Search Results (244)

Brucellosis, a zoonotic disease caused by Brucella, requires rapid, accurate, and sensitive diagnostic methods for effective prevention and control. This study presents the development of a fluorescence microsphere immunochromatographic assay (QDMs-ICA) for detecting anti-Brucella antibodies in bovine serum and milk. Lipopolysaccharide (LPS) from the Brucella abortus strain A19 was immobilized on the nitrocellulose membrane (NC membrane) as the test line (T-line), while rabbit anti-SPG polyclonal antibody was applied as the control line (C-line). Recombinant streptococcal protein G conjugated with quantum dot microspheres (QDMs-SPG) served as the detection conjugate. After optimizing the preparation parameters of QDMs-ICA, the method demonstrated sensitivities of approximately 0.98 IU/mL for bovine serum and 1.56 IU/mL for milk. No cross-reactions were observed with antibody-positive sera from Coxiella burnetii, Mycobacterium avium paratuberculosis, Mycobacterium tuberculosis, Chlamydia abortus, Bacillus anthracis, Escherichia coli O157:H7, Vibrio cholerae or Salmonella, indicating excellent specificity. In intra- and inter-batch repeatability tests, the coefficient of variation (CV) remained below 15%, confirming good reproducibility. The detection limit remained stable after storage at 37 °C for 7 days. Parallel testing of 150 bovine serum samples and 80 milk samples showed a high degree of concordance with the ID-VET commercial kit, with coincidence rates of 97.3% and 96.3%, respectively. These results demonstrate that QDMs-ICA offers high specificity, sensitivity, repeatability, and reliability, making it an effective tool for the rapid detection and epidemiological monitoring of brucellosis. Full article

Non-O1/non-O139 Vibrio cholerae strains are widely distributed in aquatic environments worldwide and are increasingly recognized as potential reservoirs of antimicrobial resistance and virulence-associated determinants. In this study, we performed an integrated phenotypic and genomic analysis of 116 V. cholerae isolates collected in 2024 from environmental and clinical sources in Jiaxing, China, including 106 non-O1/non-O139 isolates, 9 O1 isolates, and 1 O139 isolate. Antimicrobial susceptibility testing showed that most isolates remained susceptible to β-lactam/β-lactamase inhibitor combinations, third-generation cephalosporins, carbapenems, and tigecycline, whereas resistance was more frequently observed for ampicillin, streptomycin, nalidixic acid, and ciprofloxacin. Based on the non-susceptibility criteria of Maitrakas et al., 19 of 116 isolates (16.4%) were classified as multidrug-resistant, whereas none met the definition of extensively drug-resistant. Genomic analysis identified diverse resistance determinants, including plasmid-mediated quinolone resistance genes (qnrVC variants) and quinolone resistance-determining region mutations in gyrA and parC. Virulence-associated genes showed heterogeneous distributions: core regulatory and hemolysis-related genes were highly prevalent, whereas classical cholera toxin genes were largely absent. Several accessory virulence factors, including the RTX toxin operon, chxA, ninth, and makA, were detected in subsets of isolates. Core genome multilocus sequence typing revealed substantial genetic diversity, with environmental and clinical isolates distributed across multiple lineages and showing no clear clustering by isolation source. Overall, these data demonstrate the diverse antimicrobial resistance, virulence-associated gene repertoires, and population structure of the Jiaxing V. cholerae collection, with particular relevance to the predominant non-O1/non-O139 population. Full article

Aquaponic systems are increasingly recognized as sustainable technologies for integrated fish and vegetable production. However, concerns remain regarding the potential internalization of human pathogens into vegetables grown in these systems. This study assessed the risk of pathogen internalization in lettuce leaves grown in aquaponic systems with Nile tilapia challenged with Escherichia coli or Vibrio cholerae. The system comprised nine fish tanks, eighteen hydroponic pipes, and eighty-one lettuce plants, with tanks assigned to three treatments. Samples of water, fish gut, fish blood, and lettuce leaves were collected. Microbiological analyses included selective culture, biochemical assays, and molecular identification. Although colonies consistent with E. coli and V. cholerae were recovered on selective media, molecular sequencing identified other bacterial species, including Aeromonas sp., Aeromonas caviae, Aeromonas veronii, Enterobacter hormaechei, and Citrobacter freundii. The findings indicate that conventional culture-based methods may produce false-positive results and highlight the importance of molecular confirmation. Notably, pathogenic bacteria associated with tilapia were detected and appeared capable of disseminating through the system and internalizing into lettuce tissues. This result highlights the need for biosecurity measures, contamination monitoring, and the combined use of conventional and molecular diagnostic tools to ensure accurate pathogen detection and compliance with international food safety standards. Full article

