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.

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 Characterization of the φCTX-like Pseudomonas aeruginosa phage Dobby isolated from the kidney stone microbiota

2019 ◽  
Vol 1 (1) ◽  
Author(s):  
Genevieve Johnson ◽  
Alan J. Wolfe ◽  
Catherine Putonti
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Urinary Tract ◽  
Human Body ◽  
Kidney Stone ◽  
Type Species ◽  
Phage Group ◽  
Content Type ◽  
Link Type ◽  
Abundance And Diversity ◽  
Cultured Bacteria

Bacteriophages (phages) are vital members of the human microbiota. They are abundant even within low biomass niches of the human body, including the lower urinary tract. While several prior studies have cultured bacteria from kidney stones, this is the first study to explore phages within the kidney stone microbiota. Here we report Dobby, a temperate phage isolated from a strain of Pseudomonas aeruginosa cultured from a kidney stone. Dobby is capable of lysing clinical P. aeruginosa strains within our collection from the urinary tract. Sequencing was performed producing a 37 152 bp genome that closely resembles the temperate P. aeruginosa phage φCTX, a member of the P2 phage group. Dobby does not, however, encode for the cytotoxin CTX. Dobby’s genome was queried against publicly available bacterial sequences identifying 44 other φCTX-like prophages. These prophages are integrated within the genomes of P. aeruginosa strains from a variety of environments, including strains isolated from urine samples and other niches of the human body. Phylogenetic analysis suggests that the temperate φCTX phage species is widespread. With the isolation of Dobby, we now have evidence that phages are members of the kidney stone microbiota. Further investigation, however, is needed to determine their abundance and diversity within these communities.

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 An antipathogenic compound that targets the OxyR peroxide sensor in Pseudomonas aeruginosa

2021 ◽  
Vol 70 (4) ◽  
Author(s):  
Hyo-Young Oh ◽  
Shivakumar S. Jalde ◽  
In-Young Chung ◽  
Yeon-Ji Yoo ◽  
Hye-Jeong Jang ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Drug Resistance ◽  
Stress Responses ◽  
Type Species ◽  
Reduced Form ◽  
Infection Model ◽  
Redox Cycle ◽  
Content Type ◽  
Link Type ◽  
Computer Based

Introduction. Antipathogenic or antivirulence strategy is to target a virulence pathway that is dispensable for growth, in the hope to mitigate the selection for drug resistance. Hypothesis/Gap Statment. Peroxide stress responses are one of the conserved virulence pathways in bacterial pathogens and thus good targets for antipathogenic strategy. Aim. This study aims to identify a new chemical compound that targets OxyR, the peroxide sensor required for the full virulence of the opportunistic human pathogen, Pseudomonas aeruginosa . Methodology. Computer-based virtual screening under consideration of the ‘eNTRy’ rules and molecular docking were conducted on the reduced form of the OxyR regulatory domain (RD). Selected hits were validated by their ability to phenocopy the oxyR null mutant and modulate the redox cycle of OxyR. Results. We first isolated three robust chemical hits that inhibit OxyR without affecting prototrophic growth or viability. One (compound 1) of those affected the redox cycle of OxyR in response to H2O2 treatment, in a way to impair its function. Compound 1 displayed selective antibacterial efficacy against P. aeruginosa in Drosophila infection model, without antibacterial activity against Staphylococcus aureus . Conclusion. These results suggest that compound 1 could be an antipathogenic hit inhibiting the P. aeruginosa OxyR. More importantly, our study provides an insight into the computer-based discovery of new-paradigm selective antibacterials to treat Gram-negative bacterial infections presumably with few concerns of drug resistance.

scholarly journals Study of quorum-sensing LasR and RhlR genes and their dependent virulence factors in Pseudomonas aeruginosa isolates from infected burn wounds

