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International Journal of Systematic and... May 2020Two Gram-stain-negative, short rod-shaped and non-flagellated strains, designated 17-4A and L52-1-41, were isolated from the surface seawater of the Indian Ocean and...
Two Gram-stain-negative, short rod-shaped and non-flagellated strains, designated 17-4A and L52-1-41, were isolated from the surface seawater of the Indian Ocean and South China Sea, respectively. The 16S rRNA genes of the two strains shared sequence similarity of 99.45 %. Strain 17-4A shared the highest 16S rRNA gene similarity of 98.02 % with EBR-8-1, followed by BN9 (97.47 %), MJ07 (96.93 %), Ch07 (96.68 %), DCY25 (96.65 %), PB3-7B (96.63 %), 24 (96.34 %), 54Pin (96.28 %) and B201 (96.05 %). L52-1-41 shared the highest 16S rRNA gene similarity of 97.74 % with EBR-8-1, followed by BN9 (97.47 %), MJ07 (96.65 %), Ch07 (96.41 %), DCY25 (96.37 %), PB3-7B (96.35 %), B201 (96.28 %), and 24 (96.06 %). The results of phylogenetic analyses indicated that 17-4A and L52-1-41 formed a stable, distinct and highly supported lineage affiliated to the genus . The results of the digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) analyses indicated that they represented a single species. They featured similar genomic DNA G+C contents of 53.2-53.4 mol%. Activities of catalase and oxidase were negative for both strains. The fatty acids patterns of 17-4A and L52-1-41 were most similar, mostly comprised of C, Ccyclo, C, Cω9 and summed feature 8 (Cω7 and/or C ω6). The major polar lipids of the two strains were phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and unidentified aminolipids. The respiratory quinone of the two strains was Q-8. Hence, on the basis of the phenotypic, chemotaxonomic and genotypic data presented in this study, we proposed the classification of both strains as representatives of a novel species named sp. nov., with the type strain 17-4A (=MCCC 1A12670=KCTC 62121=NBRC 113794), and another strain L52-1-41 (=MCCC 1A05046=KCTC 52313).
Topics: Alcaligenaceae; Bacterial Typing Techniques; Base Composition; China; DNA, Bacterial; Fatty Acids; Indian Ocean; Nucleic Acid Hybridization; Phospholipids; Phylogeny; RNA, Ribosomal, 16S; Seawater; Sequence Analysis, DNA; Ubiquinone
PubMed: 32369004
DOI: 10.1099/ijsem.0.004202 -
Journal of Clinical Microbiology Apr 2024Whole-genome sequencing (WGS) of microbial pathogens recovered from patients with infectious disease facilitates high-resolution strain characterization and molecular...
Whole-genome sequencing (WGS) of microbial pathogens recovered from patients with infectious disease facilitates high-resolution strain characterization and molecular epidemiology. However, increasing reliance on culture-independent methods to diagnose infectious diseases has resulted in few isolates available for WGS. Here, we report a novel culture-independent approach to genome characterization of , the causative agent of pertussis and a paradigm for insufficient genomic surveillance due to limited culture of clinical isolates. Sequencing libraries constructed directly from residual pertussis-positive diagnostic nasopharyngeal specimens were hybridized with biotinylated RNA "baits" targeting fragments within complex mixtures that contained high concentrations of host and microbial background DNA. Recovery of genome sequence data was evaluated with mock and pooled negative clinical specimens spiked with reducing concentrations of either purified DNA or inactivated cells. Targeted enrichment increased the yield of sequencing reads up to 90% while simultaneously decreasing host reads to less than 10%. Filtered sequencing reads provided sufficient genome coverage to perform characterization via whole-genome single nucleotide polymorphisms and whole-genome multilocus sequencing typing. Moreover, these data were concordant with sequenced isolates recovered from the same specimens such that phylogenetic reconstructions from either consistently clustered the same putatively linked cases. The optimized protocol is suitable for nasopharyngeal specimens with diagnostic IS Ct < 35 and >10 ng DNA. Routine implementation of these methods could strengthen surveillance and study of pertussis resurgence by capturing additional cases with genomic characterization.