Freshwater ecosystems play an important role in human survival, ecosystem functioning, and biodiversity conservation, yet industrialisation and urbanisation dump over 80% of untreated sewage into them. This inadequate wastewater management leads to enteric pathogens like Escherichia coli, Salmonella, Shigella, Campylobacter, Vibrio cholerae, Pseudomonas aeruginosa, and Legionella pneumophila that are responsible for a wide range of waterborne human diseases globally with extensive morbidity and mortality. The World Health Organization (WHO) estimates that at least 2 billion individuals drink water contaminated with pathogens, resulting in illnesses like cholera, dysentery, and diarrhoea, and approximately 50,000 diarrheal deaths annually. Classical epidemiology approaches are the basis for determining disease burden in public health, but they are limited in their capacity to predict future health risks. Quantitative microbial risk assessment (QMRA) addresses this by estimating the potential health risks of any exposure to microbial pathogens in any environment using four key elements, which include the identification of the microbial hazards, human exposure to the hazard through diverse activities, dose–response relationships, and the estimated risk of the infection. This review summarises information on freshwater pathogens, their occurrence, sources and health implications. The methodological approaches of QMRA in freshwater systems are reviewed with examples drawn from recreational activities, drinking water, and wastewater-impacted environments. Global QMRA studies indicate a wide range of infection risk estimates, reflecting differences in water sources, pathogens, and exposure conditions. Thus, QMRA is known to be a valuable public health tool for freshwater ecosystems, linking microbial contamination dynamics to health risk estimates that support proactive management and policy-relevant decision-making. Full article

The highly chitinolytic marine bacterium Vibrio jasicida KMM 6832, which exhibits potent antifungal activity, possesses an atypical Glycosyl Hydrolase family 19 (GH19) chitinase (ChitVjs). This is the first report of a GH19 gene in V. jasicida, an enzyme generally absent in this species and rare within the Harveyi clade. Phylogenetically, ChitVjs-like enzymes from the genera Vibrio and Aeromonas form a distinct cluster, separate from typical plant and bacterial GH19 counterparts. Despite high sequence identity (80–94%) with characterized homologs from V. parahaemolyticus and V. cholerae, ChitVjs is distinguished by its obligate halophilicity (optimum 0.3–0.4 M NaCl), an acidic isoelectric point (pI 4.72), and a broader cation-activation profile (K⁺, Ni²⁺, Ca²⁺, Cu²⁺, Co²⁺). The recombinant ChitVjs was produced in E. coli as a soluble 63 kDa protein. It functions as a stable, salt-dependent endo-chitinase/chitosanase, exhibiting optimal activity at 40 °C and pH 7.0. The enzyme displays high affinity for colloidal chitin (K_M 0.377 mg·mL⁻¹), is activated by DTT and Tween 80, and shows moderate stability in organic solvents. Furthermore, unlike its primarily catabolic relatives, ChitVjs suppresses conidial germination in marine-derived Aspergillus strains. These findings suggest that ChitVjs significantly contributes to the competitive fitness of V. jasicida KMM 6832 in high-salinity marine environments through both nutrient acquisition and antagonism. Full article

The therapeutic efficacy of antibiotics has been significant in extending human life expectancy by combating virulent bacterial infections. Nevertheless, multidrug-resistant (MDR) microorganisms remain a global crisis as these bacteria have developed resistance to conventional antibacterial agents. An unexplored antibiotic target found exclusively in bacteria is the Na⁺-translocating NADH:ubiquinone oxidoreductase (NQR), which is an indispensable membrane-bound bacterial enzyme complex that enables cellular functionality and is present in many infectious bacterial species, including Vibrio cholerae and H. influenzae. NQR serves as an essential complex in the bacterial electron transport chain (ETC) and operates as a highly conserved primary Na⁺ pump that drives many bioenergetic functions. This six-subunit protein shuttles electrons from NADH to ubiquinone, which drives the translocation of Na⁺ ions and creates a gradient that provides the driving force for various cellular processes. We have synthesized and evaluated a series of 1,4-naphthoquinones that exhibit high potency against NQR with minimal cytotoxicity and potential to serve as new, NQR-targeting antibacterial agents for use against V. cholerae. Full article

