Cover illustration:
Biocontrol to combat Ganoderma using soil-derived fungal isolates identified strains of Talaromyces and Penicillium sp., which possess additional plant growth-promoting traits. The micrograph shows conidiophore bearing conidia of this new biocontrol agent.
(Photo: Muhammad Salahudin Kheirel Anuar, Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang, Selangor, Malaysia)
Autophagy regulates the development of Candida albicans (C. albicans) biofilms and their sensitivity to antifungals. Atg1, a serine/threonine protein kinase, recruits autophagy-related proteins for autophagosome formation. Atg9, the only transmembrane protein, is phosphorylated by Atg1 during autophagy. The specific roles of Atg1 and Atg9 in biofilm formation and resistance of C. albicans remain unclear. The study used RT-qPCR and Western blotting to assess the correlation between Atg1, Atg9 and biofilm formation, XTT reduction assays to evaluate biofilm formation and antifungal resistance, commercial kits to detect reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and autophagy activity, transmission electron microscopy (TEM) to study the morphological changes, protein-protein interaction (PPI) analysis to analyze the interaction between Atg1 and Atg9. Results demonstrated that Atg1 and Atg9 were highly expressed in biofilms than planktonic cells. Biofilm formation, antifungal resistance, MMP and autophagy activity decreased and ROS increased in atg1Δ/Δ and atg9Δ/Δ. TORC1 inhibition with rapamycin rescued the reduced biofilm formation of atg1Δ/Δ and increased antifungal resistance of atg1Δ/Δ and atg9Δ/Δ. PPI analysis and TEM observation indicated that Atg1 interacted with Atg9, which was certified by RT-qPCR and Western blotting. This study suggested that Atg1 interacts with Atg9, activates the autophagy regulating the formation and sensitivity of C. albicans biofilms.
Sugar transporters are of great importance in sensing and transporting varied sugars for cellulase biosynthesis of lignocellulolytic fungi. Nevertheless, the function and the relevant mechanism of sugar transporters in fungal cellulase biosynthesis remain to be explored. Here, putative maltose transporters Mal1, Mal2, Mal3, Mal4, and Mal5 in Trichoderma reesei were investigated. Despite that only the transcriptional abundance of Mal1 was upregulated under cellulase-generating condition, the individual deletion of Mal1, Mal2, Mal3, Mal4, and Mal5 all impaired cellulase biosynthesis. The possible reason for this is that the individual knockout of Mal2, Mal3, Mal4, and Mal5 resulted in no gene expression of Mal1 at 24 h during the cellulase production. The transcriptional analysis showed that the absence of these transporters noticeably inhibited cellulase genes at 24 h, which was then relieved. Interestingly, the individual missing of these maltose transporters significantly retarded the cellular consumption of cellobiose, rather than maltose, and they were distributed in cytoplasm, largely in endoplasmic reticulum (ER). These findings manifested that these putative maltose transporters may be in fact endomembrane cellobiose transporters, influencing fungal cellulase generation probably through Mal1 at the early stage. This research advances the knowledge of endomembrane sugar transporters in fungal cellulase biosynthesis.
This study investigates the biodegradation of methyl parathion, an organophosphate pesticide used in paddy fields. Microbial degradation transforms toxic pesticides into less harmful compounds, influenced by the microbial community in the soil. To isolate different microbial colonies, soil samples from an organophosphorus-treated groundnut field were plated on nutrient agar and MSM with 1% glucose and 0.25 mM methyl parathion. Biodegradation efficiency was determined by estimating the OP hydrolase enzyme activity spectrophotometrically. HPLC was used to quantify residual methyl parathion concentrations in the culture medium. The identified isolate effectively degraded methyl parathion in MSM with 0.25 mM methyl parathion which showed peak hydrolase activity (2.02 µmol/min/mg) after 96 h of incubation and the residual methyl parathion level was determined as 6.2 µmol by HPLC quantification. The efficient isolate was identified as Bacillus cereus by using a 16S rRNA molecular marker and the sequence was subjected to MEGA11 phylogenetic tree construction. The results show that the SM6 clade shared with B. cereus 16S rRNA sequence. B. cereus (SM6) showed substantial enzyme activity and the specific reported opdA gene-coded protein is involved in ATP hydrolysis. This OP hydrolase makes it a strong candidate for bioremediation of methyl parathion. Molecular analysis suggested that the opdA gene, likely chromosomally located, plays a key role in degradation, with potential involvement of the "Cell division protein FtsK" gene responsible for hydrolase activity. Organophosphorus compounds, widely used in agriculture, pose environmental concerns due to their persistence. This study focuses on isolating pesticide-degrading bacteria to expedite bioremediation, aiming for efficient degradation. This study highlights the cross-adaptation phenomenon, where B. cereus strains degrade similar compounds, improving bioremediation strategies.