Topics: Humans; Bordetella pertussis; Whooping Cough; Phylogeny; Genomics; DNA; Bordetella
PubMed: 38445858
DOI: 10.1128/jcm.01653-23 -
Nature Communications Dec 2021D-2-Hydroxyglutarate (D-2-HG) is a metabolite involved in many physiological metabolic processes. When D-2-HG is aberrantly accumulated due to mutations in isocitrate...
D-2-Hydroxyglutarate (D-2-HG) is a metabolite involved in many physiological metabolic processes. When D-2-HG is aberrantly accumulated due to mutations in isocitrate dehydrogenase or D-2-HG dehydrogenase, it functions in a pro-oncogenic manner and is thus considered a therapeutic target and biomarker in many cancers. In this study, DhdR from Achromobacter denitrificans NBRC 15125 is identified as an allosteric transcriptional factor that negatively regulates D-2-HG dehydrogenase expression and responds to the presence of D-2-HG. Based on the allosteric effect of DhdR, a D-2-HG biosensor is developed by combining DhdR with amplified luminescent proximity homogeneous assay (AlphaScreen) technology. The biosensor is able to detect D-2-HG in serum, urine, and cell culture medium with high specificity and sensitivity. Additionally, this biosensor is used to identify the role of D-2-HG metabolism in lipopolysaccharide biosynthesis of Pseudomonas aeruginosa, demonstrating its broad usages.
Topics: Achromobacter denitrificans; Alcohol Oxidoreductases; Bacteria; Biosensing Techniques; Gene Expression Regulation; Glutarates; HEK293 Cells; Humans; Isocitrate Dehydrogenase; Metabolic Networks and Pathways; Mutation; Neoplasms; Transcription Factors
PubMed: 34876568
DOI: 10.1038/s41467-021-27357-7 -
Microbial Genomics Jul 2021species are increasingly being detected in cystic fibrosis (CF) patients, where they can establish chronic infections by adapting to the lower airway environment. To...
species are increasingly being detected in cystic fibrosis (CF) patients, where they can establish chronic infections by adapting to the lower airway environment. To better understand the mechanisms contributing to a successful colonization by species, we sequenced the whole genome of 54 isolates from 26 patients with occasional and early/late chronic lung infection. We performed a phylogenetic analysis and compared virulence and resistance genes, genetic variants and mutations, and hypermutability mechanisms between chronic and occasional isolates. We identified five species as well as two non-affiliated genogroups (NGs). Among them were the frequently isolated and four other species whose clinical importance is not yet clear: and . While and were isolated only from chronically infected patients and only from occasionally infected patients, the other species were found in both groups. Most of the occasional isolates lacked functional genes involved in invasiveness, chemotaxis, type 3 secretion system and anaerobic growth, whereas the great majority (>60%) of chronic isolates had these genomic features. Interestingly, almost all (=22/23) late chronic isolates lacked functional genes involved in lipopolysaccharide production. Regarding antibiotic resistance, we observed a species-specific distribution of genes, confirming what has been reported in the literature and additionally identifying in some isolates and observing no genes in or NGs. No significant difference in resistance genes was found between chronic and occasional isolates. The results of the mutator genes analysis showed that no occasional isolate had hypermutator characteristics, while 60% of early chronic (<1 year from first colonization) and 78% of late chronic (>1 year from first colonization) isolates were classified as hypermutators. Although all and NG isolates presented two different genes, these seem to have a complementary rather than compensatory function. In conclusion, our results show that species can exhibit different adaptive mechanisms and some of these mechanisms might be more useful than others in establishing a chronic infection in CF patients, highlighting their importance for the clinical setting and the need for further studies on the less clinically characterized species.
Topics: Achromobacter; Cystic Fibrosis; Drug Resistance, Bacterial; Genome, Bacterial; Gram-Negative Bacterial Infections; Humans; Lung; MutS Proteins; Persistent Infection; Virulence Factors; Whole Genome Sequencing; beta-Lactamases
PubMed: 34292148
DOI: 10.1099/mgen.0.000606 -
Pest Management Science Apr 2021Drosophila suzukii (Matsumura, 1931) (spotted wing drosophila), an invasive species, has recently become a significant global pest of soft-skinned fruits such as...