Sodium-pumping NADH: ubiquinone oxidoreductase (Na⁺-NQR) is an important component of the aerobic respiratory chain of Vibrio cholerae. It oxidizes NADH, reduces ubiquinone, and uses the free energy of this redox reaction to move sodium across the cell membrane. The enzyme is a membrane complex of six subunits, two 2Fe−2S centers, and four flavins. Both the oxidized and reduced forms of Na⁺-NQR exhibit EPR signals due to flavin semiquinone radicals. It has been shown that in the oxidized form of the enzyme, the radical is a neutral flavin, while in the NADH-reduced form, the radical is an anionic flavin. Electron Spin Echo Envelope Modulation Spectroscopy (ESEEM) was used to probe the presence of the magnetic nucleus ²³Na in the immediate vicinity of the paramagnetic centers. The contribution of the ²³Na nucleus was observed only in the ESEEM spectra of the anionic flavin semiquinone previously assigned to FMN_NqrB. Analysis shows that the Na⁺ ion is within ~3–4 Å of the flavin radical. This distance is consistent with two models: (i) complexation of the Na⁺ ion with the carbonyl group of CO4; or alternatively, (ii) a “cation-π interaction,” between Na⁺ and the electron-rich π-system of the flavin aromatic rings. Full article

Biofilm formation represents a key survival strategy employed by Vibrio cholerae to adapt to the complex intestinal environment of the host. While most previous studies on V. cholerae biofilms have focused on genetic regulation and monospecies cultures, its ability to form dual-species biofilms with other intestinal pathogens is still poorly understood. In this study, using samples from both cholera patients and healthy individuals, Fusobacterium nucleatum was identified as a bacterium capable of co-aggregating with V. cholerae. Untargeted metabolomic analysis revealed that F. nucleatum-derived metabolites, specifically 6-hypoxanthine, enhance biofilm formation in V. cholerae. Further validation confirmed that these F. nucleatum-derived metabolites upregulate the biofilm-associated regulatory gene vpsT. In an adult mouse model, co-infection with F. nucleatum and V. cholerae significantly enhanced the intestinal adaptability of V. cholerae compared to infection with V. cholerae alone. Together, these findings elucidate the mechanism enabling the co-infection of F. nucleatum and V. cholerae in the host intestine, thereby shedding new light on how other pathogenic bacteria can assist in V. cholerae infection. Full article

Background: Cholera, caused by Vibrio cholerae, remains endemic in many developing countries, including Pakistan. The extensive use of antibiotics has led to the emergence of antimicrobial resistance in V. cholerae, limiting available treatment options. In this study, we performed molecular characterisation of antibiotic-resistant V. cholerae serotype Ogawa isolates from a recent cholera outbreak in Khyber Pakhtunkhwa, Pakistan. Methodology: Suspected cholera stool samples were collected from hospitalised patients at various district hospitals of Khyber Pakhtunkhwa Province (KPK), Pakistan. The samples were transported to the Public Health Reference Microbiology Laboratory at Khyber Medical University, Peshawar. V. cholerae were identified based on colonial morphology, Gram staining, and biochemical tests using EPI 10E. For serotype identification, monovalent antisera were used. Antibiotic susceptibility testing (AST) was performed using CLSI M45 and EUCAST guidelines. DNA was extracted from pure colonies of multidrug-resistant (MDR) V. cholerae and subjected to whole-genome sequencing (WGS) for genomic characterisation using an Illumina MiSeq platform. Results: Of the 350 active diarrheal cases investigated, 70 were confirmed as V. cholerae. The outbreak was initially reported in Dir and was subsequently followed by a high incidence of cholera in the Peshawar district of KPK. All strains belong to the Ogawa serotype, which shows high antibiotic resistance, particularly to ampicillin (n = 62, 88.57%), Sulfamethoxazole/Trimethoprim (n = 60, 85.71%), Erythromycin (n = 59, 84.29%), and Tetracycline (n = 53, 75.71%). The lowest resistance was against Meropenem (n = 1, 1.4%), followed by amikacin (n = 7, 10.0%) and levofloxacin (n = 13, 18.57%). Furthermore, 34 (48.57%) of the isolates were MDR, while 13 (18.57%) were extensively drug-resistant. Six samples were selected for whole-genome sequencing. The selection of six V. cholerae samples for WGS was based on their drug resistance pattern and origin of isolation. At the genomic level, all sequenced V. cholerae strains harboured multiple antimicrobial resistance determinants. Quinolone resistance was associated with mutations and genes in gyrA, gyrB, parC, and parE; resistance to sulfamethoxazole–trimethoprim with folA, folP, and dfr; tetracycline resistance with tetA and tet35; chloramphenicol resistance with catB and S10p; and aminoglycoside resistance with hns, S12p, and gigB. In addition, β-lactam resistance was linked to the presence of efflux and β-lactamase genes, including blaSHV and mox-3. Mutations were identified in gyrA at positions S83I, S177A, and S202A, and in parC at positions S85L and I231V. Collectively, the presence of these resistance determinants likely enables V. cholerae to survive exposure to high concentrations of multiple antibiotics. Conclusions: Our V. cholerae isolates showed close genetic relatedness to previously sequenced strains from Pakistan (2010 and 2022), as well as to recently reported international strains from the USA, Australia, and China. These findings highlight both the long-term persistence of these lineages within Pakistan and their international dissemination, likely facilitated by globalisation. Full article

