Use of multilocus enzyme electrophoresis to examine genetic relationships amongst isolates of Mycobacterium intracellulare and related species

Feizabadi, Mohammad M.; Robertson, Ian D.; Cousins, Debby; Dawson, David J.; Hampson, David J.

doi:10.1099/00221287-143-4-1461

Cited by 20 publications

(10 citation statements)

References 36 publications

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“…Enzymatic markers and DNA markers obtained in the bacterial population analyses are diffi cult to compare because very little is known about the dissonance between those molecular markers in bacterial domains. The value of HS reported by Feizabadi et al [9] in an enzymatic analysis of the M. tuberculosis complex was lower than that noted by other researchers who relied on DNA markers [17,26]. Many enzymatic analyses of bacteria, plants and animals produced genetic diversity values that were twoto three-fold lower in comparison with DNA marker tests [13,21].…”

Section: Discussioncontrasting

confidence: 44%

Evaluation of anthropogenic pollution in river water based on the genetic diversity of Aeromonas hydrophila

Korzekwa¹,

Gołaś²,

Harnisz³

2012

Archives of Environmental Protection

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Aeromonas hydrophila is a valuable indicator of the quality of water polluted by sewage and pathogens that pose a risk for humans and cold-blooded animals, including fi sh. The main aim of this research was to evaluate anthropogenic pollution of river water based on genetic diversity of 82 A. hydrophila strains by means of RAPD, semi-random AP-PCR (ISJ) and the rep-BOX conservative repeats test. Genetic diversity of A. hydrophila was HT = 0.28 (SD = 0.02) for all DNA markers (RAPD, semi random and rep-BOX).None of the analyzed electrophoretic patterns was identical, implying that there were many sources of strain transmission. The presence of genes for aerolysin (aerA), hemolysin (ahh1) and the cytotoxic enzyme complex (AHCYTOGEN) was verifi ed for all tested strains, and drug resistance patterns for tetracycline, enrofl oxacin and erythromycin were determined. The most diverse A. hydrophila strains isolated from river water were susceptible to enrofl oxacine (HS = 0.27), whereas less diverse strains were susceptible to erythromycin (HS = 0.24). The presence of the multidrug resistance marker (ISJ4-25; 1100 bp locus) in the examined strains (resistant to three analyzed drugs) indicates that intensive fi sh cultivation affects the microbiological quality of river water.

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Section: Discussioncontrasting

confidence: 44%

Evaluation of anthropogenic pollution in river water based on the genetic diversity of Aeromonas hydrophila

Korzekwa¹,

Gołaś²,

Harnisz³

2012

Archives of Environmental Protection

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“…(7,37,49). The gene products for argH, cya, gnd, and pgm have been used previously in the multilocus enzyme electrophoresis (MLEE) analysis of mycobacteria (20,56,57,60). Amplification was performed using 25 l of ReddyMix PCR master mix (Thermo Fisher Scientific Inc.) and 1 l of each primer (10 pmol).…”

Section: Methodsmentioning

confidence: 99%

Multilocus Sequence Analysis and rpoB Sequencing of Mycobacterium abscessus (Sensu Lato) Strains

et al. 2011

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Mycobacterium abscessus, Mycobacterium bolletii, and Mycobacterium massiliense (Mycobacterium abscessus sensu lato) are closely related species that currently are identified by the sequencing of the rpoB gene. However, recent studies show that rpoB sequencing alone is insufficient to discriminate between these species, and some authors have questioned their current taxonomic classification. We studied here a large collection of M. abscessus (sensu lato) strains by partial rpoB sequencing (752 bp) and multilocus sequence analysis (MLSA). The final MLSA scheme developed was based on the partial sequences of eight housekeeping genes: argH, cya, glpK, gnd, murC, pgm, pta, and purH. The strains studied included the three type strains (M. abscessus CIP 104536 T , M. massiliense CIP 108297 T , and M. bolletii CIP 108541 T ) and 120 isolates recovered between 1997 and 2007 in France, Germany, Switzerland, and Brazil. The rpoB phylogenetic tree confirmed the existence of three main clusters, each comprising the type strain of one species. However, divergence values between the M. massiliense and M. bolletii clusters all were below 3% and between the M. abscessus and M. massiliense clusters were from 2.66 to 3.59%. The tree produced using the concatenated MLSA gene sequences (4,071 bp) also showed three main clusters, each comprising the type strain of one species. The M. abscessus cluster had a bootstrap value of 100% and was mostly compact. Bootstrap values for the M. massiliense and M. bolletii branches were much lower (71 and 61%, respectively), with the M. massiliense cluster having a fuzzy aspect. Mean (range) divergence values were 2.17% (1.13 to 2.58%) between the M. abscessus and M. massiliense clusters, 2.37% (1.5 to 2.85%) between the M. abscessus and M. bolletii clusters, and 2.28% (0.86 to 2.68%) between the M. massiliense and M. bolletii clusters. Adding the rpoB sequence to the MLSA-concatenated sequence (total sequence, 4,823 bp) had little effect on the clustering of strains. We found 10/120 (8.3%) isolates for which the concatenated MLSA gene sequence and rpoB sequence were discordant (e.g., M. massiliense MLSA sequence and M. abscessus rpoB sequence), suggesting the intergroup lateral transfers of rpoB. In conclusion, our study strongly supports the recent proposal that M. abscessus, M. massiliense, and M. bolletii should constitute a single species. Our findings also indicate that there has been a horizontal transfer of rpoB sequences between these subgroups, precluding the use of rpoB sequencing alone for the accurate identification of the two proposed M. abscessus subspecies.

