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Environmental Microbiology Apr 2024Aphids are globally important pests causing damage to a broad range of crops. Due to insecticide resistance, there is an urgent need to develop alternative control...
Aphids are globally important pests causing damage to a broad range of crops. Due to insecticide resistance, there is an urgent need to develop alternative control strategies. In our previous work, we found Pseudomonas fluorescens PpR24 can orally infect and kill the insecticide-resistant green-peach aphid (Myzus persicae). However, the genetic basis of the insecticidal capability of PpR24 remains unclear. Genome sequencing of PpR24 confirmed the presence of various insecticidal toxins such as Tc (toxin complexes), Rhs (rearrangement hotspot) elements, and other insect-killing proteases. Upon aphids infection with PpR24, RNA-Seq analysis revealed 193 aphid genes were differentially expressed with down-regulation of 16 detoxification genes. In addition, 1325 PpR24 genes (542 were upregulated and 783 downregulated) were subject to differential expression, including genes responsible for secondary metabolite biosynthesis, the iron-restriction response, oxidative stress resistance, and virulence factors. Single and double deletion of candidate virulence genes encoding a secreted protease (AprX) and four toxin components (two TcA-like; one TcB-like; one TcC-like insecticidal toxins) showed that all five genes contribute significantly to aphid killing, particularly AprX. This comprehensive host-pathogen transcriptomic analysis provides novel insight into the molecular basis of bacteria-mediated aphid mortality and the potential of PpR24 as an effective biocontrol agent.
Topics: Animals; Aphids; Pseudomonas fluorescens; Peptide Hydrolases; Insecticides; Gene Expression Profiling
PubMed: 38561900
DOI: 10.1111/1462-2920.16604 -
MicrobiologyOpen Apr 2024Arginine-ornithine metabolism plays a crucial role in bacterial homeostasis, as evidenced by numerous studies. However, the utilization of arginine and the downstream...
Arginine-ornithine metabolism plays a crucial role in bacterial homeostasis, as evidenced by numerous studies. However, the utilization of arginine and the downstream products of its metabolism remain undefined in various gut bacteria. To bridge this knowledge gap, we employed genomic screening to pinpoint relevant metabolic targets. We also devised a targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) metabolomics method to measure the levels of arginine, its upstream precursors, and downstream products in cell-free conditioned media from enteric pathobionts, including Escherichia coli, Klebsiella aerogenes, K. pneumoniae, Pseudomonas fluorescens, Acinetobacter baumannii, Streptococcus agalactiae, Staphylococcus epidermidis, S. aureus, and Enterococcus faecalis. Our findings revealed that all selected bacterial strains consumed glutamine, glutamate, and arginine, and produced citrulline, ornithine, and GABA in our chemically defined medium. Additionally, E. coli, K. pneumoniae, K. aerogenes, and P. fluorescens were found to convert arginine to agmatine and produce putrescine. Interestingly, arginine supplementation promoted biofilm formation in K. pneumoniae, while ornithine supplementation enhanced biofilm formation in S. epidermidis. These findings offer a comprehensive insight into arginine-ornithine metabolism in enteric pathobionts.
Topics: Ornithine; Putrescine; Arginine; Escherichia coli; Chromatography, Liquid; Staphylococcus aureus; Tandem Mass Spectrometry; Bacteria; Klebsiella pneumoniae
PubMed: 38560776
DOI: 10.1002/mbo3.1408 -
Bioengineering (Basel, Switzerland) Feb 2024Acute recurrent tonsillitis is a chronic, biofilm-related infection that is a significant burden to patients and healthcare systems. It is often treated with repeated...
