scholarly journals Molecular Characterization of Cronobacter Lipopolysaccharide O-Antigen Gene Clusters and Development of Serotype-Specific PCR Assays

2011 ◽  
Vol 77 (12) ◽  
pp. 4017-4026 ◽  
Author(s):  
K. G. Jarvis ◽  
C. J. Grim ◽  
A. A. Franco ◽  
G. Gopinath ◽  
V. Sathyamoorthy ◽  
...  
Keyword(s):  
Fragment Length Polymorphism ◽  
Gene Clusters ◽  
Enterobacter Sakazakii ◽  
Polymerase Gene ◽  
Content Type ◽  
Antigen Gene ◽  
O Antigen ◽  
Specific Pcr ◽  
Pcr Assays

ABSTRACTCronobacter(formerlyEnterobacter sakazakii) is a recently defined genus consisting of six species,C. sakazakii,C. malonaticus,C. dublinensis,C. muytjensii,C. turicensis, andCronobactergenomospecies 1. In this study, MboII restriction fragment length polymorphism (RFLP) patterns of O-antigen gene clusters, located betweengalFandgnd, were used to identify serotypes inCronobacterspp. Seven O-antigen RFLP clusters were generated, including threeC. sakazakiiclusters, previously identified as serotypes O1, O2, and O3. The O-antigen regions of six strains with unique RFLP patterns, including twoC. sakazakiistrains, twoC. malonaticusstrains, oneC. turicensisstrain, and oneC. muytjensiistrain, revealed three O-antigen gene clusters shared amongCronobacterspecies. PCR assays were developed, targeting thewzxO-antigen polymerase gene, and used to screen 231Cronobacterstrains to determine the frequency of these newly identified serotypes.

Characterization of Escherichia coli O3 and O21 O antigen gene clusters and development of serogroup-specific PCR assays

2008 ◽  
Vol 75 (2) ◽  
pp. 329-334 ◽  
Author(s):  
Yi Ren ◽  
Bin Liu ◽  
Jiansong Cheng ◽  
Fenxia Liu ◽  
Lu Feng ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Gene Clusters ◽  
Antigen Gene ◽  
O Antigen ◽  
Specific Pcr ◽  
Pcr Assays

scholarly journals Molecular and Genetic Analyses of the Putative Proteus O Antigen Gene Locus

2010 ◽  
Vol 76 (16) ◽  
pp. 5471-5478 ◽  
Author(s):  
Quan Wang ◽  
Agnieszka Torzewska ◽  
Xiaojuan Ruan ◽  
Xiaoting Wang ◽  
Antoni Rozalski ◽  
...  
Keyword(s):  
Fragment Length Polymorphism ◽  
Gene Clusters ◽  
Molecular Classification ◽  
Opportunistic Pathogens ◽  
Antigen Gene ◽  
O Antigen ◽  
Tract Infections ◽  
High Degree

ABSTRACT Proteus species are well-characterized opportunistic pathogens primarily associated with urinary tract infections (UTI) of humans. The Proteus O antigen is one of the most variable constituents of the cell surface, and O antigen heterogeneity is used for serological classification of Proteus isolates. Even though most Proteus O antigen structures have been identified, the O antigen locus has not been well characterized. In this study, we identified the putative Proteus O antigen locus and demonstrated this region's high degree of heterogeneity by comparing sequences of 40 Proteus isolates using PCR-restriction fragment length polymorphism (RFLP). This analysis identified five putative Proteus O antigen gene clusters, and the probable functions of these O antigen-related genes were proposed, based on their similarity to genes in the available databases. Finally, Proteus-specific genes from these five serogroups were identified by screening 79 strains belonging to the 68 Proteus O antigen serogroups. To our knowledge, this is the first molecular characterization of the putative Proteus O antigen locus, and we describe a novel molecular classification method for the identification of different Proteus serogroups.

scholarly journals Genetic Analysis of the Cronobacter sakazakii O4 to O7 O-Antigen Gene Clusters and Development of a PCR Assay for Identification of All C. sakazakii O Serotypes

2012 ◽  
Vol 78 (11) ◽  
pp. 3966-3974 ◽  
Author(s):  
Yamin Sun ◽  
Min Wang ◽  
Quan Wang ◽  
Boyang Cao ◽  
Xin He ◽  
...  
Keyword(s):  
Gene Clusters ◽  
Cronobacter Sakazakii ◽  
Pcr Assay ◽  
Gram Negative ◽  
Content Type ◽  
Antigen Gene ◽  
O Antigen ◽  
Multiplex Pcr Assay ◽  
Food Borne ◽  
Invasive Infections

