Microbiological research最新文献

Background

This study aimed to characterize three KPC variants (KPC-33, KPC-100, and KPC-201) obtained from a clinical isolate of Pseudomonas aeruginosa (#700), along with two induced strains C109 and C108.

Methods

Genomic DNAs of #700 (ST235), C109 (ST463), and C108 (ST1076) were sequenced using Illumina and Oxford Nanopore technologies. The transferability and stability of the plasmid was assessed through conjugation experiments and plasmid stability experiments, respectively. Minimum inhibitory concentrations of bacterial strains were determined using broth microdilution methods. In vitro induction was performed using ceftazidime-avibactam (CZA) at concentrations of 6/4 µg/ml. Linear genomic alignments were visualized using Easyfig, and protein structure modeling of the novel KPC variant (KPC-201) was conducted using PyMol.

Results

The plasmids carrying the KPC variants in the three CZA-resistant strains (C109, C108, and #700) had sizes of 39,251 bp (KPC-100), 394,978 bp (KPC-201), and 48,994 bp (KPC-33). All three plasmids belonged to the IncP-like incompatibility (Inc) groups, and the plasmid exhibited relatively high plasmid stability, KPC-33 and KPC-201-harboring plasmids were successfully transferred to the recipient strain P. aeruginosa PAO1^rifR. The genetic environments of the three bla_KPC genes differed from each other. The mobile elements of the three bla_KPC genes were as follows, TnAS1-IS26-ΔISKpn27-bla_KPC-33-ISKpn6-IS26, IS6-ΔISKpn27-bla_KPC-100-ISKpn6-IS26-Tn3-IS26, and IS6100-ISKpn27-bla_KPC-201-ISKpn6-TnAS1. Notably, the length of ΔISKpn27 upstream of the bla_KPC-33 and bla_KPC-100 genes were remarkably short, measuring 114 bp and 56 bp, respectively, deviating significantly from typical lengths associated with ISKpn27 elements. Moreover, the novel KPC variant, KPC-201, featured a deletion of amino acids LDR at positions 161–163 in KPC-3, resulting in a looser pocket structure contributing to its avibactam resistance.

Conclusions

KPC-201, identified as a novel KPC variant, exhibits resistance to CZA. The presence of multiple mobile elements surrounding the bla_KPC-variant genes on stable plasmids is concerning. Urgent preventive measures are crucial to curb its dissemination in clinical settings.

Changing climate creates a challenge to agricultural sustainability and food security by changing patterns of parameters like increased UV radiation, rising temperature, altered precipitation patterns, and higher occurrence of extreme weather incidents. Plants are vulnerable to different abiotic stresses such as waterlogging, salinity, heat, cold, and drought in their natural environments. The prevailing agricultural management practices play a major role in the alteration of the Earth's climate by causing biodiversity loss, soil degradation through chemical and physical degradation, and pollution of water bodies. The extreme usage of pesticides and fertilizers leads to climate change by releasing greenhouse gases (GHGs) and depositing toxic substances in the soil. At present, there is an urgent need to address these abiotic stresses to achieve sustainable growth in agricultural production and fulfill the rising global food demand. Several types of bacteria that are linked with plants can increase plant resistance to stress and lessen the negative effects of environmental challenges. This review aims to explore the environmentally friendly capabilities and prospects of multi-trait plant growth-promoting bacteria (PGPB) in the alleviation of detrimental impacts of harsh environmental conditions on plants.

Klebsiella pneumoniae (Kp) is increasingly recognized as a reservoir for a range of antibiotic resistance genes and a pathogen that frequently causes severe infections in both hospital and community settings. In this study, we have identified a novel mechanism of conjugative transfer of a non-conjugative virulence plasmid through the formation of a fusion plasmid between the virulence plasmid and a novel 59,162 bp IncN- plasmid. This plasmid was found to be a multidrug-resistance (MDR) plasmid and carried a T4SS cluster, which greatly facilitated the efficient horizontal transfer of the fusion plasmid between Kp strains. The fused virulence plasmid conferred the resistance of serum killing and macrophage phagocytosis to the transconjugants. Importantly, this plasmid was shown to be essential for Kp virulence in a mouse model. Mechanistic analysis revealed that the virulence factors encoded by this virulence plasmid contributed to resistance to in vivo clearance and induced a high level of proinflammatory cytokine IL-1β, which acts as an inducer for more neutrophil recruitment. The transmission of the fusion plasmid in Kp has the potential to convert it into both MDR and hypervirulent Kp, accelerating its evolution, and posing a serious threat to human health. The findings of this study provide new insights into the rapid evolution of MDR and hypervirulent Kp in recent years.

