Gonotrophic discordance in Culex quinquefasciatus (Diptera: Culicidae) in the city of São Paulo, Brazil

Abstract INTRODUCTION: This study aimed to assess the occurrence of gonotrophic discordance in females of Culex quinquefasciatus in São Paulo, Brazil. METHODS: Resting females were collected monthly for 8 months. Females of Cx. quinquefasciatus were identified, and their midgut and ovaries were dissected. RESULTS: Two hundred females were dissected, out of which, 27.5% were nulliparous and 57% were parous. Most females had no blood in the midgut, but gonotrophic discordance was found in 21% females. CONCLUSIONS: Females of Cx. quinquefasciatus showed a high parity rate and gonotrophic discordance, which could favor the vector capacity of this species.

Assessment of the safety and efficacy of low pathogenic avian influenza (H9N2) virus in inactivated oil emulsion vaccine in laying hens

In Korea, several outbreaks of low pathogenic AI (H9N2) viral infections leading to decreased egg production and increased mortality have been reported on commercial farms since 1996, resulting in severe economic losses. To control the H9N2 LPAI endemic, the Korea Veterinary Authority has permitted the use of the inactivated H9N2 LPAI vaccine since 2007. In this study, we developed a killed vaccine using a low pathogenic H9N2 AI virus (A/chicken/Korea/ADL0401) and conducted safety and efficacy tests in commercial layer farms while focusing on analysis of factors that cause losses to farms, including egg production rate, egg abnormality, and feed efficiency. The egg production rate of the control group declined dramatically 5 days after the challenge. There were no changes in feed consumption of all three groups before the challenge, but rates of the control declined afterward. Clinical signs in the vaccinated groups were similar, and a slight decline in feed consumption was observed after challenge; however, this returned to normal more rapidly than the control group and commercial layers. Overall, the results of this study indicate that the safety and efficacy of the vaccine are adequate to provide protection against the AI field infection (H9N2) epidemic in Korea.

Reproductive biology of the skates Sympterygia acuta Garman, 1877 and S. bonapartii Müller & Henle, 1841 (Chondrichthyes: Rajoidei) in south Brazil

The present study analyzed the sexual development, sizes at maturity and morphometric relationships for both sexes of Sympterygia acuta and S. bonapartii, endemic of south-western Atlantic Ocean. The examined specimens were obtained through research cruises and commercial fishing trips, during 2011 and 2012, along the southern Brazilian coast, in latitudes ranging from 34°28'S to 31°29'S and at depths between 15 and 142 m. Significant differences (p<0.05) in S. bonapartii and in S acuta between sexes for the relationships total length (cm) - disc width (cm) and total length - total/eviscerated weight (g) respectively, demonstrated sexual dimorphism during the development in both species. The estimated size at maturity for males and females, respectively, were 46.1 and 44.7 cm, for S. acuta, and 58.4 and 59.9 cm, for S. bonapartii. A decrease in size at maturity for both sexes was observed in S. acuta and females of S. bonapartii of the southern coast of Brazil, respect to previous studies carried out over the last 30 years.

O presente estudo analisou o desenvolvimento sexual, os tamanhos de maturidade e as relações morfométricas para ambos os sexos das espécies Sympterygia acuta e S. bonapartii endêmica do sudoeste do Oceano Atlântico. Os indivíduos foram coletados em cruzeiros de investigação e viagens de pesca comercial durante 2011 e 2012 ao longo da costa do Sul do Brasil em latitudes que variam de 34°28'S a 31° 29'S, e em profundidades entre 15 e 142 m. Houve diferenças significativas entre sexos para S. bonapartii (p <0,05) e S acuta nas relações comprimento total (cm) - largura do disco (cm) e comprimento - peso total e comprimento total - peso eviscerado (g), demonstrando dimorfismo sexual durante o desenvolvimento em ambas as espécies.Os tamanhos estimados de maturação para machos e fêmeas, respectivamente, foram iguais a 46,1 e 44,7 cm para S. acuta; e 58,4 e 59,9 cm para S. bonapartii. Uma diminuição nos tamanhos de maturidade para ambos os sexos foi observada em S. acuta e fêmeas de S. bonapartii da costa Sul do Brasil, com relação a estudos realizados há 30 anos.

