Panagiota Gousia

Antimicrobial Resistance of Major Foodborne Pathogens from Major Meat Products

The bacterial contamination of raw and processed meat products with resistant pathogens was studied. The raw samples included sheep (40), goat (40), pork (120), beef (80), and chicken (19) meat, and the processed samples included turkey filets (33), salami (8), readymade mincemeat (16), stuffing (22), and roast-beef (50). The samples were collected from retail shops in Northwestern Greece over a period of 3 years. The isolated pathogens were evaluated for susceptibilities to 19 antimicrobial agents used in humans. Out of 428 samples, 157 strains of Escherichia coli, 25 of Yersinia enterocolitica, 57 of Staphylococcus aureus, 57 of Enterococcus spp., 4 of Salmonella spp., and 3 of Campylobacter jejuni were isolated. Among the isolates 14.6% of the E. coli, 10.5% of S. aureus, 4% of Y. enterocolitica, 25% of Salmonella spp., and 42.1% of Enterococcus spp. were susceptible to antibiotics. E. coli from chicken exhibited high rates of resistance to ciprofloxacin (62.5%) followed by lamb/goat (10.9%), pork (15.7%), and beef (27.9%) meat. Resistance to nitrofurantoin dominated in the lamb/goat isolates (60%). Resistance to tetracycline predominated in pork (68.2%) and chicken (62.5%), and resistance to aminoglycosides dominated in lamb/goat meat isolates. S. aureus resistance to clindamycin predominated in lamb/goat isolates (50%), whereas resistance to ciprofloxacin predominated in the pork strains, but no resistance to methicillin was observed. Of the enterococci isolates 21.1% were resistant to vancomycin. High resistance to ampicillin (96%) was observed in Y. enterocolitica and all of the C. jejuni isolates were resistant to ampicillin, cephalothin, and cefuroxime. These results indicate that meat can be a source of resistant bacteria, which could potentially be spread to the community through the food chain.

Prevalence, antimicrobial resistance and relation to indicator and pathogenic microorganisms of Salmonella enterica isolated from surface waters within an agricultural landscape.

During a 12 month period (June 2007-May 2008), the prevalence and susceptibility of Salmonella serovars and their relation to specific pathogenic and indicator bacteria in river and coastal waters was investigated. A total of 240 water samples were collected from selected sites in Acheron and Kalamas Rivers and the Ionian Sea coast in north western Greece. The samples were analyzed for Salmonella spp., Listeria spp., Campylobacter spp., Escherichia coli O157, Staphylococci, Pseudomonas spp., Total Coliforms, Fecal Coliforms, Fecal Streptococci, Total Heterotrophic Flora at 20°C and at 37°C, fungi and protozoa (Cryptosporidium, Giardia). Susceptibility tests to nine antimicrobials (ampicillin, amikacin, amoxicillin/clavulavic acid, cefuroxime, ciprofloxacin, cefoxitin, tetracycline, ticarcillin/clavulanic acid, ampicillin/sulbactam) were performed using the disk diffusion method for Salmonella isolates. We isolated 28 serovars of Salmonella spp. identified as Salmonella enteritidis (23), Salmonella thompson (3) and Salmonella virchow (2). Multi-drug resistant Salmonella serovars were isolated from both river and marine waters, with 34.8% of S. enteritidis and 100% of S. virchow being resistant to more than 3 antibiotics. Also we isolated 42 strains of Listeria spp. identified as L. monocytogenes (20), L. innocua (9), L. seeligeri (2) and L. ivanovii (11). All the Listeria isolates were susceptible to the tested antibiotics. No Campylobacter spp., E. coli O157, Cryptosporidium and Giardia were detected. The overall ranges (and average counts) of the indicator bacteria were: Total Coliforms 0-4×10(4)cfu/100ml (3.7×10(3)cfu/100ml), Fecal Coliforms 0-9×10(3)cfu/100ml (9.2×10(2)cfu/100ml), Fecal Streptococci 0-3.5×10(4)cfu/100ml (1.4×10(3)cfu/100ml), Total Heterotrophic Flora at 20°C 0-6×10(3)cfu/ml (10(3)cfu/ml) and at 37°C 0-5×10(3)cfu/ml (4.9×10(2)cfu/ml). Weak or non significant positive Spearman correlations (p<0.05, rs range: 0.13-0.77) were obtained between Salmonella, Listeria, fungi and indicator bacteria. The results underline the complexity of the interrelations between pathogens and indicator bacteria, and the necessity to assess the presence of resistant bacteria in the aquatic environments.

Chemical investigation and antimicrobial properties of mastic water and its major constituents.

Mastic water is a commercial flavouring obtained during the steam distillation of mastic resin (the resin of Pistacia lentiscus var. chia) for the production of mastic oil. The mastic water extracts were analysed by GC-MS. The major compounds identified were verbenone, α-terpineol, linalool and trans-pinocarveol. Overall the composition was found to be very different from that of mastic oil. Additional GC-MS revealed the enantiomeric ratio of the chiral constituents of mastic water. The antimicrobial activity of mastic water extract, as well as that of its major constituents, was examined against Escherichia coli, Staphylococcus aureus and Candida spp. including ATCC wild clinical and food-borne strains. Linalool and α-terpineol were found to be the most potent antimicrobial constituents. Finally the stability of mastic water at different temperatures was studied, showing no change in the GC-MS profile of the organic extract for a period of 4months at storage temperatures up to 4°C.

