The ethanolic extract from Rhodomyrtus tomentosa leaf exhibited good antibacterial activities against both methicillin-resistant Staphylococcus aureus (MRSA) and S. aureus ATCC 29213. Its minimal inhibitory concentration (MIC) values ranged from 31.25–62.5 µg/ml, and the minimal bactericidal concentration (MBC) was 250 µg/ml. Rhodomyrtone, an acylphloroglucinol derivative, was 62.5–125 times more potent at inhibiting the bacteria than the ethanolic extract, the MIC and MBC values were 0.5 µg/ml and 2 µg/ml, respectively. To provide insights into antibacterial mechanisms involved, the effects of rhodomyrtone on cellular protein expression of MRSA have been investigated using proteomic approaches. Proteome analyses revealed that rhodomyrtone at subinhibitory concentration (0.174 µg/ml) affected the expression of several major functional classes of whole cell proteins in MRSA. The identified proteins involve in cell wall biosynthesis and cell division, protein degradation, stress response and oxidative stress, cell surface antigen and virulence factor, and various metabolic pathways such as amino acid, carbohydrate, energy, lipid, and nucleotide metabolism. Transmission electron micrographs confirmed the effects of rhodomyrtone on morphological and ultrastructural alterations in the treated bacterial cells. Biological processes in cell wall biosynthesis and cell division were interrupted. Prominent changes including alterations in cell wall, abnormal septum formation, cellular disintegration, and cell lysis were observed. Unusual size and shape of staphylococcal cells were obviously noted in the treated MRSA. These pioneer findings on proteomic profiling and phenotypic features of rhodomyrtone-treated MRSA may resolve its antimicrobial mechanisms which could lead to the development of a new effective regimen for the treatment of MRSA infections.

The aim of this study was to develop a nanoparticle-based cell capture system combined with a lateral flow test strip (LFT) assay for rapid detection of Campylobacter jejuni from poultry samples. The developed assay was bench-marked against the standard modified Charcoal Cefoperazone Deoxycholate Agar (mCCDA) method according to ISO16140:2003 procedures. The synthesized ferromagnetic nanoparticles (FMNs) were modified with glutaraldehyde, then functionalized with polyclonal antibodies for specific C. jejuni capture and concentration from poultry samples. After lysing captured cells, DNA from C. jejuni was amplified by PCR using the primers designed to target the hipO gene, and the PCR amplicons were detected with the lateral flow test strip assay. Following the ISO16140:2003 guidelines, the relative detection limit, and the inclusivity and exclusivity tests were determined. The results showed that the limit of detection (LOD) of the assay was 10 or 1 cfu/ml with C. jejuni in pure culture and 10-10 cfu/ml with target cells spiked in poultry sample. In addition, the inclusivity and exclusivity tests were found to be 100%. Using field chicken samples (n = 60), the assay showed relative accuracy, relative specificity, and relative sensitivity of 96.67%, 100% and 93.33%, respectively. The positive predictive values (PPV) and negative predictive values (NPV), and the kappa index of concordance (k) were calculated as 100% and 93.75%, and 0.93, respectively. The developed assay required approximately 3 h to complete and gave results comparable to those analyzed by the standard culture method, which required 5-7 days. The assay is rapid, easy-to-use, and has potential to be directly applied to C. jejuni detection in various categories of poultry samples.

Culture methods for detecting virulent Yersinia enterocolitica require selective enrichment and a series of confirmatory tests that are time-consuming, costly, and laborious. The objective of this study was to evaluate a fluorogenic 5'-nuclease assay for detecting the enterotoxin yst gene of virulent Y. enterocolitica in pure cultures, inoculated ground pork samples, and naturally contaminated food samples. These results were then compared with "gold standard" methods recommended by the U.S. Food and Drug Administration in the Bacteriological Analytical Manual for detecting pathogenic Y. enterocolitica. The 5'-nuclease assay was able to identify the organism in 100% of the repetitions when 10(2) CFU/ml or more organisms were present in pure cultures and 10(3) CFU/g or more organisms were present in ground pork. Similar recovery efficiency on cefsulodin-irgasan-novobiocin (CIN) agar plates was only evident when 10(5) CFU/ml or more organisms were present in pure culture and 10(6) CFU/g or more organisms were present in inoculated ground pork. The 5'-nuclease assay indicated a contamination rate of 35.5% (94/265) in various meats and tofu, whereas the CIN plating method indicated a contamination rate of 28.3% (75/265). This resulted in 100% sensitivity and 64.5% specificity for the 5'-nuclease assay when compared with the standard culture recovery method. Only 75% (60/80) of the Yersinia spp. isolated on CIN was identified as containing a virulence plasmid by autoagglutination and crystal violet binding tests. These results indicate that the true rate of contamination of virulent Y. enterocolitica in pork and other processed meats and foods is being underestimated using current detection methods. This study demonstrates the potential of the 5'-nuclease assay for rapidly and specifically detecting virulent Y. enterocolitica in processed foods with the added advantage of being an automated detection system with high-throughput capability.

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The effect of volume of liquid medium (Hunt broth) supplemented with Oxyrase® on growth of Campylobacter jejuni was evaluated by using four sizes of containers and two concentrations of Oxyrase® (0.15 units/mL and 0.6 units/mL). We used 13×100 mm screw‐capped tubes containing 1, 3, and 6 mL of medium; 18x150 mm screw‐capped tubes containing 1, 5, 10, and 20 mL of medium; 45x45x120 mm bottles containing 25, 50, and 100 mL of medium; and Stomacher® 400 bags containing 250 mL of medium. Counts of four strains of C. jejuni were determined on blood agar plates at 0, 6, 12, 20, and 28 h of incubation. For 13×100 mm tubes, 3 mL of broth resulted the highest bacterial counts with increases of 4–5 log CFU/mL at 12 h and 7–8 log CFU/mL at 20 h. For 18x150 mm tubes, 10 mL and 20 ml of broth both provided increases of 3–4 log CFU/mL and of 6–7 log CFU/mL at 12 h and 20 h, respectively. Increases of 3.5 log CFU/mL at 12 h and of 7–8 log CFU/mL at 20 h were found in bottles containing 100 mL of broth. Overall, 0.15 units/mL Oxyrase® enhanced Campylobacter growth as effectively as 0.6 units/mL in all containers with no significant differences (p>0.05). However, supplementation with 0.6 units/mL of Oxyrase® gave slightly higher bacterial counts than addition of 0.15 units/mL in containers with high ratios of head space volume to medium volume (H:M). Ratios between 1.0 and 3.0 generally gave better results with all strains. The dissolved oxygen levels in each container were lower with high medium volume than with low medium volume. This study positively supported the efficiency of using Oxyrase® for recovery of C. jejuni under normal atmospheric conditions. In addition, various sizes of containers could be used for culturing campylobacters by maintaining the H:M ratio between 1.0 and 3.0.