scholarly journals rpoB mutations conferring rifampicin-resistance affect growth, stress response and motility in Vibrio vulnificus

Microbiology ◽  
2020 ◽  
Vol 166 (12) ◽  
pp. 1160-1170 ◽  
Author(s):  
Laura Cutugno ◽  
Jennifer Mc Cafferty ◽  
Jan Pané-Farré ◽  
Conor O’Byrne ◽  
Aoife Boyd
Keyword(s):  
Stress Response ◽  
Type Species ◽  
Vibrio Vulnificus ◽  
Growth Stress ◽  
Rifampicin Resistance ◽  
Secondary Effects ◽  
Rich Medium ◽  
Content Type ◽  
Link Type ◽  
Resistant Strains

Rifampicin is a broad-spectrum antibiotic that binds to the bacterial RNA polymerase (RNAP), compromising DNA transcription. Rifampicin resistance is common in several microorganisms and it is typically caused by point mutations in the gene encoding the β subunit of RNA polymerase, rpoB. Different rpoB mutations are responsible for various levels of rifampicin resistance and for a range of secondary effects. rpoB mutations conferring rifampicin resistance have been shown to be responsible for severe effects on transcription, cell fitness, bacterial stress response and virulence. Such effects have never been investigated in the marine pathogen Vibrio vulnificus , even though rifampicin-resistant strains of V. vulnificus have been isolated previously. Moreover, spontaneous rifampicin-resistant strains of V. vulnificus have an important role in conjugation and mutagenesis protocols, with poor consideration of the effects of rpoB mutations. In this work, effects on growth, stress response and virulence of V. vulnificus were investigated using a set of nine spontaneous rifampicin-resistant derivatives of V. vulnificus CMCP6. Three different mutations (Q513K, S522L and H526Y) were identified with varying incidence rates. These three mutant types each showed high resistance to rifampicin [minimal inhibitory concentration (MIC) >800 µg ml−1], but different secondary effects. The strains carrying the mutation H526Y had a growth advantage in rich medium but had severely reduced salt stress tolerance in the presence of high NaCl concentrations as well as a significant reduction in ethanol stress resistance. Strains possessing the S522L mutation had reduced growth rate and overall biomass accumulation in rich medium. Furthermore, investigation of virulence characteristics demonstrated that all the rifampicin-resistant strains showed compromised motility when compared with the wild-type, but no major effects on exoenzyme production were observed. These findings reveal a wide range of secondary effects of rpoB mutations and indicate that rifampicin resistance is not an appropriate selectable marker for studies that aim to investigate phenotypic behaviour in this organism.

scholarly journals Accuracy of an amplicon-sequencing nanopore approach to identify variants in tuberculosis drug-resistance-associated genes

2021 ◽  
Vol 7 (12) ◽  
Author(s):  
Carla Mariner-Llicer ◽  
Galo A. Goig ◽  
Laura Zaragoza-Infante ◽  
Manuela Torres-Puente ◽  
Luis Villamayor ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Type Species ◽  
Variant Calling ◽  
Targeted Sequencing ◽  
Clinical Samples ◽  
Drug Resistant ◽  
Content Type ◽  
Link Type ◽  
Resistant Strains ◽  
Drug Resistant Strains

A rapid and accurate diagnostic assay represents an important means to detect Mycobacterium tuberculosis , identify drug-resistant strains and ensure treatment success. Currently employed techniques to diagnose drug-resistant tuberculosis include slow phenotypic tests or more rapid molecular assays that evaluate a limited range of drugs. Whole-genome-sequencing-based approaches can detect known drug-resistance-conferring mutations and novel variations; however, the dependence on growing samples in culture, and the associated delays in achieving results, represents a significant limitation. As an alternative, targeted sequencing strategies can be directly performed on clinical samples at high throughput. This study proposes a targeted sequencing assay to rapidly detect drug-resistant strains of M. tuberculosis using the Nanopore MinION sequencing platform. We designed a single-tube assay that targets nine genes associated with drug resistance to seven drugs and two phylogenetic-determining regions to determine strain lineage and tested it in nine clinical isolates and six sputa. The study’s main aim is to calibrate MinNION variant calling to detect drug-resistance-associated mutations with different frequencies to match the accuracy of Illumina (the current gold-standard sequencing technology) from both culture and sputum samples. After calibrating Nanopore MinION variant calling, we demonstrated 100% agreement between Illumina WGS and our MinION set up to detect known drug resistance and phylogenetic variants in our dataset. Importantly, other variants in the amplicons are also detected, decreasing the recall. We identify minority variants and insertions/deletions as crucial bioinformatics challenges to fully reproduce Illumina WGS results.

