scholarly journals High-Resolution Genotyping ofCampylobacter Strains Isolated from Poultry and Humans with Amplified Fragment Length Polymorphism Fingerprinting

1999 ◽  
Vol 65 (6) ◽  
pp. 2369-2375 ◽  
Author(s):  
Birgitta Duim ◽  
Trudy M. Wassenaar ◽  
Alan Rigter ◽  
Jaap Wagenaar
Keyword(s):  
Length Polymorphism ◽  
Fragment Length ◽  
Fragment Length Polymorphism ◽  
Amplified Fragment Length Polymorphism ◽  
Pcr Primers ◽  
Epidemiological Studies ◽  
Aflp Analysis ◽  
Campylobacter Coli ◽  
Chromosomal Dna ◽  
Typing Methods

ABSTRACT For epidemiological studies of Campylobacterinfections, molecular typing methods that can differentiate campylobacters at the strain level are needed. In this study we used a recently developed genotyping method, amplified fragment length polymorphism (AFLP), which is based on selective amplification of restriction fragments of chromosomal DNA, for genetic typing ofCampylobacter jejuni and Campylobacter colistrains derived from humans and poultry. We developed an automated AFLP fingerprinting method in which restriction endonucleasesHindIII and HhaI were used in combination with one set of selective PCR primers. This method resulted in evenly distributed band patterns for amplified fragments ranging from 50 to 500 bp long. The discriminatory power of AFLP was assessed with aC. jejuni strain, an isogenic flagellin mutant, and distinct C. jejuni strains having known pulsed-field gel electrophoresis and fla PCR-restriction fragment length polymorphism genotypes. Unrelated C. jejuni strains produced heterogeneous patterns, whereas genetically related strains produced similar AFLP patterns. Twenty-five Campylobacterstrains obtained from poultry farms in The Netherlands grouped in threeC. jejuni clusters that were separate from a C. coli cluster. The band patterns of 10 C. jejunistrains isolated from humans were heterogeneous, and most of these strains grouped with poultry strains. Our results show that AFLP analysis can distinguish genetically unrelated strains from genetically related strains of Campylobacter species. However, desirable genetically related strains can be differentiated by using other genotyping methods. We concluded that automated AFLP analysis is an attractive tool which can be used as a primary method for subtyping large numbers of Campylobacter strains and is extremely useful for epidemiological investigations.

scholarly journals Identification and Molecular Epidemiology of Campylobacter coli Isolates from Human Gastroenteritis, Food, and Animal Sources by Amplified Fragment Length Polymorphism Analysis and Penner Serotyping

2005 ◽  
Vol 71 (4) ◽  
pp. 1953-1958 ◽  
Author(s):  
B. L. Siemer ◽  
E. M. Nielsen ◽  
S. L. W. On
Keyword(s):  
Molecular Epidemiology ◽  
Length Polymorphism ◽  
Fragment Length ◽  
Fragment Length Polymorphism ◽  
Amplified Fragment Length Polymorphism ◽  
Epidemiological Studies ◽  
Poor Correlation ◽  
Campylobacter Coli ◽  
Polymorphism Analysis ◽  
Food Borne

ABSTRACT Campylobacter coli is an infrequently studied but important food-borne pathogen with a wide natural distribution. We investigated its molecular epidemiology by use of amplified fragment length polymorphism (AFLP)-based genotyping and Penner serotyping. Serotype reference strains and 177 Danish isolates of diverse origin identified by routine phenotyping as C. coli were examined. Molecular tools identified some 12% of field isolates as Campylobacter jejuni, emphasizing the need for improved identification methods in routine laboratories. Cluster analysis of AFLP profiles of 174 confirmed C. coli isolates revealed a difference in the distribution of isolates from pig and poultry (chicken, duck, turkey, and ostrich) species and indicated the various poultry species, but not pigs, to be likely sources of human C. coli infection. A poor correlation was observed between serotyping and AFLP profiling, suggesting that the former method has limited value in epidemiological studies of this species.

Amplified Fragment Length Polymorphism, Serotyping, and Quinolone Resistance of Campylobacter jejuni and Campylobacter coli Strains from Chicken-Related Samples and Humans in Taiwan

2006 ◽  
Vol 69 (4) ◽  
pp. 775-783 ◽  
Author(s):  
SHAO W. FANG ◽  
CHING J. YANG ◽  
DANIEL Y. C. SHIH ◽  
CHENG C. CHOU ◽  
ROCH C. YU
Keyword(s):  
Campylobacter Jejuni ◽  
Length Polymorphism ◽  
Fragment Length ◽  
Genetic Similarity ◽  
Fragment Length Polymorphism ◽  
Genetic Relationships ◽  
Amplified Fragment Length Polymorphism ◽  
Quinolone Resistance ◽  
Aflp Analysis ◽  
Campylobacter Coli

