Tumour Growth Inhibition and Systemic Responses of ΔsopBΔsopDΔpipD Disrupted Salmonella Agona and Salmonella Typhimurium in Mice

@#Introduction: Bacteria had long been known to have tumour-targeting and tumour inhibition capabilities and have re-emerged into the limelight of cancer research as a possible alternative treatment for solid tumours. Conventional therapies for solid tumours are either by surgery, chemotherapy, radiotherapy, which are very invasive and non-specific to the tumours and results in various adverse effects on the patients. Bacterial Mediated Tumour Therapy often utilises attenuated bacteria as therapeutic agents to ensure reduced pathogenicity of the strains. However, this often results in lower invasiveness towards the tumours itself. In this study, we studied the tumour inhibition capabilities of Salmonella Pathogenicity Island (SPI) attenuated Salmonella Typhimurium (S. Typhimurium) and Salmonella Agona (S. Agona), specifically with attenuation of sopB, sopD, and pipD genes. Methods: Balb/c mice bearing CT26 tumours were inoculated with S. Typhimurium and S. Agona, both unattenuated and ΔsopBΔsopDΔpipD attenuated strains. Tumour volumes were monitored daily. Organs and blood were collected for plasma liver enzyme analysis and histopathology studies on testis, liver, kidneys and brain. Results: The ΔsopBΔsopDΔpipD S. Agona treated group showed improved inhibition of tumour growth with 51.11% tumour volume reduction compared to unattenuated S. Agona. The ΔsopBΔsopDΔpipD strains have also shown lesser systemic effects as observed in plasma and histopathological studies) compared to its unattenuated counterparts. Conclusion: The present study showed that ΔsopBΔsopDΔpipD S. Agona has a great potential to be utilised as tumour therapeutic agent as it exerts lesser systemic effect while having similar tumour inhibition capabilities as the well-studied S. Typhimurium strain.

Bacteriemia por Vibrio cholerae no-O1/no-O139 que porta una región homóloga a la isla de patogenicidad VpaI-7 / Bacteremia caused by Vibrio cholerae non-O1/non-O139 carrying a region homologous to pathogenicity island VpaI-7

Resumen Presentamos un caso de bacteriemia por Vibrio cholerae no-O1/ no-O139 en una mujer de 81 años con un cuadro de dolor abdominal, fiebre, vómitos, diarrea, coluria e ictericia, mientras visitaba una zona rural sin acceso a agua potable. La identificación se realizó por la técnica de espectrometría de masa MALDI-TOF, confirmándose una cepa no toxigénica no-O1/no-139. La caracterización molecular del aislado demostró la ausencia del gen de la toxina del cólera (CTX), y pilus TCP; sin embargo, presentó cinco de los seis genes de virulencia presentes en la isla de patogenicidad homóloga denominada VPaI-7 del V. parahaemolyticus (vcs N2+, vcs C2+, vcs V2+,toxR-, vspD+, T vopF+). Además, el aislado presentó los genes de virulencia hylA y rtxA. Este es el primer caso reportado en Chile de una cepa clínica de V. cholerae no-O1, no-O139 aislada de hemocultivos portador de un segmento homólogo de la isla de patogenicidad denominada VPaI-7 de V. parahaemolyticus, el cual codifica para un sistema de secreción tipo III (TTSS), que probablemente contribuye a su virulencia.

We report a case of V. cholerae non-O1 / non-O139 bacteremia in an 81-year-old woman with abdominal pain, fever, vomiting, liquid stools, choluria and jaundice, while visiting a rural area without access to potable water. The identification was made by the MALDI-TOF mass spectrometry technique and subsequently the non-toxigenic non-O1 / non-139 strain was confirmed in the national reference laboratory. The molecular characterization demonstrated the absence of the cholera toxin gene (CTX), and the TCP pilus, however, presented 5 of 6 virulence genes present in an island of homologous pathogenicity named VPaI-7 of V. parahaemolyticus (vcs N2 +, vcs C2 +, vcs V2 +, toxR-, vspD +, T vopF +) and in addition it was positive for hylAy rtxA virulence genes recognized outside the island. This is the first case reported in Chile of a clinical strain of V. cholerae non-O1, non-O139 isolated from blood culture that carries in its genome a homologous segment of the pathogenicity island named VPaI-7 of V. parahaemolyticus, which codifies for a type III secretion system (TTSS) that probably contributes to his virulence.

