scholarly journals Characterization of a New Plasmid-Like Prophage in a Pandemic Vibrio parahaemolyticus O3:K6 Strain

2009 ◽  
Vol 75 (9) ◽  
pp. 2659-2667 ◽  
Author(s):  
Shih-Feng Lan ◽  
Chung-Ho Huang ◽  
Chuan-Hsiung Chang ◽  
Wei-Chao Liao ◽  
I-Hsuan Lin ◽  
...  
Keyword(s):  
Vibrio Parahaemolyticus ◽  
Vibrio Vulnificus ◽  
Open Reading Frames ◽  
Host Chromosome ◽  
Food Borne ◽  
Dna Adenine Methylase ◽  
Related Group ◽  
Bacterial Genes

ABSTRACT Vibrio parahaemolyticus is a common food-borne pathogen that is normally associated with seafood. In 1996, a pandemic O3:K6 strain abruptly appeared and caused the first pandemic of this pathogen to spread throughout many Asian countries, America, Europe, and Africa. The role of temperate bacteriophages in the evolution of this pathogen is of great interest. In this work, a new temperate phage, VP882, from a pandemic O3:K6 strain of V. parahaemolyticus was purified and characterized after mitomycin C induction. VP882 was a Myoviridae bacteriophage with a polyhedral head and a long rigid tail with a sheath-like structure. It infected and lysed high proportions of V. parahaemolyticus, Vibrio vulnificus, and Vibrio cholerae strains. The genome of phage VP882 was sequenced and was 38,197 bp long, and 71 putative open reading frames were identified, of which 27 were putative functional phage or bacterial genes. VP882 had a linear plasmid-like genome with a putative protelomerase gene and cohesive ends. The genome does not integrate into the host chromosome but was maintained as a plasmid in the lysogen. Analysis of the reaction sites of the protelomerases in different plasmid-like phages revealed that VP882 and ΦHAP-1 were highly similar, while N15, ΦKO2, and PY54 made up another closely related group. The presence of DNA adenine methylase and quorum-sensing transcriptional regulators in VP882 may play a specific role in this phage or regulate physiological or virulence-associated traits of the hosts. These genes may also be remnants from the bacterial chromosome following transduction.

scholarly journals Characterization of the Urease Operon of Brucella abortus and Assessment of Its Role in Virulence of the Bacterium

2006 ◽  
Vol 75 (2) ◽  
pp. 774-780 ◽  
Author(s):  
Félix J. Sangari ◽  
Asunción Seoane ◽  
María Cruz Rodríguez ◽  
Jesús Agüero ◽  
Juan M. García Lobo
Keyword(s):  
Brucella Abortus ◽  
Urease Activity ◽  
Strong Acid ◽  
Oral Route ◽  
Open Reading Frames ◽  
Human Brucellosis ◽  
Major Route

ABSTRACT Most members of the genus Brucella show strong urease activity. However, the role of this enzyme in the pathogenesis of Brucella infections is poorly understood. We isolated several Tn5 insertion mutants deficient in urease activity from Brucella abortus strain 2308. The mutations of most of these mutants mapped to a 5.7-kbp DNA region essential for urease activity. Sequencing of this region, designated ure1, revealed the presence of seven open reading frames corresponding to the urease structural proteins (UreA, UreB, and UreC) and the accessory proteins (UreD, UreE, UreF, and UreG). In addition to the urease genes, another gene (cobT) was identified, and inactivation of this gene affected urease activity in Brucella. Subsequent analysis of the previously described sequences of the genomes of Brucella spp. revealed the presence of a second urease cluster, ure2, in all them. The ure2 locus was apparently inactive in B. abortus 2308. Urease-deficient mutants were used to evaluate the role of urease in Brucella pathogenesis. The urease-producing strains were found to be resistant in vitro to strong acid conditions in the presence of urea, while urease-negative mutants were susceptible to acid treatment. Similarly, the urease-negative mutants were killed more efficiently than the urease-producing strains during transit through the stomach. These results suggested that urease protects brucellae during their passage through the stomach when the bacteria are acquired by the oral route, which is the major route of infection in human brucellosis.

