scholarly journals Comparative Evaluation of Band-Based Genotyping Methods for Mycobacterium intracellulare and Its Application for Epidemiological Analysis

2020 ◽  
Vol 8 (9) ◽  
pp. 1315
Author(s):  
Jeong-Ih Shin ◽  
Jong-Hun Ha ◽  
Dong-Hae Lee ◽  
Jeong-Gyu Choi ◽  
Kyu-Min Kim ◽  
...  
Keyword(s):  
Minimum Spanning Tree ◽  
Variable Number ◽  
Clinical Isolates ◽  
Epidemiological Analysis ◽  
Mycobacterium Intracellulare ◽  
Fundamental Information ◽  
Tandem Repetition ◽  
Comparative Results ◽  
Geographical Characteristics ◽  
Discriminatory Index

Mycobacterium intracellulare is a leading cause of nontuberculous mycobacterial pulmonary disease, with a rapidly increasing prevalence worldwide. This bacterium, commonly distributed in soil and water, is known to be transmitted through the environment rather than between people. Therefore, it is imperative to establish distinguishable genotyping methods to understand the clinical outcome, disease relapses, and epidemiology. Therefore, in this study, representative band-based genotyping methods were performed using M. intracellualre clinical isolates, and their Hunter–Gaston discriminatory index (HGDI) was 0.947, 0.994, and 1 for variable number tandem repetition (VNTR), VNTR-mycobacterial interspersed repetitive units, pulsed field gel electrophoresis, and repetitive sequence based-PCR, respectively. Although VNTR showed relatively low HGDI, co-infection with other M. intracellualre strains could be determined by loci showing allele diversity from 0 to 0.69. Additionally, genetic distance of clinical isolates from Gyeongnam/Korea, and other regions/countries were visualized by minimum spanning tree (MST) using the globally available VNTR profiles. The results of MST revealed that M. intracellulare isolated from patients in Gyeongnam/Korea had specific VNTR genotypes, which may be evidence of the geographic distribution of M. intracellulare specific genotypes. The comparative results of genotyping techniques and geographical characteristics in this study may provide fundamental information for the epidemiology of M. intracellulare.

scholarly journals Molecular typing of Mycobacterium intracellulare using multilocus variable-number of tandem-repeat analysis: identification of loci and analysis of clinical isolates

Microbiology ◽  
2010 ◽  
Vol 156 (2) ◽  
pp. 496-504 ◽  
Author(s):  
Kazuya Ichikawa ◽  
Tetsuya Yagi ◽  
Takayuki Inagaki ◽  
Makoto Moriyama ◽  
Taku Nakagawa ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Tandem Repeat ◽  
Tandem Repeats ◽  
Variable Number ◽  
Clinical Isolates ◽  
Time Intervals ◽  
Mycobacterium Intracellulare ◽  
Epidemiological Tool ◽  
Disseminated Disease ◽  
Hiv Negative

In addition to its known status as a disseminated disease in HIV-positive patients, Mycobacterium avium complex (MAC) is increasingly recognized as a causative pathogen of respiratory disease in HIV-negative patients. MAC is divided into Mycobacterium avium, and the less-epidemiologically studied Mycobacterium intracellulare. Genetic typing for M. intracellulare using variable number of tandem repeats (VNTR) has not yet been developed. The aim of this study was to identify VNTR loci in the genome of M. intracellulare and apply them as an epidemiological tool to clinical isolates. Here, we identified 25 VNTR loci on the M. intracellulare genome, of which 16 showed variations among clinical isolates in the number of tandem repeat motifs. Among the 74 M. intracellulare isolates, 50 genotypes were distinguished using the 16 VNTR loci, resulting in a Hunter Gaston's discriminatory index of 0.988. Moreover, all 16 VNTR loci were stable in different sets of isolates recovered within time intervals ranging from 2 to 1551 days from 14 separate patients. These results indicate that for use as epidemiological markers of M. intracellulare, the loci in this VNTR assay are highly discriminating and stable over time.

scholarly journals Multiple-Locus Variable-Number Tandem Repeat Analysis of Dutch Bordetella pertussis Strains Reveals Rapid Genetic Changes with Clonal Expansion during the Late 1990s