This study presents an ultra-sensitive dual-core photonic crystal fiber-based surface plasmon resonance (PCF-SPR) sensor for the detection of waterborne pathogens through refractive index (RI) variation. The proposed sensor integrates a bimetallic coating of silver and titanium dioxide (Ag–TiO₂). Silver ensures sharp plasmonic resonance, and TiO₂ enhances chemical stability and coupling efficiency. This dual-core configuration allows for increased interaction between the core-guided modes and the plasmonic interface. As a result, the sensor’s sensitivity improves significantly. The sensor can accurately detect analytes with an RI value of 1.28 to 1.43. It demonstrates a maximum wavelength sensitivity (WS) of 107,000 nm/RIU, an amplitude sensitivity (AS) of 2209.21 RIU⁻¹, a wavelength resolution of 9.35 × 10⁻⁷ RIU, and a figure of merit (FOM) of about 520. These results support the sensor’s ability to identify the presence of different pathogenic contaminants, such as E. coli, Vibrio cholerae, and Bacillus anthracis, based on their unique RI properties. This optimized design, high resolution, and potential for real-time detection enable this sensor to be a promising solution for environmental monitoring applications. Full article

Species of the Neltuma syn Prosopis genus are known for their use in traditional medicine in America, Asia and Africa. The use of the leaves, bark and inflorescences of one species widely distributed in the arid zones of Mexico, Neltuma laevigata (Humb. & Bonpl. Ex Willd) Britton & Rose, has been reported for the treatment of ocular, gastric and skin infections. Its activities have been related to different secondary metabolites, particularly phenylpropanoids and alkaloids. In the present study, the antibacterial activity of the alkaloidal fraction of inflorescences of P. laevigata collected in Zapotitlán Salinas, Puebla, México, against Staphylococcus aureus ATCC 25,923 and Vibrio cholerae CDBB-1159 was studied by Kirby–Baüer and broth microdilution tests, and its activity on plasmatic membranes was later identified using a protein leakage assay and fluorescence microscopy. Subsequently, the alkaloidal fraction was separated via chromatographic methods, and the purified compounds were elucidated using nuclear NMR and HRESIMS analysis. The alkaloidal fraction showed an important antibacterial activity, with a possible effect on the cytoplasmic membrane of the tested strains. Julifloridine, a piperidine alkaloid previously reported in the genus, was identified for the first time in this species. Full article

The use of phytobiotics in aquafeeds is a promising strategy to enhance performance and resilience to disease. This study evaluated the protective role of Morus alba (MA) extract against Vibrio cholerae, integrating in vivo responses in Dormitator latifrons (growth, biochemical and enzymatic responses, haemato-immunology and tissue histopathology) with in vitro assessment of V. cholerae growth, virulence-associated gene expression and cellular morphology. D. latifrons juveniles were fed five diets (0, 5, 10, 15 and 20 g/kg feed; three tanks per treatment, 15 fish per tank) for eight weeks, followed by a 7-day challenge with V. cholerae. MA increased growth and feed utilisation (p < 0.05); the 20 g/kg group reached 27.57 g final weight with a feed conversion ratio of 1.24, and whole-body protein and lipid contents rose at higher doses. MA modulated plasma biochemistry and key digestive (amylase, lipase), metabolic (ALT, AST) and antioxidant (SOD, CAT, GPx) enzymes, and improved haematological profiles. Histology of the intestine, liver and spleen showed preserved architecture and reinforced mucosal features in supplemented fish, particularly at 15–20 g/kg. Post-challenge, supplemented groups exhibited higher survival/relative protection than controls, alongside lower transaminases and stronger antioxidant responses. In vitro, MA extract inhibited V. cholerae growth, attenuated virulence-associated gene (toxR, ompU) expression and induced marked morphological damage in planktonic cells. Multivariate analyses (Z-score heatmaps and PCA) linked immune–enzymatic improvements with growth and protection. Overall, 15–20 g/kg MA optimised immunophysiological status and disease resistance, supporting MA as a functional feed additive for sustainable aquaculture of D. latifrons. Full article