Salmonella Enteritidis Flagella may Enhance Attachment and Invasion of Hen Ovarian Granulosa Cells and Induce Protective Immune Response in Egg-laying Hens.

Aims: To study the effect of flagellin on bacterial attachment and invasion of avian ovary cells in vitro by comparing the attachment and invasion of wild-type S. Enteritidis with nonmotile mutants. To assess the immunogenic properties of extracted flagellin against Salmonella Enteritidis experimental infection in laying hens. Methodology: Non-flagellated mutants for wild-type S. Enteritidis (phage type 8, 13A and 28) were produced by using a strain of S. Enteritidis, SA4502, which carried an fliC::Tn 10 to transfer fliC::Tn 10 insertion into the wild type strains using phage 22 (P22)-mediated transduction with selection for antibiotic resistance encoded within the mutant alleles. Granulosa cells were harvested from Single Comb White Leghorn hens between 18-45 weeks of age. Flagellin was purified from the studied bacterial cultures of Salmonella Enteritidis following reported methods. Laying hens were immunized with the flagellin with adjuvant Results: Non-motile mutants of S. Enteritidis phage wild types were analyzed to confirm the elimination of H1 flagellin synthesis. Wild-type and fliC mutant strains were assessed for their ability to adhere to hen's ovarian granulosa cells. The adherence of the mutant strain was reduced nearly ten-fold compared with that of the wild-type phage 8. Similarly, light microscopic observation of fixed cover slips from wild-type phage types and its mutant strain revealed fewer numbers of the bacterial mutants adhered to the cultured granulosa cell monolayer. Light microscopy revealed similar findings for mutant phage types 28 and 13 A when compared to the wild-type control. There was five folds rise in the egg yolk antibody during the 2-3 weeks post-immunization. No rise was detected in the egg yolk samples from the control hens injected with the placebo mixture without flagellin. Conclusion: It was concluded that Flagellin has an important role in the attachment and invasion of Salmonella Enteritidis to avian ovary cells and that it can be used as immunogenic components to induce a protective immune response in vaccinated hens against challenge infection with the wild type strains.

Bionomics of Culex quinquefasciatus within urban areas of Rio de Janeiro, Southeastern Brazil / Bionomia de Culex quinquefasciatus em áreas urbanas no Rio de Janeiro, RJ / Bionomía de Culex quinquefasciatus en áreas urbanas en Rio de Janeiro, Brasil

OBJECTIVE: To evaluate density, parity rates, daily survival and longevity of natural populations of Culex quinquefasciatus in three neighborhoods with distinct socio-economic and infrastructure profiles. METHODS: Mosquito collections of the Culex quinquefasciatus species were performed weekly during two four month periods, from August to November 2008 (spring) and March to June 2009 (fall), in a favela (slum), a suburban area and a middle class area of Rio de Janeiro, Southeastern Brazil. Collections were performed with backpack aspirators, in 20 randomly selected houses in each area per week, during 15-20 minutes per house. Ovaries were removed from captured females and classified as initial, intermediary or final stage. Furthermore, females were dissected for determination of parity based on the condition of the tracheal system. Mosquito survival rate and longevity were estimated on a per month basis for each neighborhood. RESULTS: We collected a total of 2,062 Culex quinquefasciatus, but monthly vector density was not correlated with temperature and rainfall. We dissected the ovaries of 625 Culex quinquefasciatus, and overall, there was a higher proportion of nulliparous females during the dryer months, while gravid females were more frequent in rainy months. In the middle class neighborhood, the parity rate reached up to 93.75% with survivorship of 0.979. Lower parity and survival rates were obtained in the suburban area (as low as 36.4% parity and 0.711 daily survival). Up to 84.7% of Culex quinquefasciatus females could survive the eight day period needed to complete West Nile Virus incubation. CONCLUSIONS: The survival rate of Culex quinquefasciatus varied significantly between the neighborhoods. This suggests that vectorial capacity and disease transmission risk may vary greatly between different urban areas, which is potentially useful information for vector control programs.