The high-resolution genotyping method of amplified fragment length polymorphism (AFLP) analysis was used to study the genetic relationships between Campylobacter jejuni isolates from chicken-related samples (n = 32) and humans (n = 27) as well as between Campylobacter coli isolates from chicken-related samples (n = 27) and humans (n = 5). These isolates were collected between 1994 and 2003 in Taiwan. All C. jejuni and C. coli isolates showed highly heterogeneous fingerprints. C. jejuni isolates were separated in two distinct genetic clusters (A and B) at 40% genetic similarity and 42 different AFLP types at 90% similarity. However, three clusters at 40% genetic similarity and 33 different AFLP types at 90% similarity were observed in C. coli isolates. These results showed that AFLP analysis could be used to identify individual isolates of two Campylobacter species. Among C. jejuni isolates, the predominant AFLP type 1 was observed in five (7.9%) isolates, and types 5 and 12 in four (6.3%) isolates each. Cluster B consisted of 10 isolates, while the majority of isolates (n = 53) belonged to cluster A. In some AFLP types (1, 5, 12, 14 and 31), AFLP fingerprints of chicken-related isolates were closely related genetically to those of isolates from humans with gastroenteritis. The predominant serotypes in C. jejuni isolates were B:2 and Y:37. All isolates belonging to serotype O:19 grouped into one single AFLP type. Some chicken samples yielded multiple isolates of Campylobacter harboring simultaneously quinolone-resistant and quinolone-sensitive isolates attributed to the same species, or harboring C. jejuni and C. coli that have the characteristics of quinolone resistance.

scholarly journals High-Resolution Genotyping of Streptococcus pyogenesSerotype M1 Isolates by Fluorescent Amplified-Fragment Length Polymorphism Analysis

1999 ◽  
Vol 37 (6) ◽  
pp. 1948-1952 ◽  
Author(s):  
Meeta Desai ◽  
Androulla Efstratiou ◽  
Robert George ◽  
John Stanley
Keyword(s):  
Length Polymorphism ◽  
Fragment Length ◽  
Fragment Length Polymorphism ◽  
Geographic Origin ◽  
Amplified Fragment Length Polymorphism ◽  
Discriminatory Power ◽  
Polymorphism Analysis ◽  
Worldwide Distribution ◽  
High Discriminatory Power ◽  
Typing Methods

We have used fluorescent amplified-fragment length polymorphism (FAFLP) analysis to subtype clinical isolates of Streptococcus pyogenes serotype M1. Established typing methods define most M1 isolates as members of a clone that has a worldwide distribution and that is strongly associated with invasive diseases. FAFLP analysis simultaneously sampled 90 to 120 loci throughout the M1 genome. Its discriminatory power, precision, and reproducibility were compared with those of other molecular typing methods. Irrespective of disease symptomatology or geographic origin, the majority of the clinical M1 isolates shared a single ribotype, pulsed-field gel electrophoresis macrorestriction profile, and emm1 gene sequence. Nonetheless, among these isolates, FAFLP analysis could differentiate 17 distinct profiles, including seven multi-isolate groups. The FAFLP profiles of M1 isolates reproducibly exhibited between 1 and more than 20 amplified fragment differences. The high discriminatory power of genotyping by FAFLP analysis revealed genetic microheterogeneity and differentiated otherwise “identical” M1 isolates as members of a clone complex.

scholarly journals Genomic Relatedness of ChlamydiaIsolates Determined by Amplified Fragment Length Polymorphism Analysis

1999 ◽  
Vol 181 (15) ◽  
pp. 4469-4475 ◽  
Author(s):  
Adam Meijer ◽  
Servaas A. Morré ◽  
Adriaan J. C. Van Den Brule ◽  
Paul H. M. Savelkoul ◽  
Jacobus M. Ossewaarde
Keyword(s):  
Cluster Analysis ◽  
Length Polymorphism ◽  
Fragment Length ◽  
Fragment Length Polymorphism ◽  
Amplified Fragment Length Polymorphism ◽  
Chlamydia Psittaci ◽  
Rrna Genes ◽  
Aflp Analysis ◽  
Polymorphism Analysis ◽  
Genomic Relatedness

ABSTRACT The genomic relatedness of 19 Chlamydia pneumoniaeisolates (17 from respiratory origin and 2 from atherosclerotic origin), 21 Chlamydia trachomatis isolates (all serovars from the human biovar, an isolate from the mouse biovar, and a porcine isolate), 6 Chlamydia psittaci isolates (5 avian isolates and 1 feline isolate), and 1 Chlamydia pecorum isolate was studied by analyzing genomic amplified fragment length polymorphism (AFLP) fingerprints. The AFLP procedure was adapted from a previously developed method for characterization of clinical C. trachomatis isolates. The fingerprints of all C. pneumoniae isolates were nearly identical, clustering together at a Dice similarity of 92.6% (± 1.6% standard deviation). The fingerprints of the C. trachomatis isolates of human, mouse, and swine origin were clearly distinct from each other. The fingerprints of the isolates from the human biovar could be divided into at least 12 different types when the presence or absence of specific bands was taken into account. The C. psittacifingerprints could be divided into a parakeet, a pigeon, and a feline type. The fingerprint of C. pecorum was clearly distinct from all others. Cluster analysis of selected isolates from all species revealed groups other than those based on sequence data from single genes (in particular, omp1 and rRNA genes) but was in agreement with available DNA-DNA hybridization data. In conclusion, cluster analysis of AFLP fingerprints of representatives of all species provided suggestions for a grouping of chlamydiae based on the analysis of the whole genome. Furthermore, genomic AFLP analysis showed that the genome of C. pneumoniae is highly conserved and that no differences exist between isolates of respiratory and atherosclerotic origins.

