Appl. Microbiol., Volume 6, Issue 4 (April 2026) – 8 articles

Antimicrobial resistance (AMR) has become one of the top ten global public health threats. Many countries have recognized the societal and economic burden of AMR. AMR has reduced the effectiveness of antimicrobial therapies, and this results in high mortality, morbidity, and health care expenditure. Like all the other developing countries, South Africa (SA) falls under the same ambiguous management system of antimicrobials. A lot of research focused on the global public health threat “AMR”. However, studies on AMR in wastewater are not yet enough, even though they are beginning to gain momentum. This paper highlights the imperatives of surveying AMR pathogens in wastewater since wastewaters are consecrated as hotspots for the dissemination and propagation of AMR genes. RNA was extracted from the untreated wastewater samples collected from the Tshwane district in Gauteng province, SA. Metatranscriptomics analysis was proposed for the analysis and profiling of AMR genes present in the wastewater. A total of 39 AMR gene families and 39 AMR drug classes were detected across 17 samples. The Metatranscriptomics approach discussed in this paper demonstrates the importance of wastewater surveillance, as it can be used as an early detecting system for communicable diseases and for monitoring wastewater. Full article

This study evaluated the performance of Fourier-transform infrared (FTIR) spectroscopy for identifying spectral signatures associated with two key traits of Listeria monocytogenes: serogroup classification and biofilm-forming capacity. A total of 100 strains, previously serogrouped by PCR and categorized as high, intermediate, or low biofilm producers, were analyzed. Whole-genome sequencing was performed, and comparative genomics was conducted at core-genome, pangenome, and whole-genome (k-mer) levels to determine which genomic representation best reflected the phenotypes. Strains were typed using Fourier-Transform Infrared (FTIR Biotyper^® system from Bruker Daltonics GmbH and Co., Bremen, Germany) with five technical replicates. Spectral data from the polysaccharide region (1300–800 cm⁻¹) were extracted and used to train twelve statistical models within a machine learning pipeline combined with cross-validation to predict four serogroups and three biofilm clusters from 501 spectral variables. Genomic analyses showed strong concordance between population structure and serogroup, whereas biofilm formation displayed only weak genomic association, explaining less than 0.1% of genomic variance (PERMANOVA R² ≤ 0.001). Penalized discriminant analysis achieved the highest performance for serogroup prediction (overall accuracy 97.2%), while the k-nearest neighbor model performed best for biofilm prediction (74.8%). Two dedicated R Shiny applications were developed to facilitate model use. Overall, FTIR spectroscopy coupled with machine learning can provide a rapid and cost-effective alternative to PCR, genomic analyses, and in vitro assays for phenotypic trait prediction. Full article

Clostridium butyricum is a well-known Gram-positive, spore-forming, obligate anaerobic, and butyrate-producing bacterium with a few species of next-generation probiotic strains. By far, the most well-known strain is Clostridium butyricum CBM588 (also known as MIYAIRI 588). This strain has gained significant attention for its therapeutic potential across a variety of human health conditions. Preclinical studies have shown its ability to stabilize gut microbiota, enhance short-chain fatty acid (SCFA) production, and modulate immune responses, which contribute to its therapeutic effects in conditions such as ulcerative colitis, allergies, and cancer. We examined 28 interventional clinical trials and 7 observational studies investigating the effect of Clostridium butyricum strains. These studies have supported the findings of preclinical trials and demonstrated symptom improvement and immune modulation in diverse conditions. Clostridium butyricum CBM588 has shown efficacy in managing gastrointestinal diseases, such as acute gastroenteritis and inflammatory bowel disease, and has also proven beneficial in immune modulation, as evidenced by its positive effects in allergic rhinitis and cancer immunotherapy. Additionally, CBM588 has been reported to have a favorable safety and tolerability profile in various patient populations, including children, adults, and critically ill patients. Despite these promising results, clinical studies face limitations such as small sample sizes, varied protocols, and short study durations. Future well-designed, large-scale trials are necessary to further validate the long-term safety and efficacy of Clostridium butyricum in clinical practice. Full article

