Research in microbiology最新文献

Bacteria use type II secretion systems (T2SS) to secrete to their surface folded proteins that confer diverse functions, from nutrient acquisition to virulence. In the Klebsiella species, T2SS-mediated secretion of pullulanase (PulA) requires assembly of a dynamic filament called the endopilus. The inner membrane assembly platform (AP) subcomplex is essential for endopilus assembly and PulA secretion. AP components PulL and PulM interact with each other through their C-terminal globular domains and transmembrane segments. Here, we investigated the roles of their periplasmic helices, predicted to form a coiled coil, in assembly and function of the PulL–PulM complex. PulL and PulM variants lacking these periplasmic helices were defective for interaction in the bacterial two-hybrid (BACTH) assay. Their functions in PulA secretion and assembly of PulG subunits into endopilus filaments were strongly reduced. Interestingly, deleting the cytoplasmic peptide of PulM nearly abolished the function of variant PulMΔN and its interaction with PulG, but not with PulL, in the BACTH assay. Nevertheless, PulL was specifically proteolyzed in the presence of the PulMΔN variant, suggesting that PulM N-terminal peptide stabilizes PulL in the cytoplasm. We discuss the implications of these results for the T2S endopilus and type IV pilus assembly mechanisms.

Bacterial plant pathogens must cope with various environmental conditions and defenses from their hosts for colonization and infection. Heat shock proteins (HSPs) play critical roles in a variety of cellular processes, such as the maintenance of cellular homeostasis in response to environmental stress. However, the significance of HSP40 family protein DnaJ in virulence of plant pathogenic bacteria has not yet been explored. To elucidate the function of DnaJ in Pseudomonas cichorii JBC1 (PcJBC1) virulence, we generated dnaJ-deficient (JBC1^ΔdnaJ) mutant using CRISPR-CAS9. The disease severity by JBC1^ΔdnaJ was significantly reduced compared with wild-type (WT) and dnaJ-complemented (JBC1^ΔdnaJ + pdnaJ) strain. The defect of DnaJ suppressed siderophore production, extracellular DNA (eDNA) release, biofilm formation, and swarming motility and made the strain sensitive to stresses such as heat and H₂O₂. The supplementation of eDNA recovered the amount of biofilm formation by JBC1^ΔdnaJ. Our results indicate that DnaJ is a key player in the survival and colonization of bacterial plant pathogens on plant surfaces as well as bacterial responses to abiotic and biotic stresses, which are determinative to cause disease. These findings can broaden our understanding of plant and bacterial pathogen interactions.

Endolysins have garnered significant attention as a potential alternative to antibiotics in aquaculture, mainly for combating Vibrio spp., Gram-negative pathogens responsible for infectious outbreaks. However, endolysin effectiveness against Gram-negative bacteria is limited due to the outer membrane's poor permeability. The combat against marine pathogens poses an additional challenge of finding endolysins that retain their activity in high ionic strength conditions. Thus, this study aimed to demonstrate that certain endolysins retain muralytic activity in seawater and also evaluated outer membrane permeabilizers as endolysin adjuvants. The effectiveness of KZ144 and LysPA26 endolysins, along with EDTA and oregano essential oil, was evaluated against Vibrio parahaemolyticus ATCC-17802 in natural seawater. Results revealed the muralytic activity of both endolysins in seawater. However, the endolysins appeared to counteract the permeabilizers' effect during the initial bactericidal assays. Further investigations revealed that the observed effect was not antagonistic. After the permeabilizer action, V. parahaemolyticus likely used endolysins as a growth substrate. Endolysins may not play an indifferent role if they fail to exert a bactericidal effect. Instead, they can serve as a substrate for fast-growing bacteria, such as V. parahaemolyticus, increasing bacterial density. It should be considered a potential drawback of endolysins' proteinaceous nature as bactericidal agents.

Sphingolipids (SLs) are essential to fungal survival and represent a major class of structural and signaling lipids. Unique SL structures and their biosynthetic enzymes in filamentous fungi make them an ideal drug target. Several studies have contributed towards the functional characterization of specific SL metabolism genes, which have been complemented by advanced lipidomics methods which allow accurate identification and quantification of lipid structures and pathway mapping. These studies have provided a better understanding of SL biosynthesis, degradation and regulation networks in filamentous fungi, which are discussed and elaborated here.

