scholarly journals An Extended Multilocus Sequence Typing (MLST) Scheme for Rapid Direct Typing of Leptospira from Clinical Samples

2016 ◽  
Vol 10 (9) ◽  
pp. e0004996 ◽  
Author(s):  
Sabrina Weiss ◽  
Angela Menezes ◽  
Kate Woods ◽  
Anisone Chanthongthip ◽  
Sabine Dittrich ◽  
...  
Keyword(s):  
Multilocus Sequence Typing ◽  
Clinical Samples ◽  
Mlst Scheme

scholarly journals High-resolution multilocus sequence typing for Chlamydia trachomatis: improved results for clinical samples with low amounts of C. trachomatis DNA

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Shlomo Pilo ◽  
Gal Zizelski Valenci ◽  
Mor Rubinstein ◽  
Lea Pichadze ◽  
Yael Scharf ◽  
...  
Keyword(s):  
Chlamydia Trachomatis ◽  
Multilocus Sequence Typing ◽  
Phylogenetic Trees ◽  
Melting Curve ◽  
Pcr Amplification ◽  
Clinical Samples ◽  
Melting Curve Analysis ◽  
Whole Genome ◽  
Mlst Scheme ◽  
Low Concentrations

Abstract Background Several Multilocus Sequence Typing (MLST) schemes have been developed for Chlamydia trachomatis. Bom’s MLST scheme for MLST is based on nested PCR amplification and sequencing of five hypervariable genes and ompA. In contrast to other Chlamydia MLST schemes, Bom’s MLST scheme gives higher resolution and phylogenetic trees that are comparable to those from whole genome sequencing. However, poor results have been obtained with Bom’s MLST scheme in clinical samples with low concentrations of Chlamydia DNA. Results In this work, we present an improved version of the scheme that is based on the same genes and MLST database as Bom’s MLST scheme, but with newly designed primers for nested-1 and nested-2 steps under stringent conditions. Furthermore, we introduce a third primer set for the sequencing step, which considerably improves the performance of the assay. The improved primers were tested in-silico using a dataset of 141 Whole Genome Sequences (WGS) and in a comparative analysis of 32 clinical samples. Based on cycle threshold and melting curve analysis values obtained during Real-Time PCR of nested-1 & 2 steps, we developed a simple scoring scheme and flow chart that allow identification of reaction inhibitors as well as to predict with high accuracy amplification success. The improved MLST version was used to obtain a genovars distribution in patients attending an STI clinic in Tel Aviv. Conclusions The newly developed MLST version showed great improvement of assay results for samples with very low concentrations of Chlamydia DNA. A similar concept could be applicable to other MLST schemes.

Distribution of virulence genes and the molecular epidemiology of Streptococcus pyogenes clinical isolates by emm and multilocus sequence typing methods

2020 ◽  
Author(s):  
Siti Nur Adila Hamzah ◽  
Mohd Nasir Mohd Desa ◽  
Azmiza Syawani Jasni ◽  
Niazlin Mohd Taib ◽  
Siti Norbaya Masri ◽  
...  
Keyword(s):  
Molecular Epidemiology ◽  
Virulence Factors ◽  
Streptococcus Pyogenes ◽  
Multilocus Sequence Typing ◽  
Virulence Genes ◽  
Clinical Samples ◽  
Pcr Analysis ◽  
Gold Standard Method ◽  
Non Invasive ◽  
Typing Methods