scholarly journals Genetic Characterization of DNA Region Containing the trh and ure Genes of Vibrio parahaemolyticus

2000 ◽  
Vol 68 (10) ◽  
pp. 5742-5748 ◽  
Author(s):  
Kwon-Sam Park ◽  
Tetsuya Iida ◽  
Yoshiharu Yamaichi ◽  
Tomohito Oyagi ◽  
Koichiro Yamamoto ◽  
...  
Keyword(s):  
Vibrio Parahaemolyticus ◽  
Open Reading Frames ◽  
The Other ◽  
Close Proximity ◽  
Mutant Strains ◽  
Thermostable Direct Hemolysin ◽  
Urease Production ◽  
Reading Frames ◽  
Type Nickel

ABSTRACT We have demonstrated that possession of the gene for thermostable direct hemolysin-related hemolysin (trh) coincides with the presence of the urease gene among clinical Vibrio parahaemolyticus strains and that the location of the two genes are in close proximity on the chromosome. Here, we cloned and sequenced the 15,754-bp DNA region containing the trh gene and the gene cluster for urease production from the chromosome of clinicalV. parahaemolyticus (TH3996). We found 16 open reading frames (ORFs) and a lower G+C content (41%) compared with the total genome of this bacterium (46 to 47%). The ure cluster consisted of eight genes, namely, ureDABCEFG andureR. ureR was located 5.2 kb upstream of the other seven genes in the opposite direction. The genetic organization and sequences of the ure genes resembled those found in Proteus mirabilis. Between ureR and the other uregenes, there were five ORFs, which are homologous with the nickel transport operon (nik) of Escherichia coli. We disrupted each of the ureR, ureC, andnikD genes in TH3996 by homologous recombination and analyzed the phenotype of the mutants. In the presence of urea these mutant strains had dramatically less urease activity than the strain they were derived from. Disruption of ureR,nikD, or ureC, however, had no effect on TRH production. The DNA region containing the trh,nik, and ure genes was found in onlytrh-positive strains and not in Kanagawa phenomenon-positive and environmental V. parahaemolyticusstrains. At the end of the region, an insertion sequence-like element existed. These results suggest that the DNA region was introduced intoV. parahaemolyticus in the past through a mechanism mediated by insertion sequences. This is the first reported case that the genes for an ATP-binding cassette-type nickel transport system, which may play a role in nickel transport through bacterial cytoplasmic membrane, are located adjacent to the ure cluster on the genome of an organism.

Complete Genome of a Novel Lytic Vibrio parahaemolyticus Phage VPp1 and Characterization of Its Endolysin for Antibacterial Activities

2018 ◽  
Vol 81 (7) ◽  
pp. 1117-1125 ◽  
Author(s):  
MENGZHE LI ◽  
YANQIU JIN ◽  
HONG LIN ◽  
JINGXUE WANG ◽  
XIUPING JIANG
Keyword(s):  
Vibrio Parahaemolyticus ◽  
Foodborne Pathogen ◽  
Antibacterial Activities ◽  
Nucleotide Metabolism ◽  
Open Reading Frames ◽  
Bacterial Cells ◽  
Control Groups ◽  
Double Stranded Dna ◽  
Reading Frames

ABSTRACT Vibrio parahaemolyticus is an important foodborne pathogen that is generally transmitted via raw or undercooked seafood. Endolysins originating from bacteriophages offer a new way to control bacterial pathogens. The objectives of this study were to sequence a novel lytic V. parahaemolyticus phage VPp1 and determine the antibacterial activities of the recombinant endolysin (LysVPp1) derived from this phage. The complete VPp1 genome contained a double-stranded DNA of 50,431 bp with a total G+C content of 41.35%. The genome was predicted to encode 67 open reading frames (ORFs), which were organized as nucleotide metabolism, replication, structure, packaging, lysis, and some additional functions. Two tRNAs were encoded to carry anticodons UGG and CCA. Among the functional proteins, ORF33 was deduced to encode endolysin, whereas no holin/antiholin or Rz/Rz1 lysis gene equivalents were found in the VPp1 genome. ORF33 was cloned and expressed. The endolysin LysVPp1 could lyse 9 of 12 V. parahaemolyticus strains, showing its relatively broader host spectrum than phage VPp1, which lysed only 3 of 12 V. parahaemolyticus strains. Furthermore, for EDTA-pretreated bacterial cells, the optical density of the LysVPp1 treatment group decreased by 0.4 at 450 nm, compared with less than 0.1 in control groups, demonstrating enhanced hydrolytic properties. These results contribute to the potential for development of novel enzybiotics for controlling V. parahaemolyticus.