Antibiotic susceptibilities and virulence genes of clinically isolated Salmonella enterica serovars ;Schwarzengrund strains / 中华微生物学和免疫学杂志

Objective To investigate the antibiotic susceptibilities and the profiles of virulence genes of clinically isolated Salmonella enterica serovars Schwarzengrund ( S. Schwarzengrund) strains for bet-ter understanding the epidemiological trend of this type of non-typhoidal Salomonella and to provide guide-lines for the prevention and treatment of S. Schwarzengrund infection. Methods Stool samples and clinical data of patients with acute diarrhea who received treatment in the Second Hospital of Tianjin Medical Univer-sity during May, 2014 to October, 2014 were collected for this study. Enrichment culture and biochemical identification were used to isolate and identify the S. Schwarzengrund strains. The isolated strains were fur-ther analyzed with serotyping analysis, drug susceptibility test, pulsed field gel electrophoresis ( PFGE) and multiple locus sequence typing ( MLST ) . The representative genes carried by Salmonella pathogenicity islands (SPI) 1-5, SPI regulators and virulence plasmids were amplified by PCR. The coding genes of CdtB-islet, which were cdtB, pltA and pltB were amplified and sequenced. Results In total, 16 (14. 8%) out of 108 non-typhoidal Salmonella strains were identified as S. Schwarzengrund strains and all of them were sus-ceptible to 11 kinds of antibiotics such as fluoroquinolone, ampicillin, ceftriaxone and trimethoprim-sulfame-thoxazole. PFGE categorized the 16 S. Schwarzengrund strains into 3 clusters including A clone ( 14 strains), B clone (1 strain) and C clone (1 strain). The strains that isolated from 8 patients who ate the same food belonged to one cluster ( A clone ) , suggesting that it was an outbreak of infection. The 16 S. Schwarzengrund strains showed identical MLST type, which was ST241. The representative genes carried by SPI1-5 ( invA, sitC, hilA, sseL, sifA, mgtC, siiE and sopB) , the regulatory gene ( phoP) and the cytole-thal distending toxin islet (CdtB-islet) coding genes (cdtB, pltA and pltB) were positive, while the genes carried by virulence plasmids (pefA, prot6E and spvB) were negative. The similarities in CdtB-islet coding genes and amino acids sequences between Salmonella typhi and S. Schwarzengrund strains in this study were more than 97% and 98%, respectively. Conclusion In this study, polyclonal S. Schwarzengrund strains of ST241 type were isolated from the patients. They were susceptible to common antibiotics, but carried the virulence genes contained in SPI1-5 and CdtB-islet coding genes and might cause an outbreak of infection. Attention should be paid to the tendency and threat of clinical S. Schwarzengrund infection and continuous surveillance and investigation should be performed.

Transcriptional Analysis of the iagB within Salmonella Pathogenicity Island 1 (SPI1)

HilA is a central regulator of Salmonella pathogenicity island 1 (SPI1), which is necessary for host invasion by Salmonella and induction of gastroenteritis. The iagB lies downstream of hilA and is thought to be co-transcribed with hilA, but iagB expression has not yet been analyzed directly. In this study, iagB expression in various mutant strains was measured to determine whether the expression pattern was similar to that of hilA. A β-galactosidase assay revealed that iagB expression was greater under shaking than standing culture condition. iagB expression was decreased in relA/spoT and ihfB mutants but not in luxS mutant, in line with previous reports on hilA expression. The hilA and iagB mRNA levels decreased by approximately 2-fold in arcA mutant grown aerobically and increased by approximately 10-fold in fnr mutant grown anaerobically. Although the fold changes in hilA and iagB mRNA level differed in hfq mutant strain, the patterns of time- and Hfq-dependent regulation were similar for both genes. Thus, iagB and hilA exhibited similar expression patterns in various mutational backgrounds and under different growth condition.