ABSTRACTThe Gram-negative bacteriumCronobacter sakazakiiis an emerging food-borne pathogen that causes severe invasive infections in neonates. Variation in the O-antigen lipopolysaccharide in the outer membrane provides the basis for Gram-negative bacteria serotyping. The O-antigen serotyping scheme forC. sakazakii, which includes seven serotypes (O1 to O7), has been recently established, and the O-antigen gene clusters and specific primers for threeC. sakazakiiserotypes (O1, O2, and O3) have been characterized. In this study, theC. sakazakiiO4, O5, O6, and O7 O-antigen gene clusters were sequenced, and gene functions were predicted on the basis of homology.C. sakazakiiO4 shared a similar O-antigen gene cluster withEscherichia coliO103. The general features and anomalies of all sevenC. sakazakiiO-antigen gene clusters were evaluated and the relationship between O-antigen structures and their gene clusters were investigated. Serotype-specific genes for O4 to O7 were identified, and a molecular serotyping method for allC. sakazakiiO serotypes, a multiplex PCR assay, was developed by screening against 136 strains ofC. sakazakiiand closely related species. The sensitivity of PCR-based serotyping method was determined to be 0.01 ng of genomic DNA and 103CFU of each strain/ml. This study completes the elucidation ofC. sakazakiiO-antigen genetics and provides a molecular method suitable for the identification ofC. sakazakiiO1 to O7 strains.

scholarly journals Characterization of theEscherichia coliO59 and O155 O-antigen gene clusters: The atypicalwzxgenes are evolutionary related

2005 ◽  
Vol 248 (2) ◽  
pp. 153-161 ◽  
Author(s):  
Hongjie Guo ◽  
Qingke Kong ◽  
Jiansong Cheng ◽  
Lei Wang ◽  
Lu Feng
Keyword(s):  
Gene Clusters ◽  
Antigen Gene ◽  
O Antigen

Localization and molecular characterization of putative O antigen gene clusters of Providencia species

Microbiology ◽  
2012 ◽  
Vol 158 (4) ◽  
pp. 1024-1036 ◽  
Author(s):  
Olga G. Ovchinnikova ◽  
Bin Liu ◽  
Dan Guo ◽  
Nina A. Kocharova ◽  
Alexander S. Shashkov ◽  
...  
Keyword(s):  
Molecular Characterization ◽  
Gene Clusters ◽  
Antigen Gene ◽  
O Antigen

Escherichia coliserogroup O2 and O28ac O-antigen gene cluster sequences and detection of pathogenicE. coliO2 and O28ac by PCR

2010 ◽  
Vol 56 (4) ◽  
pp. 308-316 ◽  
Author(s):  
Pina M. Fratamico ◽  
Xianghe Yan ◽  
Yanhong Liu ◽  
Chitrita DebRoy ◽  
Brian Byrne ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Gene Cluster ◽  
Gene Clusters ◽  
Rapid Identification ◽  
Open Reading Frames ◽  
E Coli ◽  
Antigen Gene ◽  
O Antigen ◽  
Pcr Assays

The O-antigen gene clusters of Escherichia coli serogroups O2 and O28ac were sequenced, and PCR assays were developed to identify strains belonging to these 2 serogroups. Sixteen and 8 open reading frames were mapped to these loci in E. coli O2:H4 U 9-41 and E. coli O28ac:H25 96-3286, respectively. The wzx (O-antigen flippase) and wzy (O-antigen polymerase) genes in the E. coli O2 and O28ac O-antigen gene clusters were selected as targets for PCR assays for their identification. PCR assays targeting the wzx and wzy genes were specific for these serogroups, with one exception. Escherichia coli serogroup O42 strains gave positive results with wzx and wzy PCR assays targeting E. coli O28ac, and antiserum raised against O42 cross-reacted with serogroup O28ac strains. The O-antigen gene cluster of a strain of E. coli serogroup O42 was sequenced, and there were only 3 nt differences between the O-antigen gene clusters of the O28ac and O42 strains. Multiplex PCR assays targeting the O2 wzx gene, the stx1, stx2, hly, eae, and saa genes, and the O28ac wzx, ial, ipaC, and ipaH genes were developed for detecting Shiga toxin-producing E. coli O2 strains and enteroinvasive E. coli O28ac strains, respectively. The O2 and O28ac wzx and wzy genes can be used as diagnostic markers in PCR assays for rapid identification of these serogroups as an alternative to serotyping, and the multiplex PCR assays targeting serogroup-specific genes in combination with virulence genes can be used to identify and to detect pathogenic serogroup O2 and O28ac strains.

