An improved recombinase polymerase amplification assay for the visual detection of Staphylococcus epidermidis with lateral flow strips.

Staphylococcus epidermidis is an opportunistic pathogenic microorganism that is an important cause of cross-infection in hospitals. The development of rapid and effective detection techniques is important for its control. The application of traditional identification and PCR-based methods is limited by their requirements for both laboratory instrumentation and trained personnel. To overcome this issue, we developed a fast detection approach for S. epidermidis that was based on recombinase polymerase amplification (RPA) and lateral flow strips (LFS). First, five pairs of primers were designed for molecular diagnosis using the sesB gene as the target, and were screened for their amplification performance and the formation of primer dimers. Specific probes were then designed based on the best primer pairs screened, which were susceptible to primer-dependent artifacts and generated false-positive signals when used for LFS detection. This weakness of the LFS assay was overcome by modifying the sequences of the primers and probes. The efficacy of these measures was rigorously tested, and improved the RPA-LFS system. Standardized systems completed the amplification process within 25 min at a constant temperature of 37 °C, followed by visualization of the LFS within 3 min. The approach was very sensitive (with a detection limit of 8.91 CFU/µL), with very good interspecies specificity. In the analysis of clinical samples, the approach produced results consistent with PCR and 97.78% consistent with the culture-biochemical method, with a kappa index of 0.938. Our method was rapid, accurate, and less dependent on equipment and trained personnel than traditional methods, and provided information for the timely development of rational antimicrobial treatment plans. It has high potential utility in clinical settings, particularly in resource-constrained locations.

Glutathione in combination with trehalose has supplementary beneficial effects on cryopreserved red deer (cervus elaphus) sperm.

OBJECTIVES: In this study, we evaluated the effects of glutathione in combination with trehalose addition to semen extenders on the quality parameters of frozen-thawed red deer (cervus elaphus) spermatozoa. METHOD OF STUDY: The semen samples collected from six mature red deer once a week were diluted with Tris-egg yolk-based extenders. The diluted semen samples were supplemented with glutathione (8 mmol L-1 ) and or trehalose (5%, w/v), cryopreserved, thawed and then subjected to sperm quality parameter evaluation. RESULTS: Both glutathione and trehalose addition to the extender significantly improved progressive motility, acrosome integrity, membrane integrity, superoxide dismutase and glutathione peroxidase activity and decreased percentage abnormality and sperm malondialdehyde level compared with the control group (P<.05). Moreover, glutathione in combination with trehalose addition to semen extenders had higher efficiency compared with the glutathione or trehalose addition alone (P<.05). CONCLUSION: Therefore, glutathione in combination with trehalose could be a promising cryoprotectant for red deer sperm.

Effects of Ureaplasma urealyticum Infection on Sperm Quality and Concentrations of Nitric Oxide and Cytokine in the Semen of Infertile Males.

PROBLEM: The relationship between genital infection and two sperm parameters, namely, concentrations of nitric oxide (NO) and cytokines [e.g., interleukin (IL)-17 and IL-18], in the semen of infertile males remains undetermined. METHOD OF STUDY: Semen samples from 81 infertile (with or without Ureaplasma urealyticum infection) and normal males were subjected to semen analysis. RESULTS: Nitric oxide concentration in the semen of infertile males with genital infection was higher than those of infertile males without genital infection and of normal males (P < 0.05). Sperm density, pH, percentage of forward, movement of sperm, sperm activation rate, sperm survival rate, and normal rate of the sperm morphology of infertile males with U. urealyticum infection were significantly lower than those of infertile males without genital infection and of normal males (P < 0.05). NO concentrations were also positively correlated with IL-17 and IL-18 concentrations in the semen of infertile males. CONCLUSION: Increased NO concentration and abnormal IL-17 and IL-18 expression were induced by genital infection and induced damage to male reproductive capacity, thereby causing male infertility.