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Leung, Y. Tai Ida. "Infections à vibrio vulnificus, revue de la littérature à propos d'une observation de septicémie". Bordeaux 2, 1996. http://www.theses.fr/1996BOR2M053.
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Limthammahisorn, Suttinee Brady Yolanda Juanita Arias Covadonga R. "In vitro and in vivo cold shock response in Vibrio vulnificus". Auburn, Ala., 2007. http://repo.lib.auburn.edu/Send%2002-04-08/LIMTHAMMAHISORN_SUTTINEE_24.pdf.
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Meyer, Shelli Lee. "Vibrio vulnificus dynamics in a south Texas bay". [College Station, Tex. : Texas A&M University, 2007. http://hdl.handle.net/1969.1/ETD-TAMU-1587.
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Kural, Ayse G. "Temperature-assisted pressure inactivation of Vibrio parahaemolyticus and Vibrio vulnificus in oysters". Access to citation, abstract and download form provided by ProQuest Information and Learning Company; downloadable PDF file, 89 p, 2007. http://proquest.umi.com/pqdweb?did=1338870531&sid=16&Fmt=2&clientId=8331&RQT=309&VName=PQD.
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Raszl, Simone Moraes. "Vibrio vulnificus em ostras (Crassostrea gigas) em Santa Catarina". reponame:Repositório Institucional da UFSC, 2016. https://repositorio.ufsc.br/xmlui/handle/123456789/171718.
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Tese (doutorado) - Universidade Federal de Santa Catarina, Centro de Ciências Agrárias, Programa de Pós-Graduação em Ciência dos Alimentos, Florianópolis, 2016.Made available in DSpace on 2016-12-20T03:16:01Z (GMT). No. of bitstreams: 1 343109.pdf: 4273942 bytes, checksum: 0190bb7c669406b691be6cda46663d8d (MD5) Previous issue date: 2016
Vibrio vulnificus é um habitante natural de ambientes marinhos que pode infectar seres humanos através de ferimentos na pele ou pelo consumo de frutos do mar, especialmente ostras consumidas cruas ou mal cozidas. A bactéria pode atravessar a barreira intestinal e alcançar a corrente sanguínea causando septicemia, que pode levar à morte. A hipótese científica deste estudo foi a de que existem cepas potencialmente patogênicas de V. vulnificus, identificadas pelo gene vcgC, nas ostras produzidas no litoral de Santa Catarina. Os objetivos desta tese foram verificar a ocorrência cepas de V. vulnificus com potencial patogenicidade em ostras (Crassostrea gigas) produzidas em Santa Catarina; realizar estudo retrospectivo inédito sobre presença de V. vulnificus e de V. parahaemolyticus na América do Sul; e comparar a performance dos meios de cultivo microbiológico mais comuns para detecção de V. vulnificus quanto a sua capacidade de isolamento e enumeração. A hipótese foi confirmada na análise das amostras coletadas, com confirmação por PCR, indicando a presença de cepas potencialmente patogênicas de V. vulnificus. O estudo de revisão indicou que as infecções causadas por V. parahaemolyticus tem sido mais fortemente relacionadas à ingestão de alimentos de origem marinha e mais frequentes na costa do Oceano Pacífico. Por outro lado, V. vulnificus é mais frequentemente adquirido por contato de lesões de pele e sua presença é mais comum na costa do Oceano Atlântico. Além disso, o estudo indicou que o fenômeno El Niño exerce influência no aumento de casos de V. parahaemolyticus na costa do Pacífico. Entretanto, são necessários mais estudos para confirmar a importância de fatores ambientais na presença e virulência da bactéria nesta região. No Brasil, os casos de infecções causadas por V. vulnificus são restritos basicamente às regiões sul e sudeste, exceto por um caso recente em 2016, na região nordeste do Brasil. Na análise das amostras coletadas, o estudo revelou uma baixa incidência (4%) de V. vulnificus entre as amostras de ostras (C. gigas) durante os meses de outubro/2014 a janeiro/2015, confirmadas por PCR através dos genes vcg. Esta baixa incidência pode ser explicada pela alta salinidade na região, que ultrapassa os valores de 34?. Na análise da performance dos meios usando cepas puras de V. vulnificus de genótipos conhecidos vcgC e vcgE, todos os meios analisados apresentaram performance satisfatória. Com relação à avaliação da performance dos meios para isolamento eenumeração de V. vulnificus em amostras ambientais, o meio CPC+ apresentou a melhor performance. Assim, mesmo tendo em conta a ausência de relatos de casos de infecção humana em Santa Catarina e os fatores ambientais citados, é fundamental a criação de um programa de monitoramento contínuo para avaliar a presença da bactéria, assim como a adoção de um procedimento de comunicação para pessoas portadoras de condições predisponentes e para profissionais de saúde, para que possam orientar seus pacientes e que possam realizar o diagnóstico e consequente tratamento de pessoas acometidas, com a adoção de metodologias mais precisas, rápidas e específicas para detecção de cepas patogênicas, como a detecção do gene vcg por PCR, especialmente no diagnóstico de casos suspeitos, garantindo maior precisão e eficácia de tratamento.
Abstract : Vibrio vulnificus is a natural inhabitant of marine environments that can infect humans through skin wounds or seafood ingestion, especially raw or undercooked shellfish. The bacteria can cross the intestinal barrier and reach the blood stream causing sepsis that can lead to death. The scientific hypothesis of this study was that there are potentially pathogenic strains of V. vulnificus, identified by vcgC gene, on the oysters produced in the coast of Santa Catarina. The objectives of this thesis were to assess the occurrence of potentially pathogenic V. vulnificus strains in oysters (Crassostrea gigas) produced in Santa Catarina; to achieve unprecedented retrospective study on the presence of V. vulnificus and V. parahaemolyticus in South America; and compare the performance of the most common media for microbiological culture used to detect V. vulnificus as its isolation and enumeration capacity. The hypothesis was confirmed through the analysis of samples, confirmed by PCR. The review indicated that the infections caused by V. parahaemolyticus have been more closely related to the ingestion of seafood and more frequent in the Pacific Ocean. On the other hand, V. vulnificus is most often acquired by contact of skin lesions and their presence is more common in the Atlantic Ocean. In addition, the study indicated that the El Niño Southern Oscillation influences the increase in cases of V. parahaemolyticus on the Pacific coast. However, more studies are needed to confirm the importance of environmental factors in the presence and virulence of the bacteria in this region. In Brazil, cases of infection caused by V. vulnificus are basically restricted to the southern and southeastern areas, except for a recent case in 2016, in northeastern Brazil. The study revealed a low incidence (4%) of V. vulnificus between oyster samples (C. gigas) produced in Santa Catarina during the months of October/2014 to January/2015, confirmed by PCR using vcg genes. This low rate can be explained by high salinity in the region, which exceeds 34?. In the media performance research using V. vulnificus pure strains with known vcgC and vcgE genotypes, when considering the performance for total V. vulnificus, all analyzed media showed satisfactory performance. Regarding the assessment of the performance of the media for V. vulnificus isolation and enumeration in environmental samples, CPC+ had the best performance. Thus, considering the absence of reported cases of human infection in Santa Catarina and the environmental factors mentioned, the creation of a continuous monitoring program is critical to assess the presence ofbacteria, as well as the adoption of a communication procedure for people with predisposing conditions and health professionals so that they can guide their patients and can make the diagnosis and subsequent treatment of affected people, and to adopt more precise methodologies, for rapid and specific detection of pathogenic strains such as the detection of the vcg gene by PCR, especially in the case of diagnosis of suspected cases, ensuring greater accuracy and efficacy of treatment.
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Rubio, Galleguillos Felipe Andrés. "Implementación de técnica para la detección de vibrios y análisis de Vibrio vulnificus en muestras de alimentos". Tesis, Universidad de Chile, 2006. http://repositorio.uchile.cl/handle/2250/105607.
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Unidad de práctica para optar al título de Químico FarmacéuticoNo autorizada por el autor para ser publicada a texto completo en el Portal de Tesis Electrónicas
La practica prolongada fue realizada en Departamento de Microbiología de Alimentos del Instituto se Salud Pública de Chile, durante los meses de Junio a Diciembre del año 2005. Mi labor en la práctica constó de tres etapas simultáneas: La primera consistió en recibir capacitación durante todo el período de mi práctica, en detección y cuantificación de los patógenos bacterianos más frecuentes encontrados en alimentos. La segunda etapa fue implementar el último método de detección de Vibrios (principalmente Vibrio vulnificus en mi caso) según el Manual Analítico Bacteriológico (BAM) año 2004 impuesto por la Food and Drugs Administration (FDA) (1), para la cual se utilizaron cepas de diversos Vibrios tales como Vibrio vulnificus, Vibrio parahaemolyticus, Vibrio cholerae, etc. Dicha labor será necesaria para actualizar los métodos de detección en los laboratorios de la red de vigilancia epidemiológica de Chile. En la otra etapa, se evaluó según el método que se estaba desarrollando la población de Vibrio vulnificus existente en Puerto Montt, ésta etapa se comenzó a trabajar a partir aproximadamente en julio ya que primero debía estar desarrollada en alguna medida la técnica de detección, además de que estuvieran los materiales necesarios para el trabajo. Las muestras analizadas son ensayos de rutina que llegan al SEREMI de Salud de la región Metropolitana para evaluar durante todo el año la población de Vibrio parahaemolyticus y Vibrio cholerae en Puerto Montt, las cuales se reciben todos los martes y consta de 5 muestras de los cuales se analizan 12 especimenes de cada una, para luego cuantificarlas por el método de tubos múltiples según la tabla de número más probable (anexo, Tabla 1). Es importante que el ISP, como centro de referencia posea información sobre todo patógeno que pueda ser encontrado en alimentos, es por eso que esta etapa culminará con la entrega del procedimiento de detección de Vibrio vulnificus al Departamento de Microbiología de Alimentos
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Church, Selina Rebecca. "Investigating pathogenesis and virulence of the human pathogen, Vibrio vulnificus". Thesis, University of Exeter, 2015. http://hdl.handle.net/10871/17600.
