Evaluation of the nifH gene marker of Methanobrevibacter smithii for the detection of sewage pollution in environmental waters in Southeast Queensland, Australia

This study aimed at evaluating the host-specificity and -sensitivity of the nifH gene marker of Methanobrevibacter smithii by screening 272 fecal and wastewater samples from 11 animal species including humans in Southeast Queensland (SEQ), Australia. In addition, environmental water samples (n = 21) were collected during the dry and wet weather conditions and tested for the presence of the nifH marker along with other sewage-associated markers, namely, enterococci surface protein (esp) found in Enterococci faecium, Bacteroides HF183, adenoviruses (AVs), and polyomaviruses (PVs). The overall host-specificity of the nifH marker to differentiate between human and animal feces was 0.96 (maximum value of 1), while the overall sensitivity of this marker in human sourced feces and wastewater was 0.81 (maximum value of 1). Among the 21 environmental water samples tested, 2 (10%), 3 (14%), 12 (57%), 6 (29%), and 6 (29%) were positive for the nifH, esp, HF183, AVs and PVs markers, respectively. The prevalence of the nifH marker in environmental water samples, however, was low compared to other markers, suggesting that the use of this marker alone may not be sensitive enough to detect fecal pollution in environmental waters. The nifH marker, however, appears to be sewage-specific in SEQ, Australia, and therefore, it is recommended that this marker should be used as an additional marker in combination with the HF183 or viral markers such as AVs or PVs for accurate and sensitive detection of fecal pollution in SEQ waterways.     

Interlaboratory comparison of real-time PCR protocols for quantification of general fecal indicator bacteria

The application of quantitative real-time PCR (qPCR) technologies for the rapid identification of fecal bacteria in environmental waters is being considered for use as a national water quality metric in the United States. The transition from research tool to a standardized protocol requires information on the reproducibility and sources of variation associated with qPCR methodology across laboratories. This study examines interlaboratory variability in the measurement of enterococci and Bacteroidales concentrations from standardized, spiked, and environmental sources of DNA using the Entero1a and GenBac3 qPCR methods, respectively. Comparisons are based on data generated from eight different research facilities. Special attention was placed on the influence of the DNA isolation step and effect of simplex and multiplex amplification approaches on interlaboratory variability. Results suggest that a crude lysate is sufficient for DNA isolation unless environmental samples contain substances that can inhibit qPCR amplification. No appreciable difference was observed between simplex and multiplex amplification approaches. Overall, interlaboratory variability levels remained low (<10% coefficient of variation) regardless of qPCR protocol.     

Beach sands along the California coast are diffuse sources of fecal bacteria to coastal waters

Fecal indicator bacteria (FIB) are nearly ubiquitous in California (CA) beach sands. Sands were collected from 55 beaches along the CA coast. Ninety-one percent of the beaches had detectable enterococci (ENT) while 62% had detectable E. coli (EC) in their sands. The presence of a putative bacterial source (such as a river), the degree of wave shelter, and surrounding land use explained a significant (p < 0.05) fraction of the variation in both ENT and EC densities between beaches. Sand characteristics including moisture content, organic carbon, and percentfines, significantly (p < 0.05) influenced only EC densities in beach sand. We assayed 34 of 163 sand samples for salmonellae, but did not detect this bacterial pathogen. The potential for FIB to be transported from the sand to sea was investigated at a single wave-sheltered beach with high densities of ENT in beach sand: Lovers Point, CA (LP). We collected samples of exposed and submerged sands as well as water over a 24 h period in order to compare the disappearance or appearance of ENT in sand and the water column. Exposed sands had significantly higher densities of ENT than submerged sands with the highest densities located near the high tide line. Water column ENT densities began low, increased sharply during the first flood tide and slowly decreased over the remainder of the study. During the first flood tide, the number of ENT that entered the water column was nearly equivalent to the number of ENT lost from exposed sands when they were submerged by seawater. The decrease in nearshore ENT concentrations after the initial influx can be explained by ENT die-off and dilution with clean ocean water. While some ENT in the water and sand at LP might be of human origin because they were positive for the esp gene, others lacked the esp gene and were therefore equivocal with respect to their origin. Follow-up sampling at LP revealed the presence of the human specific Bacteroides marker in water and sand.     

