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PloS One 2024Evaluate the effects of five disinfection methods on bacterial concentrations in hospital sink drains, focusing on three opportunistic pathogens (OPs): Serratia...
OBJECTIVE
Evaluate the effects of five disinfection methods on bacterial concentrations in hospital sink drains, focusing on three opportunistic pathogens (OPs): Serratia marcescens, Pseudomonas aeruginosa and Stenotrophomonas maltophilia.
DESIGN
Over two years, three sampling campaigns were conducted in a neonatal intensive care unit (NICU). Samples from 19 sink drains were taken at three time points: before, during, and after disinfection. Bacterial concentration was measured using culture-based and flow cytometry methods. High-throughput short sequence typing was performed to identify the three OPs and assess S. marcescens persistence after disinfection at the genotypic level.
SETTING
This study was conducted in a pediatric hospitals NICU in Montréal, Canada, which is divided in an intensive and intermediate care side, with individual rooms equipped with a sink.
INTERVENTIONS
Five treatments were compared: self-disinfecting drains, chlorine disinfection, boiling water disinfection, hot tap water flushing, and steam disinfection.
RESULTS
This study highlights significant differences in the effectiveness of disinfection methods. Chlorine treatment proved ineffective in reducing bacterial concentration, including the three OPs. In contrast, all other drain interventions resulted in an immediate reduction in culturable bacteria (4-8 log) and intact cells (2-3 log). Thermal methods, particularly boiling water and steam treatments, exhibited superior effectiveness in reducing bacterial loads, including OPs. However, in drains with well-established bacterial biofilms, clonal strains of S. marcescens recolonized the drains after heat treatments.
CONCLUSIONS
Our study supports thermal disinfection (>80°C) for pathogen reduction in drains but highlights the need for additional trials and the implementation of specific measures to limit biofilm formation.
Topics: Serratia marcescens; Intensive Care Units, Neonatal; Disinfection; Humans; Pseudomonas aeruginosa; Infant, Newborn; Stenotrophomonas maltophilia; Serratia Infections; Cross Infection
PubMed: 38865328
DOI: 10.1371/journal.pone.0304378 -
Proceedings of the National Academy of... Jun 2024Nitrous oxide (NO), a potent greenhouse gas, can be generated by multiple biological and abiotic processes in diverse contexts. Accurately tracking the dominant sources...
Nitrous oxide (NO), a potent greenhouse gas, can be generated by multiple biological and abiotic processes in diverse contexts. Accurately tracking the dominant sources of NO has the potential to improve our understanding of NO fluxes from soils as well as inform the diagnosis of human infections. Isotopic "Site Preference" (SP) values have been used toward this end, as bacterial and fungal nitric oxide reductases (NORs) produce NO with different isotopic fingerprints, spanning a large range. Here, we show that flavohemoglobin (Fhp), a hitherto biogeochemically neglected yet widely distributed detoxifying bacterial NO reductase, imparts a distinct SP value onto NO under anoxic conditions (~+10‰) that correlates with typical environmental NO SP measurements. Using as a model organism, we generated strains that only contained Fhp or the dissimilatory NOR, finding that in vivo NO SP values imparted by these enzymes differ by over 10‰. Depending on the cellular physiological state, the ratio of Fhp:NOR varies significantly in wild-type cells and controls the net NO SP biosignature: When cells grow anaerobically under denitrifying conditions, NOR dominates; when cells experience rapid, increased nitric oxide concentrations under anoxic conditions but are not growing, Fhp dominates. Other bacteria that only make Fhp generate similar NO SP biosignatures to those measured from our Fhp-only strain. Fhp homologs in sequenced bacterial genomes currently exceed NOR homologs by nearly a factor of four. Accordingly, we suggest a different framework to guide the attribution of NO biological sources in nature and disease.
Topics: Nitrous Oxide; Oxidoreductases; Pseudomonas aeruginosa; Anaerobiosis; Bacterial Proteins; Nitric Oxide
PubMed: 38865268
DOI: 10.1073/pnas.2319960121 -
Protein Science : a Publication of the... Jul 2024Peptidoglycan is a major constituent of the bacterial cell wall. Its integrity as a polymeric edifice is critical for bacterial survival and, as such, it is a preeminent...