2021 ◽  
Vol 3 (3) ◽  
Author(s):  
Aya Ahmad Elnegery ◽  
Wafaa Kamel Mowafy ◽  
Tarek Ahmed Zahra ◽  
Noha Tharwat Abou El-Khier
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
Proteolytic Activity ◽  
Virulence Factors ◽  
Biofilm Formation ◽  
Type Species ◽  
Assay Method ◽  
Content Type ◽  
Burn Wounds ◽  
Link Type

Background. Pseudomonas aeruginosa (P. aeruginosa) is an opportunistic pathogen responsible for burn-wound infection. High incidence, infection severity and increasing resistance characterize P. aeruginosa -induced burn infection. Purpose. To estimate quorum-sensing (QS)-dependent virulence factors of P. aeruginosa isolates from burn wounds and correlate it to the presence of QS genes. Methods. A cross-sectional descriptive study included 50 P . aeruginosa isolates from burn patients in Mansoura University Plastic and Burn Hospital, Egypt. Antibiotic sensitivity tests were done. All isolates were tested for their ability to produce biofilm using a micro-titration assay method. Protease, pyocyanin and rhamnolipid virulence factors were determined using skimmed milk agar, King’s A medium and CTAB agar test, respectively. The identity of QS lasR and rhlR genes was confirmed using PCR. Results. In total, 86 % of isolates had proteolytic activity. Production of pyocyanin pigment was manifested in 66 % of isolates. Altogether, 76 % of isolates were rhamnolipid producers. Biofilm formation was detected in 96 % of isolates. QS lasR and rhlR genes were harboured by nearly all isolates except three isolates were negative for both lasR and rhlR genes and two isolates were positive for lasR gene and negative for rhlR gene. Forty-nine isolates were considered as extremely QS-proficient strains as they produced QS-dependent virulence factors. In contrast, one isolate was a QS deficient strain. Conclusions. QS affects P. aeruginosa virulence-factor production and biofilm in burn wounds. Isolates containing lasR and rhlR seem to be a crucial regulator of virulence factors and biofilm formation in P. aeruginosa whereas the lasR gene positively regulates biofilm formation, proteolytic activity, pyocyanin production and rhamnolipid biosurfactant synthesis. The QS regulatory RhlR gene affects protease and rhamnolipid production positively.

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 Lipopolysaccharide from biofilm-forming Pseudomonas aeruginosa PAO1 induces macrophage hyperinflammatory responses

2021 ◽  
Vol 70 (4) ◽  
Author(s):  
Sufei Wang ◽  
Dandan Xiang ◽  
Fangbing Tian ◽  
Ming Ni
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Cell Lines ◽  
Type Species ◽  
Macrophage Polarization ◽  
Human Macrophage ◽  
Murine Macrophage ◽  
Content Type ◽  
Link Type ◽  
Raw264.7 Macrophages

Introduction. Macrophages polarization is essential in infection control. Llipopolysaccharide (LPS) plays an essential role in host innate immune system–pathogen interaction. The LPS structure of Pseudomonas aeruginosa modifies in the adaptation of this pathogen to biofilm-related chronic infection. Gap statement. There have been several studies on LPS induced polarization of human and mouse macrophages with different results. And it was reported that the lipid A structure of the LPS derived from biofilm-forming Pseudomonas aeruginosa strain PAO1 was modified. Aim. This study aimed to investigate the effect and the involved pathway of LPS from biofilm-forming PAO1 on human and murine macrophage polarization. Methodology. LPS was isolated from biofilm-forming and planktonic PAO1 and quantified. Then the LPS was added to PMA-differentiated human macrophage THP-1 cells and Raw264.7 murine macrophage cells. The expression of iNOS, Arg-1, IL4, TNF-α, CCL3, and CCL22 was analysed in the different cell lines. The expression of TICAM-1 and MyD88 in human THP-1 macrophages was quantified by Western blot. PAO1 infected macrophages at different polarization states, and the intracellular bacterial growth in macrophages was evaluated. Results. LPS from biofilm-forming PAO1 induced more marked hyperinflammatory responses in THP-1 and Raw264.7 macrophages than LPS derived from planktonic PAO1, and these responses were related to the up-regulation of MyD88. Intracellular growth of PAO1 was significantly increased in THP-1 macrophages polarized by LPS from biofilm-forming PAO1, but decreased both in THP-1 and Raw264.7 macrophages polarized by LPS from planktonic PAO1. Conclusion. The presented in vitro study indicates that LPS derived from biofilm-forming PAO1 induces enhanced M1 polarization in human and murine macrophage cell lines than LPS from planktonic PAO1.

