AIM:To investigate the effect of in vivo environment on gene expression in Helicobacter pylori(H.pylori) as it relates to its survival in the host.METHODS:In vivo expression technology(IVET) systems are used to identi...AIM:To investigate the effect of in vivo environment on gene expression in Helicobacter pylori(H.pylori) as it relates to its survival in the host.METHODS:In vivo expression technology(IVET) systems are used to identify microbial virulence genes.We modified the IVET-transcriptional fusion vector,pIVET8,which uses antibiotic resistance as the basis for selection of candidate genes in host tissues to develop two unique IVET-promoter-screening vectors,pIVET11 and pIVET12.Our novel IVET systems were developed by the fusion of random Sau3A DNA fragments of H.pylori and a tandem-reporter system of chloramphenicol acetyltransferase and beta-galactosidase.Additionally,each vector contains a kanamycin resistance gene.We used a mouse macrophage cell line,RAW 264.7 and mice,as selective media to identify specific genes that H.pylori expresses in vivo.Gene expression studies were conducted by infecting RAW 264.7 cells with H.pylori.This was followed by real time polymerase chain reaction(PCR) analysis to determine the relative expression levels of in vivo induced genes.RESULTS:In this study,we have identified 31 in vivo induced(ivi) genes in the initial screens.These 31 genes belong to several functional gene families,including several well-known virulence factors that are expressed by the bacterium in infected mouse stomachs.Virulence factors,vacA and cagA,were found in this screen and are known to play important roles in H.pylori infection,colonization and pathogenesis.Their detection validates the efficacy of these screening systems.Some of the identified ivi genes have already been implicated to play an important role in the pathogenesis of H.pylori and other bacterial pathogens such as Escherichia coli and Vibrio cholerae.Transcription profiles of allivi genes were confirmed by real time PCR analysis of H.pylori RNA isolated from H.pylori infected RAW 264.7 macrophages.We compared the expression profile of H.pylori and RAW 264.7 coculture with that of H.pylori only.Some genes such as cag A,vac A,lpx C,mur I,tlp C,trx B,展开更多
Hydatid cyst disease is one of the most parasitic diseases transmitted from carnivores animals (such as dogs) to humans and causes a deterioration in the general health. Its transmission to herbivores animals led to a...Hydatid cyst disease is one of the most parasitic diseases transmitted from carnivores animals (such as dogs) to humans and causes a deterioration in the general health. Its transmission to herbivores animals led to a substantial economic loss in the meat productivity and reduced its quality. The increase of acquisition of dogs at homes nowadays led to an increase in the risk of infection with many parasitic diseases such as hydatid cyst disease. The present study was conducted to provide a modern view on the current status of hydatid cyst disease among slaughtered animals in Jeddah through periodic visiting to a slaughter house in North Jeddah. The mice prepared have been infected in the peritoneal membrane with 0.2 ml of hydatid cyst fluid containing ±2000 protoscoleces from fertile cysts obtained from an infected goat’s liver which resulted in a 75% infection rate. Mice infected with hydatid fluid from fertile cysts obtained from an infected camel’s lungs showed a result of 11.1% infection rate. The cysts appeared in the peritoneal and abdominal cavity.展开更多
基金Supported by Intramural Research Program of the National Institutes of Health,National Institute of Diabetes and Digestive and Kidney DiseaseThe Division of Intramural Research of the National Institute of Allergy and Infectious DiseasesAn Inter-Agency Agreement (Y3-DK-3521-07) with the National Institute on Minority Health and Health Disparities
文摘AIM:To investigate the effect of in vivo environment on gene expression in Helicobacter pylori(H.pylori) as it relates to its survival in the host.METHODS:In vivo expression technology(IVET) systems are used to identify microbial virulence genes.We modified the IVET-transcriptional fusion vector,pIVET8,which uses antibiotic resistance as the basis for selection of candidate genes in host tissues to develop two unique IVET-promoter-screening vectors,pIVET11 and pIVET12.Our novel IVET systems were developed by the fusion of random Sau3A DNA fragments of H.pylori and a tandem-reporter system of chloramphenicol acetyltransferase and beta-galactosidase.Additionally,each vector contains a kanamycin resistance gene.We used a mouse macrophage cell line,RAW 264.7 and mice,as selective media to identify specific genes that H.pylori expresses in vivo.Gene expression studies were conducted by infecting RAW 264.7 cells with H.pylori.This was followed by real time polymerase chain reaction(PCR) analysis to determine the relative expression levels of in vivo induced genes.RESULTS:In this study,we have identified 31 in vivo induced(ivi) genes in the initial screens.These 31 genes belong to several functional gene families,including several well-known virulence factors that are expressed by the bacterium in infected mouse stomachs.Virulence factors,vacA and cagA,were found in this screen and are known to play important roles in H.pylori infection,colonization and pathogenesis.Their detection validates the efficacy of these screening systems.Some of the identified ivi genes have already been implicated to play an important role in the pathogenesis of H.pylori and other bacterial pathogens such as Escherichia coli and Vibrio cholerae.Transcription profiles of allivi genes were confirmed by real time PCR analysis of H.pylori RNA isolated from H.pylori infected RAW 264.7 macrophages.We compared the expression profile of H.pylori and RAW 264.7 coculture with that of H.pylori only.Some genes such as cag A,vac A,lpx C,mur I,tlp C,trx B,
文摘Hydatid cyst disease is one of the most parasitic diseases transmitted from carnivores animals (such as dogs) to humans and causes a deterioration in the general health. Its transmission to herbivores animals led to a substantial economic loss in the meat productivity and reduced its quality. The increase of acquisition of dogs at homes nowadays led to an increase in the risk of infection with many parasitic diseases such as hydatid cyst disease. The present study was conducted to provide a modern view on the current status of hydatid cyst disease among slaughtered animals in Jeddah through periodic visiting to a slaughter house in North Jeddah. The mice prepared have been infected in the peritoneal membrane with 0.2 ml of hydatid cyst fluid containing ±2000 protoscoleces from fertile cysts obtained from an infected goat’s liver which resulted in a 75% infection rate. Mice infected with hydatid fluid from fertile cysts obtained from an infected camel’s lungs showed a result of 11.1% infection rate. The cysts appeared in the peritoneal and abdominal cavity.