scholarly journals The general stress response of Staphylococcus aureus promotes tolerance of antibiotics and survival in whole human blood

Microbiology ◽  
2020 ◽  
Vol 166 (11) ◽  
pp. 1088-1094 ◽  
Author(s):  
Nisha Ranganathan ◽  
Rebecca Johnson ◽  
Andrew M. Edwards
Keyword(s):  
Staphylococcus Aureus ◽  
Stress Response ◽  
Human Blood ◽  
Chronic Infection ◽  
Type Species ◽  
Content Type ◽  
General Stress Response ◽  
Link Type ◽  
General Stress ◽  
Immune Defences

Staphylococcus aureus is a frequent cause of invasive human infections such as bacteraemia and infective endocarditis. These infections frequently relapse or become chronic, suggesting that the pathogen has mechanisms to tolerate the twin threats of therapeutic antibiotics and host immunity. The general stress response of S. aureus is regulated by the alternative sigma factor B (σB) and provides protection from multiple stresses including oxidative, acidic and heat. σB also contributes to virulence, intracellular persistence and chronic infection. However, the protective effect of σB on bacterial survival during exposure to antibiotics or host immune defences is poorly characterized. We found that σB promotes the survival of S. aureus exposed to the antibiotics gentamicin, ciprofloxacin, vancomycin and daptomycin, but not oxacillin or clindamycin. We also found that σB promoted staphylococcal survival in whole human blood, most likely via its contribution to oxidative stress resistance. Therefore, we conclude that the general stress response of S. aureus may contribute to the development of chronic infection by conferring tolerance to both antibiotics and host immune defences.

scholarly journals Phenotypic and molecular characterization of macrolide resistance mechanisms among Streptococcus pneumoniae isolated in Tunisia

2020 ◽  
Vol 69 (4) ◽  
pp. 505-520 ◽  
Author(s):  
B. Midouni Ayadi ◽  
E. Mehiri ◽  
H. Draoui ◽  
A. Ghariani ◽  
L. Essalah ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Type Species ◽  
Resistance Mechanisms ◽  
Macrolide Resistance ◽  
23S Rrna ◽  
Serotype Distribution ◽  
Antibiotic Resistant ◽  
Content Type ◽  
Link Type ◽  
Resistant Strains

Introduction. Streptococcus pneumoniae is responsible for many community infections, with the main ones being pneumonia and meningitis. Pneumococcus has developed increased resistance to multiple classes of antibiotics. The evolution of antibiotic resistance in pneumococcus was influenced by changes in serotype distribution under vaccine selection pressure. Aim. The aim of this study was to determine the genes involved in macrolide resistance, the antimicrobial susceptibility, the serotype distribution and the spread of international antibiotic-resistant clones among clinical isolates of S. pneumoniae . Methodology. We investigated 86 erythromycin-resistant S. pneumoniae strains isolated from respiratory (n=74) or non-respiratory (n=12) samples in Tunisia. Antimicrobial susceptibility was tested using the disk diffusion method. Macrolide-resistant strains were analysed by polymerase chain reaction (PCR) for ermA, ermB, mefA and msrD. We also investigated the macrolide resistance mechanisms in eight isolates (9.3%) by sequencing the L4 and L22 riboprotein-coding genes, plus relevant segments of the three 23S rRNA genes. Capsular serotypes were detected by multiplex PCR. Sequence types (STs) were explored using multilocus sequence typing (MLST). Results. Among the 86 studied strains, 70 (81.4 %) were resistant to penicillin G. The prevalent serotypes were 19F, 14, 19A and 23F. We observed that the cMLSB phenotype (66/86, 76.7%) was the most common in these pneumococci. In addition, ermB was the most frequent resistance gene. No mutation in ribosomal protein L22 or L4 or 23S rRNA was detected. Overall, 44 STs were identified in this study, including 16 that were described for the first time. Resistance to lincomycin, tetracycline and trimethoprim/sulfamethoxazole was observed in 55 (64 %), 34 (39.5 %) and 31 (36 %) isolates, respectively. Furthermore, an increase in fluoroquinolone use in particular may lead to the emergence of levofloxacin-resistant strains. Multidrug resistance was observed in 83 isolates (96.5%). Three global antibiotic-resistant clones were identified: Denmark14 ST230, Portugal19F ST177 and Spain9V ST156. Conclusion. This study shows that macrolide resistance among S. pneumoniae isolated in Tunisia is mainly related to target site modification. Our observations demonstrate a high degree of genetic diversity and capsular types among strains resistant to macrolides.