scholarly journals Combinatorial quorum sensing in Pseudomonas aeruginosa allows for novel cheating strategies

Microbiology ◽  
2020 ◽  
Vol 166 (8) ◽  
pp. 777-784 ◽  
Author(s):  
James Gurney ◽  
Sheyda Azimi ◽  
Sam P. Brown ◽  
Stephen P. Diggle
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
Type Species ◽  
Natural Populations ◽  
Opportunistic Pathogen ◽  
Growth Media ◽  
Public And Private ◽  
Content Type ◽  
Private Goods ◽  
Link Type

In the opportunistic pathogen Pseudomonas aeruginosa, quorum sensing (QS) is a social trait that is exploitable by non-cooperating cheats. Previously it has been shown that by linking QS to the production of both public and private goods, cheats can be prevented from invading populations of cooperators and this was described by Dandekar et al. (Science 2012;338:264–266) as ‘a metabolic incentive to cooperate’. We hypothesized that P. aeruginosa could evolve novel cheating strategies to circumvent private goods metabolism by rewiring its combinatorial response to two QS signals (3O-C12-HSL and C4-HSL). We performed a selection experiment that cycled P. aeruginosa between public and private goods growth media and evolved an isolate that rewired its control of cooperative protease expression from a synergistic (AND-gate) response to dual-signal input to a 3O-C12-HSL-only response. We show that this isolate circumvents metabolic incentives to cooperate and acts as a combinatorial signalling cheat, with higher fitness in competition with its ancestor. Our results show three important principles: first, combinatorial QS allows for diverse social strategies to emerge; second, restrictions levied by private goods are not sufficient to explain the maintenance of cooperation in natural populations; and third, modifying combinatorial QS responses could result in important physiological outcomes in bacterial populations.

scholarly journals Re-identification of strains deposited as Pseudomonas aeruginosa, Pseudomonas fluorescens and Pseudomonas putida in GenBank based on whole genome sequences

2020 ◽  
Vol 70 (11) ◽  
pp. 5958-5963
Author(s):  
Yuh Morimoto ◽  
Mari Tohya ◽  
Zulipiya Aibibula ◽  
Tadashi Baba ◽  
Hiroyuki Daida ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Genome Sequencing ◽  
Dna Hybridization ◽  
Type Species ◽  
Whole Genome ◽  
Average Nucleotide Identity ◽  
Genome Sequences ◽  
Content Type ◽  
Link Type

The taxonomic classification of Pseudomonas species has been revised and updated several times. This study utilized average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH) cutoff values of 95 and 70 %, respectively, to re-identify the species of strains deposited in GenBank as P. aeruginosa , P. fluorescens and P. putida . Of the 264 deposited P. aeruginosa strains, 259 were correctly identified as P. aeruginosa , but the remaining five were not. All 28 deposited P. fluorescens strains had been incorrectly identified as P. fluorescens . Four of these strains were re-identified, including two as P. kilonensis and one each as P. aeruginosa and P. brassicacearum , but the remaining 24 could not be re-identified. Similarly, all 35 deposited P. putida strains had been incorrectly identified as P. putida . Nineteen of these strains were re-identified, including 12 as P. alloputida , four as P. asiatica and one each as P. juntendi , P. monteilii and P. mosselii . These results strongly suggest that Pseudomonas bacteria should be identified using ANI and dDDH analyses based on whole genome sequencing when Pseudomonas species are initially deposited in GenBank/DDBJ/EMBL databases.

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