Acute recurrent tonsillitis is a chronic, biofilm-related infection that is a significant burden to patients and healthcare systems. It is often treated with repeated courses of antibiotics, which contributes to antimicrobial resistance. Studying biofilms is key to understanding this disease. In vitro modelling using 3D bioprinted hydrogels is a promising approach to achieve this. A novel gelatin-PEGDA pseudomonas fluorescens-laden bioink was developed and bioprinted in a 3D hydrogel construct fabricated using computer-aided design to mimic the tonsillar biofilm environment. The bioprinted constructs were cultured at 37 °C in lysogeny broth for 12 days. Bacterial growth was assessed by spectrophotometry. Cellular viability analysis was conducted using optical fluorescence microscopy (FDA/PI staining). A biocompatible 3D-printed bacteria-laden hydrogel construct was successfully fabricated. Bacterial growth was observed using optical fluorescence microscopy. A live/dead cellular-staining protocol demonstrated bacterial viability. Results obtained after the 12-day culture period showed higher bacterial growth in the 1% gelatin concentration construct compared to the 0% control. This study demonstrates the first use of a bacteria-laden gelatin-PEGDA hydrogel for biofabrication of a 3D-printed construct designed to model acute recurrent tonsillitis. Initiating a study with clinically relevant ex vivo tonsil bacteria will be an important next step in improving treatment of this impactful but understudied disease.
PubMed: 38534476
DOI: 10.3390/bioengineering11030202 -
The Journal of Hospital Infection May 2024In recent years, hand drying has been highlighted as a key step in appropriate hand hygiene, as moisture on hands can increase the transfer of micro-organisms from hands...
BACKGROUND
In recent years, hand drying has been highlighted as a key step in appropriate hand hygiene, as moisture on hands can increase the transfer of micro-organisms from hands to surfaces and vice versa.
AIM
To understand bacterial and viral aerosolization following hand drying, and study the transfer of micro-organisms from hands to surfaces after drying using different methods.
METHODS
Groups of five volunteers had their hands pre-washed with soap, rinsed and dried, then inoculated with a concentrated mixture of Pseudomonas fluorescens and MS2 bacteriophage. Volunteers entered an empty washroom, one at a time, and rinsed their hands with water or washed their hands with soap prior to drying with a jet dryer or paper towels. Each volunteer applied one hand successively to various surfaces, while their other hand was sampled using the glove juice method. Both residual bacteria and viruses were quantified from the washroom air, surface swabs and hand samples.
FINDINGS
P. fluorescens and MS2 bacteriophages were rarely aerosolized while drying hands for any of the drying methods studied. Results also showed limited, and similar, transfer of both micro-organisms studied on to surfaces for all drying methods.
CONCLUSION
The use of jet dryers or paper towels produces low levels of aerosolization when drying hands in a washroom. Similarly, all drying methods result in low transfer to surfaces. While the coronavirus disease 2019 pandemic raised concerns regarding public washrooms, this study shows that all methods tested are hygienic solutions for dry washed hands.
Topics: Humans; Hand; Levivirus; Pseudomonas fluorescens; Aerosols; Hand Disinfection; Bacteria; Desiccation; Hand Hygiene; COVID-19; Viruses; Environmental Microbiology
PubMed: 38521417
DOI: 10.1016/j.jhin.2024.03.005 -
PloS One 2024Trichoderma uses different molecules to establish communication during its interactions with other organisms, such as effector proteins. Effectors modulate plant...
Trichoderma uses different molecules to establish communication during its interactions with other organisms, such as effector proteins. Effectors modulate plant physiology to colonize plant roots or improve Trichoderma's mycoparasitic capacity. In the soil, these fungi can establish relationships with plant growth-promoting bacteria (PGPBs), thus affecting their overall benefits on the plant or its fungal prey, and possibly, the role of effector proteins. The aim of this study was to determine the induction of Trichoderma atroviride gene expression coding for effector proteins during the interaction with different PGPBs, Arabidopsis or the phytopathogen Fusarium brachygibbosum, and to determine whether PGPBs potentiates the beneficial effects of T. atroviride. During the interaction with F. brachygibbosum and PGPBs, the effector coding genes epl1, tatrx2 and tacfem1 increased their expression, especially during the consortia with the bacteria. During the interaction of T. atroviride with the plant and PGPBs, the expression of epl1 and tatrx2 increased, mainly with the consortium formed with Pseudomonas fluorescens UM270, Bacillus velezensis AF12, or B. halotolerans AF23. Additionally, the consortium formed by T. atroviride and R. badensis SER3 stimulated A. thaliana PR1:GUS and LOX2:GUS for SA- and JA-mediated defence responses. Finally, the consortium of T. atroviride with SER3 was better at inhibiting pathogen growth, but the consortium of T. atroviride with UM270 was better at promoting Arabidopsis growth. These results showed that the biocontrol capacity and plant growth-promoting traits of Trichoderma spp. can be potentiated by PGPBs by stimulating its effector functions.