Nucleotide Binding Oligomerization Domain 1 Is an Essential Signal Transducer in Human Epithelial Cells Infected with Helicobacter pylori That Induces the Transepithelial Migration of Neutrophils

BACKGROUND/AIMS: The cytosolic host protein nucleotide binding oligomerization domain 1 (Nod1) has emerged as a key pathogen recognition molecule for innate immune responses in epithelial cells. The purpose of the study was to elucidate the mechanism by which Helicobacter pylori infection leads to transepithelial neutrophil migration in a Nod1-mediated manner. METHODS: Human epithelial cell lines AGS and Caco-2 were grown and infected with H. pylori. Interleukin (IL)-8 mRNA expression and IL-8 secretion were assessed, and nuclear factor kappaB (NF-kappaB) activation was determined. Stable transfections of AGS and Caco-2 cells with dominant negative Nod1 were generated. Neutrophil migration across the monolayer was quantified. RESULTS: Cytotoxin-associated gene pathogenicity island (cagPAI)(+) H. pylori infection upregulated IL-8 mRNA expression and IL-8 secretion in AGS and Caco-2 cells compared with controls. NF-kappaB activation, IL-8 mRNA expression and IL-8 secretion by cagPAI knockdown strains were reduced compared with those infected with the wild-type strain. NF-kappaB activation, IL-8 mRNA expression and IL-8 secretion in dominant-negative (DN)-Nod1 stably transfected cells were reduced compared with the controls. The transepithelial migration of neutrophils in DN-Nod1 stably transfected cells was reduced compared with that in controls. CONCLUSIONS: Signaling through Nod1 plays an essential role in neutrophil migration induced by the upregulated NF-kappaB activation and IL-8 expression in H. pylori-infected human epithelial cells.

Investigation of the association between clinical outcome and the cag pathogenicity-island and other virulence genes of Helicobacter pylori isolates from patients with dyspepsia in Eastern Turkey

The aims of our work were to determine the presence of the cag pathogenicity-island (cag PAI) and other virulence genes of Helicobacter pylori recovered from patients with gastritis and peptic ulcer, and to investigate the correlation of these virulence genes with clinical outcome. The presence of the cagA, the promoter regions of cagA, cagE, cagT, and the left end of cag-PAI (LEC), cag right junction (cagRJ), the plasticity region open reading frames (ORFs), vacA and oipA genes among 69 H. pylori isolates were determined by polymerase chain reaction. Intact cag PAI was detected in only one (1.4%) isolate. The cagA gene was identified in 52.1% and 76.2% of isolates from patients with dyspepsia (gastritis and peptic ulcer), respectively. The plasticity region ORFs i.e. JHP912 and JHP931 were predominantly detected in isolates from peptic ulcer. Less than 25% of the isolates carried other ORFs. Types I, II and III were the most commonly found among the isolates. None of the isolates possessed type Ib, 1c, IIIb, IV and V motifs. The most commonly vacA genotypes were s1am1a and s1m2 in isolates with peptic ulcer and gastritis, respectively. The results confirmed that the prevalence of oipA (Hp0638) gene was 75% and 85.7% in patients with gastritis and peptic ulcer, respectively. Furthermore, vacA s1am1a positivity was significantly related to peptic ulcer (p < 0.05).

Markers of pathogenicity islands in strains of Aeromonas species of clinical and environmental origin.

The aim of this study was to investigate the presence of markers of pathogenicity islands that may be informative to detect the virulent PAI carriers of clinical and environmental strains of Aeromonas spp. isolated in Mexico. virB2, virB9 and virB11 genes were found in Aeromonas strains isolated from environmental and clinical sources while cagE and tfc16 genes were only in strains of environmental origin. Having performed the wide screening presented in this study, we now have a set of strains to map and confirm the presence of a pathogenicity island in Aeromonas strains isolated in Mexico.

