Potential sources of microbial contamination in unpasteurized apple cider

A study was conducted to identify possible sources of microbial contamination and to assess the effect of good cleaning and sanitation practices on the microbial quality and safety of unpasteurized apple cider. Raw unwashed apples, washed apples, cleaning water, fresh cider, and finished cider samples were collected from five Ontario producers over 4 months and microbiologically tested. Total coliforms were found in 31, 71 and 38% of the unwashed apple, water, and washed apple samples, respectively. Escherichia coli was found in 40% of the water samples from one producer alone. The washing step was identified as a potential source of contamination, possibly due to water in the dump tanks seldom being refreshed, and because scrubbers, spray nozzles, and conveyors were not properly cleaned and sanitized. Higher total coliform counts (P < 0.0001) and prevalence (P < 0.0001) in fresh cider compared with those in unwashed apples and washed apples indicated considerable microbial buildup along the process, possibly explained by the lack of appropriate equipment sanitation procedures. Results showed that producers who had better sanitary practices in place had lower (P < 0.001) total coliform prevalence than the rest of the producers. Overall results show that good sanitation procedures are associated with improved microbial quality of fresh cider in terms of total coliforms and that operators who pasteurize and/or UV treat their product should still be required to have a sound good manufacturing practices program in place to prevent recontamination. Cryptosporidium parvum, an important pathogen for this industry, was found in different sample types, including washed apples, water, and fresh and finished cider.     

Use of hazard analysis critical control point and alternative treatments in the production of apple cider.

The purpose of this study was to evaluate the practices of Maryland cider producers and determine whether implementing hazard analysis critical control point (HACCP) would reduce the microbial contamination of cider. Cider producers (n = 11) were surveyed to determine existing manufacturing practices and sanitation. A training program was then conducted to inform operators of safety issues, including contamination with Escherichia coli O157:H7, and teach HACCP concepts and principles, sanitation procedures, and good manufacturing practice (GMP). Although all operators used a control strategy from one of the model HACCP plans provided, only one developed a written HACCP plan. None developed specific GMP, sanitation standard operating procedures, or sanitation monitoring records. Six operators changed or added production controls, including the exclusion of windfall apples, sanitizing apples chemically and by hot dip, and cider treatment with UV light or pasteurization. Facility inspections indicated improved sanitation and hazard control but identified ongoing problems. Microbiological evaluation of bottled cider before and after training, in-line apples, pomace, cider, and inoculated apples was conducted. E. coli O157:H7, Salmonella, or Staphylococcus aureus were not found in samples of in-line apple, pomace, and cider, or bottled cider. Generic E. coli was not isolated on in-coming apples but was found in 4 of 32 (13%) in-line samples and 3 of 17 (18%) bottled fresh cider samples, suggesting that E. coli was introduced during in-plant processing. To produce pathogen-free cider, operators must strictly conform to GMP and sanitation procedures in addition to HACCP controls. Controls aimed at preventing or eliminating pathogens on source apples are critical but alone may not be sufficient for product safety.     

Microbial levels in Michigan apple cider and their association with manufacturing practices

In recent decades, apple cider has been implicated in a series of outbreaks of foodborne illness. The objective of this study was to determine the presence and concentrations of pathogenic and indicator microorganisms in apple cider processed in Michigan and to evaluate the impact of thermal pasteurization, UV light radiation, and implementation of hazard analysis critical control point (HACCP) plans on these microbes. Cider samples were obtained from Michigan mills between 1997 and 2004 and analyzed for Escherichia coli O157:H7, Salmonella, generic E. coli, total coliforms, and aerobic bacteria. Neither E. coli O157:H7 nor Salmonella were detected in any tested cider samples, suggesting a very low frequency of pathogens in Michigan apple cider. The persistent and relatively high frequency of generic E. coli observed in samples obtained in all years indicates a continued risk of pathogen contamination in Michigan apple cider, especially when it is untreated. The use of thermal pasteurization or UV light radiation and reported implementation of HACCP plans were associated with lower frequency and counts of generic E. coli, total coliforms, and aerobic microorganisms. However, the relatively high counts of indicator organisms in some cider samples that were claimed to be treated according to these pathogen reduction measures indicates that some processors had inadequate practices, facilities, or equipment for pathogen reduction or did not consistently or adequately apply practices or pathogen-reduction equipment in an effective manner.     