BACKGROUND
Drosophila suzukii (Matsumura, 1931) (spotted wing drosophila), an invasive species, has recently become a significant global pest of soft-skinned fruits such as berries. Unlike other Drosophila species, female D. suzukii have evolved a specialized sharp, serrated ovipositor that pierces and penetrates ripe and ripening fruits, causing them to lose commercial value and preventing their sale. A first step for the development of biological control agents for pest management may be achieved through the identification of microbes infectious for D. suzukii in the wild.
RESULTS
We first determined that D. suzukii is susceptible to chemicals commonly used to rear Drosophilids in the laboratory and established a diet able to sustain healthy D. suzukii growth. Using this diet, we demonstrated that of 25 species of culturable bacteria and fungi isolated from field-collected D. suzukii, eight microbes decreased host survival when injected. Three of the eight bacteria (Alcaligenes faecalis, Achromobacter spanius and Serratia marcescens) were acutely pathogenic to both D. suzukii and Drosophila melanogaster adults by injection. Feeding of these bacteria resulted in susceptibility only in larvae.
CONCLUSION
We successfully identified multiple microbes from field-collected D. suzukii that are pathogenic to both larvae and adults through different routes of infection, some of which could be candidates for biocontrol of this species. © 2020 Society of Chemical Industry.
Topics: Achromobacter; Animals; Drosophila; Drosophila melanogaster; Female; Fruit
PubMed: 33342014
DOI: 10.1002/ps.6235 -
Epidemiologie, Mikrobiologie,... 2018To test clinical isolates of Bordetella pertussis from the National Reference Laboratory for Pertussis and Diphtheria for susceptibility to commonly available...
THE AIM OF STUDY
To test clinical isolates of Bordetella pertussis from the National Reference Laboratory for Pertussis and Diphtheria for susceptibility to commonly available disinfectants. Another aim was to determine the concentration and exposure time for each chemical under real conditions of use and possibly to detect the emergence of resistance to disinfectants among 34 strains of B. pertussis referred to the National Reference Laboratory for Pertussis and Diphtheria in 2014 and 2015.
MATERIAL AND METHODS
A total of 34 clinical isolates of Bordetella pertussis were tested for susceptibility to chemical disinfectants by three different methods. The microsuspension method was used for the primary screening, and the tests were carried out without protein contamination. Further testing was conducted in accordance with standard EN 14885, where the test procedure consists of several steps. Step 1 involves quantitative suspension methods (Phase 2, Step 1), and step 2 uses methods designed for practice (Phase 2, Step 2). The quantitative suspension method modified according to EN 13727+A2 was used in step 1 to confirm bactericidal activity of the test products under the dirty conditions. In step 2, clinical isolates were tested using a quantitative carrier test method under the dirty conditions modified according to EN 14561. Based on this standard, the real conditions of product use are simulated. Four disinfectants differing in composition and intended use were tested.
RESULTS
Disinfectant No. 1 showed bactericidal activity at a concentration of 0.5% after 2 min of exposure in the case of immersion or at a concentration of 5% after 2 min of exposure when treated by wiping. Disinfectant No. 2 was active at a concentration of 0.1% after 2 min of exposure or at a concentration of 1% after 2 min of exposure, respectively. Disinfectant No. 3 did not show bactericidal activity even at a concentration of 100% after 5 min of exposure. Disinfectant No. 4 showed bactericidal activity at a concentration of 10% after 5 min of exposure or at a concentration of 30% after 2 min of exposure.
CONCLUSIONS
None of the strains tested was resistant. Using the methods that simulate the real conditions of use of disinfectants Nos. 1 and 2, it was possible to determine the concentration and exposure time needed to achieve disinfection of surfaces under the dirty conditions. Disinfectants Nos. 3 and 4 are not primarily intended for the treatment of surfaces but for the treatment of the skin and mucous membranes. The results obtained with the latter two products are interesting but inconclusive as the real conditions of their use were not simulated accurately. KEYWORDS Bordetella pertussis - susceptibility - disinfectants.