Melatonin administration is an environmentally effective strategy to mitigate apple replant disease (ARD), but its mechanism of action is unknown. This study investigated the protective effect of melatonin on ARD and the underlying mechanism. In field experiments, melatonin significantly reduced phloridzin levels in apple roots and rhizosphere soil. A correlation analysis indicated that a potential antagonistic interaction between melatonin and phloridzin was crucial for improving soil physicochemical properties, increasing the diversity of endophytic bacterial communities in roots of apple seedlings, and promoting mineral element absorption by the plants. Melatonin also reduced the abundance of Fusarium in roots. The ability of melatonin to reduce phloridzin levels both in soil and in plants was also demonstrated in a pot experiment. Azovibrio were specifically recruited in response to melatonin and their abundance was negatively correlated with phloridzin levels. Fusarium species that have a negative impact on plant growth were also inhibited by melatonin. Our results show that melatonin improves the rhizosphere environment as well as the structure of the endophytic microbiota community, by reducing phloridzin levels in rhizosphere soil and roots. These regulatory effects of melatonin support its use to improve the physiological state of plants under ARD conditions and thereby overcome the barriers of perennial cropping systems.

The airborne fungus Aspergillus fumigatus is a major pathogen that poses a serious health threat to humans by causing aspergillosis. Azole antifungals inhibit sterol 14-demethylase (encoded by cyp51A), an enzyme crucial for fungal cell survival. However, the most common mechanism of azole resistance in A. fumigatus is associated with the mutations in cyp51A and tandem repeats in its promoter, leading to reduced drug-enzyme interaction and overexpression of cyp51A. It remains unknown whether post-translational modifications of Cyp51A contribute to azole resistance. In this study, we report that the Cyp51A expression is highly induced upon exposure to itraconazole, while its ubiquitination level is significantly reduced by itraconazole. Loss of the ubiquitin-conjugating enzyme Ubc7 confers resistance to multiple azole antifungals but hinders hyphal growth, conidiation, and virulence. Western blot and immunoprecipitation assays show that deletion of ubc7 reduces Cyp51A degradation by impairing its ubiquitination, thereby leading to drug resistance. Most importantly, the overexpression of ubc7 in common environmental and clinical azole-resistant cyp51A isolates partially restores azole sensitivity. Our findings demonstrate a non-cyp51A mutation-based resistance mechanism and uncover a novel role of post-translational modification in contributing to azole resistance in A. fumigatus.

The endoplasmic reticulum-mitochondrial encounter structure (ERMES) complex is known to play crucial roles in various cellular processes. However, its functional significance in filamentous fungi, particularly its impact on deoxynivalenol (DON) biosynthesis in Fusarium graminearum, remains inadequately understood. In this study, we aimed to investigate the regulatory function of the ERMES complex in F. graminearum. Our findings indicate significant changes in mitochondrial morphology of ERMES mutants, accompanied by decreased ATP content and ergosterol production. Notably, the toxisome formation in the ERMES mutant ΔFgMDM10 was defective, resulting in a substantial reduction in DON biosynthesis. This suggests a pivotal role of ERMES in toxisome formation, as evidenced by the pronounced inhibition of toxisome formation when ERMES was disrupted by boscalid. Furthermore, ERMES deficiencies were shown to diminish the virulence of F. graminearum towards host plants significantly. In conclusion, our results suggest ERMES is an important regulator of mitochondrial morphology, DON biosynthesis, and toxisome formation in F. graminearum.

This study investigates the molecular mechanisms underlying salt stress responses in plants, focusing on the regulatory roles of OsNAM2, a gene influenced by the plant growth-promoting rhizobacterium Bacillus amyloliquefaciens (SN13). The study examines how SN13-modulated OsNAM2 enhances salt tolerance in Arabidopsis through physiological, biochemical, and molecular analyses. Overexpression of OsNAM2, especially with SN13 inoculation, improves germination, seedling growth, root length, and biomass under high NaCl concentrations compared to wild-type plants, indicating a synergistic effect. OsNAM2 overexpression enhances relative water content, reduces electrolyte leakage and malondialdehyde accumulation, and increases proline content, suggesting better membrane integrity and stress endurance. Furthermore, SN13 and OsNAM2 overexpression modulates essential metabolic genes involved in glycolysis, the pentose phosphate pathway, and the tricarboxylic acid cycle, facilitating metabolic adjustments crucial for salt stress adaptation. The interaction of OsNAM2 with SUS, facilitated by SN13, suggests enhanced sucrose metabolism efficiency, providing substrates for protective responses. Additionally, OsNAM2 plays a regulatory role in the ABA signaling pathway through significant protein-protein interactions like with AFP2. This study highlights the intricate interplay between SN13-responsive OsNAM2 and key signaling pathways, suggesting strategies for enhancing crop salt tolerance through targeted genetic and microbial interventions

