scholarly journals Adaptive laboratory evolution of Pseudomonas putida and Corynebacterium glutamicum to enhance anthranilate tolerance

Microbiology ◽  
2020 ◽  
Vol 166 (11) ◽  
pp. 1025-1037 ◽  
Author(s):  
Jannis Kuepper ◽  
Maike Otto ◽  
Jasmin Dickler ◽  
Swantje Behnken ◽  
Jorgen Magnus ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Corynebacterium Glutamicum ◽  
Type Species ◽  
Product Inhibition ◽  
Adaptive Laboratory Evolution ◽  
Laboratory Evolution ◽  
Content Type ◽  
Link Type ◽  
Important Basis

Microbial bioproduction of the aromatic acid anthranilate (ortho-aminobenzoate) has the potential to replace its current, environmentally demanding production process. The host organism employed for such a process needs to fulfil certain demands to achieve industrially relevant product levels. As anthranilate is toxic for microorganisms, the use of particularly robust production hosts can overcome issues from product inhibition. The microorganisms Corynebacterium glutamicum and Pseudomonas putida are known for high tolerance towards a variety of chemicals and could serve as promising platform strains. In this study, the resistance of both wild-type strains towards anthranilate was assessed. To further enhance their native tolerance, adaptive laboratory evolution (ALE) was applied. Sequential batch fermentation processes were developed, adapted to the cultivation demands for C. glutamicum and P. putida, to enable long-term cultivation in the presence of anthranilate. Isolation and analysis of single mutants revealed phenotypes with improved growth behaviour in the presence of anthranilate for both strains. The characterization and improvement of both potential hosts provide an important basis for further process optimization and will aid the establishment of an industrially competitive method for microbial synthesis of anthranilate.

Aquabacterium soli sp. nov., a novel bacterium isolated from soil under the long-term application of bifenthrin

2021 ◽  
Vol 71 (9) ◽  
Author(s):  
Lina Sun ◽  
Wei Chen ◽  
Kaihua Huang ◽  
Weiguang Lyu ◽  
Xinhua Gao
Keyword(s):  
Type Species ◽  
Sequence Similarity ◽  
Major Fatty Acid ◽  
Rrna Gene ◽  
Aerobic Bacterium ◽  
Content Type ◽  
Link Type ◽  
Novel Bacterium ◽  
Pr China

Strain SJQ9T, an aerobic bacterium isolated from a soil sample collected in Shanghai, PR China, was characterized using a polyphasic approach. It grew optimally at pH 7.0, 30–35 °C and in the presence of 1 % (w/v) NaCl. A comparative analysis of 16S rRNA gene sequences showed that strain SJQ9T fell within the genus Aquabacterium . The closest phylogenetic relatives of strain SJQ9T were Aquabacterium citratiphilum DSM 11900T (98.6 % sequence similarity) and Aquabacterium commune DSM 11901T (96.4 %). Cells of the strain were Gram-stain-negative, motile, non-spore-forming, rod-shaped and positive for oxidase activity and negative for catalase. The chemotaxonomic properties of strain SJQ9T were consistent with those of the genus Aquabacterium : the major fatty acid was summed feature 3 (C16 : 1  ω6c and/or C16 : 1  ω7c). The isoprenoid quinone was Q-8. The major polar lipids were phosphatidylethanolamine, phosphatidylcholine, phosphatidylglycerol and diphosphatidylglycerol. The DNA G+C content was 65.7 mol%. Strain SH9T exhibited a DNA–DNA relatedness level of 34±2 % with A. citratiphilum DSM 11900T and 28±3 % with A. commune DSM 11901T. Based on the obtained data, strain SJQ9T represents a novel species of the genus Aquabacterium , for which the name Aquabacterium soli sp. nov. is proposed. The type strain is SJQ9T (=JCM 33106T=CCTCC AB 2018284T).

Olivibacter jilunii sp. nov., isolated from DDT-contaminated soil

2013 ◽  
Vol 63 (Pt_3) ◽  
pp. 1083-1088 ◽  
Author(s):  
Kai Chen ◽  
Shu-Kun Tang ◽  
Guang-Li Wang ◽  
Guo-Xing Nie ◽  
Qin-Fen Li ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Contaminated Soil ◽  
Type Species ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
The Family ◽  
The 16S Rrna Gene

