Streptococcus agalactiae critically endangers tilapia farming and causes substantial economic losses to aquaculture. In this study, a rapid and efficient recombinase polymerase amplification-lateral flow strip (RPA-LFS) assay targeting the conserved cfb gene was established for the detection of S. agalactiae. Specific RPA primers were designed for the target region of the cfb gene, and an RPA-LFS system was developed based on primer—probe amplification and colloidal-gold visualization. Conventional PCR was performed in parallel to compare detection performance. The results showed that the F3/R primer pair exhibited the highest specificity under the optimized conditions of 28 °C for 15 min. The lowest detectable DNA concentration was 2.054 × 10−6 ng/uL. Further sample testing showed that the minimum detectable bacterial copy number was 8.55 × 103 copies/µL for RPA-LFS and 8.55 × 104 copies/µL for conventional PCR. These results confirmed that the developed RPA-LFS assay had significantly higher detection sensitivity than conventional PCR and showed strong potential for the early surveillance and effective control of streptococcosis in tilapia farming.
Bacteria of the genus Bacillus attract interest for their ability to produce secondary metabolites with important biotechnological and biomedical applications. Metabolites produced by thermophilic strains, however, have received far less attention and remain poorly characterised. In the context of increasing antibiotic resistance and growing pressure to limit chemical inputs, particularly in agriculture, there is a pressing need to identify new bio-based antimicrobial compounds. Additionally, using inexpensive substrates could reduce production costs. Thermophilic B. licheniformis strain LMB3G52 was grown on two different substrates: conventional Luria–Bertani (LB) medium and an innovative medium derived from cheese waste. This strain showed broad-spectrum antimicrobial activity that was significantly stronger in LB medium. Chloroform extracts inhibited Acinetobacter baumannii (19.7 ± 0.6 mm) and Listeria monocytogenes (18 ± 1 mm). GC–MS revealed distinct substrate-dependent metabolite profiles: cheese waste medium predominantly yielded hexadecanoic acid methyl ester (31.0
Pseudomonas aeruginosa, an opportunistic Gram-negative bacterium, poses a public health challenge owing to its ability to form persistent biofilms. Combinatorial antibiotic approaches have emerged as a promising strategy for managing biofilm-associated infections caused by P. aeruginosa. In the present study, ciprofloxacin and tetracycline, two widely used antibiotics, were evaluated individually and in combination against P. aeruginosa to investigate their antimicrobial and antibiofilm activities. Checkerboard analysis revealed an additive interaction between the two antibiotics, indicating their potential for combined application. Furthermore, the combination demonstrated enhanced antimicrobial and antibiofilm activity compared with the respective individual treatments. The antimicrobial efficacy of the combination was subsequently evaluated using a zebrafish embryo infection model. The results demonstrated that P. aeruginosa could establish infection in zebrafish embryos, resulting in substantial embryonic mortality. However, treatment of infected embryos with the antibiotic combination provided preliminary evidence of a partial protective effect, as it temporarily delayed infection-associated mortality. Although the combination demonstrated encouraging in vivo activity under the experimental conditions, complete protection was not achieved, indicating the need for further optimization and evaluation. Importantly, the present study was conducted using a single standard laboratory strain (P. aeruginosa MTCC 424). Therefore, validation using diverse clinical and multidrug-resistant isolates, along with comprehensive efficacy and safety assessments, will be essential for establishing the robustness and translational potential of this combinatorial approach.
Chromium is a redox-active metal that can exert toxic effects on living systems, particularly in its hexavalent form [Cr(VI)]. This study investigated the physiological and molecular responses of chromium-resistant bacteria isolated from electroplating effluent under Cr(VI) stress and compared them with the positive control Pseudomonas aeruginosa MTCC 7812. Morphological alterations were examined using scanning electron microscopy coupled with energy-dispersive X-ray (SEM–EDX), while functional group interactions were analysed using Fourier-transform infrared (FT-IR) spectroscopy. Gene expression analysis was performed using quantitative real-time PCR (qRT-PCR). Exposure to Cr(VI) resulted in noticeable morphological changes, including cell enlargement and surface distortion, suggesting stress-induced structural modifications. FT-IR analysis indicated shifts in characteristic absorption bands, implicating carboxyl, amino and phosphate groups in chromium interaction. The isolates were tentatively assigned to Alcaligenes faecalis (UN1) and Pseudomonas aeruginosa (LH1) based on 16S rRNA gene sequencing and high sequence similarity. qRT-PCR analysis demonstrated significant up-regulation of chromate-associated genes, with ChrA expression increasing by 10.73-fold and 1.39-fold, and ChrR by 16.91-fold and 8.90-fold in Alcaligenes faecalis UN1 and Pseudomonas aeruginosa LH1, respectively, under Cr(VI) exposure. In contrast, ChrA was down-regulated in the reference strain (0.17-fold). Antioxidant genes also showed increased expression, with superoxide dismutase (Sod) up-regulated by 2.05-fold and 1.27-fold, and glutathione-S-transferase (Gst) by 1.61-fold and 3.17-fold in Alcaligenes faecalis UN1 and Pseudomonas aeruginosa LH1, respectively. These findings provide insight into the adaptive responses of chromium-resistant bacteria to Cr(VI) stress and their potential role in chromium transformation and removal.
The alarming emergence of multidrug-resistant bacterial pathogens (MDRBPs), esp.ecially associated with cancer patients, necessitates the exploration of novel biological agents as endophytic microorganisms that are natural reservoirs of bioactive substances. The purpose of the present study was to isolate endophytic bacteria and evaluate the exometabolic crude ethyl acetate extract against MDRBPs in vitro and in vivo, study their effects on cancer cells and wound recovery, and elucidate some mechanisms supporting their efficacy. In our study, a bacterium was isolated from Sp.inacia oleracea and identified as a Bacillus isolate closely related to Bacillus amyloliquefaciens and deposited in GenBank as PX736878. The exometabolic crude ethyl acetate of Bacillus sp. exhibited a strong antibacterial activity against MDRBPs, and the highest activity was against Klebsiella pneumoniae (21.2 mm). The MIC values showed its efficacy, particularly at 500 µg/ml. The extract also exhibited significant anti-biofilm activity sp.ecifically against K. pneumoniae. The cytotoxicity assays showed a selective toxicity of Bacillus sp. extract against the cancer cell lines (HCT116 and MCF7). In vivo, significant wound healing potential of the exometabolic crude ethyl acetate of Bacillus sp. was demonstrated by histological evidence of near-normal tissue regeneration. The strong activities exometabolic crude ethyl acetate of Bacillus sp. were due to the high antioxidant capacity. GC-MS analysis further supported our results and identified diverse bioactive compounds in the exometabolic crude ethyl acetate of Bacillus sp., including Diketopyrazine, fatty acids, and dipeptides. In conclusion, the extract of exometabolic crude ethyl acetate extract of Bacillus sp. is a promising multifunctional agent with strong antibacterial, anti-biofilm, antioxidant, and anticancer properties. This could be a new way to fight MDRBPs and aid damaged tissue to heal.
