Introduction: Group A streptococcus (GAS) is responsible for high morbidity and mortality globally. Hence, the need to develop sensitive, reliable and cost- effective method of detection is crucial. In this study, we developed a visual detection method for the common virulence gene, streptococcal pyrogenic exotoxin B (speB) involved in invasive GAS diseases using loop-mediated isothermal amplification (LAMP) with fluorescent detection dye (calcein). Meth-ods: The LAMP reaction was optimized at 63°C for 35 minutes using five sets of primer designed with LAMP primer V5 software. When the dye was added prior to amplification, samples with speB DNA developed a characteristic green color after the reaction, but no color reactions were observed in samples with DNAs of non-GAS isolates. De-tection of speB by LAMP assay was done among 43 clinical isolates of blood, pus, wound, tissue and throat samples and ATCCs for controls. Our findings were further reconfirmed by subjecting the LAMP products to 0.5% gel electro-phoresis. Results: The detection limit of this LAMP assay for speB was 10-7 ng/μl of genomic DNA per reaction, which was 10,000-fold more sensitive than conventional PCR 10-3 ng/μl. All 100 % samples were positive for speB gene by LAMP, and 93% by conventional PCR method. Conclusion: LAMP assay could offer remarkably high sensitivity, specificity, repeatability, reliability, affordability, and visibility; it is appropriate for rapid detection of speB in Group A streptococci (GAS) as a point of care testing.
Lactobacilli are well-documented probiotics that exert health benefits on their host. They exhibit characteristics that make them potential alternative treatments to address the antimicrobial resistance conundrum and diseases. Their mechanism of action varies with strain and species. Five lactobacilli strains previously isolated from the anogenital region were subjected to several assessments highlighted in the FAO/WHO document, ‘Guidelines for the Evaluation of Probiotics in Food’ to determine its suitability as potential probiotics. Methods: The five lactobacilli strains were subcultured onto Man de Rogosa agar (MRS). Their ability to auto- and co-aggregate was determined spectrophotometrically. Simultaneously, the cell surface hydrophobic properties of these strains towards xylene and toluene were evaluated using the microbial adhesion to hydrocarbon (MATH) test. The lactobacilli strains were also tested for their ability to withstand acid, bile and spermicide to determine their level of tolerance. Results: Lact. reuteri 29A, L. delbrueckii 45E and L. reuteri 29B exhibited the highest degree of auto- and co-aggregation properties. These lactobacilli strains also demonstrated high cell surface hydrophobicity, with the exception of L. delbrueckii 45E. Further tests to evaluate the isolated lactobacilli tolerance identified L. reuteri 29B as the most tolerant strain towards low pH (pH 2.5 for 4 h), high bile concentration (0.5% for 4 h) and high spermicides concentration (up to 10%). Conclusion: Out of the five lactobacilli strains which possessed high antimicrobial activities, L. reuteri 29B portrayed the best probiotic qualities with good auto- and co-aggregation abilities and high tolerance against acid, bile and spermicide.