Mannitol, one of the most widespread sugar alcohols, has been integral to daily human life for two centuries. Global population growth and competition for freshwater, food, and land have prompted a shift in the fermentation industry from terrestrial to marine raw materials. Mannitol is a readily available carbohydrate in brown seaweed from the ocean and possess a higher reducing power than glucose, making it a promising substrate for biological manufacturing. This has spurred numerous explorations into converting mannitol into high-value chemicals. Researchers have engineered microorganisms to utilize mannitol in various synthetic biological applications, including: (1) employing mannitol as an inducer to control the activation and deactivation of genetic circuits; (2) using mannitol as a carbon source for synthesizing high-value chemicals through biomanufacturing. This review summarizes the latest advances in the application of mannitol in synthetic biology.

Aim of review

The aim is to present a thorough and in-depth knowledge of mannitol, a marine carbon source, and then use this carbon source in synthetic biology to improve the competitiveness of biosynthetic processes. We outlined the methods and difficulties of utilizing mannitol in synthetic biology with a variety of microbes serving as hosts. Furthermore, future research directions that could alleviate the carbon catabolite repression (CCR) relationship between glucose and mannitol are also covered.

Expected contributions of review

Provide an overview of the current state, drawbacks, and directions for future study on mannitol as a carbon source or genetic circuit inducer in synthetic biology.

Rhizobia interact with leguminous plants in the soil to form nitrogen fixing nodules in which rhizobia and plant cells coexist. Although there are emerging studies on rhizobium-associated nitrogen fixation in cereals, the legume-rhizobium interaction is more well-studied and usually serves as the model to study rhizobium-mediated nitrogen fixation in plants. Rhizobia play a crucial role in the nitrogen cycle in many ecosystems. However, rhizobia are highly sensitive to variations in soil conditions and physicochemical properties (i.e. moisture, temperature, salinity, pH, and oxygen availability). Such variations directly caused by global climate change are challenging the adaptive capabilities of rhizobia in both natural and agricultural environments. Although a few studies have identified rhizobial genes that confer adaptation to different environmental conditions, the genetic basis of rhizobial stress tolerance remains poorly understood. In this review, we highlight the importance of improving the survival of rhizobia in soil to enhance their symbiosis with plants, which can increase crop yields and facilitate the establishment of sustainable agricultural systems. To achieve this goal, we summarize the key challenges imposed by global climate change on rhizobium-plant symbiosis and collate current knowledge of stress tolerance-related genes and pathways in rhizobia. And finally, we present the latest genetic engineering approaches, such as synthetic biology, implemented to improve the adaptability of rhizobia to changing environmental conditions.

2,4-dihydroxybutyric acid (DHB) and 2-keto-4-hydroxybutyrate (OHB) are non-natural molecules obtained through synthetic pathways from renewable carbon source. As they are structurally similar to lactate and pyruvate respectively, they could possibly interfere with the metabolic network of Escherichia coli. In fact, we showed that DHB can be easily oxidized by the membrane associated L and D-lactate dehydrogenases encoded by lldD, dld and ykgF into OHB, and the latter being cleaved into pyruvate and formaldehyde by several pyruvate-dependent aldolases, with YagE being the most effective. While formaldehyde was readily detoxified into formate, Escherichia coli K12 MG1655 strain failed to grow on DHB despite of the production of pyruvate. To find out the reason for this failure, we constructed a mutant strain whose growth was rendered dependent on DHB and subjected this strain to adaptive evolution. Genome sequencing of the adapted strain revealed an essential role for ygbI encoding a transcriptional repressor of the threonate operon in this DHB-dependent growth. This critical function was attributed to the derepression of ygbN encoding a putative threonate transporter, which was found to exclusively transport the D form of DHB. A subsequent laboratory evolution was carried out with E. coli K12 MG1655 deleted for ΔygbI to adapt for growth on DHB as sole carbon source. Remarkably, only two additional mutations were disclosed in the adapted strain, which were demonstrated by reverse engineering to be necessary and sufficient for robust growth on DHB. One mutation was in nanR encoding the transcription repressor of sialic acid metabolic genes, causing 140-fold increase in expression of nanA encoding N-acetyl neuraminic acid lyase, a pyruvate-dependent aldolase, and the other was in the promoter of dld leading to 14-fold increase in D-lactate dehydrogenase activity on DHB. Taken together, this work illustrates the importance of promiscuous enzymes in underground metabolism and moreover, in the frame of synthetic pathways aiming at producing non-natural products, these underground reactions could potentially penalize yield and title of these bio-based products.