A stop–gain mutation in sigma factor SigH (MAB_3543c) may be associated with tigecycline resistance in Mycobacteroides abscessus

2021 ◽  
Vol 70 (7) ◽  
Author(s):  
Col Lin Lee ◽  
Hien Fuh Ng ◽  
Yun Fong Ngeow ◽  
Zin Thaw
Keyword(s):  
Stress Response ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Sigma Factor ◽  
Type Species ◽  
Treatment Modalities ◽  
Gene Overexpression ◽  
Whole Genome ◽  
Content Type ◽  
Link Type

Introduction. Tigecycline is currently acknowledged to be one of the most effective antibiotics against infections caused by Mycobacteroides abscessus . Gap statement. The genetic determinants of tigecycline resistance in M. abscessus are not well understood. Aim. In this study, we characterized a tigecycline-resistant M. abscessus mutant, designated CL7, to identify the potential resistance mechanism. Methodology. CL7 was characterized using antimicrobial susceptibility testing, whole-genome sequencing, PCR and RT-qPCR. For biological verification, gene overexpression assays were carried out. Results. Whole-genome sequencing and the subsequent gene overexpression assays showed that CL7 harboured a stop–gain mutation in MAB_3543 c, which may be responsible for the tigecycline resistance phenotype. This gene encodes an orthologue of SigH, which is involved in the positive regulation of physiological stress response and is negatively regulated by the RshA anti-sigma factor in Mycobacterium tuberculosis . We hypothesized that the MAB_3543 c mutation may disrupt the interaction between SigH and RshA (MAB_3542 c). RT-qPCR analyses revealed the upregulation of MAB_3543 c and other key stress response genes, which has previously been shown to be a hallmark of SigH–RshA bond disruption and tigecycline resistance. Conclusion. The MAB_3543c mutation may represent a novel determinant of tigecycline resistance in M. abscessus . The findings of this study will hopefully contribute to our knowledge of potential tigecycline resistance mechanisms in M. abscessus , which may lead to better diagnostics and treatment modalities in the future.

scholarly journals Faecalicoccus acidiformans gen. nov., sp. nov., isolated from the chicken caecum, and reclassification of Streptococcus pleomorphus (Barnes et al. 1977), Eubacterium biforme (Eggerth 1935) and Eubacterium cylindroides (Cato et al. 1974) as Faecalicoccus pleomorphus comb. nov., Holdemanella biformis gen. nov., comb. nov. and Faecalitalea cylindroides gen. nov., comb. nov., respectively, within the family Erysipelotrichaceae

2014 ◽  
Vol 64 (Pt_11) ◽  
pp. 3877-3884 ◽  
Author(s):  
Celine De Maesschalck ◽  
Filip Van Immerseel ◽  
Venessa Eeckhaut ◽  
Siegrid De Baere ◽  
Margo Cnockaert ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Type Species ◽  
Gene Sequence ◽  
Sequence Similarity ◽  
16S Rrna Gene Sequence ◽  
Rrna Gene ◽  
Rrna Gene Sequence ◽  
Content Type ◽  
Link Type