Understanding the diversity of pathogenic microorganisms in wild primates is essential for assessing their health and zoonotic risks. In this study, metagenomic sequencing was applied to investigate the composition and seasonal dynamics of potential pathogenic microorganisms in the feces of François’ langurs. A total of 77 potential pathogenic taxa were identified, mainly belonging to Bacillota and Pseudomonadota. The most abundant genera were Streptococcus, Staphylococcus, Salmonella, Listeria, and Pseudomonas, while dominant species included Staphylococcus aureus, Streptococcus pneumoniae, Salmonella enterica, Listeria monocytogenes, and Escherichia coli. Significant seasonal differences were detected in both α- and β-diversity indices, with higher microbial diversity in spring and distinct community structures across seasons. Several genera and species, including Vibrio, Chlamydia, Mycobacteroides, Vibrio cholerae, Yersinia enterocolitica, Chlamydia trachomatis, and Mycobacteroides abscessus, showed marked seasonal fluctuations. The findings reveal that the pathogenic microbial community of François’ langurs is strongly shaped by seasonal environmental factors. The detection of multiple zoonotic pathogens suggests a potential risk of cross-species transmission, providing valuable baseline data for primate disease ecology and conservation health management. Full article

Intensive aquaculture and animal farming along riverbanks have emerged as significant drivers of downstream public health risks, facilitating the transmission of zoonotic pathogens and antimicrobial resistance (AMR) genes from farm effluents into natural water systems. In this study, we conducted a comprehensive 12-week water monitoring program at the Wei River in Shandong, China, using a combination of rapid detection techniques (RPA-LFD) and whole-genome sequencing to trace the origins of detected pathogens. RPA-LFD screening revealed the sequential appearance of Vibrio parahaemolyticus, Aeromonas veronii, norovirus GII, and Brucella spp. in surface water from March onward, coinciding with documented wastewater discharge events from upstream shrimp and fox farms. Subsequent isolation efforts confirmed the presence of V. parahaemolyticus and A. veronii in both river water and shrimp samples, while Brucella abortus was isolated from fox feces and water samples. Whole-genome sequencing of bacterial isolates revealed that V. parahaemolyticus strains from water and shrimp shared identical sequence types (ST150 and ST809) and resistance gene profiles, indicating a clonal relationship. Similarly, B. abortus isolates from water and fox feces differed by fewer than five SNPs, confirming farm-to-water transmission. Norovirus GII.3 and GII.6 sequences from water and fecal samples clustered phylogenetically with regional clinical strains, suggesting local circulation and environmental dissemination. Our findings highlight the critical role of river water monitoring as an early warning system for pathogen spread, emphasizing the need for integrated surveillance systems that monitor both water quality and the health of upstream farms and wildlife populations. The combined use of RPA-LFD and whole-genome sequencing provides a robust framework for real-time detection and source tracing of zoonotic pathogens, offering valuable insights for future environmental monitoring and public health interventions. Full article

Cholera remains a major public health challenge, and co-infections can complicate clinical outcomes. In a cross-sectional study, we investigated the prevalence and patterns of enteric co-infections during Zambia’s 2023–2024 cholera outbreak and evaluated their implications for disease severity. 240 suspected cholera patients were enrolled from five healthcare facilities in Lusaka. Stools were tested for 11 enteric pathogens using the Bosphore^® Gastroenteritis Panel Kit v2 on the QuantStudio 5 qPCR, with Vibrio cholerae confirmed by real-time PCR (quantitative PCR). Co-infections were highly prevalent, affecting 79.2% of participants. Campylobacter was the most frequently detected pathogen (70.0%), followed by Norovirus GI/GII (20.0%). Persons living with HIV were significantly more likely to present with co-infections than their counterparts (adjusted PR 1.27, 95% CI: 1.07–1.51; p = 0.008). Participants with confirmed V. cholerae + coinfections (N = 62) were less likely to developed moderate to severe disease compared to those with mono-infections (adjusted PR 0.59, 95% CI: 0.38–0.90; p = 0.014). These findings highlight the high prevalence and complexity of co-infections during cholera outbreaks, potentially contributing to antimicrobial resistance. They also highlight the need for targeted clinical management, particularly among persons living with HIV. Full article