Microbial ethanol production: Experimental study and multivariate evaluation §

Ethanol can be produced from all the postmortem available substrates, though with higher rates and yields from carbohydrates, during the early stages of putrefaction. The so-called higher alcohols (1-propanol, isobutanol, 2-methyl-1-butanol and 3-methyl-2-butanol) and 1-butanol could be produced, from all the available postmortem substrates. However, a quantitative relationship between the produced ethanol and the potentially produced other alcohols is still missing. The objective of this study was the development of a simple, mathematical model which could be able to approximate the microbial produced ethanol in correlation with other produced alcohols. The selected bacterial species included two Gram+ spore-forming anaerobic bacteria and two (one Gram+ one Gram-) aerobic/facultative anaerobic bacteria, all being common commensals of the digestive tract and common colonizers of the corpse. The selected bacterial strains, Escherichia coli, Clostridium perfrigens, Clostridium sporogenes and Enterococcus faecalis, were cultured separately at 25 °C, for 30 days, under controlled anaerobic conditions. The produced ethanol and the previously referred alcohols were determined in the culture medium in 24h intervals. Using partial least squares (PLS) regression, the estimation of the relevance score for the available descriptors established the statistical model to assess the ethanol concentration produced by each studied microbe. E. coli, C. perfrigens, and C. sporogenes produced different patterns of ethanol and other alcohols, while E. faecalis produced negligible amounts of ethanol and higher alcohols. In constructing the mathematical models to predict the produced ethanol, 1-propanol, 1-butanol, and isobutanol were significant for C. perfrigens and C. sporogenes, while 1-butanol, 1-propanol, and methyl-butanol were significant for E. coli. The applicability of these models was tested in microbial, anaerobic cultures of normal human blood and plasma at 25 °C. The results indicate that factors such as the type of microbe species, the glucose content and the medium composition apparently affect the procedure of microbial ethanol, and other alcohols production. However, the models can be applied with acceptable accuracy and they show potential for application in real postmortem cases.

In vitro antimicrobial activity of five essential oils on multi-drug resistant Gram-negative clinical isolates -

Aim/Background: The emergence of drug-resistant pathogens has drawn attention on medicinal plants for potential antimicrobial properties. The objective of the present study was the investigation of the antimicrobial activity of five plant essential oils on multidrug resistant Gram-negative bacteria. Materials and Methods: Basil, chamomile blue, origanum, thyme, and tea tree oil were tested against clinical isolates of Acinetobacter baumannii (n = 6), Escherichia coli (n = 4), Klebsiella pneumoniae (n = 7), and Pseudomonas aeruginosa (n = 5) using the broth macrodilution method. Results: The tested essential oils produced variable antibacterial effect, while Chamomile blue oil demonstrated no antibacterial activity. Origanum, Thyme, and Basil oils were ineffective on P. aeruginosa isolates. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration values ranged from 0.12% to 1.50% (v/v) for tea tree oil, 0.25-4% (v/v) for origanum and thyme oil, 0.50% to >4% for basil oil and >4% for chamomile blue oil. Compared to literature data on reference strains, the reported MIC values were different by 2SD, denoting less successful antimicrobial activity against multidrug resistant isolates. Conclusions: The antimicrobial activities of the essential oils are influenced by the strain origin (wild, reference, drug sensitive, or resistant) and it should be taken into consideration whenever investigating the plants’ potential for developing new antimicrobials.

Modeling microbial ethanol production by E. coli under aerobic/anaerobic conditions: Applicability to real postmortem cases and to postmortem blood derived microbial cultures

The mathematical modeling of the microbial ethanol production under strict anaerobic experimental conditions for some bacterial species has been proposed by our research group as the first approximation to the quantification of the microbial ethanol production in cases where other alcohols were produced simultaneously with ethanol. The present study aims to: (i) study the microbial ethanol production by Escherichia coli under controlled aerobic/anaerobic conditions; (ii) model the correlation between the microbial produced ethanol and the other higher alcohols; and (iii) test their applicability in: (a) real postmortem cases that had positive BACs (>0.10 g/L) and co-detection of higher alcohols and 1-butanol during the original ethanol analysis and (b) postmortem blood derived microbial cultures under aerobic/anaerobic controlled experimental conditions. The statistical evaluation of the results revealed that the formulated models were presumably correlated to 1-propanol and 1-butanol which were recognized as the most significant descriptors of the modeling process. The significance of 1-propanol and 1-butanol as descriptors was so powerful that they could be used as the only independent variables to create a simple and satisfactory model. The current models showed a potential for application to estimate microbial ethanol - within an acceptable standard error - in various tested cases where ethanol and other alcohols have been produced from different microbes.

Screening for antibiotic residues in swine and poultry tissues using the STAR test

During a 12-month period, 138 tissue samples (muscle, liver, kidney) were screened for antibiotic residues (ARs), using the five-plate STAR test. Samples positive to one or more antibiotics were detected in 33.9% of the chicken samples and in 26% of the pig samples. All the chicken liver samples, 32.2% of the chicken muscle and 14% of the swine liver samples were positive to sulphonamides and β-lactams, 25% of the kidney, 11.1% of the liver and 4% of the muscle tissue of the swine samples and 1.7% of the chicken muscle samples were positive to tetracycline, 12.5% of the kidney and 4% of the muscle swine samples were positive to aminoglycosides and macrolides. No quinolone residues were detected in any samples. The use of a microbiological method such as the STAR-test, for ARs screening in food is an effective low cost scheme indicating potential contamination with ARs.