2004 ◽  
Vol 186 (16) ◽  
pp. 5496-5505 ◽  
Author(s):  
Leo M. Schouls ◽  
Han G. J. van der Heide ◽  
Luc Vauterin ◽  
Paul Vauterin ◽  
Frits R. Mooi
Keyword(s):  
The Netherlands ◽  
Tandem Repeat ◽  
Bordetella Pertussis ◽  
Minimum Spanning Tree ◽  
Whooping Cough ◽  
Variable Number Tandem Repeat ◽  
Clonal Expansion ◽  
Variable Number ◽  
Multiple Locus ◽  
Repeat Analysis

ABSTRACT Bordetella pertussis, the causative agent of whooping cough, has remained endemic in The Netherlands despite extensive nationwide vaccination since 1953. In the 1990s, several epidemic periods have resulted in many cases of pertussis. We have proposed that strain variation has played a major role in the upsurges of this disease in The Netherlands. Therefore, molecular characterization of strains is important in identifying the causes of pertussis epidemiology. For this reason, we have developed a multiple-locus variable-number tandem repeat analysis (MLVA) typing system for B. pertussis. By combining the MLVA profile with the allelic profile based on multiple-antigen sequence typing, we were able to further differentiate strains. The relationships between the various genotypes were visualized by constructing a minimum spanning tree. MLVA of Dutch strains of B. pertussis revealed that the genotypes of the strains isolated in the prevaccination period were diverse and clearly distinct from the strains isolated in the 1990s. Furthermore, there was a decrease in diversity in the strains from the late 1990s, with a remarkable clonal expansion that coincided with the epidemic periods. Using this genotyping, we have been able to show that B. pertussis is much more dynamic than expected.

Convenient PCR method for variable-number tandem-repeat typing of Mycobacterium tuberculosis clinical isolates

2017 ◽  
Vol 139 ◽  
pp. 12-14 ◽  
Author(s):  
Junji Seto ◽  
Takayuki Wada ◽  
Yu Suzuki ◽  
Tatsuya Ikeda ◽  
Katsumi Mizuta ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Tandem Repeat ◽  
Variable Number Tandem Repeat ◽  
Variable Number ◽  
Clinical Isolates ◽  
Pcr Method

scholarly journals Microevolution ofStreptococcus agalactiaeST-261 from Australia Indicates Dissemination via Imported Tilapia and Ongoing Adaptation to Marine Hosts or Environment

2018 ◽  
Vol 84 (16) ◽  
Author(s):  
Minami Kawasaki ◽  
Jerome Delamare-Deboutteville ◽  
Rachel O. Bowater ◽  
Mark J. Walker ◽  
Scott Beatson ◽  
...  
Keyword(s):  
Streptococcus Agalactiae ◽  
Core Genome ◽  
Minimum Spanning Tree ◽  
Vaccine Development ◽  
The United States ◽  
Wild Fish ◽  
Clinical Isolates ◽  
Global Trade ◽  
Content Type ◽  
Wide Range

ABSTRACTStreptococcus agalactiae(group BStreptococcus[GBS]) causes disease in a wide range of animals. The serotype Ib lineage is highly adapted to aquatic hosts, exhibiting substantial genome reduction compared with terrestrial conspecifics. Here, we sequence genomes from 40 GBS isolates, including 25 isolates from wild fish and captive stingrays in Australia, six local veterinary or human clinical isolates, and nine isolates from farmed tilapia in Honduras, and compared them with 42 genomes from public databases. Phylogenetic analysis based on nonrecombinant core-genome single nucleotide polymorphisms (SNPs) indicated that aquatic serotype Ib isolates from Queensland were distantly related to local veterinary and human clinical isolates. In contrast, Australian aquatic isolates are most closely related to a tilapia isolate from Israel, differing by only 63 core-genome SNPs. A consensus minimum spanning tree based on core-genome SNPs indicates the dissemination of sequence type 261 (ST-261) from an ancestral tilapia strain, which is congruent with several introductions of tilapia into Australia from Israel during the 1970s and 1980s. Pangenome analysis identified 1,440 genes as core, with the majority being dispensable or strain specific, with non-protein-coding intergenic regions (IGRs) divided among core and strain-specific genes. Aquatic serotype Ib strains have lost many virulence factors during adaptation, but six adhesins were well conserved across the aquatic isolates and might be critical for virulence in fish and for targets in vaccine development. The close relationship among recent ST-261 isolates from Ghana, the United States, and China with the Israeli tilapia isolate from 1988 implicates the global trade in tilapia seed for aquaculture in the widespread dissemination of serotype Ib fish-adapted GBS.IMPORTANCEStreptococcus agalactiae(GBS) is a significant pathogen of humans and animals. Some lineages have become adapted to particular hosts, and serotype Ib is highly specialized to fish. Here, we show that this lineage is likely to have been distributed widely by the global trade in tilapia for aquaculture, with probable introduction into Australia in the 1970s and subsequent dissemination in wild fish populations. We report here the variability in the polysaccharide capsule among this lineage but identify a cohort of common surface proteins that may be a focus of future vaccine development to reduce the biosecurity risk in international fish trade.