Synthetic dyes, such as methylene blue (MB), constitute a major category of environmental pollutants due to their toxicity, persistence, and resistance to standard treatment methods. In this study, Bacillus cereus BC WW Saida was isolated from the heavily polluted Saida dumpsite in Lebanon and evaluated for its MB degradation efficiency. The isolate was identified through whole-genome sequencing, which revealed the presence of key enzymatic systems involved in azo dye degradation. Under optimized conditions, the strain achieved 82% decolorization, as determined by optical density measurements using a microplate reader. The process was further examined using High-Performance Liquid Chromatography (HPLC), which revealed a significant reduction in the original dye peak and the emergence of new intermediate products. These findings suggest the strong biodegradation capability of B. cereus BC WW Saida isolated from contaminated environments and highlight its potential application in the eco-friendly treatment of azo dye-contaminated wastewater. Full article

Background: The ability of synthetic plastics to persist in the environment and the accumulation of microplastics has intensified the need to explore biological mechanisms capable of interacting with, and possibly degrading, polymeric materials. Microbial enzymes that have extensive catalytic flexibility represent promising candidates in this context. Aim: This study set out to examine the molecular interaction patterns and dynamical stability of Pseudomonas aeruginosa elastase (LasB) with representative structural fragments of typical synthetic plastics to assess the suitability of the enzyme to polymer-derived substrates. Methods: The crystallographic structure of LasB (PDB ID: 1EZM) was retrieved from the Protein Data Bank and pre-prepared with the help of AutoDock4.2.6 Tools. Those polymer-derived ligands that were associated with the major industrial plastics such as polyamide (PA), polyvinyl chloride (PVC), polycarbonate (PC), poly-ethylene terephthalate (PET), polymethyl methacrylate (PMMA), and polyurethane (PUR) were retrieved in the PubChem database and geometrically optimized with the help of the MMFF94 force field. AutoDock Vina, with a specific grid box around the catalytic pocket, including Zn²⁺ ion, was used to perform molecular docking simulations. PyMOL and BIOVIA Discovery Studio software were used to analyze binding conformations, interaction residues and types of intermolecular contacts. Phosphoramidon, a known metalloprotease inhibitor, served as a positive control to confirm the docking protocol. Additional assessment of the structural stability and conformational behavior of the enzyme–ligand complexes was conducted by molecular dynamics (MD) simulations with the Desmond engine and explicit solvent model in a 50 ns trajectory using the OPLS4 force field. RMSD, RMSF, radius of gyration, hydrogen bonding analysis and solvent accessibility parameters were used to measure structural stability. Results: The docking experiment showed varying binding affinities with the test polymers. Polycarbonate (−5.774 kcal/mol) and polyurethane (−5.707 kcal/mol) had the highest in-teractions with the LasB catalytic pocket, polyamide (−5.277 kcal/mol) and PET (−4.483 kcal/mol) followed PMMA and PVC, which had weaker affinities. The following were the important residues involved in interaction networks: Glu141, His140, Val137, Arg198, Tyr114, and Trp115 that were implicated in interaction networks with hydrophobic interactions, π-cation interactions and van der Waals forces that were the major stabilization forces. MD simulations had stabilized complexes, and RMSD values were found to be within acceptable ranges of stability, and ligand-specific changes (around 1.0-3.2 A), which is also in line with stable protein-ligand systems. Phosphoramidon used as a positive control had an RMSD of 1.205 A which is within this stability range. PCA determined various ligand-bound conformational states of LasB with PA in com-pact state, PC and PVC in intermediate states and PUR, PMMA and PET in ex-panded conformations, indicating structur-al stability and adaptability of the binding pocket. Conclusion: These findings show that LasB has a structurally flexible catalytic pocket that can accommodate a wide range of polymer-derived ligands. These results offer an insight into the recognition of enzymes with polymers at the molecular level and also indicate that LasB might help in the interaction of microorganisms with synthetic plastics in environmental systems. Full article