OBJETIVO: Avaliar densidade, taxa de paridade, sobrevivência diária e longevidade de populações naturais de Culex quinquefasciatus de três localidades com diferentes perfis socioeconômicos e de infraestrutura. MÉTODOS: Mosquitos da espécie Culex quinquefasciatus foram coletados semanalmente em dois períodos de quatro meses, de agosto a novembro de 2008 e de março a junho de 2009, em uma comunidade, uma área de subúrbio e um bairro de classe média no Rio de Janeiro, RJ. As coletas foram realizadas com o auxílio de aspiradores costais em 20 domicílios, aleatoriamente selecionados por semana e por área, durante 15-20 minutos por domicílio. As fêmeas capturadas tiveram os ovários removidos e classificados em estágios inicial, intermediário e final. Adicionalmente, as fêmeas foram dissecadas para determinação da paridade de acordo com as condições do sistema traqueal. A taxa de sobrevivência e a longevidade das fêmeas foram estimadas por mês, para cada localidade. RESULTADOS: Foram coletados 2.062 Culex quinquefasciatus, porém a densidade mensal deste vetor não apresentou correlação com temperatura e precipitação. Dissecamos os ovários de 625 Culex quinquefasciatus e obtivemos maior proporção de fêmeas nulíparas durante os meses mais secos, enquanto fêmeas grávidas foram mais frequentes nos meses chuvosos. A taxa de paridade foi de até 93,75% no bairro de classe média, com sobrevivência de 0,979. Menores valores de paridade e sobrevivência foram obtidos no subúrbio (começando em 36,4% de paridade e 0,711 de sobrevivência diária). Até 84,7% das fêmeas de Culex quinquefasciatus poderiam sobreviver ao período de oito dias, necessário para completar o período de incubação do vírus da febre do Nilo Ocidental. CONCLUSÕES: A taxa de sobrevivência de Culex quinquefasciatus variou significativamente entre os bairros, o que sugere que a capacidade vetorial e o risco de transmissão de doenças podem variar entre diferentes áreas urbanas, informação relevante para o planejamento dos programas de controle de vetores.

OBJETIVO: Evaluar densidad, tasa de paridad, sobrevivencia diaria y longevidad de poblaciones naturales de Culex quinquefasciatus de tres localidades con diferentes perfiles socioeconómicos y de infraestructura. MÉTODOS: Mosquitos de la especie Culex quinquefasciatus fueron colectados semanalmente en dos períodos de cuatro meses, de agosto a noviembre de 2008 y de marzo a junio de 2009, en un barrio, un área de suburbio y una urbanización de clase media en Rio de Janeiro, Litoral Sudeste de Brasil. Las colectas se realizaron con ayuda de aspiradores costales en 20 domicilios, aleatoriamente seleccionados por semana y por área, durante 15-20 minutos por domicilio. A las hembras capturadas se les retiraron los ovarios y clasificaron en fases inicial, intermediario y final. Adicionalmente, las hembras fueron diseccionadas para determinar la paridad de acuerdo con las condiciones del sistema traqueal. La tasa de sobrevivencia y la longevidad de las hembras fueron estimadas por mes para cada localidad. RESULTADOS: Se colectaron 2.062 Culex quinquefasciatus, sin embargo, la densidad mensual del vector no presentó correlación con la temperatura y precipitación. Se diseccionaron los ovarios de 625 Culex quinquefasciatus y se obtuvo mayor proporción de hembras nulíparas durante los meses más secos, mientras que las hembras grávidas fueron más frecuentes en los meses de lluvia. La tasa de paridad alcanzó el 93,75% en la urbanización de clase media, con sobrevivencia de 0,979. Menores valores de paridad y sobrevivencia se obtuvieron en el suburbio (comenzando en 36,4% de paridad y 0,711 de sobrevivencia diaria). El 84,7% de las hembras de Culex quinquefasciatus podrían sobrevivir al período de ocho días, necesario para completar el período de incubación del virus de la fiebre del Oeste del Nilo. CONCLUSIONES: La tasa de sobrevivencia de Culex quinquefasciatus varió significativamente entre las urbanizaciones, sugiriendo que la capacidad vectorial y el riesgo de transmisión de enfermedades pueden variar entre diferentes áreas urbanas, información relevante para la planificación de los programas de control de vectores.