Potential use of presumptive enterococci and staphylococci as indicators of sanitary condition in plants making hard Italian-type cheese

Raw milk, pasteurized milk, unripened cheese (1 day old), and partially ripened cheese (3 months) from 42 milk lots at a plant making hard Italian-type cheese were analyzed for presumptive enterococci using kanamycin esculin azide agar pour plates. Fully ripened (> or =10 months) cheeses, derived from other milk lots, were also tested. Numbers of presumptive staphylococci (Baird-Parker agar [B-P]) were determined in the partially and fully ripened cheeses. Presumptive enterococci were ubiquitous in raw milk, usually at levels of 2.1 to 3.0 log CFU/ml. Enterococci were detected in 11 (26%) of 42 pasteurized milk samples. Enterococci and staphylococci were detected in 39 (93%) and 6 (14%) of unripened cheeses and in 33 (80%) and 4 (10%) of partially ripened cheeses, respectively. Only eight and five samples of enterococci-positive unripened and partially ripened cheese, respectively, were made from pasteurized milk in which presumptive enterococci were detected. Of 42 samples of fully ripened cheese, 35 (83%) and 8 (19%), respectively, contained presumptive enterococci and staphylococci. Results suggest either that low numbers of presumptive enterococci survive pasteurization and cheese ripening or that contamination of cheese by enterococci occurs after pasteurization. Biochemical testing confirmed 63% of presumptive enterococci isolates. None of the 20 presumptive staphylococci isolates produced colonies typical of Staphylococcus aureus on B-P agar; the isolates were identified as 1 Staphylococcus epidermidis, 1 Staphylococcus xylosus, 2 Staphylococcus saprophyticus, 1 Staphylococcus warneri, 5 Kocuria spp., and 10 unidentified gram-positive, catalase-positive cocci. Three staphylococci isolates decreased in numbers by more than 3.0 log CFU/ml in 9.9 ml of skim milk heated 30 min in a 62.8 degrees C water bath. This finding suggests that most presumptive staphylococci detected may have been prepasteurization contaminants. Unless specificity of the kanamycin esculin azide and B-P media is improved, use of presumptive enterococci and staphylococci as indicators of postpasteurization sanitation in plants making hard Italian-type cheese cannot be recommended.     

Occurrence and preliminary study of antimicrobial resistance of enterococci isolated from dairy goats in Spain

The present study performed in Spain was designed to investigate the occurrence and antimicrobial resistance of enterococci in faecal and bulk tank milk samples from 222 healthy Murciano-Granadina dairy goats reared on 12 farms. Enterococci were isolated in 54.5% and 63.6% of the faecal and bulk tank milk samples, respectively. Enterococci were detected more frequently from goat kids (70%) than from replacement animals (44.4%) and adults (56.4%). Seven species were found in the faecal samples but the most common species detected were Enterococcus faecium (32.3%), Enterococcus faecalis (27.6%) and Enterococcus hiriae (22%). In contrast, only E. faecium and E. faecalis were found in the bulk tank milk samples. According to NCCLS (2002), of the 134 strains studied, 17 (12.7%) were resistant to at least 7 antimicrobials and 14 of these strains were resistant to vancomycin.     

Tidal forcing of enterococci at marine recreational beaches at fortnightly and semidiurnal frequencies

Marine beach water quality is typically monitored in early morning once a week without respect to tidal condition. To assess the effect of tide on this public health warning system, we analyzed enterococci (ENT) data from 60 southern California marine beaches with differing geomorphology, orientation, and proximity to runoff sources. ENT concentrations during spring tides were significantly higher (p < 0.1) than those during neap tides at 50 of the beaches, and at the majority of these, water samples were also more than twice as likely to be out of compliance with the ENT single-sample standard during spring tides compared to neap tides. When tide range (spring/neap) and tide stage (ebb/flood) conditions were considered together, spring-ebb tides yielded the highest ENT concentrations and the greatest chance of exceeding the single-sample standard at the majority of beaches. The proximity to a terrestrial runoff source, the slope of the runoff source, the slope of the beach, and the orientation of the beach had minimal influence on the tidal modulation of ENT concentrations. The presence of spring and spring-ebb tide signals at such a great percentage of beaches suggests that tide should be considered in the design and interpretation of beach monitoring program data. It also suggests that ENT delivered by tidally forced sources other than terrestrial surficial runoff are widespread. Possibilities include ENT-laden groundwater (saline and fresh) from the beach aquifer as well as ENT-enriched sands, decaying wrack, and bird feces near the high water line.     

