Forensic entomological field and laboratory studies were conducted to obtain data currently missing or conflicting in the literature. The first goal of this project was to identify and qualitatively assess the major taxa of forensic importance in southwest Virginia. Carcasses of the domestic pig, Sus scrofa L., were placed in field conditions and allowed to decompose until they reached the advanced stage of decay. Over 50 taxa were collected and identified, with Phormia regina, Phaenicia coeruleiviridis, Phaenicia sericata, Calliphora spp., Sarcophaga utilis, Musca domestica, Hydrotaea leucostoma, Stearibia nigriceps, Prochyliza xanthostoma, and Meroplius minutus among the most commonly observed fly species, and Creophilis maxillosus, Platydracus maculosus, Aleochara lata, Oiceoptoma noveboracense, Necrodes surinamensis, and Necrophila americana among the most commonly observed beetle species. The second objective of this study was to analyze successional patterns of taxa collected and identified in the carrion-insect succession studies. Occurrence matrices were constructed for the successional patterns of insect taxa during 21 sampling intervals in the spring and eight intervals in the summer studies. Permutation analyses of the occurrence matrices showed that the successional patterns of insect taxa were similar between spring 2001 and 2002 (P = 0.001) and between summer 2001 and 2002 (P = 0.007). Results indicated that the successional patterns appear to be typical for the seasonal periods. The third objective of this study was to analyze the effects of antemortem ingestion of ethanol by pigs on insect successional patterns and development rates. Pigs were dosed with a mixture of 95% ethanol and saline. Blood samples were collected immediately prior to euthanasia. The carcasses then were placed at an open field site and allowed to decompose. Insect samples were collected from carcasses for ten days post-mortem and the collected data were used to develop occurrence matrices. Permutation analysis to test the null hypothesis of no similarity between successional patterns of taxa from treated and untreated pigs showed that the successional patterns were similar. Loin meat from the carcasses was used as a rearing medium for field development studies of the black blow fly, Phormia regina. Development rates of 3rd instar P. regina maggots feeding on meat from treated pigs were significantly different from development rates of maggots feeding on meat from untreated pigs. No significant differences were detected in 1st and 2nd instars. Finally, the effect of relative humidity (RH) on egg hatch time and percent hatch rate of Phormia regina and Phaenicia coeruleiviridis was tested using saturated salt solutions. The global ANOVA of mean hatch times and percent hatch was found to be significant for P. regina at 20°C and 30°C and P. coerulieviridis at 20°C among RH levels at á = 0.05. T-test (LSD comparisons) results detected significant differences between some but not all RH levels within all three data series tested. Percent hatch observed varied widely, depending on the RH level. Of the levels tested, the lowest RH where hatch rate was observed was 53% (20°C) for P. coeruleiviridis, where only 1% of the eggs hatched. At 90+% RH, almost all of the eggs hatched for both species. / Ph. D.