Abstract Background: Streptococcus pyogenes has a variety of virulence factors and the predominant invasive strains differ according to specific emm types and geographical orientation. Although emm typing is commonly used as the gold standard method for the molecular characterization, multilocus sequence typing (MLST) has become an important tool for comparing the genetic profiles globally. This study aimed to screen selected virulence genes from invasive and non-invasive clinical samples and to characterize the molecular epidemiology by emm typing and MLST methods. Methods: A total of 42 S. pyogenes isolates from invasive and non-invasive samples collected from 2014 to 2015 from two different tertiary hospitals were investigated for the distribution of virulence factors and their molecular epidemiology by emm and multilocus sequence typing methods. Detection of five virulence genes (speA , speB , speJ , ssa and sdaB) was performed using multiplex polymerase chain reaction (PCR) using the standard primers and established protocol. Results: Multiplex PCR analysis revealed that sdaB/speF (78.6%) and speB (61.9%) were the predominant virulence genes. Regardless of the type of invasiveness, diverse distribution of emm types/subtypes was noted which comprised of 27 different emm types/subtypes. The predominant emm types/subtypes were emm63 and emm18 with each gene accounted for 11.8% whereas 12% for each gene was noted for emm28, emm97.4 and emm91. The MLST revealed that the main sequence type (ST) in invasive samples was ST402 (17.7%) while ST473 and ST318 (12% for each ST) were the major types in non-invasive samples. Out of 18 virulotypes, Virulotype A (five genes, 55.6%) and Virulotype B (two genes, 27.8%) were the major virulotypes found in this study. Phylogenetic analysis indicated the presence of seven different clusters of S. pyogenes. Interestingly, Cluster VI showed that selected emm/ST types such as emm71/ST318 (n=2), emm70.1/ST318 (n=1), emm44/ST31 (n=1) and emm18/ST442 (n=1) have clustered within a common group (Virulotype A) for both hospitals studied. Conclusion: The present study showed that group A streptococci (GAS) are genetically diverse and possess virulence genes regardless of their invasiveness. Majority of the GAS exhibited no restricted pattern of virulotypes except for a few distinct clusters. Therefore, it can be concluded that virulotyping is partially useful for characterizing a heterogeneous population of GAS in hospitals.

scholarly journals Retrospective Study of Listeria Monocytogenes Isolated in the Territory of Inner Eurasia from 1947 to 1999

Pathogens ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 184 ◽  
Author(s):  
Psareva ◽  
Egorova ◽  
Liskova ◽  
Razheva ◽  
Gladkova ◽  
...  
Keyword(s):  
Retrospective Study ◽  
Listeria Monocytogenes ◽  
20Th Century ◽  
Multilocus Sequence Typing ◽  
Clonal Complex ◽  
Small Ruminants ◽  
Virulence Gene ◽  
Penicillin G ◽  
Allelic Variants ◽  
Mlst Scheme

Listeriosis is one of the most significant humans and animals foodborne infectious diseases. Here, we characterized 48 Listeria monocytogenes strains isolated in the territory of inner Eurasia during the second half of the 20th century. A total of 23 strains (52.3%) were susceptible to the nine antibiotics tested, 30.43%, 15.22%, and 8.7% were resistant penicillin G, ampicillin, and enrofloxacin, respectively. We applied the multilocus sequence typing (MLST) scheme to determine the phylogenetic positions of the strains. All but one strain belonged to the II phylogenetic lineage, and the majority of the strains belonged to one of the previously described clonal complexes (СCs). More than 60% of the strains belonged to the clonal complex CC7 that prevailed among all sources, including cattle (58%), small ruminants (64%), rodents (71%), and humans (50%). Further, CC7, CC101, and CC124 were found among human isolates. The MLST scheme was supplemented with virulence gene analysis. In total, eight inlA, six inlB, and six inlC allelic variants were found, and all but one strain carried one of the two inlE alleles. Most strains (62.5%) belonged to the same multivirulence locus sequence typing (MvLST) type, which includes CC7, inlA allele 4, inlB allele 14, inlC allele 6, and inlE allele 8.

Development of a Multilocus sequence typing (MLST) scheme for Pan-Leishmania

Acta Tropica ◽  
2020 ◽  
Vol 201 ◽  
pp. 105189
Author(s):  
Juan Jose Lauthier ◽  
Paula Ruybal ◽  
Paola Andrea Barroso ◽  
Yoshihisa Hashiguchi ◽  
Jorge Diego Marco ◽  
...  
Keyword(s):  
Multilocus Sequence Typing ◽  
Mlst Scheme

scholarly journals Genetic diversity in Italian Lactobacillus sanfranciscensis strains assessed by multilocus sequence typing and pulsed-field gel electrophoresis analyses

Microbiology ◽  
2010 ◽  
Vol 156 (7) ◽  
pp. 2035-2045 ◽  
Author(s):  
Claudia Picozzi ◽  
Gaia Bonacina ◽  
Ileana Vigentini ◽  
Roberto Foschino
Keyword(s):  
Multilocus Sequence Typing ◽  
Geographical Area ◽  
Housekeeping Genes ◽  
Processing Conditions ◽  
Clonal Population ◽  
Factors Affecting ◽  
Lactobacillus Sanfranciscensis ◽  
Mlst Scheme ◽  
Sequence Types