scholarly journals Functional Characterization of the IlpA Protein of Vibrio vulnificus as an Adhesin and Its Role in Bacterial Pathogenesis

2010 ◽  
Vol 78 (6) ◽  
pp. 2408-2417 ◽  
Author(s):  
Kyung-Jo Lee ◽  
Na Yeon Lee ◽  
Yang-Soo Han ◽  
Juri Kim ◽  
Kyu-Ho Lee ◽  
...  
Keyword(s):  
Adhesion Molecule ◽  
Vibrio Vulnificus ◽  
Polyclonal Antibodies ◽  
Functional Characterization ◽  
Intestinal Epithelial ◽  
Human Intestinal Epithelial Cells ◽  
Direct Binding ◽  
Membrane Bound

ABSTRACT Vibrio vulnificus is a Gram-negative bacterium that causes a fatal septicemia. One of its virulence factors is a membrane-bound lipoprotein, IlpA, which can induce cytokine production in human immune cells. In the present study, the role of IlpA as an adhesion molecule was investigated. An ilpA-deleted V. vulnificus mutant showed significantly decreased adherence to INT-407 human intestinal epithelial cells, which in turn resulted in reduced cytotoxicity. The ΔilpA mutant recovered the adherence ability of the wild type by complementation in trans with the intact ilpA gene. In addition, pretreatment of V. vulnificus with anti-IlpA polyclonal antibodies resulted in a significant reduction of bacterial adherence. To localize the domain of IlpA required for cytoadherence, three truncated recombinant IlpA polypeptides were constructed and tested for the ability to adhere to human cells by a ligand-binding immunoblot assay and fluorescence microscopy. The polypeptide containing the carboxy (C)-terminal hydrophilic domain exhibited direct binding to INT-407 cells. Therefore, the C-terminal domain of IlpA allows this protein to be an adhesion molecule of V. vulnificus.

scholarly journals Identification of a Group 1-Like Capsular Polysaccharide Operon for Vibrio vulnificus

2001 ◽  
Vol 69 (11) ◽  
pp. 6893-6901 ◽  
Author(s):  
Anita C. Wright ◽  
Jan L. Powell ◽  
James B. Kaper ◽  
J. Glenn Morris
Keyword(s):  
Capsular Polysaccharide ◽  
Vibrio Vulnificus ◽  
Phase Variation ◽  
Surface Expression ◽  
Open Reading Frames ◽  
Biosynthetic Genes ◽  
Virulent Phenotype ◽  
Phase Variants ◽  
Group 1

ABSTRACT Virulence of Vibrio vulnificus correlates with changes in colony morphology that are indicative of a reversible phase variation for expression of capsular polysaccharide (CPS). Encapsulated variants are virulent with opaque colonies, whereas phase variants with reduced CPS expression are attenuated and are translucent. Using TnphoA mutagenesis, we identified a V.vulnificus CPS locus, which included an upstreamops element, a wza gene (wza Vv), and several open reading frames with homology to CPS biosynthetic genes. This genetic organization is characteristic of group 1 CPS operons. The wzagene product is required for transport of CPS to the cell surface inEscherichia coli. Polar transposon mutations inwza Vv eliminated expression of downstream biosynthetic genes, confirming operon structure. On the other hand, nonpolar inactivation of wza Vv was specific for CPS transport, did not alter CPS biosynthesis, and could be complemented in trans. Southern analysis of CPS phase variants revealed deletions or rearrangements at this locus. A survey of environmental isolates indicated a correlation between deletions inwza Vv and loss of virulent phenotype, suggesting a genetic mechanism for CPS phase variation. Full virulence in mice required surface expression of CPS and supported the essential role of capsule in the pathogenesis of V.vulnificus.