Comparison of whole-genome amplifications for microsatellite genotyping of Rotylenchulus reniformis

Currently, a large number of microsatellites are available for Rotylenchulus reniformis (reniform nematode); however, two barriers exist for genotyping samples from different geographical areas. The limited amount of nucleic acids obtained from single nematodes which would require their multiplication to obtain enough DNA for testing; and the strictly regulated transport of live samples and multiplication in greenhouse for being a plant pathogen. Whole-genome amplification (WGA) of samples consisting of one and five dead gravid females with their associated egg masses was successfully performed on disrupted tissue using three commercial kits. DNA yield after WGA ranged from 0.5 to 8 ug and was used to test 96 microsatellite markers we previously developed for the reniform nematode. The results were compared to those of fingerprinting the original population (MSRR03). Out of 96 markers tested, 71 had amplicons in MSRR03. Using WGA of single gravid females with their associated egg masses, 86-93 percent of the alleles found on MSRR03 were detected, and 87-88 percent of the alleles found on MSRR03 when using WGA of samples composed of five gravid females with their associated egg masses as template. Our results indicate that reniform nematode samples as small as a single gravid female with her associated egg mass can be used in WGA and direct testing with microsatellites, giving consistent results when compared to the original population.

Effects of heat stress on some blood parameters and egg production of Shika Brown layer chickens transported by road

The effects of vitamins C and E on layer chickens transported by road for 6 h during the hot dry season were investigated. Two experimental groups consisting of thirty Shika Brown layers were separately administered vitamins C and E orally just before transportation, while another 30 layers, which were only given sterile water, served as control. Blood samples analyzed before and after transportation in the control layers showed a decrease (p<0.05) in total white blood cell, (p<0.01) lymphocyte and monocyte values, and a signifcant (p<0.05) and (p<0.001) increase in the values of eosinophils and heterophils post-transportation, respectively. In the experimental groups, post-transportation values of total white blood cells, eosinophils and monocytes were not different (p>0.05) from those obtained before transportation. Heterophil/lymphocyte values were highest in the control group. The result showed that transportation was stressful for the control layers. Post-transportation egg production was not signifcantly (p>0.05) different in the vitamin E treated group, but values recorded for the vitamin C and control groups were signifcantly (p<0.05) and (p<0.001) reduced compared to pre-transportation. In conclusion, vitamins C and E administration ameliorated the adverse effect of road transportation stress during the hot dry season.

Control of Salmonella enterica serovar Enteritidis in laying hens by inactivated Salmonella Enteritidis vaccines / "Controle de Salmonella enterica sorovar Enteritidis em poedeiras comerciais com a utilização de vacinas inativadas"

Salmonella Enteritidis is one of the agents that is responsible for outbreaks of human foodborne salmonellosis caused by Salmonella Enteritidis and is generally associated with the consumption of poultry products. Inactivated Salmonella Enteritidis cell vaccine is one of the available methods to control Salmonella Enteritidis in breeders and laying hens, however results in terms of efficacy vary. This vaccine has never been tested in Brazil, therefore, the present work was carried out to assess three commercial inactivated Salmonella Enteritidis vaccines allowed in Brazil. Four hundred white light variety commercial laying hens were obtained at one-day-of age. At eight weeks old, the birds were divided into four groups with one hundred animals each. Birds from three groups (V1, V2 and V3) received different intramuscular vaccines, followed by a booster dose at 16 weeks of age. Birds from another group (CG) were not vaccinated. When the laying hens were 20, 25 and 31 weeks old, 13 from each group were transferred to another room and were challenged by inoculating 2 mL neat culture of Salmonella Enteritidis. On the second day after each challenge, the caecal contents, spleen, liver and ovary of three birds from each group were analyzed for the presence of Salmonella Enteritidis. Twice a week a cloacal swab of each bird was taken and all eggs laid were examined for the presence of Salmonella Enteritidis. After four consecutive negative cloacal swabs in all the groups, the birds were sacrificed so as to examine the liver, caecal contents and ovaries. Overall, the inactivated vaccine used in group V3 reduced Salmonella Enteritidis in the feces and eggs. A very small amount of Salmonella was found in the spleen, liver, ovary and caeca of the birds in the four groups during the whole experiment. In general, inactivated Salmonella Enteritidis vaccines was able to decrease the presence of Salmonella Enteritidis in the birds and in the eggs as...