Bacterial strain 14-2AT, isolated from a long-term DDT-contaminated soil in China, was characterized by using a polyphasic approach to clarify its taxonomic position. Strain 14-2AT was found to be Gram-negative, aerobic, non-spore-forming, non-motile, non-flagellated and rod-shaped. The new isolate was able to grow at 4–42 °C, pH 6.0–9.0 and with 0–5 % NaCl. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the isolate belongs to the family Sphingobacteriaceae . The 16S rRNA gene sequence of strain 14-2AT showed the highest similarity with Olivibacter oleidegradans TBF2/20.2T (99.4 %), followed by Pseudosphingobacterium domesticum DC-186T (93.8 %), Olivibacter ginsengisoli Gsoil 060T (93.6 %), Olivibacter terrae Jip13T (93.1 %), Olivibacter soli Gsoil 034T (92.8 %) and Olivibacter sitiensis AW-6T (89.6 %). The DNA–DNA hybridization value between strains 14-2AT and O. oleidegradans TBF2/20.2T was 34.45±2.11 %. Strain 14-2AT contained phosphatidylethanolamine, phosphatidylmonomethylethanolamine, aminophospholipid and phosphatidylinositol mannoside as the major polar lipids. The DNA G+C content was 41.2 mol%. MK-7 is the major isoprenoid quinone. Summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c), iso-C15 : 0 and iso-C17 : 0 3-OH are the major fatty acids. The phenotypic and chemotaxonomic data confirmed the affiliation of strain 14-2AT to the genus Olivibacter . On the basis of the phylogenetic and phenotypic characteristics, and chemotaxonomic data, strain 14-2AT is considered to represent a novel species of the genus Olivibacter , for which the name Olivibacter jilunii sp. nov. is proposed; the type strain is 14-2AT ( = KCTC 23098T = CCTCC AB 2010105T).

scholarly journals Evaluation of whole-genome sequencing-based subtyping methods for the surveillance of Shigella spp. and the confounding effect of mobile genetic elements in long-term outbreaks

2021 ◽  
Vol 7 (11) ◽  
Author(s):  
Isabelle Bernaquez ◽  
Christiane Gaudreau ◽  
Pierre A. Pilon ◽  
Sadjia Bekal
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Type Species ◽  
Core Genome ◽  
Outbreak Detection ◽  
Whole Genome ◽  
Content Type ◽  
Link Type ◽  
Interpretation Criteria

Many public health laboratories across the world have implemented whole-genome sequencing (WGS) for the surveillance and outbreak detection of foodborne pathogens. PulseNet-affiliated laboratories have determined that most single-strain foodborne outbreaks are contained within 0–10 multi-locus sequence typing (MLST)-based allele differences and/or core genome single-nucleotide variants (SNVs). In addition to being a food- and travel-associated outbreak pathogen, most Shigella spp. cases occur through continuous person-to-person transmission, predominantly involving men who have sex with men (MSM), leading to long-term and recurrent outbreaks. Continuous transmission patterns coupled to genetic evolution under antibiotic treatment pressure require an assessment of existing WGS-based subtyping methods and interpretation criteria for cluster inclusion/exclusion. An evaluation of 4 WGS-based subtyping methods [SNVPhyl, coreMLST, core genome MLST (cgMLST) and whole-genome MLST (wgMLST)] was performed on 9 foodborne-, travel- and MSM-related retrospective outbreaks from a collection of 91 Shigella flexneri and 232  Shigella sonnei isolates to determine the methods’ epidemiological concordance, discriminatory power, robustness and ability to generate stable interpretation criteria. The discriminatory powers were ranked as follows: coreMLST<SNVPhyl<cgMLST<wgMLST (range: 0.970–1.000). The genetic differences observed for non-MSM-related Shigella spp. outbreaks respect the standard 0–10 allele/SNV guideline; however, mobile genetic element (MGE)-encoded loci caused inflated genetic variation and discrepant phylogenies for prolonged MSM-related S. sonnei outbreaks via wgMLST. The S. sonnei correlation coefficients of wgMLST were also the lowest at 0.680, 0.703 and 0.712 for SNVPhyl, coreMLST and cgMLST, respectively. Plasmid maintenance, mobilization and conjugation-associated genes were found to be the main source of genetic distance inflation in addition to prophage-related genes. Duplicated alleles arising from the repeated nature of IS elements were also responsible for many false cg/wgMLST differences. The coreMLST approach was shown to be the most robust, followed by SNVPhyl and wgMLST for inter-laboratory comparability. Our results highlight the need for validating species-specific subtyping methods based on microbial genome plasticity and outbreak dynamics in addition to the importance of filtering confounding MGEs for cluster detection.

scholarly journals Corynebacterium suranareeae sp. nov., a glutamate producing bacterium isolated from soil and its complete genome-based analysis