scholarly journals First description of Protoparvovirus in surface water and human Adenovirus in green leafy vegetable, Environmental Education Center - Southern Brazil

2021 ◽  
Author(s):  
Leila Elisa Gartner ◽  
Meriane Demoliner ◽  
Viviane Girardi ◽  
Kelen Gras de Oliveira ◽  
Fernanda Gil de Souza ◽  
...  
Keyword(s):  
Surface Water ◽  
Environmental Education ◽  
Water Sample ◽  
Human Adenovirus ◽  
Fecal Coliforms ◽  
Southern Brazil ◽  
Food Borne ◽  
Stool Samples

Abstract This study analyses the presence of coliforms, Mastadenovirus (AdV) and Canine protoparvovirus (CPV) in an Environmental Education Center (EEC), which includes a city kennel in Southern Brazil, in October 2017 and February 2018. In both collections were analyzed surface water at AdV (human and canine species), CPV, total (TC) and fecal coliforms (FC) presence; lettuce at AdV; and dogs stool at canine AdV (CAV) and CPV. In total, 67 samples were analyzed: 33 dog stool samples; 10 points of surface water and 24 fractions (root, stem and leaves) originated of eight lettuce. Coliforms and viral analyses were performed by Colilert and PCR assays, respectively. Amplicon was sequenced, and species/types of virus were characterized. TC and FC were detected in all surface water in at least one of the collections. Occurrence of viruses was of the 34.3% (23/67) in all samples. HAdV-C was observed in 13% [2/23] (water sample); HAdV-E in 8,7% (2/23) – water and lettuce; CAV-1 in 13% (3/23) – two stool samples and one water sample; and CPV-2a in 56.5% (13/23) – dog stools. Raccoon CPV-like and AdV (not characterized) were detected in one surface water sample and one dog stool sample, respectively. As far as we know, this is the first molecular characterization of the presence of one wildlife animal Protoparvovirus in water and AdV in lettuce reported in Brazil. Detection of HAdV-E in lettuce confirmed the role of vegetables as a potential vehicle of human food-borne viral diseases. Nonetheless, more studies are needed about Raccoon CPV-like to elucidate the potential transmission and threat of this virus in wildlife and interspecies (wild and domestic).

scholarly journals Regulatory roles of 5′ UTR and ORF-internal RNAs detected by 3′ end mapping

2020 ◽  
Author(s):  
Philip P. Adams ◽  
Gabriele Baniulyte ◽  
Caroline Esnault ◽  
Kavya Chegireddy ◽  
Navjot Singh ◽  
...  
Keyword(s):  
Efflux Pump ◽  
Open Reading Frames ◽  
Multiple Transcripts ◽  
Large Numbers ◽  
Independent Functions ◽  
Full Complement ◽  
Rna Fragments ◽  
Bacterial Genes ◽  
Reading Frames

AbstractMany bacterial genes are regulated by RNA elements in their 5′ untranslated regions (UTRs). However, the full complement of these elements is not known even in the model bacterium Escherichia coli. Using complementary RNA-sequencing approaches, we detected large numbers of 3′ ends in 5′ UTRs and open reading frames (ORFs), suggesting extensive regulation by premature transcription termination. We documented regulation for multiple transcripts, including spermidine induction involving Rho and translation of an upstream ORF for an mRNA encoding a spermidine efflux pump. In addition to discovering novel sites of regulation, we detected short, stable RNA fragments derived from 5′ UTRs and sequences internal to ORFs. Characterization of three of these transcripts, including an RNA internal to an essential cell division gene, revealed that they have independent functions as sRNA sponges. Thus, these data uncover an abundance of cis- and trans-acting RNA regulators in bacterial 5′ UTRs and internal to ORFs.