Topics: Antifungal Agents; Arabidopsis; Plant Development; Bacteria; Trichoderma; Hypocreales
PubMed: 38517906
DOI: 10.1371/journal.pone.0301139 -
Journal of Biosciences 2024Snake venom L-amino acid oxidases (LAAOs) are flavoenzymes with diverse physiological and pharmacological effects. These enzymes are found to showcase anticoagulant,...
Snake venom L-amino acid oxidases (LAAOs) are flavoenzymes with diverse physiological and pharmacological effects. These enzymes are found to showcase anticoagulant, antiplatelet, cytotoxicity and other biological effects in bite victims. However, the exact mechanism through which they exhibit several biological properties is not yet fully understood. The current study focussed on the purification of cobra venom LAAO and the functional characterization of purified LAAO. A novel L-amino acid oxidase NNLAAO70 with a molecular weight ~70 kDa was purified from the venom of an Indian spectacled cobra (). NNLAAO70 showed high substrate specificity for L-His, L-Leu, and L-Arg during its LAAO activity. It inhibited adenosine di-phosphate (ADP) and collagen-induced platelet aggregation process in a dosedependent manner. About 60% inhibition of collagen-induced and 40% inhibition of ADP-induced platelet aggregation was observed with a 40 μg/ml dose of NNLAAO70. NNLAAO70 exhibited bactericidal activity on and . NNLAAO70 also showed cytotoxicity on A549 cells . It showed severe bactericidal activity on and lysed 55% of cells. NNLAAO70 also exhibited drastic cytotoxicity on A549 cells. At 1 lg/ml dosage, it demonstrated a 60% reduction in A549 viability and induced apoptosis upon 24-h incubation. HO released during oxidative deamination reactions played a major role in NNLAAO70-induced cytotoxicity. NNLAAO70 significantly increased intracellular reactive oxygen species (ROS) levels in A549 cells by six fold when compared to untreated cells. Oxidative stress-mediated cell injury is the primary cause of NNLAAO70-induced apoptosis in A549 cells and prolonged oxidative stress caused DNA fragmentation and activated cellular secondary necrosis.
Topics: Animals; Humans; Elapidae; Naja naja; L-Amino Acid Oxidase; Hydrogen Peroxide; Elapid Venoms; Apoptosis; Necrosis; Collagen; Lung; Neoplasms
PubMed: 38516910
DOI: No ID Found -
Frontiers in Microbiology 2024Lettuce is an economically major leafy vegetable that is affected by numerous diseases. One of the most devastating diseases of lettuce is white mold caused by . Control...
Lettuce is an economically major leafy vegetable that is affected by numerous diseases. One of the most devastating diseases of lettuce is white mold caused by . Control methods for this fungus are limited due to the development of genetic resistance to commonly used fungicides, the large number of hosts and the long-term survival of sclerotia in soil. To elaborate a new and more sustainable approach to contain this pathogen, 1,210 strains previously isolated from agricultural soils in Canada were screened for their antagonistic activity against . Nine strains showed strong inhibition in dual-culture confrontational assays. Whole genome sequencing of these strains revealed their affiliation with four phylogenomic subgroups within the group, namely , , , and . The antagonistic strains harbor several genes and gene clusters involved in the production of secondary metabolites, including mycin-type and peptin-type lipopeptides, and antibiotics such as brabantamide, which may be involved in the inhibitory activity observed against . Three strains also demonstrated significant biocontrol abilities against the pathogen when either inoculated on lettuce leaves or in the growing substrate of lettuce plants grown in pots. They however did not impact populations in the rhizosphere, suggesting that they protect lettuce plants by altering the fitness and the virulence of the pathogen rather than by directly impeding its growth. These results mark a step forward in the development of biocontrol products against .
PubMed: 38516010
DOI: 10.3389/fmicb.2024.1304682 -
Angewandte Chemie (Weinheim An Der... Nov 2023Mupirocin is a clinically important antibiotic produced by a -AT Type I polyketide synthase (PKS) in . The major bioactive metabolite, pseudomonic acid A (PA-A), is...