SURVEY OF APPLE GROWING, HARVESTING, AND CIDER MANUFACTURING PRACTICES IN WISCONSIN: IMPLICATIONS FOR SAFETY

To evaluate the safety of current apple growing, harvesting and cider manufacturing practices in Wisconsin, cider manufacturers were contacted in a three-phase survey. Results revealed that seasonal, small-scale production was characteristic of the industry. Most cider mills produced less than 5,000 gal/year; only 6% produced more than 20,000 gal/year. Most cider makers used only tree picked apples (86%), inspected apples before washing (94%), washed (93%) and brushed (87%) apples, but only 16% of mills sanitized washed apples. Most mills (92%) sanitized cider making equipment after each use; however, only a few sanitized between custom pressing apples from different customers. Respondents reported that they strived to improve cider safety by pasteurization (43% of all cider), UV light treatment (4%), use of preservatives (30%), and HACCP (17%). For 31% of all cider, however, processors relied solely on refrigeration and/or freezing. These results show that most cider mills practice many steps believed to enhance cider safety, but results also identify procedures that must be addressed to further improve cider safety.     

Isolation and characterization of Escherichia coli recovered from Maryland apple cider and the cider production environment

Contaminated apple cider has been implicated in several Escherichia coli O157:H7 outbreaks. In an attempt to investigate sources and modes of entry of E. coli into apple cider, samples of fresh apple, pomace, and cider and equipment and mill floor swabs were analyzed for standard plate counts (SPC), total coliforms (TC), fecal coliforms (FC), and E. coli. E. coli was isolated from 14 (33%) of 42 samples of bottled fresh cider, from food equipment in 6 (67%) of 9 mills, and from apples, pomace, or cider in 7 (78%) of 9 mills. Seventy-five E. coli isolates were further characterized for Shiga toxin-producing E. coli (STEC)-associated virulence factors, antimicrobial susceptibility, and pulsed-field gel electrophoresis (PFGE) type. No E. coli O157:H7 or other STEC was identified. Serotyping and PFGE revealed 64 distinct profiles, suggesting that recovered E. coli arose from multiple independent sources. However, on one occasion, E. coli isolated from the source apple sample was closely related to the E. coli identified in the finished cider sample. E. coli isolates were further tested for antimicrobial susceptibility to 17 antimicrobial agents of human and veterinary importance. Fourteen (19%) of the 75 isolates were resistant to at least one of the antimicrobial agents tested, and 9 (12%) were resistant to at least two of these agents. Of the resistant isolates recovered, 64% were resistant to tetracycline and 57% were resistant to streptomycin. Overall, the level of E. coli contamination in source apple samples did not differ significantly from those in samples of pomace, cider at the press, and cider entering the bottling tank; therefore, source apples cannot be dismissed as a potential contributor of E. coli to the cider-making process.     

Inactivation of Escherichia coli O157:H7 and other naturally occurring microorganisms in apple cider by electron beam irradiation

Two Escherichia coli O157:H7 strains, SEA 13 B88 gfp 73ec and B6-914 gfp 90ec, together with two bacteria, three yeasts, and two molds that were randomly selected from a collection of microorganisms found on apples or in apple cider, were inoculated into apple cider and subjected to electron beam irradiation at several doses between 0.0 and 2.3 kGy at the Iowa State University Linear Accelerator Facility. The D-values for the E. coli O157:H7 strains ranged between 0.25 and 0.34 kGy; the D-values for most of the normal flora from apples ranged between 0.24 and 0.59 kGy. By taking into account possible variations in treatment conditions, it was calculated that irradiation at 2.47 kGy should achieve a 5-log reduction of E. coli O157:H7 in apple cider at the 95% confidence level. Naturally occurring yeasts might survive such irradiation treatment.     