Prevalence of glycopeptide and aminoglycoside resistance in Enterococcus and Listeria spp. in low microbial load diets of neutropenic hospital patients

Low microbial load diets for patients with haematological malignancy were examined for enterococci and listeria using pre-enrichment, enrichment and selective plating. Enterococci were highly prevalent and their ecology diverse; 100/211 samples yielded 132 isolates made up of 67 strains distinguishable by PFGE. Listeria monocytogenes was not found. Screening of enterococci for antibiotic resistance showed low level vancomycin resistance (6-12 microg/ml) in six isolates of E. gallinarum and high level streptomycin resistance (> or = 1000 microg/ml) in eight isolates from various foods. No strains showing high level glycopeptide or gentamicin resistance were found. The high prevalence of enterococci in food processed for safety indicates a possible route for the acquisition of antibiotic-resistant strains by vulnerable hospital patients.     

Vancomycin-resistant enterococci from animal sources in Korea

Enterococci for which the minimum inhibitory concentration (MIC) of vancomycin was >/=8 mg/l were isolated from meat, feces, and raw milk samples collected in Korea from March to November 2003. Among the 243 vancomycin-resistant enterococci (VRE) that were identified the vanA vancomycin resistance gene was carried by 51 Enterococcus faecium and one Enterococcus sp., vanC1 was carried by 151 Enterococcus gallinarum, vanC2 was carried by 39 Enterococcus casseliflavus, and one Enterococcus sp. carried no van genes. Of the isolated enterococci carrying vanA, 4% were found to be highly resistant to gentamicin and 11% were resistant to ampicillin. Further genotyping of the E. faecium isolates carrying vanA using pulsed-field gel electrophoresis (PFGE) revealed extensive heterogeneity. The vancomycin resistance transferability test revealed that only two of the 52 enterococci carrying the vanA gene were able to transfer vancomycin resistance to other enterococci. The VRE were recovered from various animal sources with a particularly high prevalence of E. faecium carrying the vanA gene being found in poultry meat.     

Enterococci predictions from partial least squares regression models in conjunction with a single-sample standard improve the efficacy of beach management advisories

Beach health advisories are issued if enterococci (ENT) densities exceed the 30-d geometric mean or single-sample water quality criteria. Current ENT enumeration procedures require 1 day of incubation; therefore, beach managers make policy decisions using 1-day-old data. This is tantamount to using a model that assumes ENT density on day t is equal to ENT density on day t-1. Research has shown that ENT densities vary over time scales shorterthan a day, calling into question the usefulness of the current model for decision-making. We created Dynamic Partial Least Square Regression (DPLSR) models for ENT at water quality monitoring stations within two adjacent marine recreational sites, Huntington State Beach (HSB) and Huntington City (HCB) Beach, California, using publicly available environmental data and tested whether these models overcome the drawbacks of the current model. The DPLSR models provide a better prediction of ENT than the current models based on comparisons of root-mean-square errors of prediction and the numbers of type 1 and 2 errors. We compared outcomes in terms of predicted illness, swimmers deterred from entering the water, and net benefits to swimmers for hypothetical management scenarios where beach advisories were issued based on (a) the previously collected sample's ENT density in conjunction with the two water quality criteria, and (b) predictions from DPLSR models in conjunction with the single-sample standard. At both HSB and HCB the DPLSR scenario produced a more favorable balance between illness prevention and recreational access. The results call into question the current method of beach management and show that model-informed decision-making and elimination of the geometric mean standard will aid beach managers in achieving more favorable outcomes in terms of illness and access than are presently achieved using 1-day-old measurements, especially at beaches where water quality problems are chronic.     