Cellulase production for hydrolyzing plant cell walls is energy-intensive in filamentous fungi during nutrient scarcity. AMP-activated protein kinase (AMPK), encoded by snf1, is known to be the nutrient and energy sensor in eukaryotes. Previous studies on AMPK identified its role in alternate carbon utilization in pathogenic fungi. However, the precise role of AMPK in cellulase production remains elusive. In the present study, we employed gene-deletion analysis, quantitative proteomics and chemical-genetic approaches to investigate the role of AMPK in cellulase synthesis in Penicillium funiculosum. Gene-deletion analysis revealed that AMPK does not promote transcription and translation but is essential for cellulase secretion in a calcium-dependent manner. Proteomic analysis of the snf1-deleted (Δsnf1) strain confirmed trapped cellulase inside the mycelia and identified HOG1 MAPK activation as the most significant Ca2+-induced signaling event during carbon stress in Δsnf1. Western blot analysis analysis revealed that the phosphorylated HOG1 (P-HOG1)/HOG1 MAPK ratio maintained by Ca2+-signaling/Ca2+-activated AMPK, respectively, forms a secretion checkpoint for cellulases, and disturbing this equilibrium blocks cellulase secretion. The proteomic analysis also indicated a massive increase in mTORC1-activated anabolic pathways during carbon stress in Δsnf1. Our study suggests that AMPK maintains homeostasis by acting as a global repressor during carbon stress.
Antimicrobial resistance is a global health problem as it contributes to high mortality rates in several infectious diseases. To address this issue, engineered nanoparticles/nano-formulations of antibiotics have emerged as a promising strategy. Nanoparticles are typically defined as materials with dimensions up to 100 nm and are made of different materials such as inorganic particles, lipids, polymers, etc. They are widely dispersed in the environment through various consumer products, and their clinical applications are diverse, ranging from contrast agents in imaging to carriers for gene and drug delivery. Nanoparticles can also act as antimicrobial agents either on their own or in combination with traditional antibiotics to produce synergistic effects, earning them the label of “next-generation therapeutics.” They have also shown great effectiveness against multidrug-resistant pathogens responsible for nosocomial infections. However, overexposure or prolonged exposure to sublethal doses of nanoparticles can promote the development of resistance in human pathogens. The resistance can arise from various factors such as genetic mutation, horizontal gene transfer, production of reactive oxygen species, changes in the outer membrane of bacteria, efflux-induced resistance, cross-resistance from intrinsic antibiotic resistance determinants, plasmid-mediated resistance, and many more. Continuous exposure to nanoparticles can also transform an antibiotic-susceptible bacterial pathogen into multidrug resistance. Considering all these, the current review focuses on the mode of action of different heavy metals and nanoparticles and possible mechanisms through which bacteria attain resistance towards these heavy metals and nanoparticles.
Despite several studies documenting secondary metabolite (SM) production by endophytes, their commercial use is often limited owing to the research lacunae in the underlying biosynthetic pathway and the corresponding metabolic flux. Combining epigenetic modulation with RNA-Seq analysis constitutes a promising approach for inducing regulatory gene(s) and thereby identifying their role in SM biosynthesis. Our earlier studies had identified the hypomethylating effects of prednisone in umbelliferone (UMB) (7-hydroxyl coumarin) producing endophytic Fusarium oxysporum isolate, ZzEF8 isolated from Zingiber zerumbet rhizomes. Hypomethylating effect of prednisone (300 μM) in ZzEF8 was validated in present experiments revealing decrease in 5-mC content (0.09 ± 0.01%) in prednisone treated ZzEF8 (PrZzEF8) compared to untreated control (UtZzEF8) (0.36 ± 0.01%). Subsequent RNA-Seq analysis detected transcriptional alterations in PrZzEF8 compared to UtZzEF8. Transcripts with significant differential expression (-2 ≥ fold change (FC) ≥ 2; q-value < 0.05) were detected for 64 transcripts, with 60 upregulated and four downregulated in PrZzEF8. Upregulated differentially expressed genes (DEGs) were annotated as transmembrane transporters, non-ribosomal peptide synthetase (NRPS), Type I and III polyketide synthase (PKS), phytoene dehydrogenase, bifunctional lycopene cyclase/phytoene synthase, geranylgeranyl pyrophosphate synthase and various genes involved in nutrient assimilation, transcription factors and transporters regulating metabolite export. Expression analysis of the selected DEGs were validated by qRT-PCR. Present study proposes UMB biosynthesis through acetate-malonate pathway from acetate units via a pentaketide intermediate in ZzEF8 instead of the phenylpropanoid pathway reported in plants. Study is of relevance as the insights gained into the UMB biosynthetic pathway in ZzEF8 will help in strategizing scale-up of UMB production.