The high incidence of persistent multidrug resistant bacterial infections is a worldwide public health burden. Alternative strategies are required to deal with such issue including the use of drugs with anti-virulence activity. The application of nanotechnology to develop advanced Nano-materials that target quorum sensing regulated virulence factors is an attractive approach. Synthesis of ascorbic acid Nano-emulsion (ASC-NEs) and assessment of its activity in vitro against the virulence factors and its protective ability against pathogenesis as well as the effect against expression of quorum sensing genes of Pseudomonas aeruginosa and Staphylococcus aureus isolates. Ascorbic acid Nano-emulsion was characterized by DLS Zetasizer Technique, Zeta potential; Transmission Electron Microscopy (TEM) and Fourier transform infrared spectroscopy (FT-IR). The antibacterial activity of ASC-NEs was tested by the broth microdilution method and the activity of their sub-MIC against the expression of quorum sensing controlled virulence was investigated using phenotypic experiments and RT-PCR. The protective activity of ASC-NEs against P. aeruginosa as well as S. aureus pathogenesis was tested in vivo. Phenotypically, ASC-NEs had strong virulence inhibitory activity against the tested bacteria. The RT-PCR experiment showed that it exhibited significant QS inhibitory activity. The in vivo results showed that ASC-NEs protected against staphylococcal infection, however, it failed to protect mice against Pseudomonal infection. These results suggest the promising use of nanoformulations against virulence factors in multidrug resistant P. aeruginosa and S. aureus. However, further studies are required concerning the potential toxicity, clearance and phamacokinetics of the nanoformulations.

Archaea are microorganisms with great ability to colonize some of the most inhospitable environments in nature, managing to survive in places with extreme characteristics for most microorganisms. Its proteins and enzymes are stable and can act under extreme conditions in which other proteins and enzymes would degrade. These attributes make them ideal candidates for use in a wide range of biotechnological applications. This review describes the most important applications, both current and potential, that archaea present in Biotechnology, classifying them according to the sector to which the application is directed. It also analyzes the advantages and disadvantages of its use.

Persister cells and biofilms are associated with chronic urinary infections which are more critical when generated by multi-drug resistant bacteria. In this context, joint administration of phages and antibiotics has been proposed as an alternative approach, since it may decrease the probability to generate resistant mutants to both agents. In this work, we exposed cultures of uropathogenic Escherichia coli conjunctly to antibiotics and phages. We determined that MLP2 combined with antibiotics eradicates persister cells. Similarly, MLP1 and MLP3 impact viability of biofilm-forming cells when administered with ampicillin. Our findings suggest a feasible prophylactic and therapeutic use of these non-transducing phages.

The Resistance-nodulation-division (RND)-type AcrAB-TolC efflux pump contributes to multidrug resistance in Gram-negative bacteria. Recently, the bacterium Photorhabdus laumondii TT01 has emerged as a goldmine for novel anti-infective drug discovery. Outside plants, Photorhabdus is the only Gram-negative known to produce stilbene-derivatives including 3,5-dihydroxy-4-ethyl-trans-stilbene and 3,5-dihydroxy-4-isopropyl-trans-stilbene (IPS). IPS is a bioactive polyketide which received considerable attention, mainly because of its antimicrobial properties, and is currently in late-stage clinical development as a topical treatment for psoriasis and dermatitis. To date, little is known about how Photorhabdus survives in the presence of stilbenes. We combined genetic and biochemical approaches to assess whether AcrAB efflux pump exports stilbenes in P. laumondii. We demonstrated that the wild-type (WT) exerts an antagonistic activity against its derivative ΔacrA mutant, and that is able to outcompete it in a dual-strain co-culture assay. The ΔacrA mutant also showed high sensitivity to 3,5-dihydroxy-4-ethyl-trans-stilbene and IPS as well as decreased IPS concentrations in its supernatant comparing to the WT. We report here a mechanism of self-resistance against stilbene derivatives of P. laumondii TT01, which enables these bacteria to survive under high concentrations of stilbenes by extruding them out via the AcrAB efflux pump.

Myxobacteria are Gram-negative eubacteria and they thrive in a variety of habitats including soil rich in organic matter, rotting wood, animal dung and marine environment. Myxobacteria are a promising source of new compounds associated with diverse bioactive spectrum and unique mode of action. The genome information of myxobacteria has revealed many orphan biosynthetic pathways indicating that these bacteria can be the source of several novel natural products. In this review, we highlight the biology of myxobacteria with emphasis on their habitat, life cycle, isolation methods and enlist all the bioactive secondary metabolites purified till date and their mode of action.

Cells have evolved strategies to safeguard their genome integrity. We describe a mechanism to counter double strand breaks in the chromosome that involves the protection of an essential housekeeping enzyme from external agents. YacG is a DNA gyrase inhibitory protein from Escherichia coli that protects the bacterium from the cytotoxic effects of catalytic inhibitors as well as cleavage-complex stabilizers of DNA gyrase. By virtue of blocking the primary DNA binding site of the enzyme, YacG prevents the accumulation of double strand breaks induced by gyrase poisons. It also enables the bacterium to resist the growth-inhibitory property of novobiocin. Gyrase poison-induced oxidative stress upregulates YacG production, probably as a cellular response to counter DNA damage. YacG-mediated protection of the genome is specific for gyrase targeting agents as the protection is not observed from the action of general DNA damaging agents. YacG also intensifies the transcription stress induced by rifampicin substantiating the importance of gyrase activity during transcription. Although essential for bacterial survival, DNA gyrase often gets entrapped by external inhibitors and poisons, resulting in cell death. The existence of YacG to specifically protect an essential housekeeping enzyme might be a strategy adopted by bacteria for competitive fitness advantage.