Peptidoglycan is a major constituent of the bacterial cell wall. Its integrity as a polymeric edifice is critical for bacterial survival and, as such, it is a preeminent target for antibiotics. The peptidoglycan is a dynamic crosslinked polymer that undergoes constant biosynthesis and turnover. The soluble lytic transglycosylase (Slt) of Pseudomonas aeruginosa is a periplasmic enzyme involved in this dynamic turnover. Using amber-codon-suppression methodology in live bacteria, we incorporated a fluorescent chromophore into the structure of Slt. Fluorescent microscopy shows that Slt populates the length of the periplasmic space and concentrates at the sites of septation in daughter cells. This concentration persists after separation of the cells. Amber-codon-suppression methodology was also used to incorporate a photoaffinity amino acid for the capture of partner proteins. Mass-spectrometry-based proteomics identified 12 partners for Slt in vivo. These proteomics experiments were complemented with in vitro pulldown analyses. Twenty additional partners were identified. We cloned the genes and purified to homogeneity 22 identified partners. Biophysical characterization confirmed all as bona fide Slt binders. The identities of the protein partners of Slt span disparate periplasmic protein families, inclusive of several proteins known to be present in the divisome. Notable periplasmic partners (K < 0.5 μM) include PBPs (PBP1a, K = 0.07 μM; PBP5 = 0.4 μM); other lytic transglycosylases (SltB2, K = 0.09 μM; RlpA, K = 0.4 μM); a type VI secretion system effector (Tse5, K = 0.3 μM); and a regulatory protease for alginate biosynthesis (AlgO, K < 0.4 μM). In light of the functional breadth of its interactome, Slt is conceptualized as a hub protein within the periplasm.
Topics: Pseudomonas aeruginosa; Bacterial Proteins; Periplasm; Periplasmic Proteins; Glycosyltransferases; Peptidoglycan
PubMed: 38864725
DOI: 10.1002/pro.5038 -
Microbiology Spectrum Jul 2024This study aimed to characterize the composition of intestinal and nasal microbiota in septic patients and identify potential microbial biomarkers for diagnosis. A total...
This study aimed to characterize the composition of intestinal and nasal microbiota in septic patients and identify potential microbial biomarkers for diagnosis. A total of 157 subjects, including 89 with sepsis, were enrolled from the affiliated hospital. Nasal swabs and fecal specimens were collected from septic and non-septic patients in the intensive care unit (ICU) and Department of Respiratory and Critical Care Medicine. DNA was extracted, and the V4 region of the 16S rRNA gene was amplified and sequenced using Illumina technology. Bioinformatics analysis, statistical processing, and machine learning techniques were employed to differentiate between septic and non-septic patients. The nasal microbiota of septic patients exhibited significantly lower community richness ( = 0.002) and distinct compositions ( = 0.001) compared to non-septic patients. , , , and were identified as enriched genera in the nasal microbiota of septic patients. The constructed machine learning model achieved an area under the curve (AUC) of 89.08, indicating its efficacy in differentiating septic and non-septic patients. Importantly, model validation demonstrated the effectiveness of the nasal microecological diagnosis prediction model with an AUC of 84.79, while the gut microecological diagnosis prediction model had poor predictive performance (AUC = 49.24). The nasal microbiota of ICU patients effectively distinguishes sepsis from non-septic cases and outperforms the gut microbiota. These findings have implications for the development of diagnostic strategies and advancements in critical care medicine.IMPORTANCEThe important clinical significance of this study is that it compared the intestinal and nasal microbiota of sepsis with non-sepsis patients and determined that the nasal microbiota is more effective than the intestinal microbiota in distinguishing patients with sepsis from those without sepsis, based on the difference in the lines of nasal specimens collected.