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V. vulnificus is a Gram negative opportunistic pathogen that is ubiquitous in the marine environment. Of the three main biotypes, biotype 1 is most commonly associated with human infection and is the causative agent of septicaemia, gastroenteritis and wound infection. In the United States V. vulnificus is the leading cause of seafood related deaths and is commonly associated with ingestion of raw or undercooked oysters. However, despite the abundant prevalence of this bacterium in the environment, the number of severe human infections is low. This has led to the hypothesis that not all strains of this pathogen are equal in virulence, with some strains better adapted to causing human disease than others. Therefore the current study tested a panel of 10 V. vulnificus strains in several phenotypic experiments that assayed the strains for known virulence factors, with the aim of identifying a marker for strains hazardous to human health. However, not one assay correlated with either virulence potential of the strains, as determined by an in vivo mouse model of virulence, or source of isolation. As the study hypothesised that the varying virulence potentials displayed by the strains may be due to genetic differences, whole genome sequencing (WGS) was performed. Bioinformatic comparison of the strains demonstrated many genetic differences between the strains. However, in unison with the WGS comparison, WGS gene annotation was also performed. This identified the presence of two previously undescribed type 6 secretion systems (T6SS). Therefore the current study continued investigation into the T6SSs. The two T6SSs identified were termed T6SS1 and T6SS2. T6SS2 was found in all sequenced isolates, whereas T6SS1 was only present in a sub-set of strains. As T6SS1 shared synteny with the previously described T6SS in V. cholerae, T6SS1 was chosen for further investigation. During this study T6SS1 was shown to be functional and displayed thermoregulation. Further investigation into T6SS1 by construction and characterising of a T6SS1 mutant, demonstrated that T6SS1 contained anti-prokaryotic properties.
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Paranjpye, Rohinee. "The role of a Vibrio vulnificus type IV pilin in pathogenesis and in persistence in oysters /". Thesis, Connect to this title online; UW restricted, 2005. http://hdl.handle.net/1773/5372.
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McDougald, S. Diane School of Microbiology & Immunology UNSW. "Regulation of starvation and nonculturability in the marine pathogen, Vibrio vulnificus". Awarded by:University of New South Wales. School of Microbiology and Immunology, 2000. http://handle.unsw.edu.au/1959.4/19118.
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Vibrio vulnificus is a model environmental organism exhibiting a classical starvation response during nutrient limitation as well as a non-culturable state when exposed to low temperatures. In addition to these classic global responses, this organism is an opportunistic pathogen that exhibits numerous virulence factors. This organism was chosen as the model organism for the identification of regulators of the viable but nonculturable response (VBNC) and the starvation-induced maintenance of culturability (SIMC) that occurs when cells are starved prior to low temperature incubation. In order to accomplish this, three indirect approaches were used; proteomics, investigation of intercellular signalling pathways and genetic analysis of regulators involved in these responses. Two-dimensional gel electrophoresis was used to identify proteins expressed under conditions that induced SIMC. It was determined that carbon and long-term phosphorus starvation were important in the SIMC response. V. vulnificus was shown to possess genes, luxS and smcR, that are homologues of genes involved in signalling system system 2 in Vibrio harveyi. Signal molecules were produced upon starvation and the entry to stationary phase in V. vulnificus. Furthermore, a null mutation in smcR, a transcriptional regulator was shown to have pleiotropic effects in V. vulnificus, including up-regulation of numerous virulence factors and a defect in starvation survival and development of the SIMC response. We propose that V. vulnificus possesses a signalling system analogous to that of system 2 in V. harveyi, and that this system is involved in the regulation of stationary phase and starvation adaptation in this organism.
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Gordon, Katrina V. "Multilocus Virulence Typing of Clinical and Environmental Vibrio vulnificus Isolates". [Tampa, Fla] : University of South Florida, 2008. http://purl.fcla.edu/usf/dc/et/SFE0002557.
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Brenton, Carolyn E. "Effect of storage temperatures on the microbiological quality, safety, and viability of quahog clams, Mercenaria mercenaria". Thesis, This resource online, 1996. http://scholar.lib.vt.edu/theses/available/etd-06112009-063149/.
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Ramos, Roberta Juliano. "Vibrio sp. em ostras e águas de áreas de cutivo da Baía Sul de Santa Catarina". Florianópolis, 2012. http://repositorio.ufsc.br/xmlui/handle/123456789/99318.
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Tese (doutorado) - Universidade Federal de Santa Catarina, Centro de Ciências Agrárias. Programa de Pós-graduação em Ciências dos AlimentosMade available in DSpace on 2013-03-04T19:09:44Z (GMT). No. of bitstreams: 1 309352.pdf: 1330978 bytes, checksum: 53f32bc5f3243f33f297d2c1954775e2 (MD5)
Esta pesquisa teve por objetivo identificar e quantificar vibrios marinhos potencialmente patogênicos em ostras (Crassostrea gigas) e águas provenientes da Baía Sul de Santa Catarina, Brasil. Assim como avaliar a patogenicidade e susceptibilidade destas cepas frente a diferentes antibióticos e verificar a eficácia da depuração de ostras (Crassostrea gigas) contaminadas com Vibrio parahaemolyticus ou Vibrio vulnificus usando luz ultravioleta (UV) isoladamente e efeito sinérgico com cloro. Das 60 amostras de ostras 48,3% estavam contaminadas por uma ou mais espécies de vibrio. As espécies mais frequentes foram V. parahaemolyticus e V. vulnificus. As contagens de V. parahaemolyticus e V. vulnificus nas amostras variaram entre < 0,5 log10 NMP g-1 a 2,3 log10 NMP g-1 de ostra, e < 0,5 log10 NMP g-1 a 2,1 log10 NMP g-1 de ostra, respectivamente. Das 60 amostras de água 73,3% estavam contaminadas por uma ou mais espécies de vibrios. As contagens de V. parahaemolyticus e V. vulnificus nas amostras variaram entre < 0,3 log10 NMP 100 mL-1 a 1,7 log10 NMP 100 mL-1 de água marinha, e < 0,3 log10 NMP 100 mL-1 a 2,0 log10 NMP 100 mL-1 de água marinha, respectivamente. Das 120 cepas enviadas para tipificação, foram avaliadas as características de 106 cepas reativadas. Destas 83 (78,3%) foram previamente identificadas como V. parahaemolyticus, 20 (18,9%) como V. vulnificus, três (2,8%) como V. cholerae. Foram caracterizados 20 diferentes sorotipos de V. parahaemolyticus, e identificados genes de patogenicidade tanto de V. parahaemolyticus quanto de V. cholerae. A resistência à gentamicina e cefuroxima foi observada em 84,3% das cepas testadas para ambos antibióticos, e a resistência a ampicilina foi observada em 80,0% das cepas de Vibrio spp. O cloranfenicol foi o antibiótico ao qual as cepas de Vibrio spp. foram mais sensíveis, seguido da tetraciclina, diferente do observado para V. cholerae que apresentou maior sensibilidade a Amoxicilina + clavulanato. Para o experimento de depuração as ostras foram contaminadas com um coquetel de cinco cepas de V. parahaemolyticus ou V. vulnificus ao nível de 104 a 105 UFC mL-1. A depuração foi realizada num sistema fechado de recirculação. As contagens de V. parahaemolyticus ou V. vulnificus foram realizadas no tempo zero, 6, 18, 24 e 48 h. Três tratamentos foram realizados: T1 (controle), T2 (luz UV) e T3 (luz UV mais cloro). Após 48 h de depuração de V. parahaemolyticus, T3 reduziu 3,1 log NMP g-1 e T2 reduziu 2,4 log NMP g-1, enquanto T1 reduziu apenas 2,0 log NMP g-1. Após 48 h de depuração de V. vulnificus, tanto T2, como T3 foram eficientes na redução das contagens, reduzindo a população em 2,5 e 2,4 log NMP g-1, respectivamente, enquanto que T1 reduziu apenas 1,4 log NMP g-1. O tratamento utilizando Luz UV mais cloro foi mais eficiente para controlar V. parahaemolyticus em ostras. Tanto o tratamento utilizando luz UV, como o tratamento utilizando luz UV mais cloro foram eficientes para reduzir as contagens de V. vulnificus. Os resultados desta pesquisa servem de subsídio para analise de risco de vibrios potencialmente patogênicos em ostras.
The purpose of this study was to assess the incidence and level of contamination for Vibrio spp. in seawater and oysters (Crassostrea gigas) harvested in Southern Brazil, as to evaluate the pathogenicity of strains and test the susceptibility of these strains to different antibiotics, and to evaluate efficacy of depuration using UV light and UV light plus chlorinated seawater for decontaminating Vibrio parahaemolyticus and Vibrio vulnificus in oysters. Of the 60 oysters# samples analyzed, 29 (48.3%) contained one or more vibrio species. The most frequently isolated species were Vibrio parahaemolyticus and Vibrio vulnificus. The counts of Vibrio parahaemolyticus and Vibrio vulnificus in the samples ranged between < 0.5 log10 MPN g-1 to 2.3 log10 MPN g-1 of oyster, and < 0.5 log10 MPN g-1 to 2.1 log10 MPN g-1of oyster, respectively. Of the 60 seawater samples analyzed, 44 (73.3%) contained one or more species vibrios. The counts of Vibrio parahaemolyticus and Vibrio vulnificus in the samples ranged between < 0.3 log10 MPN 100 mL-1 to 1.7 log10 MPN 100 mL-1 of seawater, and < 0.3 log10 MPN 100 mL-1 to 2.0 log10 MPN 100 mL-1 of seawater, respectively. Of the 120 strains sent for typing were evaluated the characteristics of the strains 106 reactivated, 83 (78.3%) were previously identified as V parahaemolyticus, 20 (18.9%) as V vulnificus, three (2.8%) and V. cholerae. Were characterized 20 different serotypes of V. parahaemolyticus, and isolated genes of patogenicity for V. parahaemolyticus and V. cholerae. The gentamycin and cefuroxime resistance was observed in 84.3% of the strains to both antibiotics, and the ampicillin resistance was observed in 80.0% of the strains of Vibrio spp. Chloramphenicol is the antibiotic to which the strains of Vibrio spp. were more susceptible. The tetracycline antibiotic was the second to which the strains of V. parahaemolyticus and V. vulnificus were more susceptible than the observed for V. cholerae that showed greater susceptibility to amoxicillin + clavulanate. For depuration the oysters were contaminated with five-strain cocktail of V. parahaemolyticus or V. vulnificus to level of 104 to 105 CFU mL-1. The depuration was conducted in a recirulated closed system. Counts of V. parahaemolyticus or V. vulnificus were determined at zero, 6, 18, 24 and 48 h. Three treatments were conducted: T1 (control treatment); T2 (UV treatment); and T3 (UV plus chlorine treatment). After 48 h of depuration of Vibrio parahaemolyticus, T3 reduced 3.1 log MPN g-1 and T2 reduced 2.4 log MPN g-1, while T1 reduced only 2.0 log MPN g-1. After 48 h of depuration of Vibrio vulnificus, T2 as well as T3 were efficient reducing counts in 2.5 and 2.4 log MPN g-1, respectively; while T1 reduced only 1.4 log MPN g-1. UV light plus chlorine treatment was more efficient for controlling V. parahaemolyticus in oysters. Both UV light and UV light plus chlorine were efficient for V. vulnificus. These results serve as input for risk analysis studies of potentially pathogenic vibrios in oysters in southern Brazil.
