die laboratorium het afgeneem met 'n toename in predatordigtheid, maar toegeneem met 'n toename in verhongering. Verhongering het nie 'n invloed gehad op die predasietempo van M. muscaedomesticae nie. 'n Tegniek met Drosophila melanogaster Meig. as bron van prooi in die teling van C pumilio is suksesvol ontwikkel en dit kan van kommersiele waarde wees. Dit blyk dat D. melanogaster ideaal kan wees as 'n bron van prooi in die massateling van Cpumilio vanwee sy kort ontwikkkelingstyd van ongeveer tien dae. Die spesies is ook goedkoop en maklik om te teel, en het 'n baie hoe biotiese potensiaal.Care inops pumilio wat op 'n kunsmatige dieet gevoed het, het 'n verlengde ontwikkelingstyd en gewigstoename gehad, en het minder eiers gele as die wat op 'n natuurlike dieet gevoed het. C. pumilio het sy lewensloop op die kunsmatige dieet voltooi en beide die FJ en F2 generasies wat op die kunsmatige dieet gevoed het, was in staat om eiers te Ie. Dit kan die eerste stap wees in 'n poging om 'n kunsmatige dieet te vind wat dit sal moontlik maak om C. pumilio aaneenlopend te teel sodat dit deurlopend beskikbaar kan wees vir gebruik in die biologiese beheer van huisvliee,

Blow flies (Diptera: Calliphoridae) are among the most dominant and conspicuous insects in the decomposition process. They are important in forensic entomology to determine time of death and, in certain situations, cause of death or relocation of a body. Insects are now included as standard operating procedures in crime scene investigations in many countries, however, this is not standard procedure in the Caribbean area due to lack of knowledge of insects involved in cadaveric decomposition. Successful application of forensic entomology depends on solid underlying data. Our main goal is to advance the knowledge of Calliphoridae in the Caribbean to enable forensic entomology studies. We performed a mega-transect across the Caribbean and extensively collected flies attracted to rotten meat baits during five years from 2011 to 2015. Overall we collected 61,332 flies of which 34,650 were Calliphoridae. We sampled 16 of the 18 species of forensically important Caribbean Calliphoridae and three continental species. We determine the diversity and distribution of Calliphoridae in the Caribbean. We also present a thorough DNA barcode dataset, covering the geographic range of most species in the region. Finally we established phylogenetic relationships among Calliphoridae species and test biogeographical hypotheses, and patterns of diversification and endemism in the Caribbean. In sum, this is the most comprehensive study of the family Calliphoridae from the Caribbean that will open the door for future research on forensic entomology in the region.

Six species of insects were found attacking limber pine cones from July 26, 1968, through October 4, 1969, in Cache National Forest. The three species considered of major importance are: Conophthorus flexilis Hopkins, Dioryctria abietella (D. & S.), and D. sp. near or disclusa Heinrich. The three minor species encountered are; Bradysia sp., Trogoderma parabile Beal, and Asynapta keeni (Foote). In addition to the major and minor cone pests three parasites, Apanteles sp . prob. starki Mason, Elacherus sp., and Hypopteromalus percussor Girault were found associated with the cone pests. C. flexilis, which completely destroys the cone, was ranked as the number one pest on the basis of numbers present plus severity of damage. During 1968 and 1969 C.flexilis destroyed 11.47 percent of the 1500 cones examined, with a mean of 5.87 larvae per infested cone. The cone moths, D. abietella and D. sp. near or disclusa, were ranked second and third in importance respectively. D. sp . near or disclusa was potentially the more important cone moth, as it caused a total destruction of the seed bearing portion of the cones. However, D. abietella infested 15.40 percent of the cones, in contrast to 2.00 percent by D. sp. near or disclusa. There were no significant statistical differences in insect populations between 1968 and 1969, although the percent infestation of C. flexilis and D. sp. near or disclusa increased slightly and D. abietella decreased.

Indiana University-Purdue University Indianapolis (IUPUI) / Forensic entomology is a subdiscipline of entomology that involves the use of insect behavior and developmental data to aid in criminal investigations. Genetic data has become increasingly important to the field as there has been a push for DNA-based species identification methods of forensically relevant insects. Genetic data can also elucidate population structure and relatedness of these insects, and such knowledge can contribute to the development of more specific datasets for insects in different regions. The first study presented here investigated the phylogenetics of sister species Lucilia cuprina and Lucilia sericata to identify possible subspecies divisions and issues with DNA-based identifications in the United States. The initial aim of this study was to identify genetic differences between specimens of L. cuprina that preferred live versus carrion flesh. Flies collected from Indiana, USA and South Africa were sequenced and analyzed. Upon sequencing of the genes COI, Period, and 28s, our results indicated that L. cuprina from Indiana possess a unique combination of nuclear and mitochondrial haplotypes that suggest a unique lineage, possibly indicating modern hybridization with L. sericata. The inability of both nuclear and mitochondrial genes to distinguish between L. cuprina and L. sericata raises questions about the capabilities of DNA-based species identifications within this genus. Additionally, the inability of these genes to distinguish between specimens that preferred live versus carrion flesh highlights a need for continued research of these behavioral differences. The second study presented here investigated the population structure and relatedness of house flies in the American southwest in relation to a civil lawsuit where neighbors of a poultry farm alleged that flies were emanating from the farm to their homes. Musca domestica (house fly) specimens were collected from the chicken farm and from locations in varying directions and distances from the farm. Amplified fragment length polymorphism (AFLP) analysis was performed and the data were used in a number of analyses. Population reallocation simulations generally indicated that samples from different locations were not genetically different enough from other locations to allocate to their true origin population over others. Kinship analysis showed differences in samples collected in a later season that indicate a genetic bottleneck over time. Population structure analysis indicated the presence of two intermixing genetic populations in the dataset. AMOVA revealed that the majority of genetic variation laid within, rather than among, populations. A Mantel test revealed no significant correlation between genetic and geographic distances. These results indicate that the M. domestica population in this region of southwestern America is large and intermixing, with no clear genetic distinctions between specimens collected at the poultry farm versus the surrounding locations. In regard to the civil lawsuit, it was not possible to conclude that the flies did not emanate from the poultry farm. In a broader perspective, these data can be utilized to develop pest management strategies in this region. Overall, the data from both studies presented here will be useful to forensic investigations, development of more specific and detailed data and identification techniques, and pest control measures.