Strains LMG 27428T and LMG 27427 were isolated from the caecal content of a chicken and produced butyric, lactic and formic acids as major metabolic end products. The genomic DNA G+C contents of strains LMG 27428T and LMG 27427 were 40.4 and 38.8 mol%. On the basis of 16S rRNA gene sequence similarity, both strains were most closely related to the generically misclassified Streptococcus pleomorphus ATCC 29734T. Strain LMG 27428T could be distinguished from S. pleomorphus ATCC 29734T based on production of more lactic acid and less formic acid in M2GSC medium, a higher DNA G+C content and the absence of activities of acid phosphatase and leucine, arginine, leucyl glycine, pyroglutamic acid, glycine and histidine arylamidases, while strain LMG 27428 was biochemically indistinguishable from S. pleomorphus ATCC 29734T. The novel genus Faecalicoccus gen. nov. within the family Erysipelotrichaceae is proposed to accommodate strains LMG 27428T and LMG 27427. Strain LMG 27428T ( = DSM 26963T) is the type strain of Faecalicoccus acidiformans sp. nov., and strain LMG 27427 ( = DSM 26962) is a strain of Faecalicoccus pleomorphus comb. nov. (type strain LMG 17756T = ATCC 29734T = DSM 20574T). Furthermore, the nearest phylogenetic neighbours of the genus Faecalicoccus are the generically misclassified Eubacterium cylindroides DSM 3983T (94.4 % 16S rRNA gene sequence similarity to strain LMG 27428T) and Eubacterium biforme DSM 3989T (92.7 % 16S rRNA gene sequence similarity to strain LMG 27428T). We present genotypic and phenotypic data that allow the differentiation of each of these taxa and propose to reclassify these generically misnamed species of the genus Eubacterium formally as Faecalitalea cylindroides gen. nov., comb. nov. and Holdemanella biformis gen. nov., comb. nov., respectively. The type strain of Faecalitalea cylindroides is DSM 3983T = ATCC 27803T = JCM 10261T and that of Holdemanella biformis is DSM 3989T = ATCC 27806T = CCUG 28091T.

scholarly journals Pseudoseohaeicola caenipelagi gen. nov., sp. nov., isolated from a tidal flat

2015 ◽  
Vol 65 (Pt_6) ◽  
pp. 1819-1824 ◽  
Author(s):  
Sooyeon Park ◽  
Ji-Min Park ◽  
Chul-Hyung Kang ◽  
Song-Gun Kim ◽  
Jung-Hoon Yoon
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Tidal Flat ◽  
Type Strain ◽  
Type Species ◽  
Phylogenetic Trees ◽  
Sequence Similarity ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type

A Gram-stain-negative, non-motile, aerobic and pleomorphic bacterium, designated BS-W13T, was isolated from a tidal flat on the South Sea, South Korea, and its taxonomic position was investigated using a polyphasic approach. Strain BS-W13T grew optimally at 25 °C, at pH 7.0–8.0 and in the presence of 1.0–2.0 % (w/v) NaCl. Neighbour-joining and maximum-parsimony phylogenetic trees based on 16S rRNA gene sequences showed that strain BS-W13T clustered with the type strain of Seohaeicola saemankumensis , showing the highest sequence similarity (95.96 %) to this strain. Strain BS-W13T exhibited 16S rRNA gene sequence similarity values of 95.95, 95.91, 95.72 and 95.68 % to the type strains of Sulfitobacter donghicola , Sulfitobacter porphyrae , Sulfitobacter mediterraneus and Roseobacter litoralis , respectively. Strain BS-W13T contained Q-10 as the predominant ubiquinone and C18 : 1ω7c as the major fatty acid. The polar lipid profile of strain BS-W13T, containing phosphatidylcholine, phosphatidylglycerol, phosphatidylethanolamine, one unidentified aminolipid and one unidentified lipid as major components, was distinguishable from those of some phylogenetically related taxa. The DNA G+C content of strain BS-W13T was 58.1 mol%. The phylogenetic data and differential chemotaxonomic and other phenotypic properties revealed that strain BS-W13T constitutes a novel genus and species within family Rhodobacteraceae of the class Alphaproteobacteria , for which the name Pseudoseohaeicola caenipelagi gen. nov., sp. nov. is proposed. The type strain is BS-W13T ( = KCTC 42349T = CECT 8724T).