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Cañigral, Cárcel Irene. "Desarrollo de métodos moleculares para la detección y caracterización de bacterias patógenas emergentes del género Vibrio en aguas y alimentos". Doctoral thesis, Universitat Politècnica de València, 2011. http://hdl.handle.net/10251/11799.
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Vibrio parahaemolyticus y Vibrio vulnificus son microorganismos patógenos pertenecientes a la familia Vibrionaceae y al género Vibrio. Son microorganismos con morfología ligeramente curvada, gram negativos y oxidasa positivos. En cuanto a sus aspectos ecológicos son halófilos, por tanto suelen estar presentes en aguas marinas costeras y en el interior de moluscos, crustáceos y peces, y tienden a encontrarse en aguas cálidas.
La patología que producen está asociada al consumo de mariscos, moluscos y pescado crudo o poco cocinado y a la exposición a aguas contaminadas. Ambas especies son consideradas patógenos emergentes debido al aumento en su incidencia y a la mayor distribución geográfica de los casos de intoxicación alimentaria, sobre todo en países asiáticos.
La detección e identificación de ambas especies en alimentos de origen marino y agua, presenta gran dificultad, ya que los procedimientos convencionales son largos y tediosos, y pueden proporcionar falsos negativos. Los métodos de detección molecular pueden suponer una alternativa más rápida, sensible y fiable.
Objetivos:
Debido a esto, en este trabajo se han desarrollado métodos de detección y cuantificación de Vibrio spp., Vibrio parahaemolyticus y Vibrio vulnificus mediante PCR tradicional, PCR a tiempo real e hibridación in situ con sondas fluorescentes (FISH) para su aplicación en matrices alimentarias y en agua.
En este estudio se han aplicado los distintos métodos a matrices ambientales (agua de playa y agua residual) y a alimentos de origen marino.
Resultados:
Los resultados muestran que la PCR a tiempo real es el método más rápido y sensible para la detección de las diferentes especies estudiadas, además de permitir la cuantificación del microorganismo en las muestras de forma fiable.
En este trabajo hemos demostrado la presencia de bacterias del género Vibrio en ambos tipos de matrices y el potencial de aguas y alimentos de origen marino para actuar como vehículos de transmisión de V. parahaemolyticus y V. vulnificusCañigral Cárcel, I. (2011). Desarrollo de métodos moleculares para la detección y caracterización de bacterias patógenas emergentes del género Vibrio en aguas y alimentos [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/11799
Palancia
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La, Valley Kenneth John. "Aspects of bacterial community ecology within the American oyster (Crassostrea virginica), and surveillance of post-harvest Vibrio vulnificus occurrence /". View online ; access limited to URI, 2005. http://0-wwwlib.umi.com.helin.uri.edu/dissertations/dlnow/3188062.
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Ostrander, Vicki C. "Survival of Vibro vulnificus and other Vibrios in raw oysters (Crassostrea virginica) during processing in Virginia and cold storage". Thesis, This resource online, 1996. http://scholar.lib.vt.edu/theses/available/etd-11012008-063712/.
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Jones, Melissa Kolsch. "Regulation of phase variation and deletion mutation in the Vibrio vulnificus group 1 CPS operon". [Gainesville, Fla.] : University of Florida, 2006. http://purl.fcla.edu/fcla/etd/UFE0013839.
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Chang, Wan-Hsui, i 張莞詡. "Analysis of cytotoxins of Vibrio vulnificus". Thesis, 2013. http://ndltd.ncl.edu.tw/handle/17013026506293525000.
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碩士嘉南藥理科技大學
生物科技系
101
Vibrio vulnificus is a human pathogen. It causes serious wound infection and fatal septicemia. The cytotoxicity is regarded as the important mechanism in the pathogenesis of this bacterium infection. V. vulnificus RtxA toxin is composed of 4701 amino acids wiht a molecuar mass of ~501 kDa. The rtxA inactivation resulted in decrease in cytotoxicity of Vibrio vulnificus in host cells. In previous studies, we found that the h contributes cytotoxicity to V. vulnificus in macrophages. To examine the cytotoxic effect of RtxA and H toxins of V. vulnificus on macrophages, macrophages were treated with V. vulnificus strains lacking rtxA or h or rtxA/h or culture supernatants of the mutants. The mutants lacking rtxA, h or rtxA/h induced a significant decrease of cytotoxicity as compared with wild type. In addition, the culture supernatant of h or rtxA/h mutants did not cause a significant cytotoxicity, while the culture supernatant of rtxA mutant induced similar levels of cytotoxicity as the wild type. These results indicated that H is a major cytotoxin in culture supernatant of V. vulnificus in this study.
18
Gilbert, Leslie Deanne. "The Association of Virulent Vibrio Spp. Bacteria on Gafftopsail and Hardhead Catfish in Galveston Bay". Thesis, 2010. http://hdl.handle.net/1969.1/ETD-TAMU-2010-08-8529.
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Vibrio vulnificus (Vv) and V. parahaemolyticus (Vp) are gram negative, halophilic bacteria that occur naturally in estuarine waters of Galveston Bay. Both bacteria have the potential to cause infections in humans either via consumption or direct contact. Finfish are a potential vector for these bacteria. Previous work by Brinkmeyer determined that these bacteria are present on the benthic dwelling catfish, Ariopsis felis and Bagre marinus, using a conventional microbial method. The present work focused on using Quantitative Polymerase Chain Reaction (QPCR) and Terminal Restriction Fragment Length Polymorphism (T-RFLP) to not only determine presence of these bacteria, but also to quantify them and look at community structure.
QPCR was able to detect bacteria presence in 34 percent, 31.6 percent, and 0 percent for V.vulnificus, V.parahaemolyticus. thermolabile hemolysin (tlh) and thermostable direct hemolysin (tdh) genes, respectively. Statistical analysis of the QPCR results found that there was no significant difference between the length of fish, location of catch or species of fish in relation to the abundance of bacteria.
T-RFLP was able to detect the presence of bacteria in approximately 70 percent of the samples surveyed. Bands produced from T-RFLP were able to be grouped into five different ranges. The most frequently occurring band fell in the range of 213-219 base pairs, and the most common number of bands per sample was 1 band.
This study found that both QPCR and T-RFLP were better assays than conventional microbial methods for detecting the presence of V. vulnificus and V. parahaemolyticus on catfish fins. QPCR proved to be the most rapid detection method. Based on this study, it was determined that these Vibrio spp. bacteria have some type of relationship with A. felis and B. marinus. This information may be useful to the medical community for determining when there is a greater risk of infection via catfish puncture wounds.
19
Neiman, Jana. "Evolution of cps Loci in Vibrio vulnificus". Thesis, 2011. http://hdl.handle.net/1807/31364.
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Vibrio vulnificus is an opportunistic human and animal pathogen with the highest death rate of any foodborne disease agent. The capsular polysaccharide (CPS) is essential for virulence. Over 100 CPS types (carbotypes) have been identified among natural isolates, yet little is known about the genetic mechanisms that drive such diversity. Chitin, the second most abundant polysaccharide in nature, induces competence in Vibrio species. We found that transformation frequency varies by strain and (GlcNAc)2 was the shortest chitin-derived polymer capable of inducing competence. We confirmed that V. vulnificus can undergo chitin-dependent carbotype conversion following the uptake and recombination of complete cps loci from exogenous genomic DNA. The acquisition of a partial locus was also demonstrated when internal regions of homology between the endogenous and exogenous loci existed. Thus, the same mechanism governing the transfer of complete cps loci also contributes to their evolution by generating novel combinations of CPS biosynthesis genes.
20
Wu, Hui-Chi, i 吳蕙綺. "Characterization of the nuclease of Vibrio vulnificus". Thesis, 2001. http://ndltd.ncl.edu.tw/handle/44615870464268202546.
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碩士國立中山大學
生物科學系研究所
89
The periplasmic nuclease of Vibrio vulnificus, Vvn, has been purified to homogeneity by a one step purification procedure using chromatography on a SP Sepharose column. The purified enzyme showed different mobilities on reducing and non-reducing SDS-PAGE, suggesting that disulfide bonds are involved in the maintenance of a stable tertiary conformation of the protein. Vvn randomly cleaved single and double stranded DNA and RNA, and possessed endonucleolytic activity. The enzyme exhibited an optimal activity between pH 8.0 and pH 10.0, and the optimal temperatures for the DNase and RNase activity were 40 oC – 60 oC and 40 oC – 50 oC, respectively. The enzymatic activity was inhibited by EDTA and EGTA, indicating that Vvn was a metalloenzyme. The DNase and RNase activity of Vvn had different requirements for divalent cations. Chemical modification studies on Vvn revealed the involvement of lysine, arginine, tryptophan and carboxylate residues in the catalytic activity of the enzyme. The extents of inactivation of the DNase and RNase activity of Vvn by modification of the carboxylate group with EDC were different. Substrate DNA and RNA protected the DNase and RNase activity of Vvn from inactivation by PLP, PGO, NBS and EDC which modified lysine, arginine, tryptophan and the carboxylate group. Mg2+ could not protect the DNase and RNase activity of Vvn against the inactivation by PLP and PGO. Whereas Mg2+ protection was observed in NBS- and EDC-mediated inactivation of the DNase but not the RNase activity of Vvn . From these results, it is postulate that there may be two distinct but overlapping active sites, for the DNase and RNase activity, respectively.