干酪中肠球菌种群的遗传差异及万古霉素抗性基因分析

通过种、属引物特异性扩增、重复序列PCR（rep-PCR）技术和万古霉素抗性基因检测对新疆北疆地区干酪样品中球菌的遗传结构差异进行分析。结果表明,15份样品共分离52株肠球菌,包括31株耐久肠球菌(Enterococcus durans)、18株粪肠球菌(Enterococcus faecalis)和3株屎肠球菌(Enterococcus faecium)。依据rep-PCR遗传指纹带谱分析,52株菌可以聚类成7个群,其中4个由E. durans构成。24株肠球菌检测到了万古霉素抗性基因,17株E. durans为VanC2/C3型,4株E. faecalis为VanC1型,2株E. faecium为VanB型,只有1株E. faecium为VanA型。新疆北疆地区干酪中肠球菌种群分布较为广泛,地域之间优势种群和基因型不同,同种肠球菌菌株之间存在广泛的遗传差异。     

Prevalence of Escherichia coli in apple cider manufactured in Connecticut.

Cider samples obtained from 11 cider mills operating in Connecticut during the 1997 to 1998 production season were tested for the presence of Escherichia coli. Cider production began in mid August and continued through March, with peak production in September and October. Of 314 cider samples tested, 11 (4%) were found to contain E. coli. Of the 11 mills, 6 (55%) tested positive for E. coli in the cider at least once during the production year. E. coli was first observed in cider samples produced in mid to late October and was not detected in samples made after January. A trend was observed for cider to decrease in acidity and increase in Brix (soluble sugars) throughout the production season. No correlation between pH and soluble sugars of cider and the presence of E. coli was detected. Eight mills used both dropped apples and tree-picked apples, whereas three mills used tree-picked apples only. The use of dropped apples in cider production began 5 weeks before the first detection of E. coli in cider. E. coli was isolated from cider samples produced using dropped apples and from samples produced using only tree-picked apples. No direct correlation between the use of dropped apples or tree-picked apples and the presence of E. coli in the cider was observed. An association between the time of apple harvest and the appearance of E. coli in cider was noted. For mills providing adequate records, all contaminated cider was produced from apples harvested between mid October and mid November.     

Location of Escherichia coli O157:H7 on and in apples as affected by bruising, washing, and rubbing

Confocal scanning laser microscopy (CSLM) was used to determine the location of Escherichia coli O157:H7 cells on the surface and in tissue of bruised Red Delicious cv. apples. Undamaged and bruised apples were inoculated by immersing in a suspension of E. coli O157:H7 cells transformed with a plasmid that encodes for the production of a green fluorescent protein. Apples were then washed in 0.1% (wt/vol) peptone water and/or rubbed with a polyester cloth and examined to determine if these treatments removed or introduced cells into lenticels, cutin, and cracks on the skin surface. Optical slices of the apples obtained using CSLM were examined to determine the depth at which colonization or attachment of cells occurred. Populations of E. coli O157:H7 on the surface of apples were determined to assess the effectiveness of washing and rubbing in physically removing cells. The location of cells on or in undamaged and bruised areas of apples that were not washed or rubbed did not differ significantly. However, washing apples resulted in an approximate 2-log reduction in CFU of E. coli O157:H7 per cm2 of apple surface. On unwashed apples, cells were detected at depths up to 30 microm below the surface. No E. coli O157:H7 cells were detected at locations more than 6 microm below the surface of washed apples. Cells that remained on the surface of rubbed apples appeared to be sealed within naturally occurring cracks and crevices in waxy cutin platelets. These cells may be protected from disinfection and subsequently released when apples are eaten or pressed for cider production.     