2020 ◽  
Vol 70 (3) ◽  
pp. 1903-1911 ◽  
Author(s):  
Nawarat Nantapong ◽  
Minenosuke Matsutani ◽  
Pawina Kanchanasin ◽  
Naoya Kataoka ◽  
Pawantree Paisrisan ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Corynebacterium Glutamicum ◽  
Complete Genome ◽  
Type Species ◽  
Gene Sequence ◽  
Rrna Gene ◽  
Rrna Gene Sequence ◽  
Content Type ◽  
Link Type

Strain N24T was isolated from soil contaminated with starling’s feces collected from Roi-Et province, Thailand. Cells of N24T were Gram-stain-positive rods, aerobic and non-spore-forming. N24T was positive for catalase, urease, citrate utilization, nitrate reduction and Methyl Red (MR) test but negative for oxidase, casein, gelatin liquefaction, tyrosine, Voges–Proskauer (VP) reaction and starch hydrolysis. Meso-diaminopimelic acid, rhamnose, ribose, arabinose and galactose were detected in its whole-cell hydrolysates. The results of the 16S rRNA gene sequence analysis indicated that N24T represented a member of the genus Corynebacterium . N24T was closely related to Corynebacterium glutamicum ATCC 13032T, with 99.0 % 16S rRNA gene sequence similarity. According to results obtained using in silico DNA–DNA hybridization approaches, N24T showed highest DNA–DNA relatedness (27.6 %) and average nucleotide identity (84.1 %) to Corynebacterium glutamicum ATCC 13032T. The DNA G+C content of N24T was 51.8 mol% (genome based). The major cellular fatty acids of N24T were C16 : 0, and C18 : 1ω9c. N24T had the nine isoprenes unit, MK-9(H2) as the predominant menaquinone. The predominant polar lipids were phosphatidylglycerol, phosphatidylinositol and diphosphatidylglycerol. Mycolic acids were also present. According to the complete genome sequence data, strain N24T and C. glutamicum ATCC 13032T are close phylogenetic neighbours, but have different genome characteristics. On the basis of the results of the genotypic and genomic studies and phenotypic characteristics including chemotaxonomy, strain N24T should be classified as representing a novel species of the genus Corynebacterium , for which the name Corynebacterium suranareeae sp. nov. is proposed. The type strain is N24T (TBRC 5845T=NBRC 113465T).

scholarly journals Construction and evaluation of a bioluminescent Pseudomonas aeruginosa reporter for use in preservative efficacy testing

Microbiology ◽  
2021 ◽  
Vol 167 (8) ◽  
Author(s):  
Laura Rushton ◽  
Denise Donoghue ◽  
Matthew Bull ◽  
Peter Jay ◽  
Eshwar Mahenthiralingam
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type Species ◽  
Random Mutagenesis ◽  
Light Output ◽  
Rapid Decline ◽  
Content Type ◽  
Link Type ◽  
Term Evaluation ◽  
Positive Attribute

Preservative efficacy testing (PET) is a fundamental practice in industrial microbiology used to ensure product shelf-life and quality. To improve on current growth-based PET, bioluminescence was evaluated as a real-time bacterial viability indicator using Pseudomonas aeruginosa . Random mutagenesis of an industrial P. aeruginosa strain with a promoter-less luxCDABE mini-Tn5 was used to select a stable reporter (LUX12H5) with an un-altered growth and preservative susceptibility phenotype. Bioluminescence and viability were measured with and without preservatives (isothiazolinones, phenoxyethanol, and dimethyl dimethylol hydantoin) and an antibiotic comparator (ciprofloxacin). In the absence of antimicrobials, a good correlation between bioluminescence and viability (r2=0.92) was established. However, metabolic inhibition by isothiazolinone preservatives caused a rapid decline in light output that did not correlate to a reduced viability. Conversely, after ciprofloxacin exposure, the decline in viability was greater than that of bioluminescence. A positive attribute of the bioluminescence was the early detection of metabolic recovery and re-growth of preservative injured bacteria. Overall, while initial bioluminescence read-outs were less suited to current PET requirements, it shows promise as an early, direct indicator of bacterial regrowth in the context of long-term evaluation of preservative efficacy.

scholarly journals Dynamics of extended-spectrum beta-lactamase-producing Enterobacterales colonization in long-term carriers following travel abroad

2021 ◽  
Vol 7 (7) ◽  
Author(s):  
Laurence Armand-Lefèvre ◽  
Emilie Rondinaud ◽  
Dimitri Desvillechabrol ◽  
Jimmy Mullaert ◽  
Olivier Clermont ◽  
...  
Keyword(s):  
Type Species ◽  
Beta Lactamase ◽  
Content Type ◽  
Extended Spectrum Beta Lactamase ◽  
E Coli ◽  
Link Type ◽  
Extended Spectrum ◽  
Travel Abroad