Gastroenteritis aguda causada por Vibrio cholerae no-O1, no-O139 que porta una región homologa a la isla de patogenicidad VpaI-7 / Acute gastroenteritis caused by a Vibrio cholerae non-O1, non-O139 strain harboring a genetic region homologous to the VpaI-7 pathogenicity island

Pathogenic Vibrio cholerae isolates, the etiologic agents of cholera, generally express one of two O antigens (O1 or O139). Most environmental isolates are nonpathogenic and are referred to as "non-O1, non-O139". However some V. cholerae non-O1, non-O139 strains are clearly pathogenic and have caused outbreaks or sporadic cases of gastroenteritis and extraintestinal infections in humans. We report a case of acute gastroenteritis by a V. cholerae non-O1, non-O139 harboring a genetic region homologous to a segment of the VpaI-7 V. parahaemolyticus pathogenicity island.

Cepas patogénicas de Vibrio cholerae, el agente causal del cólera, expresan generalmente uno de dos antígenos O (denominados O1 u O139). La mayoría de las cepas ambientales son no patogénicas y corresponden al tipo denominado "no-O1, no-O139". Sin embargo, algunas cepas de este tipo son claramente patogénas y han causado brotes de gastroenteritis e infecciones extra-intestinales en humanos. Se reporta un caso clínico de gastroenteritis aguda causado por una cepa de V. cholerae no-O1, no-O139 que contiene en su genoma una región homóloga a un segmento de la isla de patogenicidad VpaI-7 descrita previamente en V. parahaemolyticus.

The complexity of ToxT-dependent transcription in Vibrio cholerae.

Vibrio cholerae is the causative agent of the disease cholera, characterized by profuse watery diarrhoea. Two of the main virulence factors associated with the disease are cholera toxin (CT) and toxin-coregulated pilus (TCP). Expression of CT and TCP is regulated via a complex cascade of factors that respond to environmental signals, but ultimately ToxT is the direct transcriptional activator of the genes encoding CT and TCP. Recent studies have begun to unveil the mechanisms behind ToxT-dependent transcription. We review current knowledge of transcriptional activation by ToxT and the environmental stimuli that allow ToxT to regulate virulence gene expression, resulting in cholera pathogenesis.

Helicobacter pylori infection in relation to gastric cancer progression.

Gastric cancer is a major cause of cancer death worldwide, especially in developing countries. The incidence of gastric cancer varies from country to country, probably as a result of genetic, epigenetic, and environmental factors. H. pylori infection is considered as a major risk factor in the development of gastric cancer. However, the scenario varies in Asian countries, exhibiting a higher rate of H. pylori infection and low incidence of gastric cancer, which could be attributed to strain-specific virulence factors and host genetic makeup. In this review, we discuss the various virulence factors expressed by this bacterium and their interaction with the host factors, to influence pathogenesis.

Mecanismos moleculares utilizados por Salmonella para causar infecciones del tracto digestivo / Molecular mechanisms used by Salmonella to cause infections in the digestive tract

Salmonella enterica es uno de los principales causantes de gastroenteritis infecciosa en el mundo, debido al consumo de alimentos y aguas contaminadas con esta bacteria. Este grupo de bacterias Gram negativas se caracteriza por tener la capacidad de invadir células eucariontes y sobrevivir en el interior de ellas, evento fundamental para que la bacteria cause una enfermedad tanto localizada como sistémica. Las proteínas de virulencia que este agente utiliza para invadir y sobrevivir en células eucariontes son codificadas por genes presentes en islas de patogenicidad, que corresponden a grandes bloques de material genético integrados en el cromosoma bacteriano y que fueron posiblemente adquiridos a través de transferencia lateral de genes desde otros microorganismos. Estudios recientes han permitido identificar el rol que poseen estas proteínas de virulencia en el proceso infectivo de Salmonella y su impacto en el funcionamiento de la célula eucarionte. De este modo, ha sido posible entender de mejor manera los mecanismos moleculares utilizados para infectar a su hospedero y se han identificado posibles blancos terapéuticos para el tratamiento de los cuadros infecciosos causados por este patógeno.