A study of U.S. orchards to identify potential sources of Escherichia coli O157:H7

The association of unpasteurized apple cider with Escherichia coli O157:H7 foodborne illness has led to increased interest in potential reservoirs of this pathogen in the orchard. Fourteen U.S. orchards were surveyed in autumn 1999 to determine the incidence and prevalence of E. coli O157:H7, E. coli, total aerobic microflora, and yeasts and molds. Fruit samples (n = 63) (eight apple and two pear varieties) and soil, water, and fecal samples were collected. Samples were plated on (i) tryptic soy agar for total mesophilic aerobic count, (ii) E. coli and coliform Petrifilm for total coliforms and E. coli, and (iii) yeast and mold Petrifilm. Samples positive for coliforms and E. coli were enriched and tested for E. coli O157:H7. Fruit was also tested for internalization of microflora by aseptically removing the core, stem, and calyx areas, and the individual sections were assessed for the categories of microflora listed above. E. coli was detected in soil and water and in 6% of fruit samples (three pear samples and one apple sample), generally collected from areas previously designated as high risk in this study. However, no E. coli O157:H7 was found. Coliforms were found in 74% of fruit samples and were internalized in the cores of 40% of fruit tested. Yeasts and molds were internalized in 96.7% of samples and aerobic bacteria in 89.6%. E. coli was not found to be internalized. Total aerobic counts and total coliforms were higher in dropped and damaged fruit (P < 0.05). Findings suggest that dropped or damaged fruit should not be included in fruit designated for the production of unpasteurized juice or for the fresh or fresh-cut market. In addition, orchards should be located away from potential sources of contamination, such as pastures.     

Survival of Escherichia coli O157:H7 and Salmonella in apple cider and orange juice as affected by ozone and treatment temperature

Inactivation of Escherichia coli O157:H7 and Salmonella in apple cider and orange juice treated with ozone was evaluated. A five-strain mixture of E. coli O157:H7 or a five-serovar mixture of Salmonella was inoculated (7 log CFU/ml) into apple cider and orange juice. Ozone (0.9 g/h) was pumped into juices maintained at 4 degrees C, ambient temperature (approximately 20 degrees C), and 50 degrees C for up to 240 min, depending on organism, juice, and treatment temperature. Samples were withdrawn, diluted in 0.1% peptone water, and surface plated onto recovery media. Recovery of E. coli O157:H7 was compared on tryptic soy agar (TSA), sorbitol MacConkey agar, hemorrhagic coli agar, and modified eosin methylene blue agar; recovery of Salmonella was compared on TSA, bismuth sulfite agar, and xylose lysine tergitol 4 (XLT4) agar. After treatment at 50 degrees C, E. coli O157:H7 populations were undetectable (limit of 1.0 log CFU/ml; a minimum 6.0-log CFU/ml reduction) after 45 min in apple cider and 75 min in orange juice. At 50 degrees C, Salmonella was reduced by 4.8 log CFU/ml (apple cider) and was undetectable in orange juice after 15 min. E. coli O157:H7 at 4 degrees C was reduced by 4.8 log CFU/ml in apple cider and by 5.4 log CFU/ml in orange juice. Salmonella was reduced by 4.5 log CFU/ml (apple cider) and 4.2 log CFU/ml (orange juice) at 4 degrees C. Treatment at ambient temperature resulted in population reductions of less than 5.0 log CFU/ml. Recovery of E. coli O157:H7 and Salmonella on selective media was substantially lower than recovery on TSA, indicating development of sublethal injury. Ozone treatment of apple cider and orange juice at 4 degrees C or in combination with mild heating (50 degrees C) may provide an alternative to thermal pasteurization for reduction of E. coli O157:H7 and Salmonella in apple cider and orange juice.     

HIGH PREVALENCE OF EXTENDED-SPECTRUM β-LACTAMASES ESCHERICHIA COLI AND VANCOMYCIN-RESISTANT ENTEROCOCCI ISOLATES FROM CHICKEN PRODUCTS. A PROBLEM OF PUBLIC HEALTH