Travel to tropical regions is associated with high risk of acquiring extended-spectrum beta-lactamase-producing Enterobacterales (ESBL-E) that are typically cleared in less than 3 months following return. The conditions leading to persistent carriage that exceeds 3 months in some travellers require investigation. Whole-genome sequencing (Illumina MiSeq) was performed on the 82 ESBL-E isolates detected upon return and 1, 2, 3, 6 and 12 months later from the stools of 11 long-term (>3 months) ESBL-E carriers following travel abroad. One to five different ESBL Escherichia coli strains were detected per traveller upon return, and this diminished to one after 3 months. Long-term carriage was due to the presence of the same ESBL E. coli strain, for more than 3 months, in 9 out of 11 travellers, belonging to epidemic sequence type complexes (STc 10, 14, 38, 69, 131 and 648). The mean carriage duration of strains belonging to phylogroups B2/D/F, associated with extra-intestinal virulence, was higher than that for commensal-associated A/B1/E phylogroups (3.5 vs 0.5 months, P=0.021). Genes encoding iron capture systems (fyuA, irp), toxins (senB, sat), adhesins (flu, daaF, afa/nfaE, pap, ecpA) and colicin (cjrA) were more often present in persistent strains than in transient ones. Single-nucleotide polymorphism (SNP) analysis in persistent strains showed a maximum divergence of eight SNPs over 12 months without signs of adaptation. Genomic plasticity was observed during the follow-up with the loss or gain of mobile genetic elements such as plasmids, integrons and/or transposons that may contain resistance genes at different points in the follow-up. Long-term colonization of ESBL-E following travel is primarily due to the acquisition of E. coli strains belonging to epidemic clones and harbouring ‘virulence genes’, allowing good adaptation to the intestinal microbiota.

scholarly journals The putative phosphate transporter PitB (PP1373) is involved in tellurite uptake in Pseudomonas putida KT2440

Microbiology ◽  
2020 ◽  
Author(s):  
Rafael Montenegro ◽  
Sofía Vieto ◽  
Daniela Wicki-Emmenegger ◽  
Felipe Vásquez-Castro ◽  
Carolina Coronado-Ruiz ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Type Strain ◽  
Type Species ◽  
Wild Type Strain ◽  
Phosphate Transporter ◽  
Transport Systems ◽  
Wild Type ◽  
Pseudomonas Putida Kt2440 ◽  
Content Type ◽  
Link Type

Tellurium oxyanions are chemical species of great toxicity and their presence in the environment has increased because of mining industries and photovoltaic and electronic waste. Recovery strategies for this metalloid that are based on micro-organisms are of interest, but further studies of the transport systems and enzymes responsible for implementing tellurium transformations are required because many mechanisms remain unknown. Here, we investigated the involvement in tellurite uptake of the putative phosphate transporter PitB (PP1373) in soil bacterium Pseudomonas putida KT2440. For this purpose, through a method based on the CRISPR/Cas9 system, we generated a strain deficient in the pitB gene and characterized its phenotype on exposing it to varied concentrations of tellurite. Growth curves and transmission electronic microscopy experiments for the wild-type and ΔpitB strains showed that both were able to internalize tellurite into the cytoplasm and reduce the oxyanion to black nano-sized and rod-shaped tellurium particles, although the ΔpitB strain showed an increased resistance to the tellurite toxic effects. At a concentration of 100 μM tellurite, where the biomass formation of the wild-type strain decreased by half, we observed a greater ability of ΔpitB to reduce this oxyanion with respect to the wild-type strain (~38 vs ~16 %), which is related to the greater biomass production of ΔpitB and not to a greater consumption of tellurite per cell. The phenotype of the mutant was restored on over-expressing pitB in trans. In summary, our results indicate that PitB is one of several transporters responsible for tellurite uptake in P. putida KT2440.

scholarly journals Nocardioides soli sp. nov., a carbendazim-degrading bacterium isolated from soil under the long-term application of carbendazim

2014 ◽  
Vol 64 (Pt_6) ◽  
pp. 2047-2052 ◽  
Author(s):  
Li-Na Sun ◽  
Jun Zhang ◽  
Fen-Fen Gong ◽  
Xiang Wang ◽  
Gang Hu ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Type Strain ◽  
Type Species ◽  
Diaminopimelic Acid ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
Degrading Bacterium