Studies were conducted on the effects of alcohol and opioids on the development of forensically important flies. In addition different methods of degree-day calculations and development thresholds were used to determine the effects on PMI estimates. The first study determined the effects of ethanol on the development of Phormia regina in vitro. Ground pork loin was treated with a 1, 5, or 10% ethanol solution to give an equivalent Blood Alcohol Concentration (BAC) of 0.01, 0.04, and 0.8 % w/v. A significant difference in the time for second instars to complete the stage was seen between the 1% treated and control. Significant differences were also found among pupal and adult weights between all treatments and the control. A significant difference was shown between growth curves of the 5% treated and control for third instar larvae using the Kolmogorov-Smirnov test. The ethanol content of Phormia regina in migrating third instar larvae that fed on treated meat was examined using headspace-gas chromatography (HSGC). All larvae had a content of 0.01% w/v, including the control. The effects of ante-mortem injection of oxycodone in pigs were examined with respect to insect succession patterns and the development rates of Phormia regina. Pigs were given a subcutaneous injection of oxycodone hydrochloride (3 mg/kg by weight) and antemortem blood samples were collected prior to and following drug injection. Shortly after death the carcasses were placed at an open field site and allowed to decompose in a field cage. Insect samples were collected from carcasses for seven days post-mortem and the collected data were used to develop occurrence matrices. The Simple Matching Coefficient showed that successional patterns were similar between treated and untreated animals. Loin and liver from the carcasses were used as rearing media for in vitro development studies of Phormia regina. Kolmogorov-Smirnov test showed that third instar P. regina maggots from treated loin tissue were significantly longer in length than maggots feeding on untreated loin tissue. A significant difference in time was found among larvae on loin for the time from eclosion to completion of the second instar. Significant differences were seen in the weight of adults reared on liver and loin. A chi-square for homogeneity showed that adults were biased towards males (2:1) from untreated loin tissue. A final study compared weather data sources, Accumulated Degree-Day (ADD) methods, and postmortem interval (PMI) estimations based on threshold and developmental data source. Four pigs were used for statistical comparisons. Pigs were taken to a test site and allowed to decompose in an experimental cage. Probes recorded ambient temperatures and body temperatures. Maggot sampling was completed every day for each pig. A three way factorial linear fit model was used to test for statistical differences. Significant differences were seen in the calculated ADD based on probe location and the development threshold used. The ADD calculated from local weather station locations: Kentland Farm, Moore Farm, and Blacksburg Airport were also compared. A significant difference in ADD was found in the main effects among locations (Airport 44.1 ADD, Kentland 37.5 ADD, Moore 48.6 ADD), as well as among the thresholds used (10,12.2, and 14°C). Different PMI estimations also resulted when using development data from different development studies on Phormia regina. / Ph. D.

Orientador: Aricio Xavier Linhares / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituo de Biologia / Made available in DSpace on 2018-08-15T15:49:24Z (GMT). No. of bitstreams: 1 Souza_CarinaMarade_M.pdf: 1457552 bytes, checksum: 6cd38fc74da441a111a04348ee418d6d (MD5) Previous issue date: 2010 / Resumo: Devido ao aumento no número de mortes relacionadas ao abuso de drogas, o conhecimento da interferência destas substâncias sobre o desenvolvimento de certas espécies de insetos necrófagos torna-se de grande importância, especialmente quando a estimativa do intervalo pós-morte (IPM) usa como base a idade do inseto. Além disso, esta metodologia é uma valiosa alternativa para determinar a causa da morte, quando o corpo encontra-se em avançado estágio de decomposição. Assim, o presente estudo objetivou avaliar o efeito de diferentes concentrações de esteróides anabólico-androgênicos (EAA), comumente usados para aumento da massa muscular, sobre a taxa de desenvolvimento de Chrysomya putoria (Wiedemann), C. albiceps (Wiedemann) e C. megaacephala (Fabricius) (Diptera: Calliphoridae). Dois delineamentos experimentais foram feitos: um somente com a substância decanoato de nandrolona (Deca-Durabolin®) e outro associando decanoato aos ésteres de testosterona (Durateston®). Imaturos obtidos