Comparative Fingerprinting Analysis ofCampylobacter jejuni subsp. jejuni Strains by Amplified-Fragment Length Polymorphism Genotyping

2000 ◽  
Vol 38 (9) ◽  
pp. 3379-3387 ◽  
Author(s):  
Bjørn-Arne Lindstedt ◽  
Even Heir ◽  
Traute Vardund ◽  
Kjetil K. Melby ◽  
Georg Kapperud
Keyword(s):  
Length Polymorphism ◽  
Fragment Length ◽  
Fragment Length Polymorphism ◽  
Amplified Fragment Length Polymorphism ◽  
Epidemiological Surveillance ◽  
Aflp Analysis ◽  
Polymorphism Analysis ◽  
Pcr Rflp ◽  
Pcr Products ◽  
Ofcampylobacter Jejuni

Amplified-fragment length polymorphism (AFLP) analysis with the endonucleases BglII and MfeI was used to genotype 91 Campylobacter jejuni subsp. jejunistrains from outbreaks and sporadic cases. AFLP-generated fragments were labeled with fluorescent dye and separated by capillary electrophoresis. The software packages GeneScan and GelCompar II were used to calculate AFLP pattern similarities and to investigate phylogenetic relationships among the genotyped strains. The AFLP method was compared with two additional DNA-based typing methods, pulsed-field gel electrophoresis (PFGE) using SmaI and restriction fragment length polymorphism analysis on PCR products (PCR-RFLP) of theflaA and flaB genes. We found that AFLP analysis of C. jejuni strains is a rapid method that offers better discriminatory power than do both PFGE and PCR-RFLP. AFLP and, to a lesser extent, PCR-RFLP could differentiate strains within the same PFGE profiles, which also makes PCR-RFLP an alternative to PFGE. We were able to clearly distinguish 9 of 10 recognized outbreaks by AFLP and to identify similarities among outbreak and sporadic strains. Therefore, AFLP is suitable for epidemiological surveillance ofC. jejuni and will be an excellent tool for source identification in outbreak situations.

Genetic Variability in the Potato Pathogen Colletotrichum coccodes as Determined by Amplified Fragment Length Polymorphism and Vegetative Compatibility Group Analyses

2006 ◽  
Vol 96 (10) ◽  
pp. 1097-1107 ◽  
Author(s):  
Larry J. Heilmann ◽  
Nadav Nitzan ◽  
Dennis A. Johnson ◽  
Julie S. Pasche ◽  
Curt Doetkott ◽  
...  
Keyword(s):  
Length Polymorphism ◽  
Fragment Length ◽  
Fragment Length Polymorphism ◽  
Amplified Fragment Length Polymorphism ◽  
Vegetative Compatibility ◽  
Colletotrichum Coccodes ◽  
Aflp Analysis ◽  
Aflp Data ◽  
Bootstrap Analysis ◽  
Nit Mutants

Amplified fragment length polymorphism (AFLP) using three primer sets was used to characterize 211 Colletotrichum coccodes isolates from North America, 112 of which were assigned to six vegetative compatibility groups (VCGs) using nitrate nonutilizing (nit) mutants. These isolates clustered into five corresponding groups by unweighted pairgroup method with arithmetic means-based cluster analysis of AFLP banding patterns. Isolates of C. coccodes belonging to NA-VCG1 and NA-VCG3 were closely related, as were isolates belonging to NA-VCG2 and NA-VCG5. Based on bootstrap analysis of AFLP data, the two isolates originally assigned to NA-VCG4 clustered with isolates belonging to NA-VCG2 and NA-VCG5. C. coccodes isolates that clustered with two isolates belonging to NA-VCG6 were the most diverged from other groups, including seven isolates collected from hosts other than potato. As opposed to the bootstrap analysis, a quadratic discriminant analysis (QDA) of AFLP data correctly categorized the two isolates of NA-VCG4. Furthermore, in isolates where VCG determinations had been made, this model correctly classified isolates of all VCGs. QDA classifications were identical to those made by the bootstrap analysis, with the exception of VCG4. Overall, classifications made by the QDA model were strongly correlated (r = 0.970, P < 0.001) to the VCGs assigned by traditional methods. All 99 C. coccodes isolates evaluated only by AFLP also were subjected to QDA, leading to the assignment of a presumptive VCG for each isolate. No isolates of VCG4 or VCG6 were identified by QDA within this population. Symptoms of black dot developed in plants inoculated with isolates collected from both potato and non-potato hosts. However, total yield was not significantly reduced by infection with non-potato isolates. The lack of any additional groups identified by AFLP analysis may be an indicator of a limited level of genetic variation among North American C. coccodes isolates. AFLP is a much more efficient technique for subspecific characterization in C. coccodes than VCG analysis utilizing nit mutants and will provide an effective means by which the population biology of this pathogen can be further investigated worldwide.

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