Vancomycin resistance and antibiotic susceptibility of enterococci in raw meat

The purpose of this study was to investigate antimicrobial resistance, in particular to vancomycin, of enterococci in samples (100) of meat (beef, chicken, turkey, lamb, and pork) sold in retail outlets of Catanzaro (Italy). Enterococci were identified to the species level. Antimicrobial susceptibility tests for a large spectrum of antibiotics including glycopeptides were performed by the disk diffusion method. Kappa statistic was used to evaluate associations of resistance to vancomycin with other antimicrobials. Enterococci were isolated from 45% of the samples, mostly from chicken meat (65.4%). Overall, 29% of samples were contaminated by vancomycin-resistant enterococci (VRE), whereas among those positive they represented 64.4% of isolates. Higher prevalence of vancomycin resistance was found in chicken samples (76.5%). The overall resistance to teicoplanin (TRE) was 30%, whereas among those positive, TRE represented 66.7% of isolates. The most frequent isolates were Enterococcus faecium (35.6%) and Enterococcus faecalis (33.3%). Resistance to vancomycin and teicoplanin was observed in 75% and 78.5% of E. faecium, and in 40% and 46.7% of E. faecalis, respectively. Most strains were susceptible to ampicillin (80%), while 88.9% were resistant to methicillin. The most effective antimicrobials were imipenem (73.3% susceptible) and rifampin (80%). The highest prevalence of resistance was for streptomycin (88.9%), tetracycline (84.4%), and erythromycin (75.6%). Resistance to vancomycin was significantly associated to methicillin, teicoplanin, erythromycin, tetracycline, and chloramphenicol. Further investigations about enterococcal colonization and infections in community and hospital subjects are needed.     

Enterococci concentrations in diverse coastal environments exhibit extreme variability

Fecal indicator bacteria (FIB) concentrations in a single grab sample of water are used to notify the public about the safety of swimming in coastal waters. If concentrations are over a single-sample standard, waters are closed or placed under an advisory. Previous work has shown that notification errors occur often because FIB vary more quickly than monitoring results can be obtained (typically 24 h). Rapid detection technologies (such as quantitative polymerase chain reaction) that allow FIB quantification in hours have been suggested as a solution to notification errors. In the present study, I explore variability of enterococci (ENT) over time scales less than a day that might affect interpretation of FIB concentrations from a single grab sample, even if obtained rapidly. Five new data sets of ENT collected at 10 and 1 min periodicities for 24 and 1 h, respectively, are presented. Data sets are collected in diverse marine environments from a turbulent surf zone to a quiescent bay. ENT vary with solar and tidal cycles, as has been observed in previous studies. Over short time scales, ENT are extremely variable in each environment even the quiescent bay. Changes in ENT concentrations between consecutive samples (1 or 10 min apart) greater than the single-sample standard (104 most probable number per 100 mL) are not unusual. Variability, defined as the change in concentration between consecutive samples, is not distinct between environments. ENT change by 60% on average between consecutive samples, and by as much as 700%. Spectral analyses reveal no spectral peaks, but power-law decline of spectral density with frequency. Power-law exponents are close to 1 suggesting ENT time series share properties with 1/f noise and are fractal in nature. Since fractal time series have no characteristic time scale associated with them, it is not obvious how the fractal nature of ENT can be exploited for adaptive sampling or management. Policy makers, as well as scientists designing field campaigns for microbial source tracking and epidemiology studies, are cautioned that a single sample of water reveals little about the true water quality at a beach. Multiple samples must be taken to gain a snapshot into the patchy structure of microbial water quality and associated human health risk.     

Prevalence and characterization of Enterococcus spp. isolated from Brazilian foods

Enterococci can be used in the food industry as starter or probiotic cultures. However, enterococci are also implicated in severe multi-resistant nosocomial infections. In this study, the prevalence of enterococci in selected Brazilian foodstuffs (raw and pasteurized milk, meat products, cheeses and vegetables) was evaluated. Phenotypic and PCR protocols were used for species identification. Tests for production of gelatinase, haemolysin, bacteriocin and bile salt hydrolysis were done with all enterococci isolates, whereas molecular determination of virulence markers (genes esp, gel, ace, as, efaA, hyl and cylA) and antibiotic resistance was checked only for Enterococcus faecium and Enterococcus faecalis isolates. The antibiotic-resistant isolates were assayed for biofilm formation and adhesion to mammalian cells. From the 120 food samples analyzed, 52.5% were positive for enterococci, meat and cheese being the most contaminated. E. faecium was the predominant species, followed by E. faecalis, E. casseliflavus and Enterococcus gallinarum. Phenotypic tests indicated that 67.7% of isolates hydrolyzed bile salts, 15.2% produced bacteriocin, 12.0% were beta-hemolytic and 18.2% produced gelatinase. Antibiotic resistance (gentamicin, tetracycline and erythromycin) and genes encoding for virulence traits were more frequent in E. faecalis than in E. faecium. Three E. faecium isolates were resistant to vancomycin. Among antibiotic-resistant isolates, 72.4% of E. faecalis were able to form biofilm and 13.8% to adhere to Caco-2 cells. Antibiotic-resistant E. faecalis and E. faecium isolates were grouped by RAPD-PCR and a scattered distribution was noted, indicating that resistance was not related to a particular clone. The spread of virulence/resistance traits in isolates of the two species and different RAPD-types suggest the pathogenic potential of both species. By contrast, the recovery of bacteriocinogenic E. faecium isolates with no virulence traits suggests their potential for biotechnological applications. In conclusion, our results showed that enterococci from Brazilian foods present important dualist aspects for food safety.     