de colônias de moscas adultas estabelecidas em laboratório foram depositados em potes plásticos contendo dieta artificial para montagem dos grupos experimentais, sem (grupo controle) e com adição de esteróides. Cada tratamento foi repetido seis vezes. O ganho de massa foi registrado para grupos de 10 larvas, em intervalos de 12 h, até que atingissem o estágio de pupa. Além disso, foram medidos a viabilidade larval, a taxa e o intervalo de emergência. ANOVA de um critério e teste de comparações múltiplas de Duncan foram feitos para comparar o efeito dos EAA, associados ou não em diferentes e crescentes concentrações, sobre o desenvolvimento larval usando o ganho de massa como variável resposta. Diferenças significativas não foram observadas quanto ao tempo de desenvolvimento e intervalo de emergência para todas espécies tratadas, mas houve nítida heterogeneidade quanto às curvas de desenvolvimento, quando associados os dois EAA para C. putoria e C. albiceps. A viabilidade larval e a emergência de C. putoria foram significativamente afetadas pela associação de EAA, com valores inferiores aos apresentados pelo controle. Frente ao decanoato, C. putoria apresentou inicialmente maior ganho de massa, contudo, em idades mais avançadas, os imaturos expostos à droga obtiveram menor média de massa corpórea que o controle. Para C. albiceps a maior concentração de decanoato afetou negativamente o ganho de massa durante todo o desenvolvimento larval. A análise do ganho de massa, por hora, indicou diferenças significativas no período compreendido entre 36 e 84 h entre grupos controle e com adição de EAA associados, o que levaria a erro no cálculo do IPM, se esta condição não fosse considerada. / Abstract: Due to increase of deaths related to drug abuse, the knowledge of the interference of these substances on the development of certain necrophagous insects species can be very important, especially when the estimative of the postmortem interval (PMI) is based on insect age. Furthermore, they are valuable alternatives to determine the cause of death, when the body is in an advanced stage of decomposition. The present study aimed to evaluate the effect of different concentrations of anabolic-androgenic steroids (AAS), commonly used to increase muscle mass, on the developmental rate of Chrysomya putoria (Wiedemann), C. albiceps (Wiedemann) and C. megaacephala (Fabricius) (Diptera: Calliphoridae). Two experimental designs were made: one only with nandrolone decanoate (Deca-Durabolin®) and another with decanoate associated to ester of testosterone (Durateston®). Immatures obtained from adult flies colonies established in lab were placed in plastic vials containing an artificial rearing diet for assembly of experimental groups, without (control group) and with addition of steroids. Each treatment was replicated six times. The mass gain was recorded for groups of 10 larvae at 12h-intervals until they reached the pupal stage. In addition, we measured the larval viability, the rate and interval of emergency. ANOVA one way and Duncan's multiple comparison test were made to compare the effect of the EAA, associated or not in different and increased concentrations on the developmental larval using the mass gain as dependent variable. Significant differences were observed regarding the developmental time and interval of emergency for all threatened species, but there was a clear heterogeneity on the developmental curves for C. putoria and C. albiceps by the EAA association. The larval viability and the emergence rates of C. putoria were significantly affected by the EAA association, with rates lower than control group. Initially, C. putoria rearing in decanoate showed greater weight gain, however, at older ages, immatures obtained a lower average weight than control. For C. albiceps the highest concentration of decanoate negatively affected mass gain during the larval development. The mass gain analysis, per hour, indicated significant differences from 36 to 84 h between control and EAA groups, which could lead to incorrect estimation of the PMI, if this condition was not considered. / Mestrado / Mestre em Parasitologia