Salmonella enterica is one of the main ethiological agents of infectious gastroenteritis in the world, due the consumption of food and water contaminated with these bacteria. This group of Gram negative bacterials characterized by its capacity to invade eukaryotic cells and survive inside them, an event that is fundamental for the bacteria to cause a localized as well as a systemic disease. The virulence proteins that this bacterium uses to invade and survive within eukaryotic cells are encoded by genes found in pathogenicity islands, big blocks of genetic material integrated in the bacterial chromosome, that were probably acquired through lateral gene transfer from other microorganisms. Recent studies have identified the role that these virulence proteins play in the infective process of Salmonella, and their impact in the function of the eukaryotic cell. This way, it has been possible to better understand the molecular mechanisms used by Salmonella to infect their hosts, and potential therapeutic targets have been identified to improve the treatment of the infection caused by this pathogen.

Study on the classification of gonococcal island of different genotypes and effects of sac-4 gene on serum resistance of Neisseria gonorrhoeae / 中华流行病学杂志

Objective Using molecular methods to study the relationship between genotypes and serum resmtance of Neisseria gonorrhoeae in Wuhan area.Methods NG-mast and serum bactericidal assays at the molecular level were used to differentiate the 46 strains which were isolated from the outpatients of sexually transmitted disease clinics and the relationship between different genotypes while phenotypes was also studied.Results 80.43% of the 46 strains contained the island and we were able to define three dillerent combinations of genes in the isolates.Results from serum bactericidal assays showed that all 9 sac-4 stralns did not provide any serum resistance.Conclusion Different isolates carried clifferent gonococcal genetm islands(pathogenicity island)and certain phenotypes.There were no sobious relationship between sac-4 gene and serum resistance of Neisseria gonorrhoeae.

Analysis of cag Pathogenicity Island of Helicobacter pylori Korean Isolate

It is commonly believed in the Western World that the more severe forms of gastroduodenal diseases like peptic ulcer are associated with infection by specific Helicobacter pylori strains classified as type I being considered to be more virulent than type II strains. However, in Korea, most of H. pylori isolates belong to type 1 strains regardless of virulence. Type I H. pylori strains differ from type II strains by the presence of the cag pathogenicity island (cag PAI) composed of a block of genes. In this study, the nucleotide sequence of cag PAI of the H. pylori Korean strain 51 was determined and compared with those of strains 26695 and J99 to assess the structural variation in the region and to evaluate its implication in the virulence of the H. pylori. The cag PAI of H. pylori strain 51 was smaller in size and in the number of constituting ORFs in comparison with 26695 and J99 strains. Although many cag orthologues were nearly identical one another with the similarity of 90% or more at the nucleotide and amino acid levels, there were some remarkable and significant differences in several cag genes among the three cag PAIs. Surprisingly, the percent similarities at amino acid level were lower than those at nucleotide level in one third of the ORFs. The two genes (cag7 and cagA) of strain 51 differed in sizes and deduced amino acid sequences from the corresponding genes of the other two strains. When comparing cagA ORF of H. pylori strain 51 with that of 8 non-Korean strains, phylogenetic tree revealed that the strain 51 formed a separate branch with the most far distances from the other strains except for a Japanese strain. The Cag7 protein of, strain 51 had a deletion in the repeat region II, suggesting a major change in the conformation and function of the protein.