Lactobacillus sanfranciscensis is a lactic acid bacterium that characterizes the sourdough environment. The genetic differences of 24 strains isolated in different years from sourdoughs, mostly collected in Italy, were examined and compared by PFGE and multilocus sequence typing (MLST). The MLST scheme, based on the analysis of six housekeeping genes (gdh, gyrA, mapA, nox, pgmA and pta) was developed for this study. PFGE with the restriction enzyme ApaI proved to have higher discriminatory power, since it revealed 22 different pulsotypes, while 19 sequence types were recognized through MLST analysis. Notably, restriction profiles generated from three isolates collected from the same firm but in three consecutive years clustered in a single pulsotype and showed the same sequence type, emphasizing the fact that the main factors affecting the dominance of a strain are correlated with processing conditions and the manufacturing environment rather than the geographical area. All results indicated a limited recombination among genes and the presence of a clonal population in L. sanfranciscensis. The MLST scheme proposed in this work can be considered a useful tool for characterization of isolates and for in-depth examination of the strain diversity and evolution of this species.

scholarly journals Simplified MLST scheme for direct typing of Leptospira human clinical samples

2018 ◽  
Vol 112 (4) ◽  
pp. 203-209 ◽  
Author(s):  
Vanina Varni ◽  
Yosena Chiani ◽  
Ariel Nagel ◽  
Paula Ruybal ◽  
Norma Bibiana Vanasco ◽  
...  
Keyword(s):  
Clinical Samples ◽  
Mlst Scheme

Is Multilocus Sequence Typing Approach Useful in Identification of Commensal Neisseria from Clinical Samples?

2018 ◽  
Vol 33 (4) ◽  
pp. 248-253
Author(s):  
Arij Mechergui ◽  
Wafa Achour ◽  
Dario Giorgini ◽  
Rekaya Baaboura ◽  
Muhamed-Kheir Taha ◽  
...  
Keyword(s):  
Multilocus Sequence Typing ◽  
Clinical Samples ◽  
Commensal Neisseria

scholarly journals Determination of Molecular Phylogenetics of Vibrio parahaemolyticus Strains by Multilocus Sequence Typing

2008 ◽  
Vol 190 (8) ◽  
pp. 2831-2840 ◽  
Author(s):  
Narjol González-Escalona ◽  
Jaime Martinez-Urtaza ◽  
Jaime Romero ◽  
Romilio T. Espejo ◽  
Lee-Ann Jaykus ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Population Structure ◽  
Multilocus Sequence Typing ◽  
Vibrio Parahaemolyticus ◽  
Molecular Phylogenetics ◽  
Clonal Complex ◽  
Housekeeping Genes ◽  
The United States ◽  
Health Concern ◽  
Mlst Scheme

ABSTRACT Vibrio parahaemolyticus is an important human pathogen whose transmission is associated with the consumption of contaminated seafood. There is a growing public health concern due to the emergence of a pandemic strain causing severe outbreaks worldwide. Many questions remain unanswered regarding the evolution and population structure of V. parahaemolyticus. In this work, we describe a multilocus sequence typing (MLST) scheme for V. parahaemolyticus based on the internal fragment sequences of seven housekeeping genes. This MLST scheme was applied to 100 V. parahaemolyticus strains isolated from geographically diverse clinical (n = 37) and environmental (n = 63) sources. The sequences obtained from this work were deposited and are available in a public database (http://pubmlst.org/vparahaemolyticus ). Sixty-two unique sequence types were identified, and most (50) were represented by a single isolate, suggesting a high level of genetic diversity. Three major clonal complexes were identified by eBURST analysis. Separate clonal complexes were observed for V. parahaemolyticus isolates originating from the Pacific and Gulf coasts of the United States, while a third clonal complex consisted of strains belonging to the pandemic clonal complex with worldwide distribution. The data reported in this study indicate that V. parahaemolyticus is genetically diverse with a semiclonal population structure and an epidemic structure similar to that of Vibrio cholerae. Genetic diversity in V. parahaemolyticus appears to be driven primarily by frequent recombination rather than mutation, with recombination ratios estimated at 2.5:1 and 8.8:1 by allele and site, respectively. Application of this MLST scheme to more V. parahaemolyticus strains and by different laboratories will facilitate production of a global picture of the epidemiology and evolution of this pathogen.

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