Development of multiplex HRM-based loop-mediated isothermal amplification method for specific and sensitive detection of Treponema pallidum.

Syphilis is a sexually transmitted disease caused by the spirochaete bacterium Treponema pallidum. This study has developed a multiplex High-Resolution Melt-curve Loop-mediated isothermal amplification (multiplex HRM-LAMP) assay targeting the marker genes polA and tprL to detect T. pallidum. The multiplex HRM-LAMP assay conditions were optimized at 65 °C for 45 min. Real-time melt-curve analysis of multiplex HRM-LAMP shows two melt-curve peaks corresponding to polA and tprL with a Tm value of 80 ± 0.5 °C and 87 ± 0.5 °C, respectively. The detection limit of multiplex HRM-LAMP was found to be 6.4 × 10-4 ng/µL (3.79 copies/µL) of T. pallidum. The specificity was evaluated using seven different bacterial species, and the developed method was 100% specific in detecting T. pallidum. A total of 64 blood samples of T. pallidum suspected cases were used to validate the assay method. The clinical validation showed that the assay was 96.43% sensitive and 100% specific in detecting syphilis. Thus, the developed method was more rapid and sensitive than other available methods and provides a multigene-based diagnostic approach to detect T. pallidum.

Exploring antifungal activities of acetone extract of selected Indian medicinal plants against human dermal fungal pathogens.

A broad spectrum of medicinal plants was used as traditional remedies for various infectious diseases. Fungal infectious diseases have a significant impact on public health. Fungi cause more prevalent infections in immunocompromised individuals mainly patients undergoing transplantation related therapies, and malignant cancer treatments. The present study aimed to investigate the in vitro antifungal effects of the traditional medicinal plants used in India against the fungal pathogens associated with dermal infections. Indian medicinal plants (Acalypha indica, Lawsonia inermis Allium sativum and Citrus limon) extract (acetone/crude) were tested for their antifungal effects against five fungal species isolated from skin scrapings of fungal infected patients were identified as including Alternaria spp., Curvularia spp., Fusarium spp., Trichophyton spp. and Geotrichum spp. using well diffusion test and the broth micro dilution method. All plant extracts have shown to have antifungal efficacy against dermal pathogens. Particularly, Allium sativum extract revealed a strong antifungal effect against all fungal isolates with the minimum fungicidal concentration (MFC) of 50-100 µg/mL. Strong antifungal activity against Curvularia spp., Trichophyton spp., and Geotrichum spp. was also observed for the extracts of Acalypha indica, and Lawsonia inermis with MFCs of 50-800 µg/mL respectively. The extracts of Citrus limon showed an effective antifungal activity against most of the fungal strains tested with the MFCs of 50-800 µg/mL. Our research demonstrated the strong evidence of conventional plants extracts against clinical fungal pathogens with the most promising option of employing natural-drugs for the treatment of skin infections. Furthermore, in-depth analysis of identifying the compounds responsible for the antifungal activity that could offer alternatives way to develop new natural antifungal therapeutics for combating resistant recurrent infections.

Restoration of energy metabolism in leukemic mice treated by a siddha drug--Semecarpus anacardium Linn. nut milk extract.

Chronic myeloid leukemia (CML) is a clonal disorder characterized by proliferation of hematopoietic cells that possess the BCR-ABL fusion gene resulting in the production of a 210 kDa chimeric tyrosine kinase protein. CML, when left untreated, progresses to a blast phase during which the disease turns aggressive and shows poor response to known treatment regimens. We have studied a Siddha herbal agent, Semecarpus anacardium Linn. nut milk extract (SA) for its antileukemic activity and its effect on the changes in energy metabolism in leukemic mice. Leukemia was induced in BALB/c mice by tail vein injection of BCR-ABL(+) 12B1 murine leukemia cell line. This resulted in an aggressive leukemia, similar to CML in blast crisis, myeloid subtype, confirmed by histopathological study and RT-PCR for the p210 mRNA in the peripheral blood, spleen and liver. Leukemia-bearing mice showed a significant increase in lipid peroxides, glycolytic enzymes, a decrease in gluconeogenic enzymes and significant decrease in the activities of TCA cycle and respiratory chain enzymes as compared to control animals. SA treatment was compared with standard drug imatinib mesylate. SA administration to leukemic animals resulted in clearance of the leukemic cells from the bone marrow and internal organs on histopathological examination and this was confirmed by RT-PCR for the p210 mRNA. Treatment with SA significantly reversed the changes seen in the levels of the lipid peroxides, the glycolytic enzymes, the gluconeogenic enzymes and the mitochondrial enzymes. These effects are probably due to the flavonoids, polyphenols and other compounds present in SA which result in total regression of leukemia and correction of the alterations in energy metabolism. Study of animals treated with SA alone did not reveal any adverse effects. On the basis of the observed results, SA can be considered as a readily accessible, promising and novel antileukemic chemotherapeutic agent.