Mupirocin is a clinically important antibiotic produced by a -AT Type I polyketide synthase (PKS) in . The major bioactive metabolite, pseudomonic acid A (PA-A), is assembled on a tetrasubstituted tetrahydropyran (THP) core incorporating a 6-hydroxy group proposed to be introduced by α-hydroxylation of the thioester of the acyl carrier protein (ACP) bound polyketide chain. Herein, we describe an in vitro approach combining purified enzyme components, chemical synthesis, isotopic labelling, mass spectrometry and NMR in conjunction with in vivo studies leading to the first characterisation of the α-hydroxylation bimodule of the mupirocin biosynthetic pathway. These studies reveal the precise timing of hydroxylation by MupA, substrate specificity and the ACP dependency of the enzyme components that comprise this α-hydroxylation bimodule. Furthermore, using purified enzyme, it is shown that the MmpA KS shows relaxed substrate specificity, suggesting precise spatiotemporal control of MupA recruitment in the context of the PKS. Finally, the detection of multiple intermodular MupA/ACP interactions suggests these bimodules may integrate MupA into their assembly.
PubMed: 38515435
DOI: 10.1002/ange.202312514 -
Colloids and Surfaces. B, Biointerfaces May 2024Biofilms are complex porous materials formed by microorganisms, polysaccharides, proteins, eDNA, inorganic matter, and water. They are ubiquitous in various...
Biofilms are complex porous materials formed by microorganisms, polysaccharides, proteins, eDNA, inorganic matter, and water. They are ubiquitous in various environmental niches and are known to grow at solid-liquid, solid-air and air-liquid interfaces, often causing problems in several industrial and sanitary fields. Their removal is a challenge in many applications and numerous studies have been conducted to identify promising chemical species as cleaning agents. While these substances target specific components of biofilm structure, the role of water content in biofilm, and how it can influence wettability and detergent absorption have been quite neglected in the literature. Estimating water content in biofilm is a challenging task due to its heterogeneity in morphology and chemical composition. In this study, we controlled water content in Pseudomonas fluorescens AR 11 biofilms grown on submerged glass slides by regulating environmental relative humidity after drying. Interfacial properties of biofilm were investigated by measuring wetting of water and soybean oil. The morphology of biofilm structure was evaluated using Confocal Laser Scanning Microscopy and Scanning Electron Microscopy. The results showed that biofilm water content has a significant and measurable effect on its wettability, leading to the hypothesis that a preliminary control of water content can play a crucial role in biofilm removal process.
Topics: Wettability; Pseudomonas fluorescens; Humidity; Biofilms; Water
PubMed: 38508084
DOI: 10.1016/j.colsurfb.2024.113831 -
Proceedings of the National Academy of... Mar 2024Biofilms of sulfate-reducing bacterium (SRB) like Hildenborough (DvH) can facilitate metal corrosion in various industrial and environmental settings leading to...
Biofilms of sulfate-reducing bacterium (SRB) like Hildenborough (DvH) can facilitate metal corrosion in various industrial and environmental settings leading to substantial economic losses. Although the mechanisms of biofilm formation by DvH are not yet well understood, recent studies indicate the large adhesin, DvhA, is a key determinant of biofilm formation. The gene neighborhood resembles the biofilm-regulating Lap system of but is curiously missing the c-di-GMP-binding regulator LapD. Instead, DvH encodes an evolutionarily unrelated c-di-GMP-binding protein (DVU1020) that we hypothesized is functionally analogous to LapD. To study this unusual Lap system and overcome experimental limitations with the slow-growing anaerobe DvH, we reconstituted its predicted SRB Lap system in a strain lacking its native Lap regulatory components (ΔΔ). Our data support the model that DvhA is a cell surface-associated LapA-like adhesin with a N-terminal "retention module" and that DvhA is released from the cell surface upon cleavage by the LapG-like protease DvhG. Further, we demonstrate DVU1020 (named here DvhD) represents a distinct class of c-di-GMP-binding, biofilm-regulating proteins that regulates DvhG activity in response to intracellular levels of this second messenger. This study provides insight into the key players responsible for biofilm formation by DvH, thereby expanding our understanding of Lap-like systems.
Topics: Pseudomonas fluorescens; Sulfates; Adhesins, Bacterial; Biofilms; Carrier Proteins; Cyclic GMP; Bacterial Proteins; Gene Expression Regulation, Bacterial
PubMed: 38498718
DOI: 10.1073/pnas.2320410121