Topics: Bordetella pertussis; Disinfectants; Microbial Sensitivity Tests
PubMed: 30602279
DOI: No ID Found -
International Journal of Systematic and... Apr 2022A novel pale white-pigmented bacterial strain designated YC-7-48 was isolated from activated sludge in China. Cells of the strain, which grew at 15-37 °C (optimum at 30...
A novel pale white-pigmented bacterial strain designated YC-7-48 was isolated from activated sludge in China. Cells of the strain, which grew at 15-37 °C (optimum at 30 °C) and pH 6.0-9.0 (optimum at 7.0), were Gram-stain-negative, rod-shaped and motile. Strain YC-7-48 had 97.4-97.1% 16S rRNA gene sequence similarity to type strains of eight species in the genera , , , and of the family . Phylogenetic analysis based on 16S rRNA gene sequencing placed the strain on a separate branch in the genus and showed that it exhibited 97.4, 97.3 and 96.6% similarity to EBR-8-1, BN9 and 17-4A, respectively. The genome size of strain YC-7-48 was 3202438 bp, with 54.3 mol% G+C content. According to the genome analysis, YC-7-48 encodes several heavy metal resistance proteins and enzymes related to the metabolism of nicotine and aromatic compounds. The results of digital DNA-DNA hybridization and average nucleotide identity analyses based on whole genome sequences between strain YC-7-48 and the closely related strains indicated that the strain represented a new species of the genus . The chemotaxonomic results identified Q-8 as the predominant respiratory quinone, phosphatidylethanolamine, phosphatidylmonomethylethanolamine, diphosphatidylglycerol and two unidentified aminolipids as the major polar lipids, and C (27.4 %), C cyclo (22.0 %), C (11.7 %) and C cyclo 8 (9.5 %) as the major fatty acids. Thus, based on morphological, chemotaxonomic and phylogenetic characterization and genomic data, we proposed that the isolate is a representative of a novel species named sp. nov., with the type strain YC-7-48 (=CGMCC 1.17466=KACC 21349).
Topics: Alcaligenaceae; Bacterial Typing Techniques; Base Composition; DNA, Bacterial; Fatty Acids; Phospholipids; Phylogeny; RNA, Ribosomal, 16S; Sequence Analysis, DNA; Sewage
PubMed: 35451948
DOI: 10.1099/ijsem.0.005323 -
Journal of Clinical Microbiology Feb 2019Molecular methods offer superior sensitivity and specificity and reduce testing turnaround time from days to hours for detection of and In this study, we evaluated the...
Molecular methods offer superior sensitivity and specificity and reduce testing turnaround time from days to hours for detection of and In this study, we evaluated the performance of the automated PCR-based Aries Assay, which detects both and directly from nasopharyngeal swab specimens. The limits of detection (LoDs) were 1,800 CFU·ml for and 213 CFU·ml for The assay detected 16/18 unique / strains. Of 71 potentially cross-reacting organisms, 5 generated false positives in 1/6 replicates; none of 6 additional spp. were erroneously detected. Specimens were stable at 20 to 25°C for at least 10 h, at 4 to 8°C for 10 days, and at temperatures not exceeding -70°C for 6 months. Of 1,052 nasopharyngeal specimens from patients with suspected pertussis, 3.0% ( = 32) were positive and 0.2% ( = 2) were positive. Combining these data with Aries Assay data from 57 nasopharyngeal samples with previously confirmed or data and with data from 50 contrived samples, the proportions of positive and negative agreement of the respective Aries assays with the reference assays were 97.1% and 99.0% for and 100% and 99.7% for The Aries Assay provides accurate detection and distinction of and infections within 2 h. (This study has been registered at ClinicalTrials.gov under registration no. NCT02862262.).
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Automation, Laboratory; Bordetella Infections; Bordetella parapertussis; Bordetella pertussis; Child; Child, Preschool; Female; Humans; Infant; Infant, Newborn; Male; Middle Aged; Molecular Diagnostic Techniques; Nasopharynx; Polymerase Chain Reaction; Prospective Studies; Sensitivity and Specificity; Time Factors; Young Adult
PubMed: 30518543
DOI: 10.1128/JCM.01471-18 -
Epidemiology and Infection Aug 2020Bordetella bronchiseptica is a potential zoonotic pathogen, which mainly causes respiratory diseases in humans and a variety of animal species. B. bronchiseptica is one...
