Identification of aquatic mycobacteria based on sequence analysis of the 16S–23S rRNA internal transcribed spacer region

Pourahmad, Fazel; Adams, Alexandra; Thompson, Kim D.; Richards, R. H.

doi:10.1099/jmm.0.000891

Cited by 6 publications

(6 citation statements)

References 32 publications

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“…Moreover, previous studies have reported discordant groupings when single genes are used in phylogenetic analyses of genus Mycobacterium . A phylogenetic analysis of ITS1 alone was not able to separate the complexes between slowly growing and rapidly growing mycobacteria (Zerihun et al 2011b; Pourahmad et al 2019). Similar disagreements have been reported when hsp65 is used alone (Whipps et al 2007).…”

Section: Resultsmentioning

confidence: 99%

“…The 16S rRNA gene was amplified and sequenced using published primers (Weisburg et al 1991) and modified protocols (Pierezan et al 2020). Similarly, the amplification process for the ITS1, rpoB , and hsp65 genes was conducted by using 2X DreamTaq Master Mix (Thermo Fisher Scientific) and an estimated 1–15 ng of gDNA template for each PCR, following the protocols that are outlined by Pourahmad et al (2019). All of the PCRs were performed on a SimpliAmp Thermal Cycler (Applied Biosystems, Foster City, California, USA) using the cycling conditions that are outlined by Pourahmad et al (2019).…”

Section: Methodsmentioning

confidence: 99%

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Mycobacterium salmoniphilum and M. chelonae in Captive Populations of Chinook Salmon

et al. 2021

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Chinook Salmon Oncorhynchus tshawytscha is a keystone fish species in the Pacific Northwest. In 2019, unusual mortalities occurred in two different populations of cultured fingerlings from the same facility in California, USA. The systems consist of outdoor, enclosed, flow‐through freshwater tanks that are maintained at 18 ± 1°C. Clinical signs and gross findings were only observed in one population and included abnormal swimming, inappetence, lethargy, skin discoloration, and the presence of multifocal nodular and ulcerative skin lesions. Microscopic lesions were infrequent and consisted of severe, locally extensive granulomatous dermatitis and myositis and mild, multifocal, granulomatous branchitis, myocarditis, and hepatitis. Intracellular acid‐fast organisms were observed within areas of granulomatous myositis. Posterior kidney swabs were collected and inoculated in nutrient‐rich and selective agar media and incubated at 25°C for 2 weeks. Visibly pure bacterial colonies were observed 7–10 d postinoculation. Partial sequences of 16S rRNA initially identified the recovered bacteria as members of the genus Mycobacterium. However, marked variability was observed among Mycobacterium spp. isolates by using repetitive extragenic palindromic polymerase chain reaction fingerprinting. Amplification and sequencing of the ribosomal RNA internal transcribed spacer, 65‐kDa heat shock protein, and RNA polymerase β‐subunit gene of the cultured isolates identified M. salmoniphilum and M. chelonae, discrete members of the M. chelonae‐abscessus complex, isolated from diseased Chinook Salmon fingerlings.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Mycobacterium salmoniphilum and M. chelonae in Captive Populations of Chinook Salmon

et al. 2021

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“…(Choudhary et al, 2015) ITS due to more variation properties than ribosomal sequences have been commonly used for molecular markers to figure out phylogenetic analysis and/or identification of bacteria, plant and animal strains and/or species (Lee et al, 2017;Rocha et al, 2019;Ali et al, 2019;Fazeli-Nasab et al, 2020). Pourahmad et al (2019) performed sequence analysis of the mycobacterial 16S-23S ITS region to determine aquatic mycobacteria species, reporting effectiveness of this marker. In another study, Yu et al (2020) evaluated the species specificity of ITS.…”

Section: Its (Internal Transcribed Spacer)mentioning

confidence: 99%

Application of different molecular markers in biotechnology

Kocaman

Toy

Marakli

2020

International Journal of Science Letters

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“…It has also been utilized in bacterial phylogenetic analysis and identification [ 26 ]. As a non-coding sequence, the ITS is more specific than the 16S rRNA gene [ 27 – 30 ]. In our previous study, further analysis of BLAST results based on ITS sequences alignment were utilized to identified Vibrio and Streptococcus species-specifically [ 31 – 33 ].…”

Section: Introductionmentioning

confidence: 99%

“…It has also been utilized in bacterial phylogenetic analysis and identification [26]. As a non-coding sequence, the ITS is more specific than the 16S rRNA gene [27][28][29][30].…”

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confidence: 99%

Species-specific identification of Pseudomonas based on 16S–23S rRNA gene internal transcribed spacer (ITS) and its combined application with next-generation sequencing

Yin

et al. 2022

BMC Microbiol

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Background Pseudomonas species are widely distributed in the human body, animals, plants, soil, fresh water, seawater, etc. Pseudomonas aeruginosa is one of the main pathogens involved in nosocomial infections. It can cause endocarditis, empyema, meningitis, septicaemia and even death. However, the Pseudomonas classification system is currently inadequate and not well established. Results In this study, the whole genomes of 103 Pseudomonas strains belonging to 62 species available in GenBank were collected and the specificity of the 16S–23S ribosomal RNA internal transcribed spacer (ITS) sequence was analysed. Secondary structures of ITS transcripts determining where the diversity bases were located were predicted. The alignment results using BLAST indicated that the ITS sequence is specific for most species in the genus. The remaining species were identified by additional frequency analyses based on BLAST results. A double-blind experiment where 200 ITS sequences were randomly selected indicated that this method could identify Pseudomonas species with 100% sensitivity and specificity. In addition, we applied a universal primer to amplify the Pseudomonas ITS of DNA extracts from fish samples with next-generation sequencing. The ITS analysis results were utilized to species-specifically identify the proportion of Pseudomonas species in the samples. Conclusions The present study developed a species-specific method identification and classification of Pseudomonas based on ITS sequences combined NGS. The method showed its potential application in other genera.

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Identification of aquatic mycobacteria based on sequence analysis of the 16S–23S rRNA internal transcribed spacer region

Cited by 6 publications

References 32 publications

Mycobacterium salmoniphilum and M. chelonae in Captive Populations of Chinook Salmon

Mycobacterium salmoniphilum and M. chelonae in Captive Populations of Chinook Salmon

Application of different molecular markers in biotechnology

Species-specific identification of Pseudomonas based on 16S–23S rRNA gene internal transcribed spacer (ITS) and its combined application with next-generation sequencing

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