Twenty-nine chicken products were acquired from different supermarkets in Portugal during September to December 2007 and were analyzed for extended-spectrum β-lactamases (ESBL) Escherichia coli and vancomycin-resistant enterococci (VRE). Cefotaxime-resistant E. coli isolates were recovered in 27 of 29 chicken samples representing 93% of the analyzed samples. The highest percentages of resistance (more than 50% of the isolates) were detected for ampicillin, nalidixic acid and tetracycline. VRE isolates were detected in 17 of 29 samples of chicken origin (59%) and were identified as Enterococcus durans (n  =  15) and E. faecium (n = 2) with the highest percentages of resistance being detected for erythromycin, tetracycline, ampicillin and ciprofloxacin. Seven E. durans and the two E. faecium isolates recovered from chicken wings, gizzard and skin show gelatinase activity.
The high rate of colonization of chicken products by these bacteria supports other studies suggesting that the food chain could be a source of ESBL and VRE colonization in humans representing a public health problem.

PRACTICAL APPLICATIONS

The data indicate that chicken products may be contaminated with a high prevalence of extended-spectrum β-lactamases Escherichia coli and vancomycin-resistant enterococci (VRE). It is important to mention that the isolates present a diversity of phenotypes of antimicrobial resistance, and half of the VRE isolates show also gelatinase activity, indicating that these animals may be a reservoir of bacteria showing virulence and increased resistance to antimicrobial agents, raising special concerns about their transmission to humans through the food chain.     

Reduction of microbial pathogens during apple cider production using sodium hypochlorite, copper ion, and sonication

Sodium hypochlorite (100 ppm), copper ion water (1 ppm), and sonication (22 to 44 kHz and 44 to 48 kHz) were assessed individually and in combination for their ability to reduce populations of Escherichia coli O157:H7 and Listeria monocytogenes on apples and in apple cider. Commercial unpasteurized cider was inoculated to contain approximately 10(6) CFU/ml of either pathogen and then sonicated at 44 to 48 kHz, with aliquots removed at intervals of 30 to 60 s for up to 5 min and plated to determine numbers of survivors. Subsequently, whole apples were inoculated by dipping to contain approximately 10(6) CFU/g E. coli O157:H7 or L. monocytogenes, held overnight, and then submerged in 1 ppm copper ion water with or without 100 ppm sodium hypochlorite for 3 min with or without sonication at 22 to 44 kHz and examined for survivors. Treated apples were also juiced, with the resulting cider sonicated for 3 min. Populations of both pathogens decreased 1 to 2 log CFU/ml in inoculated cider following 3 min of sonication. Copper ion water alone did not significantly reduce populations of either pathogen on inoculated apples. However, when used in combination with sodium hypochlorite, pathogen levels decreased approximately 2.3 log CFU/g on apples. Sonication of this copper ion-sodium hypochlorite solution at 22 to 44 kHz did not further improve pathogen reduction on apples. Numbers of either pathogen in the juice fraction were approximately 1.2 log CFU/ml lower after being juiced, with sonication (44 to 48 kHz) of the expressed juice decreasing L. monocytogenes and E. coli O157:H7 populations an additional 2 log. Hence, a 5-log reduction was achievable for both pathogens with the use of copper ion water in combination with sodium hypochlorite followed by juicing and sonication at 44 to 48 kHz.     

Influence of fruit variety, harvest technique, quality sorting, and storage on the native microflora of unpasteurized apple cider

Apple variety, harvest, quality sorting, and storage practices were assessed to determine their impact on the microflora of unpasteurized cider. Seven apple varieties were harvested from the tree or the ground. The apples were used fresh or were stored at 0 to 4 degrees C for < or = 5 months and were pressed with or without quality selection. Cider yield, pH, Brix value, and titratable acidity were measured. Apples, postpressing apple pomace, and cider samples were analyzed for aerobic bacteria, yeasts, and molds. Aerobic bacterial plate counts (APCs) of ciders from fresh ground-picked apples (4.89 log CFU/ml) were higher than those of ciders made from fresh, tree-picked apples (3.45 log CFU/ml). Quality sorting further reduced the average APC to 2.88 log CFU/ml. Differences among all three treatment groups were significant (P < 0.0001). Apple and pomace microbial concentrations revealed harvest and postharvest treatment-dependent differences similar to those found in cider. There were significant differences in APC among apple varieties (P = 0.0001). Lower counts were associated with varieties exhibiting higher Brix values and higher titratable acidity. Differences in APC for stored and fresh apples used for cider production were not significant (P > 0.05). Yeast and mold counts revealed relationships similar to those for APCs. The relationship between initial microbial load found on incoming fruit and final cider microbial population was curvilinear, with the weakest correlations for the lowest apple microflora concentrations. The lack of linearity suggests that processing equipment contributed to cider contamination. Tree-picked quality fruit should be used for unpasteurized cider production, and careful manufacturing practices at cider plants can impact both safety and quality of the final product.     