Siderophores are high-affinity iron-chelating metabolites that underpin microbial survival in iron-limited environments and play central roles in metal homeostasis, ecological competition, and pathogenesis. Traditionally viewed as dedicated Fe(III) scavengers, siderophores are now recognized as structurally and functionally versatile coordination agents whose donor-set architectures—particularly catecholate and α-hydroxycarboxylate motifs—permit conditional interactions beyond iron. In iron- and boron-rich niches, especially marine and mildly alkaline systems where borate availability increases, certain siderophores are chemically capable of forming reversible borate complexes through cis-diol coordination. Although Fe(III) exhibits substantially higher thermodynamic affinity and remains the primary biological target, boron binding represents a predictable secondary property arising from shared oxygen-donor chemistry. This dynamic interplay allows siderophores to cycle between iron-bound, boron-bound, and apo states depending on local redox conditions, pH, and metal availability. Here, we synthesize current knowledge on the structural classes of microbial siderophores, their transport and regulatory mechanisms, and emerging evidence for boron coordination within catecholate and carboxylate systems. By integrating coordination chemistry with microbial ecology, we propose an expanded model in which siderophores function not only as iron acquisition molecules but also as modulators of boron speciation and environmental sensing. This functional plasticity positions siderophores at the intersection of iron and boron biogeochemical cycles and highlights new directions for understanding microbial adaptation in complex metal-rich environments. Full article

Quorum sensing (QS) represents a promising target for anti-virulence therapy; however, effective pharmacological intervention requires a detailed understanding of regulatory network architecture and environmental context. In Klebsiella pneumoniae, the orphan LuxR-type receptor SdiA lacks a cognate LuxI synthase and instead detects exogenous acyl-homoserine lactones (AHLs), positioning it as an inter-species signal integrator. Here, we demonstrate that SdiA functions as a context-dependent regulator whose impact on biofilm formation and virulence gene expression is gated by environmental AHL availability. Using isogenic ΔluxS, ΔsdiA, and ΔluxSΔsdiA mutants in a clinical bloodstream isolate, we show that under AHL-limited conditions, SdiA promotes baseline biofilm development, whereas in the presence of exogenous C6-HSL, it restrains excessive biofilm maturation. Two-way ANOVA confirmed significant genotype, treatment, and interaction effects, establishing that SdiA-mediated regulation is signal contingent. We further investigated the halogenated thiolactone meta-bromo-thiolactone (mBTL), previously described as a QS inhibitor in Pseudomonas aeruginosa. In K. pneumoniae, mBTL acts as a context-selective modulator rather than a simple inhibitor. Under AHL-limited conditions, mBTL phenocopied ΔsdiA, reducing biofilm formation and inducing overlapping transcriptional profiles. In contrast, under AHL-replete conditions, mBTL opposed SdiA-dependent gene expression, consistent with competitive antagonism of ligand-bound receptor. RNA-seq analysis revealed substantial concordance between ΔsdiA and WT + mBTL under AHL-free conditions, with the inversion of transcriptional directionality in the presence of C6-HSL. The findings redefine SdiA as a conditional quorum-sensing integrator and identify mBTL as a ligand-context-dependent modulator of LuxR-type signaling. Our results highlight the necessity of evaluating anti-virulence compounds across relevant signal environments and introduce receptor state-selective modulation as a strategic framework for targeting hybrid quorum-sensing systems in polymicrobial pathogens. Full article

Helicobacter pylori is a highly prevalent pathogen associated with chronic gastritis, peptic ulcers, and gastric cancer. Treatment is increasingly challenging due to antibiotic resistance and adverse effects that can reduce adherence. These limitations have encouraged the exploration of complementary strategies. This study evaluated the in vitro antibacterial activity of selected probiotic strains and synbiotic formulations containing inulin against clinical isolates of H. pylori. Isolates obtained from gastric biopsies were identified by MALDI-TOF. Four probiotic strains (Lacticaseibacillus casei, Lacticaseibacillus rhamnosus, Limosilactobacillus fermentum, and Streptococcus thermophilus) were tested individually and as a mixed culture, both alone and combined with inulin. Antibacterial activity was assessed using the agar well diffusion method under microaerophilic conditions after 72 h of incubation at 37 °C. Variable inhibitory effects were observed, with L. fermentum (8.08 ± 1.98 mm) and the probiotic mixture (7.92 ± 0.90 mm) showing greater activity, while S. thermophilus exhibited limited inhibition. The addition of low-dose inulin (3 mg/mL) was associated with increased inhibition by the probiotic mixture (9.58 ± 1.51 mm), whereas higher concentrations did not enhance this effect. These findings indicate that certain probiotic and synbiotic formulations exhibit in vitro activity against H. pylori and warrant further investigation as complementary approaches. Full article