21
CIOU, MAN-CHUN, i 邱曼淳. "Signals analysis of Vibrio vulnificus-infected macrophages". Thesis, 2016. http://ndltd.ncl.edu.tw/handle/b9j5pt.
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碩士嘉南藥理大學
生物科技系
104
Vibrio vulnificus is a halophilic Gram-negative bacteria living in tropical or subtropical waters. V. vulnificus is an opportunistic pathogen, the mortality rate is as high as 50% once the infection progresses into sepsis. Macrophages of the human innate immune defense system is the first line of defense for infection. V. vulnificus infection frequently causes severe infection since it is able to kill macrophages to reduce the immune response , leading to infection and sepsis. Our previous studies found that the V. vulnificus toxins have the ability to kill macrophages. Here we want to investigate how V. vulnificus infection causes protein expression within the macrophages by using a Western blot. Our results showed that V. vulnificus infection yields increase HMGB1 expression within macrophages in time-dependent manner. In addition, NF-B expression increased between 30 and 60 min within macrophages after infection with V. vulnificus.
22
Chen, Yu-Chung, i 陳昱仲. "Study of Virulence Factor from Vibrio vulnificus". Thesis, 2004. http://ndltd.ncl.edu.tw/handle/80909777649844346663.
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博士國立中興大學
食品科學系
92
Vibrio vulnificus, a highly virulent marine bacterium, is the causative agent of seafood-related diseases, such as primary septicemia and wound infection in human, particularly in those with certain underlying diseases especially in patients with chronic liver disease. Cases of V. vulnificus infections have been reported from many areas of the world. Over the last decade, there has been a dramatic increase in the number of cases due to V. vulnificus in the southern part of Taiwan. Because V. vulnificus often contaminates the seafood and the consumption of seafood especially raw seafood is very popular, V. vulnificus infection is apparently more prevalent in this island than in many areas of the world. The bactericidal effect of serum is an important defense by the host against invading microorganisms. To identify the genes required for resistance to human serum, we constructed a library of transposon mutants of V. vulnificus and screened them for hypersensitivity to human serum. Here we report that one of the isolated serum-susceptible mutants had a mutation in an open reading frame identified as trkA, a gene encoding an amino acid sequence showing high identity to that of TrkA of Vibrio alginolyticus, a protein required for the uptake of potassium. The other mutant had a mutation in a DNA fragment that was found to potentially encode three open reading frames, two (polS and polR) of which appeared to be arranged in an operon and shared sequence similarity with members of two-component signal transduction systems. PolS shows sequence homology to sensor proteins and also contains a histidine residue highly conserved in the histidine kinase family of sensor protein. PolR is homologous to other regulatory proteins that bind to specific upstream activating elements to enhance transcription of genes with σ54 promoter. Both trkA or polR isogenic mutants were constructed via insertional inactivation, and they were significantly more easily killed by human serum, protamine, or polymyxin B than was the wild type. In addition, the polR isogenic mutant was more sensitive than parental strain to killing by neutrophil. Reverse transcription-PCR and Northern blot analysis indicated that polR could be expressed as a bicistronic transcript from polS in normal grow condition; nevertheless polR could also be markedly induced as a monocistonic transcript from its own promoter located in C-terminal coding region of polS during exposure of bacteria to serum or polymyxin B. In addition, a novel gene hlyIII of V. vulnificus located in downstream of trkA was identified and its recombinant protein exhibited hemolytic activity. Furthermore, BALB/c mice inoculated with trkA, polR or hlyIII isogenic mutant demonstrated virulence attenuation compared to the wild-type strain. Taken together, these results showed that the three genes (trkA, polR and hlyIII) play a significant role in pathogenesis of V. vulnificus.
23
LIN, YONG-ZHU, i 林勇助. "Characterization of monoclonal antibodies against Vibrio vulnificus". Thesis, 1990. http://ndltd.ncl.edu.tw/handle/73872579503450190970.
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Jen, Yu-Chin, i 任佑君. "Isolation and charactrization of Vibrio vulnificus acapsular mutants". Thesis, 1994. http://ndltd.ncl.edu.tw/handle/81925529265511125195.
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碩士國立成功大學
微生物及免役學研究所
82
Vibrio vulnificus, a marine bacerium, causes serious wound infection and septicemia with a high mortality rate in persons with underlying diseases that result in immunocompromised conditions and/or high serum iron levels, such as chronic cirrohosis and hemochromatosis. It is important to understand the pathogenesis of this bacterium in order to be able to properly prevent and treat its infectious diseases. Most V. vulnificus are encapsulated. To study the role of the capsular polysaccharide in the pathogenesis of this microorganism, spontaneous or U.V mutagenized translucent (morphology of acapsular strains) mutants were isolated from three clinical isolates and characterized for their virulence in mice and properties relating to pathogenesis. It was shown that the LD50 of acapsular mulants is higher than their parental strains by 3 logs or more, either by intraperitoneal injection or force feeding, in the mice pretreated with iron or CCl4. This result indicates that the capsule is an important virulence factor of V. vulnificus in susceptible hosts. The growth rates of acapsular mutants were similar to their parental strains, in either rich or iron-limitng media. Adherence of the acapsular mutans to an epithelial cell line HEp-2 was less efficient, but the hemagglutination ability was slightly higher than their parental strains indicating that the capsule but not the hemagglutinin(s) is involved in adherence to the epithelial cells. The ability of the acapsular mutants to resist killing by human serum was also reduced in two of the three strains. The third one, although exhibited equal resistance to human serum with its parental strain, was rapidly cleared from the blood of an infected mouse, suggesting that the capsule may protect the bacteria from other host defense mechanisms, such as phagocytosis. In another study, we tried to develop transposon mutagenesis in V. vulnificus. We used two transposon (Tn5 or TnphoA)-containing plasmids to mutagenize this bacterium.
25
Hsu, Li-Fang, i 徐立芳. "Characterization of a plasmid in Vibrio vulnificus YJ016". Thesis, 2003. http://ndltd.ncl.edu.tw/handle/81072138132763739677.
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碩士國立成功大學
微生物暨免疫學研究所
91
Vibrio vulnificus is a gram-negative, marine bacterium that causes septicemia and serious wound infection in persons who are immunocompromised or have underlying diseases such as liver cirrhosis. V. vulnificus is divided into three distinct biotypes based on the phenotypic and host-range differences. Typically, the biotype 1 strains are associated with human illness. A plasmid (designated pYJ016) of 48.5 kb has been identified in the completed genome sequence of a biotype 1 clinical isolate, YJ016. pYJ016 was uniquely present in strain YJ016 among the various clinical or environmental V. vulnificus strains tested. To examine the role of the plasmid in bacterial growth and virulence, this plasmid was cured from the organism by acridine orange treatment. The growth rate of the cured strain was not distinctly different from the plasmid-harboring strain, and the virulence of the cured strain in mice is comparable to the wild-type strain YJ016. These results indicate that pYJ016 is not required for the growth and virulence of YJ016. On the other hand, the organization and nucleotide sequences of some of the open reading frames (ORFs) in pYJ016 are strikingly like those of the F plasmid, except that pYJ016 lacks some genes for the formation of the sex pilus, like traM, traJ, traP, traV, traR, traE, traQ, traS, and traT. Nevertheless, pYJ016 could still be transferred to other biotype 1 strains by conjugation at a frequency of approximately 10-3. However, pYJ016 could not be transferred to Escherichia coli DH5α by either conjugation or transformation, suggesting that this plasmid may not replicate in E. coli.
26
Lin, Mei-Ping, i 林美萍. "Genetic Study of virulent factor from Vibrio vulnificus". Thesis, 1998. http://ndltd.ncl.edu.tw/handle/85694004644886762337.
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碩士國立成功大學
生物化學研究所
86
Vibrio vulnificus is an opportunistic human pathogen causing wound infection and septicemia. The extracellular protease of V. vulnificus, one of the putative virulence factor , has been demonstrated to be capable of increasing vascular permeability and edema formation. In order to study the virulence role of an extracellular protease in the clinical infection of V. vulnificus , we knockout the protease structural gene of V. vulnificus by homologous recombinant. The results showed that the deficiency of the protease in this organism could affect the survival rate of mouse, indicating the protease plays an important role in the pathogenesis of V. vulnificus. Light microscopy of mouse skin damage caused by subcutaneous injection of the mutant strain or wild type strain of V. vulnificus revealed three probable mechanisms of protease in pathogenesis of clinical infection: (1) Destruction of connective tissue matrices such as collagen contributes to the invading health tissue by V. vulnificus. (2) Protease can help V. vulnificus to resist the phagocytosis of polymorphonuclear leukocytes. (3) Enhancement of vascular permeability and edema formation which result from the activation of kinin generating cascade such as Hageman factor by protease. On the other hand, the plasmid carried the major part of the protease gene was subcloned into pET vector and transformed into Escherichia coli BL21 for overproduction of the protease. A 55 kD biologically inactive recombinant protein accumulated in the insoluble inclusion body was obtained. The inclusion body was purified by His-tag resin under the denature condition and followed with renaturation by the urea gradient concentration dialysis . The renartured protease was caseinolytically and collagenolytically. The polyclonal antibody against the purified 55 kD protease was prepared. Antibody against the culture supernatant and cytoplasmic protein of V. vulnificus was found to recognize the near 47.5-50 kD and 50-62 kD respectively by Western immunoblot analysis. According to the amino sequence and the above deta suggest the V. vulnificus should be belong to the thermolysin family. The protease of the thermolysin family are synthesized as inactive precursors with several domains: a signal peptide, an N-terminal propeptide and a mature protease domain. Studies of enzyme activity of the protease without C terminal 10 kD domain , indicating C-terminal 10 kD polypeptide may be essential for elastase activity.