Efficacy of sanitation and cleaning methods in a small apple cider mill

The efficacy of cleaning and sanitation in a small apple cider processing plant was evaluated by surface swab methods as well as microbiological examination of incoming raw ingredients and of the final product. Surface swabs revealed that hard-to-clean areas such as apple mills or tubing for pomace and juice transfer may continue to harbor contaminants even after cleaning and sanitation. Use of poor quality ingredients and poor sanitation led to an increase of approximately 2 logs in aerobic plate counts of the final product. Reuse of uncleaned press cloths contributed to increased microbiological counts in the finished juice. Finally, using apples inoculated with Escherichia coli K-12 in the plant resulted in an established population within the plant that was not removed during normal cleaning and sanitation. The data presented in this study suggest that current sanitary practices within a typical small cider facility are insufficient to remove potential pathogens.     

Effect of Enterococcus faecium on microbiological,physicochemical and sensory characteristics of Greek Feta cheese

Greek Feta cheese was prepared using as adjunct starter cultures Enterococcus faecium FAIR-E 198, E. faecium FAIR-E 243, and their combination. Numbers of enterococci in the control and in the batches containing E. faecium strains as adjunct starters rapidly increased until day 15 of ripening, and then remained constant. Both E. faecium strains positively affected the counts of non-starter lactic acid bacteria (NSLAB), micrococci and coliforms, while thermophilic cocci were not influenced. Moreover. E. faecium FAIR-E 243 enhanced the growth of mesophilic cocci and thermophilic bacilli. Physicochemical characteristics, such as pH, moisture, ash, salt in moisture and fat in dry matter (FDM) were not influenced by the addition of the E. faecium strains. The most pronounced effect was observed in the case of proteolysis. Both E. faecium strains, either as sole adjunct starter or in combination, increased the proteolytic index and the free amino groups concentration, and enhanced degradation of alpha(s1)- and beta-caseins in comparison to the control. Furthermore, the reverse-phase (RP)-HPLC peptide profiles of the water-soluble nitrogen (WSN) fractions were significantly affected by the addition of enterococci. The main volatile compounds produced were ethanol, acetate, acetone, acetaldehyde, acetoin and diacetyl, with highest amounts determined for ethanol, followed by acetate. Both E. faecium strains positively affected taste, aroma, colour and structure of the full-ripened cheeses, as well as the overall sensory profile. The present work emphasizes the technological significance of E. faecium strains and supports their use as adjunct cultures in the manufacture of Feta cheese.     

Survival differences of Escherichia coli O157:H7 strains in apples of three varieties stored at various temperatures

Differences in survival and growth among five different Escherichia coli O157:H7 strains in three apple varieties were determined at various temperatures. Jonathan, Golden Delicious, and Red Delicious apples were wounded and inoculated with E coli O157:H7 strains C7929 (apple cider isolate), 301C (chicken isolate), 204P (pork isolate), 933 (beef isolate), and 43890 (human isolate) at an initial level of 6 to 7 log CFU/g. The inoculated apples were stored at a constant temperature of 37, 25, 8, or 4 degrees C or at 37 degrees C for 24 h and then at 4 degrees C, and bacterial counts were determined every week for 28 days. By day 28, for Jonathan apples at 25 degrees C, the apple isolate counts were significantly higher than the chicken and human isolate counts. At 4 degrees C for 28 days, the human isolate inoculated into Jonathan, Golden Delicious, and Red Delicious apples was present in significantly smaller numbers than the other strains. The apple isolate survived significantly better at 4 degrees C, yielding the highest number of viable cells. By days 21 and 28, for apples stored at 37 degrees C for the first 24 h and then at 4 degrees C, the counts of viable E. coli O157:H7 apple and human isolates were 6.8 and 5.8 log CFU/g at the site of the wound, whereas for apples kept at 4 degrees C for the duration of storage, the respective counts were 5.6 and 1.5 log CFU/g. Our study shows that E. coli O157:H7 strains responded differentially to their ability to survive in these three apple varieties at 25 or 4 degrees C and produced higher viable counts when apples were temperature abused at 37 degrees C for 24 h and then stored at 4 degrees C for 27 days.     