scholarly journals Genotyping of ClinicalMycobacterium tuberculosisIsolates Based on IS6110and MIRU-VNTR Polymorphisms

2013 ◽  
Vol 2013 ◽  
pp. 1-6 ◽  
Author(s):  
Anna Żaczek ◽  
Anna Brzostek ◽  
Arkadiusz Wojtasik ◽  
Jarosław Dziadek ◽  
Anna Sajduda
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Variable Number Tandem Repeat ◽  
Pcr Amplification ◽  
Detailed Comparison ◽  
Variable Number ◽  
Apparent Difference ◽  
Typing Method ◽  
Banding Patterns ◽  
Typing Technique ◽  
Discriminatory Index

In this study, 155 clinicalMycobacterium tuberculosisisolates were subject to genotyping with fast ligation-mediated PCR (FLiP). This typing method is a modified mixed-linker PCR, a rapid approach based on the PCR amplification ofHhaI restriction fragments of genomic DNA containing the 3′ end of IS6110and resolving the amplicons by polyacrylamide gel electrophoresis. The results were compared with previous data of the more commonly used methods, 15-locus mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) typing and, to verify combined FLiP/MIRU-VNTR clusters, the reference IS6110restriction fragment length polymorphism (RFLP). FLiP banding patterns were highly reproducible and polymorphic. This method differentiated 119 types among the study set compared to 108 distinct MIRU-VNTR profiles. The discriminatory power of FLiP was slightly higher than that of MIRU-VNTR analysis (Hunter-Gaston Discriminatory Index = 0.991 and 0.990, resp.). Detailed comparison of the clusters defined by each of the methods revealed, however, a more apparent difference in the discriminatory abilities that favored FLiP. Clustering of strains by using combined results of these two PCR-based methods correlated well with IS6110RFLP-defined clusters, further confirming high discriminatory potential of FLiP typing. These results indicate that FLiP could be an attractive and valuable secondary typing technique for verification of MIRU-VNTR clusters ofM. tuberculosisstrains.

scholarly journals Feature of Adhesins Produced by Human Clinical Isolates of Mycobacterium intracellulare, Mycobacterium intracellulare subsp. chimaera and Closely Related Species

2020 ◽  
Vol 8 (8) ◽  
pp. 1154
Author(s):  
Louise H. Lefrancois ◽  
Thierry Cochard ◽  
Maxime Branger ◽  
Olivia Peuchant ◽  
Cyril Conde ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Related Species ◽  
Fluorescent Protein ◽  
Severe Disease ◽  
A549 Cells ◽  
Clinical Isolates ◽  
Heparin Binding ◽  
Closely Related Species ◽  
Mycobacterium Intracellulare ◽  
Green Fluorescent

The Mycobacterium avium complex includes two closely related species, Mycobacterium avium and Mycobacterium intracellulare. They are opportunistic pathogens in humans and responsible for severe disease in a wide variety of animals. Yet, little is known about factors involved in their pathogenicity. Here, we identified, purified and characterized adhesins belonging to the heparin-binding hemagglutinin (HBHA) and laminin-binding protein (LBP) family from M. intracellulare ATCC13950 and examined clinical isolates from patients with different pathologies associated with M. intracellulare infection for the presence and conservation of HBHA and LBP. Using a recombinant derivative strain of M. intracellulare ATCC13950 producing green fluorescent protein and luciferase, we found that the addition of heparin inhibited mycobacterial adherence to A549 cells, whereas the addition of laminin enhanced adherence. Both HBHA and LBP were purified by heparin-Sepharose chromatography and their methylation profiles were determined by mass spectrometry. Patients with M. intracellulare infection mounted strong antibody responses to both proteins. By using PCR and immunoblot analyses, we found that both proteins were highly conserved among all 17 examined clinical M. intracellulare isolates from patients with diverse disease manifestations, suggesting a conserved role of these adhesins in M. intracellulare virulence in humans and their potential use as a diagnostic tool.

