Synergistic activity and mechanism of action of Stephania suberosa Forman extract and ampicillin combination against ampicillin-resistant Staphylococcus aureus.

BACKGROUND: Ampicillin-resistant S. aureus (ARSA) now poses a serious problem for hospitalized patients, and their care providers. Plant-derived antibacterial that can reverse the resistance to well-tried agents which have lost their original effectiveness are the research objectives of far reaching importance. To this aim, the present study investigated antibacterial and synergistic activities of Stephania suberosa extracts (SSE) against ARSA when used singly and in combination with ampicillin. RESULTS: The majority chemical compounds of SSE were alkaloid (526.27 ± 47.27 mg/1 g of dried extract). The Minimum inhibitory concentration (MICs) for ampicillin and SSE against all ARSA strains were >512 µg/ml and 4 mg/ml, respectively. Checkerboard assay revealed synergistic activity in the combination of ampicillin (0.15 µg/ml) and SSE (2 mg/ml) at fractional inhibitory concentration index (FICI) <0.5. The killing curve assay had confirmed that the viability of ARSA was dramatically reduced from 5 x 10(5) cfu/ml to 10(3) cfu/ml within 6 h after exposure to SSE (2 mg/ml) plus ampicillin (0.15 µg/ml) combination. Electron microscopic study clearly revealed that these ARSA cells treated with this combination caused marked morphological damage, peptidoglycan and cytoplasmic membrane damage, and average cell areas significant smaller than control. Obviously, Immunofluorescence staining and confocal microscopic images confirmed that the peptidoglycan of these cells were undoubtedly disrupted by this combination. Furthermore, the CM permeability of ARSA was also increased by this combination. Enzyme assay demonstrated that SSE had an inhibitory activity against ß-lactamase in concentrations manner. CONCLUSIONS: So, these findings provide evidence that SSE has the high potential to reverse bacterial resistance to originate traditional drug susceptibility of it and may relate to three modes of actions of SSE: (1) inhibits peptidoglycan synthesis, resulting in morphological damage, (2) inhibits ß-lactamases activity, and (3) increases CM permeability. It is widely recognized that many types of drugs are derived from alkaloids. So, this SSE offers the prominent potential to develop a novel adjunct phytopharmaceutical to ampicillin for the treatment of ARSA. Further active ingredients study, toxicity of it, and the synergistic effect on blood and tissue should be performed and confirmed in an animal test or in humans.

Induction of RNA-mediated resistance to papaya ringspot virus type W.

Transformation of cantaloupes with the coat protein (cp) gene of papaya ringspot virus type W (PRSV-W), Thai isolate, was used to introduce virus resistance. Binary vectors containing either the full length coat protein coding region under control of the 35S CaMV promoter(pSA1175), or the inverted-repeat of a coat protein coding region (pSA1304), were constructed and used for Agrobacterium-mediated transformation of cotyledonary explants of the cantaloupe cultivar Sun Lady. Four independent transgenic lines were obtained using pSA1304 and one using pSA1175. Integration of the PRSV-W cp gene into the genome of these transgenic lines was verified by PCR amplification, GUS assays and Southern blot hybridization. In vitro inoculation of these lines with PRSV-W revealed that whereas the line containing pSA1175 remained sensitive, the four lines containing pSA1304 were resistant. The presence of small RNA species, presumably siRNA, corresponding to regions of the viral cp gene in transgenic lines resistant to PRSV-W supports the involvement of post-transcriptional gene silencing in the establishment of resistance.