27
鍾文彬. "Interaction of Vibrio vulnificus with cultured epithelial cells". Thesis, 1997. http://ndltd.ncl.edu.tw/handle/85601712935131118247.
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Chen, Ming-Hsiang, i 陳銘祥. "Establishment of an inducible system in Vibrio vulnificus". Thesis, 2006. http://ndltd.ncl.edu.tw/handle/18481873348482127141.
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碩士國立成功大學
微生物及免疫學研究所
94
Many genes that encode essential products for bacterial growth may exert toxicity to the host cells when overexpressed. To study this kind of genes, the inducible systems are more desirable. In our laboratory, we have been focusing on the identification of virulence factors in Vibrio vulnificus, which is a gram-negative halophilic marine bacterium causing severe wound infection and septicemia. One of our targets was the tolC gene, which encodes an outer membrane protein for transport of a variety of molecules. We have previously isolated a TolC-deficient mutant strain, MW021, which is attenuated in resistance to bile and erythromycin, non-cytotoxic to the HEp-2 cells, and much less virulent in mice. However, when we tried to clone the entire tolC gene into E. coli to further complement the mutant, we were not able to obtain a clone without mutation in tolC. This suggests that overexpression of TolCvv might be toxic for the host cells. Therefore, we intended to establish an inducible system in V. vulnificus to solve this problem. In this study, I have cloned the araC-PBAD DNA fragment, contained the PBAD promoter together with the regulator AraC whose actively is controlled by L-arabinose, into a derivative of the broad-host-range plasmid, pJRD215. The first plasmid we obtained, designated pMH01, was found to harbor three mutations in araC. We then cloned a short DNA fragment and finally obstained a construct, pMH02, that contained no mutation in araC. pMH01 was shown to express vvn (V. vulnificus nuclease gene) and lacZvv cloned downstream of the pBAD promoter in the presence of arabinose in a dose-dependent manner. I then cloned tolCvv into pMH01, and for the first time that this gene can be readily cloned into a plasmid without any mutation. We found that TolCvv was expressed at a basal level in E. coli DH5α without induction by arabinose and the basal level expression could make DH5α resistant to bile. However, addition of arabinose to induce TolCvv resulted in growth retardation and reduced resistance to bile of the host cell. When pMH01-tolCvv was transferred to a ΔtolC V. vulnificus mutant, MW021 by conjugation. The expression of TolCvv in the outer membrane, and the defects of the mutant in resistance to bile salt and erythromycin, cytotoxicity to the HEp-2 cells, and the virulence to mice was restored without arabinose induction. On the other hand, we found that compared to pMH02, the expression of gene cloned downstream of PBAD in pMH02 was induced to a much higher level by arabinose. However, the expression of tolCvv from pMH02 also restored the ability of growth on TCBS agar plate without arabinose induction. These results suggest that pMH01 and pMH02 can be selectively used for overexpression of the cloned gene or performing the complementation experiments.
29
Chen, Yi-Cheng, i 陳奕成. "Analysis of the virulence factor of Vibrio vulnificus". Thesis, 2012. http://ndltd.ncl.edu.tw/handle/2687ns.
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碩士嘉南藥理科技大學
生物科技系暨研究所
100
Vibrio vulnificus is a Gram-negative bacterium causing both serious wound infections and fatal septicemia in human. A gene (spoOM) was found in V. vulnificus. The spoOM was predicted to encode a 267 amino acid polypeptide and the SpoOM was predicted to belong to a putative regulator. A spoOM mutant was constructed via insertional inactivation and showed a similar cytotoxicity compared with the wild-type strain to macrophages. The results showed that SpoOM may not be an important regulator responsible for bacterial cytotoxicity to macrophages in V. vulnificus.
30
Lin, Pei-Ting, i 林珮婷. "Functional Characterization of RND Homologs in Vibrio Vulnificus". Thesis, 2016. http://ndltd.ncl.edu.tw/handle/73259f.
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31
Chiu, Te-Pin, i 邱德賓. "Analysis of epidemiological survey of Vibrio vulnificus infection". Thesis, 2016. http://ndltd.ncl.edu.tw/handle/axf7fc.
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碩士嘉南藥理大學
生物科技系
104
Vibrio vulnificus is a halophilic gram-negative pathogen that infections through open wound or contaminated seafood, can cause serious skin damage and sepsis. Clinical symptoms are primary septicemia, wound infection, and gastrointestinal discomfort. This study is to investigate primary septicemia and wound infection of demographic data and mortality in Taiwan, United States, Japan, South Korea, and China. V. vulnificus infection mainly occused the summer during April to October in Japan and Taiwan, and highly correlated with increased seawater temperatures. The primary septicemia that a significant higher mortality compared to those with wound infection. In prensent, among the classes of antibiotics fluoroquinolone plus a third-generation cephalosporin and tetracycline are effective against V. vulnificus infection.
32
HUNG, WEI-SHUN, i 洪偉舜. "Effect of Vibrio vulnificus cytotoxin on host cell". Thesis, 2018. http://ndltd.ncl.edu.tw/handle/qk2ec7.
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碩士嘉南藥理大學
生物科技系
106
Vibrio vulnificus is a halophilic, motile, curved Gram-negative bacterium living in ocean. Infection caused by this bacterium progresses very fast, and high mortality rate up to 50%. In recent years, more and more clinicians have begun to pay attention to study this bacterium. Several groups experience serious complications after being infected with Vibrio vulnificus, including alcoholics, liver disease and chronic hepatitis, or immune insufficiency. Taiwan is surrounded by the sea, and many people prefer to eat seafood food, so prevention and treatment of Vibrio vulnificus infection deserves our attention. Previous studies have shown that Vibrio vulnificus can use one of their extracellular toxins (H toxin) to kill host cell. This study was to determine the effect of H toxin of Vibrio vulnificus on bacterial mobility and phagocytosis by macrophages. The results showed that H toxin plays an important role in increasing the mobility of Vibrio vulnificus and the bacterial survival rate after phagocytosis
33
Lin, Mai-Wei, i 林美薇. "Role of TolC in virulence of Vibrio vulnificus". Thesis, 2002. http://ndltd.ncl.edu.tw/handle/4d9hr8.
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碩士國立成功大學
微生物及免疫學研究所
90
Vibrio vulnificus, a gram-negative marine bacterium, is an opportunistic pathogen that causes severe wound infection and septicemia in immunocompromised patients. V. vulnificus strains produce a number of potential virulence factors including capsular polysaccharide, siderophores, protease, cytolysin and phospholipase. Our previous data showed that the protease, cytolysin and phospholipase are not the major virulence factors, because disruption of all the three genes encoding these factors did not reduce the virulence in mice or cytotoxicity to HEp-2 cells. However, a non-cytotoxic mutant, NY303, isolated fortuitously from a phospholipase-cytolysin double-deficient mutant was shown to be much less virulent in mice, indicating that cytotoxicity is an important virulence factor. We suspected that NY303 might have lost the activity of an unidentified cytotoxin. We have tried to construct a library in E. coli strain S17-1lpir and will screen for clones that complement the defect of NY303 in cytotoxicity. However, we have not had sufficient numbers of clones in this library for doing this complementation experiment. On the other hand, a homologue of V. cholerae rtx gene cluster, which has been shown to cause the morphological change of cultured cells and seemed to serve as a key role in the emergence of the El Tor strain, was identified in V. vulnificus. We designed primers to amplify the RTX gene cluster in the wild type strain and NY303. Although there was no distinct difference in the length of the PCR products between these two strains, we could not rule out the possibility of point mutation in the rtx gene cluster in NY303. Meanwhile, tolC, an outer membrane protein involved in the export of diverse molecules ranging from large protein toxin to antibiotics, was shown to be required for the secretion of RTX toxin and colonization of V. cholerae. A tolC homologue was also identified in V. vulnificus. To determine the role of tolC in the virulence of V. vulnificus, a tolC-deficient mutant, MW021, was isolated by allelic exchange. MW021 was more sensitive to bile and erythomycin compared with the parental strain. In addition, MW021, was non-cytotoxic to the HEp-2 cells and much less virulent in mice, suggesting that tolC may be involved in the pathogenesis of V. vulnificus, probably by secreting an unidentified cytolysin. The tolC gene was suspected to be an essential gene because it was extremely difficult to obtain a tolC-deficient mutant. To test this, a plasmid carrying tolC was introduced into MW021 and the frequency of isolating the ΔtolC mutant was determined. No increase in the isolation frequency ofΔtolC mutant was observed in the presence of tolC in trans. Whether tolC is essential for V. vulnificus viability awaits further investigation.
34
Yi-HsuanChen i 陳怡璇. "Role of Vibrio vulnificus RTX toxin in antiphagocytosis". Thesis, 2010. http://ndltd.ncl.edu.tw/handle/68735040383101270478.