Antibiotic resistance is one of the major health threat for humans, animals, and the environment, according to the World Health Organization (WHO) and the Global Antibiotic-Resistance Surveillance System (GLASS). In the last several years, wastewater/sewage has been identified as potential hotspots for the dissemination of antibiotic resistance and transfer of resistance genes. However, systematic approaches for mapping the antibiotic resistance situation in sewage are limited and underdeveloped. The present review has highlighted all possible perspectives by which the dynamics of ARBs/ARGs in the environment may be tracked, quantified and assessed spatio-temporally through surveillance of wastewater. Moreover, application of advanced methods like wastewater metagenomics for determining the community distribution of resistance at large has appeared to be promising. In addition, monitoring wastewater for antibiotic pollution at various levels, may serve as an early warning system and enable policymakers to take timely measures and build infrastructure to mitigate health crises. Thus, by understanding the alarming presence of antibiotic resistance in wastewater, effective action plans may be developed to address this global health challenge and its associated environmental risks.
The fact that free-living amoebae of the genus Acanthamoeba can live in many different environments causes these protozoa to have different interactions with other microorganisms. Investigation of Acanthamoeba-pathogenic bacteria interaction is important for the discovery of new antibacterial agents that can be used against pathogenic bacteria. In this study, it was aimed to investigate the antibacterial effect of cell-free supernatants obtained from Acanthamoeba against some pathogenic bacteria. One standard strain (Acanthamoeba castellanii ATCC 50373) and one environmental strain (B1) of the genus Acanthamoeba were used in the study. Cell-free supernatants were obtained by centrifuging the axenic cultures (3000 rpm, 5 min) and passing through a sterile filter with a pore diameter of 0.22 µm. The antibacterial effect of cell-free supernatants against five different pathogenic bacteria (Acinetobacter baumannii, Klebsiella pneumoniae, Enterococcus faecalis, Salmonella Typhi, and Salmonella enterica) was investigated by colony counting method. As a result of the study, it was determined that the standard Acanthamoeba cell-free supernatant showed the highest antibacterial effect against E. faecalis (75.79%), while B1 cell-free supernatant showed the highest antibacterial effect against K. pneumoniae (8.5%). The content of the tested Acanthamoeba cell-free supernatants was analyzed by gas chromatography/mass spectrometry in our previous study and was also found to contain major compounds with antibacterial properties. Therefore, it is thought that the metabolites produced by Acanthamoeba can be used as an alternative to existing antimicrobial drugs in the fight against infections caused by some important pathogenic bacteria.
DNA replication origins play a crucial role in cellular division and are evolutionarily conserved across domains. This study investigated the evolutionary transitions of replication origins between archaea and bacteria by analyzing 2733 bacterial and 257 archaeal genomes. Our findings revealed that certain methanogens and bacteria share phylogenetic proximity, suggesting evolutionary interactions across diverse ecological systems. Evolutionary transitions in replication origins may have occurred between gut methanogens and bacteria, haloarchaea (Halogeometricum borinquense DSM 11551 and Halovivax ruber XH-70), halobacteria, and sulfur-reducing archaea. Methanosarcina barkeri (M. barkeri), Methanosaeta thermophila, and Methanococcoides burtonii (M. burtonii) were closely related to respiratory tract bacteria in humans. Methanohalobium evestigatum (M. evestigatum) is strongly linked to the animal gut pathogen Mycoplasma putrefaciens (M. putrefaciens). Several thermophilic hydrogenotrophic methanogens clustered with oral and fish pathogens. Pyrococcus furiosus (P. furiosus) was evolutionarily related to the replication origin of plant pathogens. This study sheds light on the ecological drivers of DNA replication origin evolution and their role in microbial speciation and adaptation. Our findings highlight the influence of mutualistic and parasitic relationships on these evolutionary transitions. It could have significant implications in biotechnology and medicine, such as developing novel antimicrobial strategies and understanding host-pathogen dynamics.
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