Litorilinea aerophila gen. nov., sp. nov., an aerobic member of the class Caldilineae , phylum Chloroflexi , isolated from an intertidal hot spring

2013 ◽  
Vol 63 (Pt_3) ◽  
pp. 1149-1154 ◽  
Author(s):  
Varsha Kale ◽  
Snædís H. Björnsdóttir ◽  
Ólafur H. Friðjónsson ◽  
Sólveig K. Pétursdóttir ◽  
Sesselja Ómarsdóttir ◽  
...  
Keyword(s):  
Type Species ◽  
Hot Spring ◽  
Sequence Similarity ◽  
Rrna Gene ◽  
Content Type ◽  
Phenotypic Differences ◽  
Link Type ◽  
Fermentative Growth ◽  
Distinct Lineage ◽  
Ph Range

A thermophilic, aerobic, Gram-stain-negative, filamentous bacterium, strain PRI-4131T, was isolated from an intertidal hot spring in Isafjardardjup, NW Iceland. The strain grew chemo-organotrophically on various carbohydrates. The temperature range for growth was 40–65 °C (optimum 55 °C), the pH range was pH 6.5–9.0 (optimum pH 7.0) and the NaCl range was 0–3 % (w/v) (optimum 0.5 %). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain PRI-4131T represented a distinct lineage within the class Caldilineae of the phylum Chloroflexi. The highest levels of sequence similarity, about 91 %, were with Caldilinea aerophila STL-6-O1T and Caldilinea tarbellica D1-25-10-4T. Fermentative growth was not observed for strain PRI-4131T, which, in addition to other characteristics, distinguished it from the two Caldilinea species. Owing to both phylogenetic and phenotypic differences from the described members of the class Caldilineae , we propose to accommodate strain PRI-4131T in a novel species in a new genus, Litorilinea aerophila gen. nov., sp. nov. The type strain of Litorilinea aerophila is PRI-4131T ( = DSM 25763T  = ATCC BAA-2444T).

Catenulispora graminis sp. nov., a rhizobacterium from bamboo (Phyllostachys nigro var. henonis) rhizosphere soil

2012 ◽  
Vol 62 (Pt_11) ◽  
pp. 2589-2592 ◽  
Author(s):  
Hyo-Jin Lee ◽  
Song-Ih Han ◽  
Kyung-Sook Whang
Keyword(s):  
Type Species ◽  
Rhizosphere Soil ◽  
Major Fatty Acid ◽  
Rrna Gene ◽  
Phenotypic Data ◽  
Content Type ◽  
Dna Relatedness ◽  
Link Type ◽  
Isoprenoid Quinones ◽  
Type Strains

A novel actinobacterium, designated strain BR-34T, was isolated from rhizosphere soil of bamboo (Phyllostachys nigro var. henonis) sampled in Damyang, Korea. The strain was found to have morphological and chemotaxonomic characteristics typical of the genus Catenulispora . The strain contained iso-C16 : 0 as the major fatty acid and MK-9(H4), MK-9(H6) and MK-9(H8) as major isoprenoid quinones. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain BR-34T formed a cluster separate from members of the genus Catenulispora and was related most closely to Catenulispora acidiphila ID139908T (97.4 % similarity), Catenulispora rubra Aac-30T (97.3 %), Catenulispora yoronensis TT N02-20T (97.3 %) and Catenulispora subtropica TT 99-48T (97 %). However, the level of DNA–DNA relatedness between strain BR-34T and C. acidiphila ID139908T was only 45.32 %. Based on DNA–DNA relatedness, morphological and phenotypic data, strain BR-34T could be distinguished from the type strains of phylogenetically related species. It is therefore considered to represent a novel species of the genus Catenulispora , for which the name Catenulispora graminis sp. nov. is proposed. The type strain is BR-34T ( = KACC 15070T = NBRC 107755T).