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碩士國立成功大學
微生物及免疫學研究所
98
Vibrio vulnificus, an invasive bacterial pathogen, causes severe skin lesions and septicemia in humans via wound infection or ingestion of contaminated seafood. Previous studies in this laboratory have shown that the RTX toxin of V. vulnificus promotes bacterial colonization at the infection site and subsequent invasion into the bloodstream by countering the infiltrating phagocytes. In this study we compared the total surviving and phagocytosed bacterial numbers, and the intracellular survival rate in phagocytes between the wild type strain, YJ016, and ΔrtxA mutant, HL128, to understand how RTX contributes to antiphagocytosis. We found that when coincubated with the mouse macrophage cell line RAW 264.7 at a moi of 1 for 90 min, conditions in which the cells remained viable, YJ016 survived about four-fold better than HL128. On the other hand, compared to YJ016, nearly four-fold more of HL128 were ingested into the RAW 264.7 cells in the gentamicin protection assay, which is consistent with the results of acridine orange-crystal violet stain for detecting the intracellular bacteria. Nevertheless, the phagocytosed bacteria of both YJ016 and HL128 were killed at similar rates. Inhibition of phagocytosis of RAW 264.7 cells by low temperature or cytochalasin D treatment resulted in reduced intracellular number of HL128, and the total survival of HL128 increased to the wild-type level after cytochalasin D treatment of the phagocytes. These results suggest that RTX may prevent phagocytosis of V. vulnificus by the macrophages but have little contribution to the survival of phagocytosed bacteria. The phagocytosis of various V. vulnificus strains by the peritoneal macrophages was similar to that by the RAW264.7 cells when examined by acridine orange-crystal violet stain. The acapsular mutant, JF046, and HL128 survived equally well in the presence of RAW 264.7 cells, but much lower number of JF046 was phagocytosed. We further deleted rtxA from JF046 to obtain an acapsular, ΔrtxA mutant, YH001. The survival of this double mutant in the presence of RAW 264.7 was significantly reduced, and the phagocytosed bacterial number of this mutant was even higher than that of HL128. These results suggest that RTX is more important than the capsule in protecting the unopsonized V. vulnificus from phagocytosis.
35
Teng, Yj-Fang, i 鄧怡芳. "Study on the virulent factors of Vibrio vulnificus". Thesis, 2004. http://ndltd.ncl.edu.tw/handle/urz98t.
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碩士國立成功大學
生物科技研究所碩博士班
92
Vibrio vulnificus is a marine Gram-negative bacteria that is recognized as a cause of fulminant primary septicemia, wound infections and acute self-limiting diarrhea. In our primary study, two genes (polS and polR) were cloned from vibrio vulnificus CKM-1. These two genes appeared to be arranged in an operon and shared sequence similarity with members of two-component signal transduction systems. PolS shows sequence homologous to sensor proteins and also contains a histidine residue highly conserved in the histidine kinase family of sensor protein. PolR is homologous to other regulatory proteins that bind to specific upstream activating elements to enhance transcription of genes with σ54 promoter. Our previous northern blot analysis also indicated that PolR could be expressed as a bicistronic transcript from PolS; nevertheless PolR could also be markedly induced as a monocistronic transcript from its own promoter during exposure of bacteria to serum. In this study, reverse transcription polymerase chain reaction experiments were performed to confirm that PolR but not PolS could be specifically induced during exposure of bacteria to serum. In addition, whether PolR could autoregulate itself by binding its upstream region, which locates in PolS gene structure region, or σ54 could bind the expected σ54 promtor would be examined by electrophoresis mobility shift assay. Both PolS and PolR would be overexpressed and purified, and whether PolR could be phosphorylated by PolS or not would be investigated.
36
Kuo, L.-L., i 郭莉莉. "Evidences for H+and Na+-cycle of Vibrio vulnificus". Thesis, 1994. http://ndltd.ncl.edu.tw/handle/07635661686056420141.
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37
Hsieh, Chai-Fan, i 謝彩凡. "Genetic Study of the Virulent Factor from Vibrio vulnificus". Thesis, 1999. http://ndltd.ncl.edu.tw/handle/82045924492650505706.
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碩士國立成功大學
生物化學研究所
87
Vibrio vulnificus is a marine Gram-negative bacillus, that is recognized as a cause of fulminant primary septicemia, wound infections and acute self-limiting diarrhea. Vibrio vulnificus septicemia is associated with a mortality greater than 50%. With septic shock, mortality approaches 100%. Aggressive wound care must include administration of antibiotics (cefotaxime and minocycline), supportive care, debridement of nonviable or necrotic tissues, as well as consideration of amputation of affected extremities. Many extracellular proteins and virulent factors of V. vulnificus, including phospholipase and lipase, are associated with pathogenic mechanism. In this study, a clone (V4P), which exhibited a clear zone on egg yolk plate, was isolated from a genomic library of V. vulnificus. The crude cell extract obtained from V4P is toxic to bladder cancer cell line TSGH8301. Nucleotide sequence analysis of the cloned DNA fragment in V4P predicted that a gene (lipA) encoding extracellular lipase consists of an open reading frame (ORF) of 906-bp. Downstream from the lipA gene an ORF was identified and the corresponding was named lipB. Further experments revealed that no lipase activity is produced by lipA in E. coli unless the lipB is also present. Immunoblot analysis confirmed that LipA is an extracellular lipase. In the course of determing the nucleotide sequence of the lipB region, an additional ORF was found. This ORF is 1341-bp in length and would encode a polypeptide, that showed high homology to 54 transcriptional activator of V. cholerea, and therefore named CT4. Transcritional activity of the non-housekeeping protein 54 transcriptional activator is usually modulated in response to environmental signals. A number of important virulent genes are co-ordinately regulated through the action of the 54 transcriptional activator. There are three functional different domains in 54 transcriptional activator：COOH-terminal domain contains a helix-turn-helix DNA binding motif involved in binding to the enhancer-like element, a conserved central domain contains an ATP binding site responsible for the activation of transcription, and a nonconserved NH2-terminal domain presumably consists of phosphorylation site to be the target for certain regulatory signals. To investigate whether the CT4 of V. vulnificus is responsive to enviromental signals or not, the CT4 PCR product was inserted into pET system and the 49.8 kDa CT4 protein was purified, and then anti-CT4 polyclonal antibodies were prepared. The results of immunoblot analysis showed that no CT4 protein was detected when V. vulnificus was clutured in the general growth condotion, however, when the organism was clutured in the sheep blood medium after 4.5 hours, significant amount of CT4 was detected. The data suggest that CT4 seems to be associated with V. vulnificus invasion to human blood system.
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Kuo, Tsai-Nung, i 郭周彩濃. "Prognostic Factors of Mortality in Vibrio Vulnificus Infected Patients". Thesis, 2010. http://ndltd.ncl.edu.tw/handle/48262332156764780293.
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博士中山醫學大學
醫學研究所
98
Vibrio vulnificus infection is extremely invasive. Even with prompt diagnosis and aggressive therapy, the case-fatality rate is still very high. Therefore, it is important to identify the prognostic factors for V. vulnificus infected patients. Initially, we investigated the predictive factors for mortality in primary septicemia or wound infections caused by V. vulnificus. A retrospective review of 90 patients 18 years and older who were hospitalized due to V. vulnificus infection between January 2000 and December 2006 was performed. Clinical characteristics, laboratory studies, treatments, and outcomes retrieved from medical records were analyzed. Of 90 patients identified as V. vulnificus infections, 39 had primary septicemia and 51 had wound infection. The mean Acute Physiology and Chronic Health Evaluation (APACHE II) and Mortality in Emergency Department Sepsis (MEDS) scores on admission were 11.1 ± 4.9 and 5.5 ± 3.8, respectively. Fifteen patients died, yielding an in-hospital mortality rate of 17%. Multivariate analysis revealed that higher APACHE II (odds ratio [OR] = 1.5; 95% confidence interval [CI] = 1.2-1.8) and MEDS (OR = 1.3; 95% CI = 1.1-1.6) scores on admission were significantly associated with mortality. The area under the receiver operating characteristic curves values for APACHE II and MEDS in predicting in-hospital mortality were 0.928 (95% CI = 0.854-0.972) and 0.830 (95% CI = 0.736-0.901), respectively. Vibrio vulnificus infection can progress rapidly in skin or soft tissue, and it is potentially life-threatening. Therefore, we collected more study subjects, and subsequently explored the predictors of mortality in patients with V. vulnificus infections of skin or soft tissue. The medical records of 119 consecutive patients aged 18 years and older, hospitalized for V. vulnificus infections of skin or soft tissue between January 2000 and December 2007 were reviewed. Twenty-four patients died, yielding an overall case fatality rate of 20%. Of the 24 deaths, 20 (83%) occurred within 72 h after hospital admission. Multivariate analysis revealed that hemorrhagic bullous skin lesions/necrotizing fasciitis (P = 0.003), primary septicemia (P = 0.042), a greater organ dysfunction and/or infection score (P = 0.005), absence of leukocytosis (P = 0.0001), and hypoalbuminemia (P = 0.003) were associated with mortality. Treatment with surgical intervention plus antibiotics (P = 0.038) and surgical intervention within 24 h after admission (P = 0.017) were protective factors. Our results showed that the APACHE II and MEDS scores on admission are significant prognostic indicators in primary septicemia or wound infections caused by V vulnificus. In addition, the presence of hemorrhagic bullous skin lesions/necrotizing fasciitis, primary septicemia, a greater severity-of-illness, absence of leukocytosis, and hypoalbuminemia were the significant risk factors for mortality in V vulnificus infected patients. Moreover, patients treated with surgery plus antibiotics, especially those receiving a prompt surgical evaluation within 24 h after hospital admission, may have a better prognosis. However, a further prospective study to strengthen our findings is required.
39
Horng-RenLo i 羅宏仁. "The role of RTX toxin in Vibrio vulnificus pathogenesis". Thesis, 2010. http://ndltd.ncl.edu.tw/handle/33760587094107465179.
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博士國立成功大學
基礎醫學研究所
99
Vibrio vulnificus is a halophilic gram-negative bacterium causing wound infection and bacteremia in human. An RTX toxin produced by this pathogen can lyse a variety of cells and is required for bacterial virulence. To investigate the role of RTX in V. vulnificus pathogenesis, an RTX-deficient mutant, ?rtxA1, was constructed. This mutant showed about 100-fold reduced virulence in mice challenged via various routes. The mouse infected subcutaneously by the wild type strain showed severely damaged adipose, muscle and connective tissue; however, infection by the ?rtxA1 mutant resulted in phagocytes infiltration but no obvious tissue destruction. In an air-sac infection model, the ?rtxA1 mutant was much less able to colonize at the infection site and invade into the bloodstream and internal organs than the parent and revertant strains. The colonization and virulence of??rtxA1 mutant in the mouse were improved when the neutrophils were depleted by cyclophosphamide treatment. The ability of ?rtxA1 mutant to cause tissue damage and spread into bloodstream and internal organs were also comparable to that of the parent strain in the neutropenic mouse. Furthermore, the RTX-deficient mutants survived much poorly in the phagocyte-enriched peritoneum and were more readily internalized by the mouse macrophage cell line, RAW264.7 than the RTX-proficient strains. These suggest that the RTX toxin could promote bacterial colonization probably by impairing the phagocytic ability of the phagocytes.