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“…The amplification profile consisted of 10 min at 95°C, followed by 30 cycles, with 1 cycle consisting of 30 s at 95°C, 30 s at 63°C, and 45 s at 72°C. We extended the established method to 85 clinical strains independently identified by a multiple-target sequence analysis of rpoB, hsp65, gnd, and glpK as previously described (Table 1) (2,5,12).…”

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confidence: 99%

Efficient Differentiation of Mycobacterium abscessus Complex Isolates to the Species Level by a Novel PCR-Based Variable-Number Tandem-Repeat Assay

Choi¹,

Chang

Whang

et al. 2011

J Clin Microbiol

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A novel duplex PCR method based on variable-number tandem-repeat targets to discriminate among Mycobacterium abscessus complex isolates was developed and evaluated in 85 clinical isolates. The assay accuracy was confirmed by a multiple-target sequence analysis. The duplex PCR assay is a one-step, reliable, and accurate assay for discriminating M. abscessus species.Mycobacterium abscessus belongs to a group of rapidly growing mycobacteria (RGM) that cause a wide spectrum of infections in humans (6,17). Recently, M. abscessus complex was divided into three closely related Mycobacterium species: M. abscessus (sensu stricto) (hereafter referred to as M. abscessus), M. massiliense, and M. bolletii (1, 4).M. abscessus is the most drug-resistant mycobacterial species known and exhibits unsatisfactory treatment response rates, especially for patients with pulmonary disease (6, 17). Inducible resistance of M. abscessus to clarithromycin has been suggested as an explanation for the lack of efficacy of clarithromycin-based treatments against the bacterium (14). In contrast, M. massiliense shows marked susceptibility to clarithromycin due to the absence of inducible resistance to macrolides (8). Therefore, treatment response rates to clarithromycin-based antibiotic therapy are higher in patients with M. massiliense than in those with M. abscessus lung disease (10). M. bolletii, a rare pathogen at present, has also been shown to be highly resistant to clarithromycin (3, 9, 11).Since recent studies suggest that antibiotic susceptibility and treatment outcomes differ significantly among these species, the species-level identification of M. abscessus complex isolates has been strongly recommended in order to determine the clinical significance and to assist in managing patients (8,10).Despite progress in our ability to identify mycobacterial species using molecular methods, M. abscessus complex isolates cannot be reliably identified at the species level by sequencing a single genetic locus such as rpoB or hsp65 (13). Currently, the species-level identification of M. abscessus complex solely depends on multiple sequencing analysis targeting several genes, including rpoB, hsp65, gnd, glpK, secA, and sodA (2, 13, 15). This process requires a number of additional, laborious steps (e.g., cloning) to obtain the sequences of the products, leading to expensive and complicated procedures that must be done in reference laboratories.Increasingly, variable-number tandem-repeat (VNTR) analyses have been used for the molecular typing of isolated mycobacterial strains as well as for mycobacterial species identification (7, 16). In this study, a novel duplex PCR method based on variable-number tandem-repeat targets to discriminate among Mycobacterium abscessus complex isolates was developed and evaluated in 85 clinical isolates.A total of 85 clinical isolates were collected between

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Use of multilocus enzyme electrophoresis to examine genetic relationships amongst isolates of Mycobacterium intracellulare and related species

Cited by 20 publications

References 36 publications

Evaluation of anthropogenic pollution in river water based on the genetic diversity of Aeromonas hydrophila

Evaluation of anthropogenic pollution in river water based on the genetic diversity of Aeromonas hydrophila

Multilocus Sequence Analysis and rpoB Sequencing of Mycobacterium abscessus (Sensu Lato) Strains

Efficient Differentiation of Mycobacterium abscessus Complex Isolates to the Species Level by a Novel PCR-Based Variable-Number Tandem-Repeat Assay

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