Bordetella bronchiseptica is a potential zoonotic pathogen, which mainly causes respiratory diseases in humans and a variety of animal species. B. bronchiseptica is one of the important pathogens isolated from rabbits in Fujian Province. However, the knowledge of the epidemiology and characteristics of the B. bronchiseptica in rabbits in Fujian Province is largely unknown. In this study, 219 B. bronchiseptica isolates recovered from lung samples of dead rabbits with respiratory diseases in Fujian Province were characterised by multi-locus sequencing typing, screening virulence genes and testing antimicrobial susceptibility. The results showed that the 219 isolates were typed into 11 sequence types (STs) including five known STs (ST6, ST10, ST12, ST14 and ST33) and six new STs (ST88, ST89, ST90, ST91, ST92 and ST93) and the ST33 (30.14%, 66/219), ST14 (26.94%, 59/219) and ST12 (16.44%, 36/219) were the three most prevalent STs. Surprisingly, all the 219 isolates carried the five virulence genes (fhaB, prn, cyaA, dnt and bteA) in the polymerase chain reaction screening. Moreover, the isolates were resistant to cefixime, ceftizoxime, cefatriaxone and ampicillin at rates of 33.33%, 31.05%, 11.87% and 3.20%, respectively. This study showed the genetic diversity of B. bronchiseptica in rabbits in Fujian Province, and the colonisation of the human-associated ST12 strain in rabbits in Fujian Province. The results might be useful for monitoring the epidemic strains, developing preventive methods and preventing the transmission of epidemic strains from rabbits to humans.
Topics: Animals; Anti-Bacterial Agents; Bordetella Infections; Bordetella bronchiseptica; China; Drug Resistance, Bacterial; Genetic Variation; Phylogeny; Rabbits; Respiratory Tract Diseases
PubMed: 32829720
DOI: 10.1017/S0950268820001879 -
MBio Aug 2022Copper is essential to most living beings but also highly toxic and as such is an important player at the host-pathogen interface. Bacteria have thus developed...
Copper is essential to most living beings but also highly toxic and as such is an important player at the host-pathogen interface. Bacteria have thus developed homeostatic mechanisms to tightly control its intracellular concentration. Known Cu export and import systems are under transcriptional control, whereas posttranscriptional regulatory mechanisms are yet to be characterized. We identified a three-gene operon, , downregulated by copper and notably encoding a TonB-dependent transporter in Bordetella pertussis. We show here that the protein encoded by the first gene, which is a member of the DUF2946 protein family, represents a new type of upstream Open Reading Frame (uORF) involved in posttranscriptional regulation of the downstream genes. In the absence of copper, the entire operon is transcribed and translated. Perception of copper by the nascent -coded protein via its conserved CXXC motif triggers Rho-dependent transcription termination between the first and second genes by relieving translation arrest on a conserved C-terminal RAPP motif. Homologs of are widespread in bacterial genomes, where they head operons predicted to participate in copper homeostasis. This work has thus unveiled a new mode of genetic regulation by a transition metal and identified a regulatory function for a member of an uncharacterized family of bacterial proteins that we have named CruR, for copper-responsive upstream regulator. Copper is a transition metal necessary for living beings but also extremely toxic. Bacteria thus tightly control its homeostasis with transcriptional regulators. In this work, we have identified in the whooping cough agent Bordetella pertussis a new control mechanism mediated by a small protein called CruR, for copper-responsive upstream regulator. While being translated by the ribosome CruR is able to perceive intracellular copper, which shuts down the transcription of downstream genes of the same operon, coding for a copper uptake system. This mechanism limits the import of copper in conditions where it is abundant for the bacterium. This is the first report of "posttranscriptional regulation" in response to copper. Homologs of CruR genes head many operons harboring copper-related genes in various bacteria, and therefore the regulatory function unveiled here is likely a general property of this new protein family.
Topics: Bacterial Proteins; Bordetella pertussis; Copper; Gene Expression Regulation; Gene Expression Regulation, Bacterial; Open Reading Frames; Operon; Ribosomes
PubMed: 35862763
DOI: 10.1128/mbio.00912-22