40
Chang, Tsong-Min, i 張聰民. "Genetic study of protease and hemolysin from Vibrio vulnificus". Thesis, 1997. http://ndltd.ncl.edu.tw/handle/40021056229402549293.
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博士國立成功大學
基礎醫學研究所
85
A gene (empV) encoding the extracellular metalloprotease of Vibrio vulnificus have been cloned and sequenced. When the empV gene was expressed in minicells, a unique peptide of approx. 46 kDa was identified. Protease activity staining experiment also indicated a similar Mr for the protease. The empV gene product (EmpV) is secreted to the periplasm of Escherichia coli, but not out of it. The crude enzyme prepared from periplasmic fraction of recombinant E. coli was inhibited by a metalloprotease inhibitor and Zn2+ is essential for its protease activity. Nucleotide sequence analysis predicted a single open reading frame (ORF) of 1818 bp encoding a 606-amino acid (aa) polypeptide, with a potential 24-aa signal peptide followed by a long ''pro'' sequence consisting of 172 aa. The N-terminal 20-aa sequence for the elastolytic protease (EepV) purified from the culture supernatant of V. vulnificus 29307, completely identified the beginning of the predicted mature protein within the deduced aa sequence except for one aa residue difference. The estimated pI and molecular weight weight of the predicted mature protein was 5.86 and 44.3 kDa, respectively, which are nearly identical to those of V. vulnificus L-180 extracellular neutral metalloprotease (EnmV) and of strain ATCC 29307 EepV. The estimated molecular weight also agrees closely with that determined by SDS-PAGE analysis of the minicells and by protease activity staining. The deduced amino acid sequence of EmpV showed high homology to V. anguillarum metalloprotease (EmpA), V. cholerae HA/protease (HprC), and V. proteolyticus neutral protease (NprP), particularly with respect to active-site residues, zinc-binding residues, and cystein residues. A gene (vllY) encoding a novel hemolysin of Vibrio vulnificus CKM-1 has been cloned and sequenced. When the vllY gene was expressed in minicells, a unique peptide of approximately 40 kDa was identified. Subcellular fractionation of Escherichia coli carrying the vllY gene indicated that the VllY protein was distributed in both cytoplasmic and periplasmic fraction, with the noted ability to appear in the latter compartment. Nucleotide sequence analysis predicted a single open reading frame (ORF) of 1,071 bp encoding a 357 amino acid polypeptide with an estimated pI of 5.02. The deduced amino acid sequences of VllY showed high similarity to the sequences of legiolysin responsible for hemolysis, pigment production, and fluorescence in Legionella pneumophila. The enzyme also exhibited sequence homology to the MelA protein sequences of Shewanella colwelliana and the sequences of 4-hydroxyphenylpyruvate dioxygenase family proteins from various organisms. PCR screening and Southern blotting experiments of V. vulnificus strains revealed that all the 41 V. vulnificus clinical isolates contain vllY-like genes. A gene (eprA1) encoding the extracellular protease of Aeromonas hydrophila AH1 has been cloned and sequenced. Nucleotide sequence analysis of eprA1 predicted a single open reading frame (ORF) of 1038 bp encoding a 346 amino acid (aa) polypeptide, with a potential 21-aa signal peptide. When the eprA1 gene was expressed in minicells, one major band of approx. 37 kDa was identified, while protease activity staining experiments identified a caseinolytic band of approx. 29 kDa determined by SDS-PAGE analysis of the minicells. The deduced C-terminal aa region (Arg-290 to Gly-313) showed sequence homology to partial C-terminal sequences of other zinc metalloproteases including Penicillium citrinum metalloprotease (PlnC), Aspergillus oryzae metalloprotease (NpII), Aspergillus flavus metalloprotease (MepA), and Aspergillus fumigatus metalloprotease (Mep20), particularly with respect to zinc-binding residues.
41
Chen, Ying-Chou, i 陳盈州. "Study the nuclease of Vibrio vulnificus by DNA shuffling". Thesis, 2001. http://ndltd.ncl.edu.tw/handle/94680979718682604432.
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碩士國立中山大學
生物科學系研究所
89
The nuclease gene of Vibrio vulnificus, vvn, is 696 bp long encoding a protein（Vvn）of 232 amino acids. Vvn is a periplasmic protein and is active in the oxidized form. DNA shuffling is a powerful method for in vitro mutational mechanism by homologous recombination with a low level of point mutation . DNA shuffling consists of four steps：（1）preparation of genes to be shuffled, （2）random fragmentation with DNase I, （3）fragment reassembly by primerless polymerase chain reaction（PCR）, and（4）amplification of reassembled products by a conventional PCR. The advantage of this process is that it can be used to rapidly evolve any protein, without any knowledge of its structure. The goal of this work was using DNA shuffling to generate a diversity of mutation in vvn within a short time. Followed by analyzing the DNase activity of periplasmic protein or in vivo, the mutants were divided into three groups for increase, decrease or no change in DNase activity. Randomly DNA sequencing vvn gene of fourteen transformed clones from the three groups showed only one clone has one base change with comparison to wild-type sequence. The mutation is at amino acid 22 of the N-terminus of Vvn, the change is from serine to isoleucine. The relative activity of mutant Vvn was 82 % in DNase and 59 % in RNase. The effect of a single amino acid change on the DNase and RNase activity of Vvn is different. It supports the postulation that there are two distinct but overlapping active sites exist in Vvn.
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Liu, Shu-hui, i 劉淑惠. "Study on the low salinity adaptation in Vibrio vulnificus". Thesis, 2005. http://ndltd.ncl.edu.tw/handle/d2udh8.
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碩士東吳大學
微生物學系
93
Vibrio vulnificus is an estuarine bacterium that causes serious wound infection and septicemia with high mortality. It is also transmitted through the consumption of raw contaminated seafood and caused the death. V. vulnificus is a global concerned foodborne pathogen and also becomes an important food poisoning pathogen in Taiwan. During food processing, pathogenic bacteria are challenged by various environmental stresses, such as heat, acid, low temperature, and dry treatments, and these bacteria will generate adaptive tolerance under sublethal stresses and thus enhances its risk in food hygiene. In this study, physiological aspects of V. vulnificus under lethal or sublethal hypoosmotic stresses were analyzed. Sensitiveness of the environment and clinical bacterial strains to low salinity and protective effects of osmoprotectants were also examed. The cross-protection of adaptive hypoosmotic tolerance against other stresses, such as acid, heat, bile and food preservative treatment, were examined. Bacteria adapted to acid, heat or bile were also challenged by hypoosmotic stress. Our data showed that decrease of the salt concentration increased the death of V. vulnificus and the clinical strains exhibited higher tolerance to the low salinity than those environmental strains. Osmoprotectants, such as glycine betaine or sucrose, showed obvious protection to V. vulnificus under low salinity, and also showed obvious recovery of the V. vulnificus injured under low salinity. Adaptive hypoosmotic tolerance protected V. vulnificus against low salinity but not against the high temperature and acid treatments. V. vulnificus adapted to non-lethal heat or acid treatment was not protected against low salinity, whereas cells adapted to bile treatment significantly cross-protected against low salinity. These results confirmed the adaptive hypoosmotic tolerance in V. vulnificus and partially clarify the cross-protective effects of different environmental stresses. These phenomena are important in the process of seafood.
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Lu, Kai-Hsi, i 盧凱熙. "Cloning, Sequencing, and Expression of HSP60 Protein Gene of Vibrio parahaemolyticus and Vibrio vulnificus". Thesis, 2001. http://ndltd.ncl.edu.tw/handle/80875351758448414719.
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碩士東吳大學
微生物學系
89
Vibrio parahaemolyticus, a gastrointestinal pathogen of human, shows significant adaptive acid tolerance response and heat shock response. Current studies suggest that V. parahaemolyticus can enhance or de novo synthesize some kinds of heat shock proteins (HSP) during heat shock response. These HSPs, such as GroEL, GroES and DnaJ, are generally observed in many bacteria. HSP have been shown to elicit a strong immune response of host during infection by a variety of pathogens. In this study, HSP60 gene of V. parahaemolyticus was amplified using oligonucleotide primers based on the Salmonella typhi groEL sequence. The nucleotide sequence of the amplified fragment was determined and used to predict the amino acid sequence of V. parahaemolyticus GroEL. The deduced GroEL proteins in V.cholerae and V. parahaemolyticus were highly similar, however, several non-conservative substitutions near the carboxy-termini of the two polypeptides were found. By the same approach, GroEL of Vibrio vulnificus was studied, and to be highly similar to that of GroEL of V.cholerae. We constructed the plasmids that can express each GroEL for further characterizations. Knowing the amino acid sequence of the V. parahaemolyticus and V.vulnificus HSP60 homologue will enhance our knowledge of host immune recognition of HSP produced by bacterial pathogens.
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Chae, Minjung. "Low-temperature post-harvest processing for reducing Vibrio parahaemolyticus and Vibrio vulnificus in raw oysters". Thesis, 2007. http://hdl.handle.net/1957/5705.