Topics: Humans; Sepsis; Intensive Care Units; Male; Female; Middle Aged; Aged; Microbiota; RNA, Ribosomal, 16S; Biomarkers; Bacteria; Feces; Adult; Machine Learning; Gastrointestinal Microbiome; Nose; Corynebacterium; Acinetobacter; Aged, 80 and over; Staphylococcus; Pseudomonas
PubMed: 38864649
DOI: 10.1128/spectrum.03441-23 -
Microbial Biotechnology Jun 2024Treating plant bacterial diseases is notoriously difficult because of the lack of available antimicrobials. Pseudomonas syringae pathovar syringae (Pss) is a major...
Treating plant bacterial diseases is notoriously difficult because of the lack of available antimicrobials. Pseudomonas syringae pathovar syringae (Pss) is a major pathogen of cherry (Prunus avium) causing bacterial canker of the stem, leaf and fruit, impacting productivity and leading to a loss of trees. In an attempt to find a treatment for this disease, naturally occurring bacteriophage (phage) that specifically target Pss is being investigated as a biocontrol strategy. However, before using them as a biocontrol treatment, it is important to both understand their efficacy in reducing the bacterial population and determine if the bacterial pathogens can evolve resistance to evade phage infection. To investigate this, killing curve assays of five MR phages targeting Pss showed that phage resistance rapidly emerges in vitro, even when using a cocktail of the five phages together. To gain insight to the changes occurring, Pss colonies were collected three times during a 66-h killing curve assay and separately, Pss and phage were also coevolved over 10 generations, enabling the measurement of genomic and fitness changes in bacterial populations. Pss evolved resistance to phages through modifications in lipopolysaccharide (LPS) synthesis pathways. Bacterial fitness (growth) and virulence were affected in only a few mutants. Deletion of LPS-associated genes suggested that LPS was the main target receptor for all five MR phages. Later generations of coevolved phages from the coevolution experiment were more potent at reducing the bacterial density and when used with wild-type phages could reduce the emergence of phage-resistant mutants. This study shows that understanding the genetic mechanisms of bacterial pathogen resistance to phages is important for helping to design a more effective approach to kill the bacteria while minimizing the opportunity for phage resistance to manifest.
Topics: Pseudomonas syringae; Plant Diseases; Pseudomonas Phages; Bacteriophages
PubMed: 38864499
DOI: 10.1111/1751-7915.14489 -
Microbial Biotechnology Jun 2024Antimicrobial resistance (AMR) is an escalating global health crisis, driven by the overuse and misuse of antibiotics. Multidrug-resistant Gram-negative bacteria, such...
Antimicrobial resistance (AMR) is an escalating global health crisis, driven by the overuse and misuse of antibiotics. Multidrug-resistant Gram-negative bacteria, such as Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae, are particularly concerning due to their high morbidity and mortality rates. In this context, endolysins, derived from bacteriophages, offer a promising alternative to traditional antibiotics. This study introduces LysJEP8, a novel endolysin derived from Escherichia phage JEP8, which exhibits remarkable antimicrobial activity against key Gram-negative members of the ESKAPE group. Comparative assessments highlight LysJEP8's superior performance in reducing bacterial survival rates compared to previously described endolysins, with the most significant impact observed against P. aeruginosa, and notable effects on A. baumannii and K. pneumoniae. The study found that LysJEP8, as predicted by in silico analysis, worked best at lower pH values but lost its effectiveness at salt concentrations close to physiological levels. Importantly, LysJEP8 exhibited remarkable efficacy in the disruption of P. aeruginosa biofilms. This research underscores the potential of LysJEP8 as a valuable candidate for the development of innovative antibacterial agents, particularly against Gram-negative pathogens, and highlights opportunities for further engineering and optimization to address AMR effectively.
Topics: Endopeptidases; Drug Resistance, Multiple, Bacterial; Anti-Bacterial Agents; Gram-Negative Bacteria; Pseudomonas aeruginosa; Biofilms; Microbial Sensitivity Tests; Bacteriophages; Klebsiella pneumoniae; Hydrogen-Ion Concentration; Acinetobacter baumannii; Microbial Viability
PubMed: 38864495
DOI: 10.1111/1751-7915.14483 -
Environmental Microbiology Jun 2024The environmental bacterium, Pseudomonas putida, possesses a broad spectrum of metabolic pathways. This makes it highly promising for use in biotechnological production...
