Antibiotic resistant Vibrio harveyi isolated from shrimp pond and anti-Vibrio activity of Combretum quadrangulare Kurz. and Mimosa pudica extracts

Aims@#The occurrence of bacterial disease in shrimp ponds is a major problem faced in shrimp farming. Thus, the aims of this study were to isolate and evaluate antibiotic resistant profile of Vibrio harveyi strain isolated from shrimp pond water, as well as to study the potential anti-Vibrio activity of Combretum quadrangulare Kurz. (CQ) and Mimosa pudica (MP) leaves extracts.@*Methodology and results@#Vibrio harveyi WSC103 was isolated from water in white shrimp (Litopenaeus vannamei) culture pond and identified using 16S rRNA gene sequencing analysis. This strain showed characteristics of multidrug-resistant (7 antibiotics). It had become more sensitive to antibiotics (9 out of 10 antibiotics) after plasmid curing. It is showed CQ and MP leaves extracts contain potent bioactive compounds (tannins, flavonoids, steroids, cardiac glycosides and alkaloids) against V. harveyi WSC103. The aqueous, 95% ethanolic and 75% acetone extracts of CQ (MIC value of 3.13-12.50 mg/mL) and MP (MIC value of 3.13-25.00 mg/mL) leaves revealed strong vibriostatic activity, but aqueous and 95% ethanolic extracts in both plants showed vibriocidal activity. The 95% ethanolic extract of both CQ and MP leaves displayed the excellent vibriocidal property with MBC value of 100 mg/mL with zone of inhibition at 11.44 ± 1.01 and 11.78 ± 1.01 mm by agar disc diffusion.@*Conclusion, significance and impact of study@#The isolated Vibrio harveyi WSC103 was successfully characterized as a novel multidrug-resistant strain. The ethanolic C. quadrangulare Kurz. and M. pudica extracts exhibited prominent vibriostatic and vibriocidal capacities. These finding is proven that C. quadrangulare Kurz. and M. pudica extracts would be an alternative anti-Vibrio agent for aquaculture infectious treatment.

Antibiotic resistance reversal of multiple drug resistant bacteria using Piper longum fruit extract.

Present investigation was aimed to identify natural products of plant-origin as novel antibiotic resistance reversal agents. Aqueous and methanol extracts of Piper longum (fruits) were tested against multiple drug resistant (MDR) clinical isolates of Enterococcus faecalis, Staphylococcus aureus, Salmonella typhi, Shigella sonnei, as well as reference-plasmid-harboring strains of Escherichia coli (RP4) and Bacillus subtilis (pUB110). The crude methanol extract showed significant antibacterial activity with a minimal inhibitory concentration of 400 μg/mL against Bacillus subtilis (harboring pUB110 plasmid). Methanol extract could reverse the antibiotic resistance in clinical isolates of Shigella sonnei, with a curing efficiency of 42%. In comparison with methanol extract, aqueous extract showed antibiotic resistance reversal efficiency against wider range of clinical isolates. Aqueous extract showed strong antibiotic resistance reversal activities against R-plasmid harboring strains of clinical origin- Enterococcus faecalis, Staphylococcus aureus, Salmonella typhi with curing efficiencies of 64%, 50% and 32% respectively. This antibiotic resistance reversal may be attributed to the elimination of R-plasmids as the multiple antibiotic resistance genes are usually located on R-plasmids. Active biomolecules from P. longum may prove to be a source to develop MDR reversal agents of natural origin to contain the development and spread of plasmid borne multiple antibiotic resistance.

Changes of characterization of Salmonella Typhimurium isolate following sequential exposures to porcine neutrophil / 대한수의학회지

To develop a live vaccine candidate using an attenuated strain of Salmonella Typhimurium (ST), biochemical properties, plasmid profile, PFGE patterns and pathogenic analysis of the ST isolate were carried out after sequential passage of the ST isolate in porcine neutrophils. By the passage, the ability of the neutrophil-adapted isolate to utilize d-xylose was lost, while the ability of the strain to ferment trehalose was delayed after 2 or more days of the culture. Also, changes including deletion of the gene fragments were observed in PFGE analysis of the neutrophil-adapted isolates. Two plasmids, 105kb and 50kb, were cured in the strain passaged over 15 times in porcine neutrophils. The 50% of lethal dose (LD50) of the parent strain was changed from 1 x 10(5) LD50 to 6 x 10(6) LD50 by the passage in intraperitoneal injection of the strains into mice. These results suggested that bacterial genotypic and phenotypic responses might be globally altered depending on the inside environment of neutrophils.

Antibacterial & antiplasmid activities of Helicteres isora L.

Background & objectives: The multiple drug resistance (MDR) is a serious health problem and major challenge to the global drug discovery programmes. Most of the genetic determinants that confer resistance to antibiotics are located on R-plasmids in bacteria. The present investigation was undertaken to investigate the ability of organic extract of the fruits of Helicteres isora to cure R-plasmids from certain clinical isolates. Methods: Active fractions demonstrating antibacterial and antiplasmid activities were isolated from the acetone extracts of shade dried fruits of H. isora by bioassay guided fractionation. Minimal inhibitory concentration (MIC) of antibiotics and organic extracts was determined by agar dilution method. Plasmid curing activity of organic fractions was determined by evaluating the ability of bacterial colonies (pre treated with organic fraction for 18 h) to grow in the presence of antibiotics. The physical loss of plasmid DNA in the cured derivatives was further confirmed by agarose gel electrophoresis. Results: The active fraction did not inhibit the growth of either the clinical isolates or the strains harbouring reference plasmids even at a concentration of 400 μg/ml. However, the same fraction could cure plasmids from Enterococcus faecalis, Escherichia coli, Bacillus cereus and E. coli (RP4) at curing efficiencies of 14, 26, 22 and 2 per cent respectively. The active fraction mediated plasmid curing resulted in the subsequent loss of antibiotic resistance encoded in the plasmids as revealed by antibiotic resistance profile of cured strains. The physical loss of plasmid was also confirmed by agarose gel electrophoresis. Interpretation & conclusions: The active fraction of acetone extract of H. isora fruits cured R-plasmids from Gram-positive and Gram-negative clinical isolates as well as reference strains. Such plasmid loss reversed the multiple antibiotic resistance in cured derivatives making them sensitive to low concentrations of antibiotics. Acetone fractions of H. isora may be a source to develop antiplasmid agents of natural origin to contain the development and spread of plasmid borne multiple antibiotic resistance.

Studies with RP1 deletion plasmids: Incompatibility properties, plasmid curing and the spontaneous loss of resistance.

Four deletion plasmids, pHH301, pHH302, pHH303 and pHH401, obtained from RP1 DNA-transformed bacterial clones, were shown to be incompatible with three Ρ plasmids in Escherichia coli K12 strains. Kinetic experiments and colony tests were used to verify the position of these R plasmids. Pseudomonas aeruginosa and Ε. coli strains, harbouring deletion plasmids, could be cured by using two mutagens, acriflavine and mitomycin C, which affect a percentage of the cell population. The deletion plasmid-positive strains could also be induced at an elevated temperature to spontaneously loose their plasmids.