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Oysters are filter-feeding bivalves, which filter water for nutrients and often accumulate contaminants and human pathogens such as Vibrio parahaemolyticus and Vibrio vulnificus naturally occurring in the marine environment. These naturally occurring pathogens have been frequently isolated from raw shellfish, particularly oyster, in the United States and are recognized as the leading causes of human gastroenteritis associated with seafood consumption. Human illness caused by consumption of raw oyster contaminated with V. parahaemolyticus and Vibrio vulnificus typically results in reduced sales of oysters and a consequent significant financial burden for the producers. The United States produces more than 27 million pounds of oysters each year with a large portion of them being produced from the coastal water of the Gulf of Mexico. It is estimated that 20 million Americans eat raw shellfish and consumption of raw oyster
is responsible for about 95% of all deaths associated with seafood consumption in the U.S., making raw oysters one of the most hazardous seafoods. Several post-harvest processes, including low temperature pasteurization, freezing, high pressure processing and irradiation, have been reported capable of reducing Vibrio contamination in raw oysters. However, most of them require either a significant amount of initial investment or operation costs, and oysters are often killed during processing. Cost-effective post-harvest processing for reducing V. parahaemolyticus in raw oysters without significant adverse effects on the oysters remains to be developed. This study was conducted to determine impacts of low-temperature (15, 10 and 5°C) depuration and frozen storage on reducing V. parahaemolyticus and V. vulnificus in raw oysters. Depuration of the Gulf oyster (Crassostrea virginica) with electrolyzed oxidizing (EO) water (chlorine, 30 ppm; pH 2.82; oxidation-reduction potential, 1,131mV) containing 3% NaCl was found ineffective on reducing both V. parahaemolyticus and V. vulnificus in the oysters. Reductions of V. parahaemolyticus and V. vulnificus in oyster after 48 h of EO water depuration at 22°C were limited to 0.7 and 1.4 log MPN/g, respectively. Depuration with EO water at lower temperatures did not enhance reductions of Vibrio in the oysters. Greater reductions of V. parahaemolyticus (1.2 log MPN/g) and V. vulnificus (2.0 log MPN/g) were observed when the oysters were depurated with artificial seawater (ASW) at room temperature (22°C) for 48 h. Decreasing temperature of ASW to 15°C for depuration significantly increased the reductions of V. parahaemolyticus and V. vulnificus to 2.1 and 2.9 log MPN/g, respectively, after 48 h of process. However, depuration of oyster in ASW at 10 and 5°C were found less effective than at 15°C in reducing Vibrio in
the Gulf oysters. An extended depuration with ASW at 15°C for 96 h was capable of achieving 2.6 and 3.3 log MPN/g of reductions of V. parahaemolyticus and V. vulnificus, respectively, in the Gulf oysters. Study of effects of frozen storage at -10, -23 and -30°C on reducing V. parahaemolyticus in raw half-shell Pacific oyster (Crassostrea gigas) found that the population of the bacterium decreased faster in oysters stored at -10 than at -23 or -30°C. Holding half-shell Pacific oyster at -10°C for three months or at -23°C for four months was capable of achieving a greater than 3-log (MPN/g) reduction of V. parahaemolyticus in the Pacific oyster.Graduation date: 2008
45
McDougald, S. Diane. "Regulation of starvation and nonculturability in the marine pathogen, Vibrio vulnificus /". 2000. http://www.library.unsw.edu.au/~thesis/adt-NUN/public/adt-NUN20030926.134228/index.html.
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46
Nakhamchik, Alina. "Characterization of Polysaccharide Biosynthesis, Structure and Regulation in Vibrio vulnificus". Thesis, 2008. http://hdl.handle.net/1807/16776.
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Vibrio vulnificus are marine bacteria causing fatal septicemia through wound infections or consumption of contaminated seafood. V. vulnificus is an excellent model for the study of surface polysaccharides, as it is capable of synthesizing capsular polysaccharide (CPS), lipopolysaccharide (LPS) and exopolysaccharide (EPS). V. vulnificus strains exhibit a multitude of carbotypes that evolve through unknown mechanisms. CPS is a confirmed virulence factor, but the genetics of its biosynthesis are unknown. The main objective of these experiments was to gain insight into the biosynthesis, regulation and evolution of ATCC 27562 outer surface polysaccharides. A miniTn10 transposon (Tn) system was used for mutagenesis and single insertions were confirmed through Southern analysis. A novel 25 kb CPS biosynthesis locus was identified through sequencing of regions surrounding Tn insertions; a region encoding putative LPS core biosynthetic functions was identified adjacent to the CPS cluster. The CPS locus contained features of O-antigen biosynthetic loci and was unusual in carrying characteristics of both group I and IV capsular biosynthetic loci. Mutations in this region resulted in elimination of CPS and LPS, and both were shown to be dependent on the activity of the polymerase Wzy. Evidence is presented here supporting horizontal transfer (HT) as a contributor to V. vulnificus CPS evolution. CPS regions of V. vulnificus 27562, YJ016 and CMCP6 contain strain specific genes surrounded by conserved regions, suggestive of HT. Moreover, a CPS locus virtually identical to that of 27562 was discovered in Shewanella putrefaciens strain 200. 27562 CPS is distinctive as it contains N-acetylmuramic acid. Genes encoding murA and murB activities were identified within the cluster and shown to be functionally redundant, supporting HT acquisition of this region. A screen of V. vulnificus gDNA library using CPS biosynthesis and transport mutants identified a cyclic diguanylate cyclase, dcpA. dcpA-mediated increase in cyclic diguanylate lead to EPS production, rugosity phenotypes and enhanced biofilm formation. Interestingly, virulence and motility were not affected suggesting complexity of cyclic diguanylate regulation in V. vulnificus, supported by the large number of cyclic diguanylate related proteins in Vulnificus strains.
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Shi-Kan, Lo, i 羅仕淦. "Isolation and characterization of Vibrio vulnificus mutant deficient in nuclease". Thesis, 2000. http://ndltd.ncl.edu.tw/handle/14765988094691671401.
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碩士國立成功大學
微生物暨免疫學研究所
88
Vibrio vulnificus is a halophilic gram-negative bacterium that causes septicemia and severe wound infection in persons who are immunocompromised or having underlying diseases, such as liver cirrhosis or hemochromatosis. This organism produces a cell-associated nuclease the gene of which has been cloned in our laboratory, shown to be a thermostable enzyme capable of digesting both DNA and RNA. Expression of V. vulnificus nuclease (designated Vvn) in Escherichia coli resulted in a dramatic decrease of transformation efficiency indicating that this nuclease may be responsible for failure of introducing foreign DNA into V. vulnificus by conventional transformation or electroporation. Bacterial nucleases have also been proposed to be involved in providing carbon and nitrogen sources for the microorganisms, and promoting infection of enteric pathogens by facilitating their colonization in the mucosa of the small intestinal. To determine the role of Vvn in bacterial virulence and in preventing the bacterial cell from uptake of foreign DNA, a V. vulnificus mutant with a deletion in the nuclease gene (vvn) was isolated and characterized in this study. This V. vulnificus mutant, SK005, was generated by allelic exchange in which a deletion of 335 bp that resulted in apparent abolishment of the nuclease activity of Vvn was introduced into the vvn gene in the chromosome of a clinical V. vulnificus isolate, YJ016. SK005 exhibited a wild-type growth rate in rich medium and virulence in mice, indicating that Vvn is probably not an important virulence factor. Furthermore, it showed only a slight increase in the frequency of transformation and virtually no difference in conjugation, compared with its parental strain. When tested with DNase test agar plate or by the nuclease assay with various DNA substrates, substantial nuclease activity could still be detected in the periplasmic preparation of SK005. These data suggested that nuclease(s) other than Vvn might be present in the periplasm of V. vulnificus and take part in preventing uptake of foreign nucleic acids by th
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Lu, Pei-Jhen, i 盧姵禎. "Study of Vibrio vulnificus cytotoxin – induced cell death in macrophages". Thesis, 2015. http://ndltd.ncl.edu.tw/handle/p4s9gs.
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碩士嘉南藥理大學
生物科技系
103
Vibrio vulnificus is an opportunistic pathogen, and the bacteria can cause fatal septicemia and wound infections. V. vulnificus infections can lead to death in 2-3 days after hospitalization, with mortality rates as high as 50%. Our previous studies found that V.vulnificus H toxin has the ability to cause cell necrosis in macrophages. Therefore, the goal of this thesis is the analysis of pathway of V.vulnificus H toxin-induced macrophage death. Our previous studies showed that calcium ions cause cell death in V.vulnificus-treated macrophages. In this study, we showed that the V.vulnificus H toxin cause extracelluar calcium influx into cytoplasm of macrophages. By using inhibition analysis, inhibition of NADPH oxidase leaded to a reduction in V.vulnificus H toxin-induced cytotoxicity in macrophages. The H toxin also caused ROS generation and mitochondria damage in V.vulnificus-infected macrophages. These findings suggest that H toxin plays an important role in V.vulnificus infection.
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Wu, Chia-Ju, i 吳佳儒. "The Role of Arginine Biosynthesis Gene in Vibrio vulnificus Infection". Thesis, 2015. http://ndltd.ncl.edu.tw/handle/mx42n9.
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碩士嘉南藥理大學
生物科技系
103
Vibrio vulnificus is a halophilic Gram-negative bacterium and always occurs in tropical and subtropical oceans. The bacteria can result in a serious, fulminant infection. Therefore, patients are dead within 2 days after hospitalization. V. vulnificus infection can cause primary septicemia via eating raw seafood with fever, shock and skin lesions, with the fatality rate about 50%. Wound infection can cause secondary septicemia and cellulitis, with the fatality rate about 25%. In our previous study, we found argD gene has cytotoxicity to macrophage, and its function was related to arginine synthesis from sequence analysis. Therefore, in this study, we focus on the influence of argD gene in V. vulnificus infection. Analysis of cytotoxicity, argD mutant had lower virulence to macrophage than the wild type and argD-complemented strains. The argD gene showed increased growth under arginine-limited conditions. In H2O2 resistance assay, argD mutant showed decreased resistance to H2O2. The intracellular survival rate of argD mutant inside macrophages also was lower than those of wild type and argD-complemented strains. In mouse experiment, argD mutant displayed reduced virulence. In conclusion, argD plays an important role in V. vulnificus infection.
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Drake, Stephenie Lynn. "The ecology Vibrio vulnificus and Vibrio parahaemolyticus from oyster harvest sites in the Gulf of Mexico". 2008. http://www.lib.ncsu.edu/theses/available/etd-10312008-090733/unrestricted/etd.pdf.
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