Orientador: Aricio Xavier Linhares / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-10T18:18:35Z (GMT). No. of bitstreams: 1 Soto_DoraAmparoEstrada_D.pdf: 2535722 bytes, checksum: 550a33ae626a5a9a30ff32773e785904 (MD5) Previous issue date: 2008 / Resumo: A Entomologia Forense utiliza dados de desenvolvimento e aspectos ecológicos de insetos necrófagos com o objetivo de auxiliar as investigações criminais, entre outros, no cálculo da estimativa do intervalo pós-morte (IPM). Igualmente, pode-se caracterizar a causa da morte quando essas drogas são detectadas em insetos imaturos que se alimentam dos corpos em decomposição. Além disso, os insetos podem servir como uma ferramenta alternativa para análises toxicológicas na ausência de tecidos e fluídos coletados habitualmente para este fim. Trabalhos recentes têm demonstrado que a presença de drogas ou toxinas em tecidos em decomposição pode alterar a taxa de desenvolvimento de insetos que usam este recurso como fonte alimentar. A ausência de conhecimentos dos possíveis interferentes no ciclo biológico pode gerar dados imprecisos sobre a idade do inseto, o que prejudicaria a estimativa do IPM. O ramo da Entomologia Forense que estuda os aspectos acima mencionados é denominado Entomotoxicologia. Rotineiramente, é necessário fazer testes preliminares em laboratório com dieta artificial e com certas substâncias para criar um banco de dados que possa ser útil como um padrão para investigar e quantificar as modificações que podem ocorrer com insetos coletados numa cena criminal. O objetivo principal deste estudo foi observar o efeito dos barbitúricos, fenobarbital (Gardenal®) e tiopental (Thiopentax®) em diferentes dosagens no desenvolvimento de imaturos de três espécies de califorídeos ¿ Chrysomya albiceps (Wiedemann), Chrysomya megacephala (Fabricius) e Chrysomya putoria (Wiedemann) (Diptera). Foi observado que os dois barbitúricos afetaram significativamente o desenvolvimento larval das três espécies, sendo que na maior dosagem a taxa de mortalidade foi elevada e na menor, o desenvolvimento retardou em até duas vezes, quando comparado ao grupo controle (sem adição de substância química). O conhecimento da ação destes barbitúricos no desenvolvimento dos imaturos dessas espécies de interesse forense pode contribuir para evitar erros no cálculo da estimativa do IPM. Também foi avaliada a taxa de desenvolvimento de Chrysomya albiceps (Wiedemann) e Chrysomya megacephala (Fabricius) (Diptera) criadas em diferentes dietas artificiais contendo tecidos de origem animal e comparadas a uma dieta já descrita em literatura, a qual não contém tecido animal, e a um grupo controle, onde foi usada somente a carne moída. A eficiência de cada dieta foi observada levando em conta o ganho de massa dos imaturos, o tempo de desenvolvimento larval, a taxa de sobrevivência de pupas e adultos, o intervalo de emergência e o tamanho dos adultos. Assim, foi proposta uma nova dieta artificial proporcionando uma melhor uniformidade para a adição de drogas e sobrevivência das espécies estudadas, tornando mais eficiente a criação e os testes biológicos com os dípteros necrófagos em laboratório / Abstract: The Forensic entomology utilizes developmental data and ecological aspects of necrophagic insects with the aim of helping criminal investigations, being able to, for instance, estimate the post mortem interval (PMI). Likewise, the death cause may be detected when these drugs are detected in immature insects from the same substrate. Besides, the insects can work as an alternative tool for toxicological analyses in the absence of tissues and fluids usually collected for this purpose. Recent investigations have been demonstrating that the presence of drugs or toxins in decomposing tissues may alter the development rate of insects that use such resource as a food substrate. The non-observance of this knowledge may lead to imprecise conclusions on the insect age, producing mistakes on the PMI estimative. This field of forensic entomology is named entomotoxicology. On routine basis, laboratory preliminary tests using artificial diet and certain substances are necessary to create a data bank that could be useful as a pattern to investigate and quantify the modifications that may occur with insects collected from death scenes. The present study aimed to verify the effect of the barbiturics phenobarbital (Gardenal®) and thiopental (Thiopentax®) in three concentrations, in the development of immatures of three Calliphoridae species ¿ Chrysomya albiceps (Wiedemann), Chrysomya megacephala (Fabricius) and Chrysomya putoria (Wiedemann) (Diptera). The barbiturics significantly affected the larval development of the three species, when the highest mortality rate occurred with the highest concentrations, and in the other concentrations the development retarded twice as much compared to the control group (without chemic substance). The knowledge on the action of these barbiturics in the immature development of these species of forensic interest can contribute to minimize the errors in the estimative of the PMI. A comparison of the developmental rate of the blowflies Chrysomya albiceps (Wiedemann), and Chrysomya megacephala (Fabricius) (Diptera) reared on: a) artificial diets containing animal tissues; b) artificial diet without animal tissue; and c) a control group (which contained only meat) was thus carried out. The efficiency of each diet was evaluated by observing the gain of immatures¿ weight, larvae developmental time, viability percentage obtained from pupae and adults, emergence interval and adult size. To make the rearing and biological tests with the necrophagic dipterans in laboratory more efficient, a new artificial diet was proposed, leading to a better uniformity and increased survivorship of the species studied. / Doutorado / Parasitologia / Doutor em Parasitologia