Flavobacterium akiainvivens sp. nov., from decaying wood of Wikstroemia oahuensis, Hawai‘i, and emended description of the genus Flavobacterium

2013 ◽  
Vol 63 (Pt_9) ◽  
pp. 3280-3286 ◽  
Author(s):  
Iris Kuo ◽  
Jimmy Saw ◽  
Durrell D. Kapan ◽  
Stephanie Christensen ◽  
Kenneth Y. Kaneshiro ◽  
...  
Keyword(s):  
Type Species ◽  
Original Description ◽  
Gliding Motility ◽  
Rrna Gene ◽  
Content Type ◽  
Phenotypic Differences ◽  
Whole Cells ◽  
Link Type ◽  
Decaying Wood ◽  
Type Strains

Strain IK-1T was isolated from decaying tissues of the shrub Wikstroemia oahuensis collected on O‘ahu, Hawai‘i. Cells were rods that stained Gram-negative. Gliding motility was not observed. The strain was oxidase-negative and catalase-positive. Zeaxanthin was the major carotenoid. Flexirubin-type pigments were not detected. The most abundant fatty acids in whole cells of IK-1T grown on R2A were iso-C15 : 0 and one or both of C16 : 1ω7c and C16 : 1ω6c. Based on comparisons of the nucleotide sequence of the 16S rRNA gene, the closest neighbouring type strains were Flavobacterium rivuli WB 3.3-2T and Flavobacterium subsaxonicum WB 4.1-42T, with which IK-1T shares 93.84 and 93.67 % identity, respectively. The G+C content of the genomic DNA was 44.2 mol%. On the basis of distance from its nearest phylogenetic neighbours and phenotypic differences, the species Flavobacterium akiainvivens sp. nov. is proposed to accommodate strain IK-1T ( = ATCC BAA-2412T = CIP 110358T) as the type strain. The description of the genus Flavobacterium is emended to reflect the DNA G+C contents of Flavobacterium akiainvivens IK-1T and other species of the genus Flavobacterium described since the original description of the genus.

scholarly journals Nesterenkonia alkaliphila sp. nov., an alkaliphilic, halotolerant actinobacteria isolated from the western Pacific Ocean

2015 ◽  
Vol 65 (Pt_2) ◽  
pp. 516-521 ◽  
Author(s):  
Gaiyun Zhang ◽  
Yubian Zhang ◽  
Xijie Yin ◽  
Shuang Wang
Keyword(s):  
Pacific Ocean ◽  
Type Species ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Western Pacific ◽  
Rrna Gene ◽  
Western Pacific Ocean ◽  
Content Type ◽  
Link Type ◽  
The Western Pacific

A Gram-staining-positive, aerobic, motile and non-spore-forming actinobacteria, designated strain F10T, was isolated from a deep-sea sediment of the western Pacific Ocean. Phylogenetic and phenotypic properties of the organism supported that it belonged to the genus Nesterenkonia . Strain F10T shared highest 16S rRNA gene sequence similarity of 96.8 % with Nesterenkonia aethiopica DSM 17733T, followed by Nesterenkonia xinjiangensis YIM 70097T (96.7 %) and Nesterenkonia alba CAAS 252T (96.6 %). The organism grew at 4–50 °C, at pH 7.0–12.0 and in the presence of 0–12 % (w/v) NaCl, with optimal growth occurring at 40 °C, at pH 9.0 and in the presence of 1 % (w/v) NaCl. The peptidoglycan type was A4(alpha), l-Lys–Gly–l-Glu. The polar lipid profile of strain F10T consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, two unknown glycolipids and two unknown lipids. The isolate contained MK-9 (92 %) and MK-8 (5.8 %) as the major components of the menaquinone system, and anteiso-C17 : 0 (50.9 %) and anteiso-C15 : 0 (29.8 %) as the predominant fatty acids. The G+C content of the genomic DNA of strain F10T was 66.2 mol%. Based on phenotypic, genotypic and phylogenetic analyses, strain F10T represents a novel species of the genus Nesterenkonia for which the name Nesterenkonia alkaliphila sp. nov. is proposed. The type strain is F10T ( = LMG 28112T = CGMCC 1.12781T = JCM 19766T = MCCC 1A09946T).

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