scholarly journals Diversity of O Antigens within the Genus Cronobacter: from Disorder to Order

2015 ◽  
Vol 81 (16) ◽  
pp. 5574-5582 ◽  
Author(s):  
Martina Blažková ◽  
Barbora Javůrková ◽  
Jiří Vlach ◽  
Sandra Göselová ◽  
Ludmila Karamonová ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Genomic Analysis ◽  
Rflp Analysis ◽  
Clinical History ◽  
Bacterial Strains ◽  
Content Type ◽  
Nucleotide Variability ◽  
Antigen Gene ◽  
O Antigen ◽  
Specific Pcr

ABSTRACTCronobacterspecies are Gram-negative opportunistic pathogens that can cause serious infections in neonates. The lipopolysaccharides (LPSs) that form part of the outer membrane of such bacteria are possibly related to the virulence of particular bacterial strains. However, currently there is no clear overview of O-antigen diversity within the variousCronobacterstrains and links with virulence. In this study, we tested a total of 82 strains, covering each of theCronobacterspecies. The nucleotide variability of the O-antigen gene cluster was determined by restriction fragment length polymorphism (RFLP) analysis. As a result, the 82 strains were distributed into 11 previously published serotypes and 6 new serotypes, each defined by its characteristic restriction profile. These new serotypes were confirmed using genomic analysis of strains available in public databases: GenBank and PubMLSTCronobacter. Laboratory strains were then tested using the current serotype-specific PCR probes. The results show that the current PCR probes did not always correspond to genomic O-antigen gene cluster variation. In addition, we analyzed the LPS phenotype of the reference strains of all distinguishable serotypes. The identified serotypes were compared with data from the literature and the MLST database (www.pubmlst.org/cronobacter/). Based on the findings, we systematically classified a total of 24 serotypes for theCronobactergenus. Moreover, we evaluated the clinical history of these strains and show thatCronobacter sakazakiiO2, O1, and O4,C. turicensisO1, andC. malonaticusO2 serotypes are particularly predominant in clinical cases.

scholarly journals Development of PCR Assays Targeting Genes in O-Antigen Gene Clusters for Detection and Identification of Escherichia coli O45 and O55 Serogroups

2005 ◽  
Vol 71 (8) ◽  
pp. 4919-4924 ◽  
Author(s):  
Chitrita DebRoy ◽  
Pina M. Fratamico ◽  
Elisabeth Roberts ◽  
Michael A. Davis ◽  
Yanhong Liu
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Gene Cluster ◽  
Real Time Pcr ◽  
Gene Clusters ◽  
E Coli ◽  
Antigen Gene ◽  
O Antigen ◽  
Detection And Identification ◽  
Pcr Assays

ABSTRACT The Escherichia coli O45 O-antigen gene cluster of strain O45:H2 96-3285 was sequenced, and conventional (singleplex), multiplex, and real-time PCR assays were designed to amplify regions in the wzx (O-antigen flippase) and wzy (O-antigen polymerase) genes. In addition, PCR assays targeting the E. coli O55 wzx and wzy genes were designed based on previously published sequences. PCR assays targeting E. coli O45 showed 100% specificity for this serogroup, whereas by PCR assays specific for E. coli O55, 97/102 strains serotyped as E. coli O55 were positive for wzx and 98/102 for wzy. Multiplex PCR assays targeting the E. coli O45 and the E. coli O55 wzx and wzy genes were used to detect the organisms in fecal samples spiked at levels of 106 and 108 CFU/0.2 g feces. Thus, the PCR assays can be used to detect and identify E. coli serogroups O45 and O55.

Characterization of E. coli O24 and O56 O Antigen Gene Clusters Reveals a Complex Evolutionary History of the O24 Gene Cluster

2006 ◽  
Vol 53 (6) ◽  
pp. 470-476 ◽  
Author(s):  
Jiansong Cheng ◽  
Quan Wang ◽  
Wei Wang ◽  
Ying Wang ◽  
Lei Wang ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Evolutionary History ◽  
Gene Clusters ◽  
E Coli ◽  
Antigen Gene ◽  
O Antigen ◽  
History Of ◽  
Complex Evolutionary History
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