Salmonella Enteritidis é um dos agentes responsáveis por toxinfecção alimentar em humanos e tem sido associada a alimentos de origem avícola. Entre os métodos disponíveis para o seu controle está a vacinação de poedeiras e matrizes com vacinas inativadas (bacterinas). Os resultados a respeito da proteção das bacterinas contra Salmonella Enteritidis em aves são variados. Face à inexistência de dados referentes ao uso dessas vacinas no Brasil, realizou-se o presente trabalho. Foram utilizadas 400 pintinhas de uma linhagem de postura leve. Na 8º semana de idade, as aves foram divididas em quatro grupos (V1, V2, V3 e CG). Três diferentes bacterinas comerciais foram administradas às aves do V1, V2 e V3 em duas doses, na 8º e 16º semanas de vida; as do CG não receberam vacina. Treze aves por grupo foram infectadas com Salmonella Enteritidis nas 20º, 25º e 31º semanas. No 2º dia após cada desafio foram sacrificadas três aves por grupo, para contagem de Salmonella Enteritidis em fígado, baço, conteúdo cecal e pesquisa do microrganismo no ovário. Suabes de cloaca foram realizados dois dias pós-infecção (dpi) e duas vezes por semana. Todos os ovos foram examinados. Após a ausência de Salmonella Enteritidis em quatro suabes de cloaca consecutivos, esse microrganismo foi pesquisado em fígado, conteúdo cecal e ovário. Não houve diferença da contagem de Salmonella Enteritidis nos órgãos. O conteúdo cecal das aves do V1 teve menos Salmonella que as do CG. As aves do V3 excretaram menos Salmonella em fezes e ovos. Conforme os resultados observados no V3, é possível reduzir excreção de Salmonella Enteritidis com o uso de bacterinas; contudo, deve ser utilizado de forma complementar a boas práticas de manejo, limpeza e desinfecção.

Biological activities of chamomile (Matricaria chamomile) flowers' extract against the survival and egg laying of the cattle fever tick (Acari Ixodidae) / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

In the present work, the potential of acaricidal activity of chamomile flowers' extract was studied against engorged Rhipicephalus annulatus tick under laboratory condition. For this purpose, the engorged females of Rhipicephalus annulatus were exposed to two-fold serial dilutions of chamomile flowers' extract (0.5%, 1.0%, 2.0%, 4.0% and 8.0%) using "dipping method" in vitro. The engorged ticks were immersed in different plant dilutions (five ticks for each dilution) for 1 min and they were immediately incubated in separate Petri dishes for each replicate at 26 degrees C and 80% relative humidity. Mortality rate for each treatment was recorded 5 d after incubation. The mortality rate caused by different dilutions of chamomile flower' extract ranged from 6.67% to 26.7%, whereas no mortality was recorded for non-treated control group. The mass of produced eggs varied form 0.23 g (in 8.0% solutions) to 0.58 g (in control), with no statistical differences between the treatments and control (P>0.05). Also the chamomile flowers' extract in highest concentration used (8.0%) caused 46.67% failure in egg laying in engorged females while non failure was observed for non-treated control group. Macroscopic observations indicated that in effective concentrations of plant (4.0% and 8.0%), patchy hemorrhagic swelling appeared on the skin of treated ticks. The results presented for the first time in this study imply that chamomile may be considered as a promising plant for biocontrol of cattle fever tick disease in the field condition.