scholarly journals Detection and characterization of clustered regularly interspaced short palindromic repeat-associated endoribonuclease gene variants in Vibrio parahaemolyticus isolated from seafoods and environment

2019 ◽  
Vol 12 (5) ◽  
pp. 689-695
Author(s):  
Pallavi Baliga ◽  
Malathi Shekar ◽  
Moleyur Nagarajappa Venugopal
Keyword(s):  
Polymerase Chain Reaction ◽  
Vibrio Parahaemolyticus ◽  
Genotypic Variation ◽  
Gene Variants ◽  
Chain Reaction ◽  
Pcr Products ◽  
Short Palindromic Repeat ◽  
Polymerase Chain

Aim: In Vibrio parahaemolyticus, the clustered regularly interspaced short palindromic repeat (CRISPR)-associated cas6 endoribonuclease gene has been shown to exhibit sequence diversity and has been subtyped into four major types based on its length and composition. In this study, we aimed to detect and characterize the cas6 gene variants prevalent among V. parahaemolyticus strains isolated from seafoods and environment. Materials and Methods: Novel primers were designed for each of the cas6 subtypes to validate their identification in V. parahaemolyticus by polymerase chain reaction (PCR). In total, 38 V. parahaemolyticus strains isolated from seafoods and environment were screened for the presence of cas6 gene. Few representative PCR products were sequenced, and their phylogenetic relationship was established to available cas6 gene sequences in GenBank database. Results: Of the 38 V. parahaemolyticus isolates screened, only about 40% of strains harbored the cas6 endoribonuclease gene, among which 31.6% and 7.9% of the isolates were positive for the presence of the cas6-a and cas6-d subtypes of the gene, respectively. The subtypes cas6-b and cas6-c were absent in strains studied. Sequence and phylogenetic analysis also established the cas6 sequences in this study to match GenBank sequences for cas6-a and cas6-d subtypes. Conclusion: In V. parahaemolyticus, the Cas6 endoribonuclease is an associated protein of the CRISPR-cas system. CRISPR-positive strains exhibited genotypic variation for this gene. Primers designed in this study would aid in identifying the cas6 genotype and understanding the role of these genotypes in the CRISPR-cas immune system of the pathogen.

Prevalence and characterization of quinolone resistance in Laribacter hongkongensis from grass carp and Chinese tiger frog

2013 ◽  
Vol 62 (10) ◽  
pp. 1559-1564 ◽  
Author(s):  
Ding-Qiang Chen ◽  
Ling Yang ◽  
Yu-Ting Luo ◽  
Min-Jie Mao ◽  
Yong-Ping Lin ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Efflux Pump ◽  
Quinolone Resistance ◽  
Efflux Pump Inhibitor ◽  
Electrophoretic Patterns ◽  
Food Borne ◽  
Laribacter Hongkongensis ◽  
Resistant Strains

Laribacter hongkongensis is a food-borne bacterium associated with community-acquired gastroenteritis and diarrhoea. Quinolone resistance was recently reported in bacterial isolates from aquatic products, but the molecular mechanisms for resistance were still unknown. In this study, a total of 157 L. hongkongensis strains were isolated from grass carps (n = 443) and Chinese tiger frogs (n = 171). Twenty-one ciprofloxacin-resistant strains were analysed for mutations in quinolone resistance-determining regions (QRDR), acquired quinolone resistance (AQR) genes and the role of efflux pumps in resistance. All QRDR mutations in gyrA (codons 85 and 89) and parC (codons 83 and 231) were found to be closely associated with ciprofloxacin resistance. The AQR gene aac(6′)-Ib-cr was found in 42.9 % (9/21) of the resistant strains, but qnrA, qnrB, qnrC, qnrD, qnrS and qepA were not detected. No significant change of MICs to ciprofloxacin was observed in the presence of an efflux pump inhibitor, indicating the role of efflux pump was probably absent. All 21 ciprofloxacin-resistant strains showed different electrophoretic patterns, which suggested they were not genetically related. These data highlight the importance of QRDR mutations and the AQR gene aac(6′)-Ib-cr during the development of quinolone resistance in a heterogeneous population of L. hongkongensis.

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