scholarly journals Clonal Associations among Staphylococcus aureus Isolates from Various Sites of Infection

2001 ◽  
Vol 69 (1) ◽  
pp. 345-352 ◽  
Author(s):  
Mary C. Booth ◽  
Lisa M. Pence ◽  
Param Mahasreshti ◽  
Michelle C. Callegan ◽  
Michael S. Gilmore
Keyword(s):  
Staphylococcus Aureus ◽  
High Frequency ◽  
Gel Electrophoresis ◽  
Pulsed Field Gel Electrophoresis ◽  
Clinical Isolates ◽  
Epidemiological Analysis ◽  
Specific Pcr ◽  
Novel Approaches ◽  
Phylogenetic Lineages ◽  
Infection Types

ABSTRACT A molecular epidemiological analysis was undertaken to identify lineages of Staphylococcus aureus that may be disproportionately associated with infection. Pulsed-field gel electrophoresis analysis of 405 S. aureus clinical isolates collected from various infection types and geographic locations was performed. Five distinct S. aureus lineages (SALs 1, 2, 4, 5, and 6) were identified, which accounted for 19.01, 9.14, 22.72, 10.12, and 4.69% of isolates, respectively. In addition, 85 lineages which occurred with frequencies of <2.5% were identified and were termed “sporadic.” The most prevalent lineage was methicillin-resistant S. aureus (SAL 4). The second most prevalent lineage, SAL 1, was also isolated at a high frequency from the anterior nares of healthy volunteers, suggesting that its prevalence among clinical isolates may be a consequence of high carriage rates in humans. Gene-specific PCR was carried out to detect genes for a number of staphylococcal virulence traits. tstand cna were found to be significantly associated with prevalent lineages compared to sporadic lineages. When specific infection sites were examined, SAL 4 was significantly associated with respiratory tract infection, while SAL 2 was enriched among blood isolates. SAL 1 and SAL 5 were clonally related to SALs shown by others to be widespread in the clinical isolate population. We conclude from this study that at least five phylogenetic lineages of S. aureus are highly prevalent and widely distributed among clinical isolates. The traits that confer on these lineages a propensity to infect may suggest novel approaches to antistaphylococcal therapy.

scholarly journals MUC Gene Abnormalities in Sporadic and Hereditary Mucinous Colon Cancers with Microsatellite Instability

2005 ◽  
Vol 21 (3) ◽  
pp. 121-126 ◽  
Author(s):  
Chiara Pastrello ◽  
Manuela Santarosa ◽  
Mara Fornasarig ◽  
Roberto Sigon ◽  
Tiziana Perin ◽  
...  
Keyword(s):  
Microsatellite Instability ◽  
Southern Blotting ◽  
Repetitive Sequences ◽  
Variable Number ◽  
Molecular Pathway ◽  
Colon Cancers ◽  
Tandem Repetition ◽  
Indirect Consequence ◽  
Muc Genes ◽  
Gene Alterations

Aim of this study was verifying whether mucin producing colon cancers (CRCs) could develop through a molecular pathway involving microsatellite instability (MSI) and MUC gene alterations. Out of 49 CRCs expressing variable amounts of mucin, 22 (44.9%) were MSI-H and 5 (10.2%) were MSI-L. MUC genes were analyzed by Southern blotting and extra bands were evident in the Variable Number Tandem Repetition (VNTR) regions of MUC2 (5 cases) and MUC5AC (2 cases), but not MUC1 and MUC4 genes. Since the somatic VNTR abnormalities were detected in 6 MSI-H and in 1 MSI-L tumors, they seem to be peculiar of mismatch repair defective CRCs. Our finding suggests that alteration and/or loss of structurally normal MUC genes may be an important step in the neoplastic molecular pathway of a subset of CRCs and that mutations involving VNTR repetitive sequences may exist in MSI tumors as a direct and/or indirect consequence of an inefficient MMR system.

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