Enterococcus populations in Pecorino Abruzzese cheese: biodiversity and safety aspects

The presence of enterococci in Pecorino Abruzzese cheese during ripening was evaluated. Counts were high, especially in fully ripened summer batches. Seventy strains were isolated and identified based on phenotypical and genotypical features as Enterococcus faecium (48.5%), Enterococcus faecalis (40%), and Enterococcus durans (11.5%), with the first species predominant in spring batches and the second predominant in summer batches. High biodiversity was revealed by random amplification of polymorphic DNA and a PCR assay, suggesting the presence of autochthonous strains. E. faecium isolates were the most resistant to the tested antibiotics, especially to erythromycin, chloramphenicol, and penicillin, but all strains were susceptible to vancomycin, as confirmed by the absence of vanA and vanB genes. The presence of some virulence determinants was investigated, revealing the diffusion of aggregation substance (asal) and gelatinase (gelE) genes in 37.5% of E. faecalis strains. However, none of the isolates produced gelatinase in vitro, suggesting the presence of silent genes. The virulence genes were absent in E. durans. Among E. faecium strains, only Lab 41/1 possessed gelE and asal, whose presence previously has been reported only in E. faecalis. Decarboxylating activity was revealed for phenylalanine (27% of the strains) and tyrosine (96%) but not histidine. The presence of a tyrosine decarboxylase-encoding gene was observed for all strains. A comparison of these results with those of previous studies of clinical and food isolates indicates that enterococci from Pecorino Abruzzese cheese have low pathogenic potential.     

Species distribution and antibiotic resistance patterns of enterococci isolated from food of animal origin in Germany

Presently, enterococci take the third place of bacterial pathogens associated with nosocomial infections, after staphylococci and Escherichia coli. Especially, the resistances of enterococci to several available antibiotics are threatening. We attempted to determine which species of enterococci could be found in food of animal origin and their significance according to their antibiotic resistances for human beings. From November 2000 to May 2002 we investigated 155 samples of food of animal origin bought in retail outlets in Germany: 27 samples of sausages, 19 of ham, 83 of minced meat, 26 of cheese. From these food samples we isolated 416 enterococcal strains. The most frequent species was Enterococcus faecalis (299 strains); furthermore, we found Enterococcus faecium (54 strains), Enterococcus durans together with Enterococcus hirae (24 strains), Enterococcus casseliflavus (22 strains), Enterococcus avium (9 strains) and Enterococcus gallinarum (8 strains). We focused on the resistance patterns of 118 selected E. faecium and E. faecalis strains to 13 antimicrobial active agents (ampicillin, amoxicillin/clavulanic acid, avilamycin, chloramphenicol, enrofloxacin, erythromycin, flavomycin, gentamicin, penicillin, quinupristin/dalfopristin, teicoplanin, tetracycline and vancomycin). From the clinical point of view, the situation of antibiotic resistance to the examined antimicrobial agents seemed to be favourable. The investigated strains were sensitive to ampicillin and amoxicillin/clavulanic acid. These antibiotics are, in combination with an aminoglycoside, for example gentamicin, agents of choice for the treatment of enterococcal infections in human medicine. Only one E. faecium strain was resistant to penicillin, while all strains were sensitive to the glycopeptide antibiotics, vancomycin and teicoplanin. Resistances found against the antibiotics, tetracycline, quinupristin/dalfopristin and erythromycin, are causes for concern.