The transcription of interleukin-8 in gastric epithelial cells downregulated by ORF10 of cag Ⅱ of Helicobacter pylori / 中华消化杂志

Objective Helicobacter pylori (H. pylori )contains more than 30 genes and many of them in cag pathogenicity island (cag PAI) composed of cag Ⅰ and cag Ⅱ are involved in eliciting the transcription of interleikin 8 (IL 8) and the induction of IL 8 protein secretion in gastric epithelial cells, although some genes are not involved. This study was designed to observe the effect of ORF10 of H. pylori cag Ⅱ on the transcription of IL 8 in gastric epithelial cells and to test the possibility that certain genes in the cag PAI also downregulate (modulate) the transcription and expression of IL 8 in gastric epithelial cells. Methods Three strains of H. pylori mutants with deletion of ORF10 gene (ORF10, namely 28 1, 28 2, 28 3) and L5F11 cells containing IL 8 reporter gene were constructed. The constructed L5F11 cells were co cultured with the above strains and the luciferase activity (IL 8 transcription) was measured in a scintillation counter and the concentration of IL 8 protein was assayed by enzyme linked immunosorbent assay (ELISA). Results The activities of luciferase induced by 3 strains of H. pylori mutants with deletion of ORF10 gene were much higher than that of the mother strain 26695 [ 28 1:(1.49 ? 0.27)?10 6 cpm vs. (0.67 ? 0.08 )? 10 6 cpm, P

CONSTRUCTION AND IDENTIFICATION OF THE VECTOR FOR CHINESE Hp MUTANT STRAINS DELETING PATHOGENICITY ISLAND / 解放军医学杂志

A vector for a mutant Chinese Helicobacter pylori (Hp) strains knock out pathogenicity island (PAI) was constructed, which was the basis to establish a Chinese Hp mutant deleting pathogenicity island. Genetic engineering techniques such as polymerase chain reaction, plasmid extraction, agarose gel electrophoresis, restriction analysis, ligation, preparation of competence cell and transformation were used to make two cloning fragments containing two end regions of PAI and a selectable chloramphenicol resistance marker between them, and then engineering the recombined fragments into pBluescript plasmid. The result showed that restriction analysis demonstrated that the engineering mutant vector had been recombined. It suggested that we constructed an engineering mutant vector which targeted to delete the PAI in Chinese Hp strains. It will be useful for addressing the role of PAI in the pathogenesis of Hp infection.

The characteristic structure and classification of cag pathogenicity island in Chinese Helicobacter pylori strains / 中华消化杂志

0.05). The products of conjunction of cagⅠ and cagⅡ were found only in 5 strains. The detectable rate of continuous cag PAI was much higher in duodenal ulcer than in chronic gastritis ( P

Prevalence of cagA, cagE and cagT genes in cag pathogenicity island of Helicobacter pylori strains isolated from Shanghai patients and their clinical implications / 中华消化杂志

0.05). Of 98 cagE positive isolates, 14(14.3%) were cagA negative. Conclusion Most of the H. pylori isolates in Shanghai region may have intact cag PAI. cagE, could be taken as the marker for the presence of cag PAI. There is no correlation between the integrality of cag PAI and the clinical outcome of H. pylori of infection.

THE YERSINIA HIGH-PATHOGENICITY ISLAND / 微生物学通报

There have been many studies on the structure and function of the high pathogenicity island since it was first discovered to be located on Yersinia . And it was detected to be present in a few different members of the family Enterobacteriaceae that are pathogenic to human. This review describes the Yersinia high pathogenicity island: structure,function and prevalence among pathogenic enterobacteriaceae.

THE DISTRIBUTION AND SIGNIFICANCE OF cag Ⅰ IN HELICOBACTER PYLORI ISOLATED FROM CHINESE PATIENTS / 解放军医学杂志

To investigate the distrbution of cag Ⅰin Helicobacter pylori(Hp) isolated from Chinese patients, and its relationship to gastroduodenal diseases. Fragment in cag Ⅰ was amplified by polymerase chain reaction(PCR) in 107 Hp strains from Chinese patients. Results The amplicom to cag Ⅰ was positive in 93 strains, of which 57 strains came from patients with chronic gastritis, 31 strains from peptic ulcer, and 5 strains from gastric carcinoma.Conclusion Existance of cag Ⅰ of Hp strains was popular in the Chinese population, but no data had proven the relation ship between the existence of cag 1and occurrence of Hp related diseases and inflammatory infiltration in gastric mucosa.