Orientador: Patricia Jacqueline Thyssen / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-27T11:10:49Z (GMT). No. of bitstreams: 1 Alonso_MarcelaAquiyama_M.pdf: 1564049 bytes, checksum: 75bec9069782a8cb90070bdbf5ad0c93 (MD5) Previous issue date: 2015 / Resumo: Os insetos constituem o grupo mais diversificado e abundante do Reino Animal, com ampla diversidade morfológica, fisiológica e de hábitos alimentares e por isso podem ser encontrados em vários habitats e ecossistemas. Insetos necrófagos podem ser vestígios e fontes de informações de interesse forense, como para a estimativa do intervalo pós-morte (IPM), baseada, por exemplo, na idade dos imaturos que se criam em corpos em decomposição. O desenvolvimento desses insetos pode ser afetado pela variação de temperatura e presença de substâncias tóxicas nos tecidos de um cadáver, entre outros fatores. Chrysomya megacephala (F.) e Chrysomya putoria (W.) (Diptera: Calliphoridae), introduzidas no Brasil, são consideradas de importância forense, médico e veterinária, devido aos seus comportamentos sinantrópicos e necrófagos. No presente estudo foi avaliado o tempo de desenvolvimento de imaturos na fase embrionária e pós-embrionária de C. megacephala e C. putoria sob diferentes temperaturas e / ou presença de cloridrato de fluoxetina, um antidepressivo, em fígado de coelho, o substrato alimentar. A relação entre temperatura e desenvolvimento, na fase embrionária, foi similar entre ambas as espécies. O tempo de desenvolvimento dos ovos para C. megacephala variou aproximadamente de 64-7h a 13 e 35 °C, respectivamente, e para C. putoria de 69-8h a 13 e 35 °C, respectivamente. Houve eclosão de larvas a 13 °C, mas as mesmas não completaram o desenvolvimento. A temperatura e o cloreto de fluoxetina afetaram o desenvolvimento dos imaturos, na fase na pós-embrionária. Para ambas as espécies, as larvas do grupo controle completaram seu desenvolvimento 24h mais rápido que o grupo com fluoxetina a 17 °C, mas apresentaram o desenvolvimento 12h mais lento a 35 °C. Estudos considerando tempo real de desenvolvimento dos ovos e avaliando como a combinação de duas ou mais variáveis podem influenciar o desenvolvimento de insetos de interesse forense são de grande valia para aumentar a acurácia da estimativa do IPM / Abstract: Insects are the most diverse and abundant group of the Animal Kingdom, with great diversity of morphological, physiological and feeding habits and are found in nearly all habitats and ecosystems. Scavengers species can provide important information of forensic interest, as the post-mortem interval (PMI) estimate, based on, e.g., the age of the larvae reared in decomposing bodies. The development of these insects can be affected by temperature oscillation and presence of toxic substances, among other factors, on the rearing media. Chrysomya megacephala (F.) and Chrysomya putoria (W.) (Diptera: Calliphoridae), introduced in Brazil, are considered of forensic, medical and veterinary importance, due to their necrophagous and synanthropic behaviour. This study evaluated the developmental time of C. megacephala and C. putoria under different temperatures and / or with fluoxetine hydrochloride, an antidepressant drug, in rabbit liver, the rearing substrate. The relationship between temperature and development, on the embryonic phase, was similar for both species. Egg developmental time for C. megacephala was approximately of 64-7h at 13 and 35 °C, respectively, and for C. putoria was 69-8h at 13 and 35 °C, respectively. The larval hatching occurred at 13 °C, but, at this temperature, the larval development was not completed. Both temperature and fluoxetine hydrochloride, when present, affected the development of the larvae. For both species, the larvae of control group completed their development 24h faster than the fluoxetine hydrochloride group at 17 °C, but the development was 12h slower at 35 °C. Studies considering real egg developmental time and evaluating, simultaneously, the insects' response for two or more variables that might influence their development are of great value to increase the accuracy of PMI estimate / Mestrado / Relações Antrópicas, Meio Ambiente e Parasitologia / Mestra em Biologia Animal