The taxonomic status of a carbendazim-degrading strain, mbc-2T, isolated from soil under the long-term application of carbendazim in China was determined by means of a polyphasic study. The cells were Gram-stain-positive, motile and rod-shaped. Strain mbc-2T grew optimally at pH 7.0, 30–35 °C and in the presence of 1 % (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain mbc-2T fell within the genus Nocardioides , forming a coherent cluster with the type strain of Nocardioides hankookensis , with which it exhibited 16S rRNA gene sequence similarity values of 97.9 %. The chemotaxonomic properties of strain mbc-2T were consistent with those of the genus Nocardioides : the cell-wall peptidoglycan type was based on ll-2,6-diaminopimelic acid, the predominant menaquinone was MK-8 (H4) and the major fatty acid was iso-C16 : 0. The major polar lipids were phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, unknown phospholipids and an unknown aminolipid. The DNA G+C content was 72 mol%. Strain mbc-2T exhibited DNA–DNA relatedness values of 12.5±1.5 %, 23.7±2.7 % and 26.3±3.2 % with respect to Nocardioides hankookensis DS-30T, Nocardioides aquiterrae GW-9T and Nocardioides pyridinolyticus OS4T. On the basis of the data obtained, strain mbc-2T represents a novel species of the genus Nocardioides , for which the name Nocardioides soli sp. nov. is proposed. The type strain is mbc-2T ( = KACC 17152T = CCTCC AB 2012934T).

scholarly journals Thermal and Solvent Stress Cross-Tolerance Conferred to Corynebacterium glutamicum by Adaptive Laboratory Evolution

2015 ◽  
Vol 81 (7) ◽  
pp. 2284-2298 ◽  
Author(s):  
Shinichi Oide ◽  
Wataru Gunji ◽  
Yasuhiro Moteki ◽  
Shogo Yamamoto ◽  
Masako Suda ◽  
...  
Keyword(s):  
Thermal Stress ◽  
Stress Tolerance ◽  
Corynebacterium Glutamicum ◽  
Parental Strain ◽  
Cross Tolerance ◽  
Adaptive Laboratory Evolution ◽  
Laboratory Evolution ◽  
Content Type ◽  
Positive Effects ◽  
Solvent Stress

ABSTRACTReinforcing microbial thermotolerance is a strategy to enable fermentation with flexible temperature settings and thereby to save cooling costs. Here, we report on adaptive laboratory evolution (ALE) of the amino acid-producing bacteriumCorynebacterium glutamicumunder thermal stress. After 65 days of serial passage of the transgenic strain GLY3, in which the glycolytic pathway is optimized for alanine production under oxygen deprivation, three strains adapted to supraoptimal temperatures were isolated, and all the mutations they acquired were identified by whole-genome resequencing. Of the 21 mutations common to the three strains, one large deletion and two missense mutations were found to promote growth of the parental strain under thermal stress. Additive effects on thermotolerance were observed among these mutations, and the combination of the deletion with the missense mutation onotsA, encoding a trehalose-6-phosphate synthase, allowed the parental strain to overcome the upper limit of growth temperature. Surprisingly, the three evolved strains acquired cross-tolerance for isobutanol, which turned out to be partly attributable to the genomic deletion associated with the enhanced thermotolerance. The deletion involved loss of two transgenes,pfkandpyk, encoding the glycolytic enzymes, in addition to six native genes, and elimination of the transgenes, but not the native genes, was shown to account for the positive effects on thermal and solvent stress tolerance, implying a link between energy-producing metabolism and bacterial stress tolerance. Overall, the present study provides evidence that ALE can be a powerful tool to refine the phenotype ofC. glutamicumand to investigate the molecular bases of stress tolerance.

scholarly journals Ribosome hibernation: a new molecular framework for targeting nonreplicating persisters of mycobacteria

Microbiology ◽  
2021 ◽  
Vol 167 (2) ◽  
Author(s):  
Yunlong Li ◽  
Manjuli R. Sharma ◽  
Ravi K. Koripella ◽  
Nilesh K. Banavali ◽  
Rajendra K. Agrawal ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Type Species ◽  
Drug Regimen ◽  
Content Type ◽  
Link Type ◽  
A Minor ◽  
Molecular Framework

Treatment of tuberculosis requires a multi-drug regimen administered for at least 6 months. The long-term chemotherapy is attributed in part to a minor subpopulation of nonreplicating Mycobacterium tuberculosis cells that exhibit phenotypic tolerance to antibiotics. The origins of these cells in infected hosts remain unclear. Here we discuss some recent evidence supporting the hypothesis that hibernation of ribosomes in M. tuberculosis, induced by zinc starvation, could be one of the primary mechanisms driving the development of nonreplicating persisters in hosts. We further analyse inconsistencies in previously reported studies to clarify the molecular principles underlying mycobacterial ribosome hibernation.

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