The environmental bacterium, Pseudomonas putida, possesses a broad spectrum of metabolic pathways. This makes it highly promising for use in biotechnological production as a cell factory, as well as in bioremediation strategies to degrade various aromatic pollutants. For P. putida to flourish in its environment, it must withstand the continuous threats posed by bacteriophages. Interestingly, until now, only a handful of phages have been isolated for the commonly used laboratory strain, P. putida KT2440, and no phage defence mechanisms have been characterized. In this study, we present a new Collection of Environmental P. putida Phages from Estonia, or CEPEST. This collection comprises 67 double-stranded DNA phages, which belong to 22 phage species and 9 phage genera. Our findings reveal that most phages in the CEPEST collection are more infectious at lower temperatures, have a narrow host range, and require an intact lipopolysaccharide for P. putida infection. Furthermore, we show that cryptic prophages present in the P. putida chromosome provide strong protection against the infection of many phages. However, the chromosomal toxin-antitoxin systems do not play a role in the phage defence of P. putida. This research provides valuable insights into the interactions between P. putida and bacteriophages, which could have significant implications for biotechnological and environmental applications.
Topics: Pseudomonas putida; Host Specificity; Prophages; Pseudomonas Phages; Estonia; Bacteriophages
PubMed: 38863081
DOI: 10.1111/1462-2920.16671 -
The AAPS Journal Jun 2024Addressing the intertwined challenges of antimicrobial resistance and impaired wound healing in diabetic patients, an oil/water emulsion-based nano-ointment integrating...
Addressing the intertwined challenges of antimicrobial resistance and impaired wound healing in diabetic patients, an oil/water emulsion-based nano-ointment integrating phenylpropanoids-Eugenol and Cinnamaldehyde-with positively-charged silver nanoparticles was synthesized. The process began with the synthesis and characterization of nano-silver, aimed at ensuring the effectiveness and safety of the nanoparticles in biological applications. Subsequent experiments determined the minimum inhibitory concentration (MIC) against pathogens such as Streptococcus aureus, Pseudomonas aeruginosa and Candida albicans. These MIC values of all three active leads guided the strategic formulation of an ointment base, which effectively integrated the bioactive components. Evaluations of this nano-ointment revealed enhanced antimicrobial activity against both clinical and reference bacterial strains and it maintained stability after freeze-thaw cycles. Furthermore, the ointment demonstrated superior in-vitro diabetic wound healing capabilities and significantly promoted angiogenesis, as shown by enhanced blood vessel formation in the Chorioallantoic Membrane assay. These findings underscore the formulation's therapeutic potential, marking a significant advance in the use of nanotechnology for topical wound care.
Topics: Silver; Wound Healing; Microbial Sensitivity Tests; Metal Nanoparticles; Animals; Ointments; Acrolein; Candida albicans; Anti-Infective Agents; Pseudomonas aeruginosa; Administration, Topical; Humans; Anti-Bacterial Agents; Staphylococcus aureus
PubMed: 38862870
DOI: 10.1208/s12248-024-00936-4 -
Scientific Reports Jun 2024The Increase in infections caused by resistant strains of Pseudomonas aeruginosa poses a formidable challenge to global healthcare systems. P. aeruginosa is capable of...
The Increase in infections caused by resistant strains of Pseudomonas aeruginosa poses a formidable challenge to global healthcare systems. P. aeruginosa is capable of causing severe human infections across diverse anatomical sites, presenting considerable therapeutic obstacles due to its heightened drug resistance. Niosomal drug delivery systems offer enhanced pharmaceutical potential for loaded contents due to their desirable properties, mainly providing a controlled-release profile. This study aimed to formulate an optimized niosomal drug delivery system incorporating stearylamine (SA) to augment the anti-bacterial and anti-biofilm activities of quercetin (QCT) against both standard and clinical strains of P. aeruginosa. QCT-loaded niosome (QCT-niosome) and QCT-loaded SA- niosome (QCT-SA- niosome) were synthesized by the thin-film hydration technique, and their physicochemical characteristics were evaluated by field emission scanning electron microscopy (FE-SEM), zeta potential measurement, entrapment efficacy (EE%), and in vitro release profile. The anti-P. aeruginosa activity of synthesized niosomes was assessed using minimum inhibitory and bactericidal concentrations (MICs/MBCs) and compared with free QCT. Additionally, the minimum biofilm inhibitory and eradication concentrations (MBICs/MBECs) were carried out to analyze the ability of QCT-niosome and QCT-SA-niosome against P. aeruginosa biofilms. Furthermore, the cytotoxicity assay was conducted on the L929 mouse fibroblasts cell line to evaluate the biocompatibility of the formulated niosomes. FE-SEM analysis revealed that both synthesized niosomal formulations exhibited spherical morphology with different sizes (57.4 nm for QCT-niosome and 178.9 nm for QCT-SA-niosome). The EE% for cationic and standard niosomal formulations was reported at 75.9% and 59.6%, respectively. Both formulations showed an in vitro sustained-release profile, and QCT-SA-niosome exhibited greater stability during a 4-month storage time compared to QCT-niosome. Microbial experiments indicated that both prepared formulations had higher anti-bacterial and anti-biofilm activities than free QCT. Also, the QCT-SA-niosome exhibited greater reductions in MIC, MBC, MBIC, and MBEC values compared to the QCT-niosome at equivalent concentrations. This study supports the potential of QCT-niosome and QCT-SA-niosome as effective agents against P. aeruginosa infections, manifesting significant anti-bacterial and anti-biofilm efficacy alongside biocompatibility with L929 cell lines. Furthermore, our results suggest that optimized QCT-niosome with cationic lipids could efficiently target P. aeruginosa cells with negligible cytotoxic effect.
Topics: Pseudomonas aeruginosa; Liposomes; Quercetin; Animals; Biofilms; Mice; Anti-Bacterial Agents; Drug Delivery Systems; Pseudomonas Infections; Microbial Sensitivity Tests; Cations; Humans; Lipids; Cell Line; Amines
PubMed: 38862754
DOI: 10.1038/s41598-024-64416-7 -
PLoS Genetics Jun 2024Bacteria use diverse strategies and molecular machinery to maintain copper homeostasis and to cope with its toxic effects. Some genetic elements providing copper...
Bacteria use diverse strategies and molecular machinery to maintain copper homeostasis and to cope with its toxic effects. Some genetic elements providing copper resistance are acquired by horizontal gene transfer; however, little is known about how they are controlled and integrated into the central regulatory network. Here, we studied two copper-responsive systems in a clinical isolate of Pseudomonas paraeruginosa and deciphered the regulatory and cross-regulation mechanisms. To do so, we combined mutagenesis, transcriptional fusion analyses and copper sensitivity phenotypes. Our results showed that the accessory CusRS two-component system (TCS) responds to copper and activates both its own expression and that of the adjacent nine-gene operon (the pcoA2 operon) to provide resistance to elevated levels of extracellular copper. The same locus was also found to be regulated by two core-genome-encoded TCSs-the copper-responsive CopRS and the zinc-responsive CzcRS. Although the target palindromic sequence-ATTCATnnATGTAAT-is the same for the three response regulators, transcriptional outcomes differ. Thus, depending on the operon/regulator pair, binding can result in different activation levels (from none to high), with the systems demonstrating considerable plasticity. Unexpectedly, although the classical CusRS and the noncanonical CopRS TCSs rely on distinct signaling mechanisms (kinase-based vs. phosphatase-based), we discovered cross-talk in the absence of the cognate sensory kinases. This cross-talk occurred between the proteins of these two otherwise independent systems. The cusRS-pcoA2 locus is part of an Integrative and Conjugative Element and was found in other Pseudomonas strains where its expression could provide copper resistance under appropriate conditions. The results presented here illustrate how acquired genetic elements can become part of endogenous regulatory networks, providing a physiological advantage. They also highlight the potential for broader effects of accessory regulatory proteins through interference with core regulatory proteins.
Topics: Copper; Pseudomonas; Gene Expression Regulation, Bacterial; Operon; Bacterial Proteins; Drug Resistance, Bacterial; Signal Transduction
PubMed: 38861577
DOI: 10.1371/journal.pgen.1011325