Visual function,ocular surface integrity and symptomatology of a new extended depth-of-focus and a conventional multifocal contact lens

PurposeTo evaluate visual function, ocular surface integrity and dry eye symptoms with an extended depth-of-focus (EDOF) design and a conventional multifocal (MF) contact lens (CL) after 15 days of wear.MethodsA crossover single mask randomised clinical trial was conducted including 30 presbyopes who used an EDOF and a conventional MF CL (Biofinity MF) for 15 days each. Defocus curves, depth-of-focus range, contrast sensitivity (CS) under photopic and mesopic conditions (with and without glare) and subjective perception of halos and glare were evaluated. The ocular surface was evaluated through non-invasive Keratograph tear breakup time (NIKBUT), averaged tear breakup time (NIKBUT-avg), tear meniscus height (TMH), bulbar and limbal redness, and conjunctival and corneal staining. Dry eye symptoms were assessed with the OSDI questionnaire.ResultsNo statistically significant differences were found for defocus curves or depth-of-focus between the two CLs (both p > 0.05). Subjective perception of halos and glare was not significantly different between CLs. Statistically significant differences were observed for CS under mesopic conditions for low spatial frequencies (p = 0.008). None of the CL produced significant changes in NIKBUT, NIKBUT-avg, TMH or redness. No change in conjunctival staining was observed in 76.7 % and 73.3 % of participants for EDOF and Biofinity MF, respectively. No change in corneal staining was observed in 86.7 % and 83.3 % of participants for EDOF and Biofinity MF, respectively. No changes were observed in the symptomatology measured with OSDI questionnaire (p > 0.05).ConclusionsBoth CL for presbyopia offer good visual quality, preserve the ocular surface integrity and provide the patient with similar symptomatology levels after 15 days of lens wear.     

Assessing the severity of rainfall-derived infiltration and inflow and sewer deterioration based on the flux stability of sewage markers

This study investigated the flux stability of select chemical and biological sewage markers, including caffeine, total nitrogen (TN), total suspended solids (TSS), E. coli, and enterococci, and their suitability in assessing the severity of rainfall-derived infiltration and inflow (RDII) in a residential sewershed. To quantify and compare marker flux stability, concentrations of the candidate markers in two dry-weather periods were determined and the one-day lag autocorrelation coefficients (r) of their mass fluxes were calculated. TN (r = 0.82-0.88) exhibited higher and more consistent flux stability than TSS (r = 0.49-0.82), caffeine (r = 0.56-0.58), E. coli (r = 0.36-0.87), and enterococci (by culture; r = 0.40-0.52), all of which except enterococci by qPCR (r = -0.10-0.21) showed significant autocorrelation. Sewage flows and marker concentrations were also monitored in two wet-weather periods, and the severity of RDII (R(RDII)) were calculated using either flow measurements or marker concentrations independently. Corresponding to its outstanding flux stability, R(RDII) values estimated by TN predicted all severe RDII instances and gave the highest and most consistent correlation (r = 0.74-0.78) among the different sewage markers. Overall, the study illustrated the feasibility of using the flux stability of sewage markers in assessing the severity of RDII and thereby deterioration levels in sewer systems.     

和西牛与西杂牛股二头肌肉品品质及脂肪酸营养特性比较分析

本实验旨在研究西门塔尔杂种牛（西杂牛）与和西牛（日本和牛♂×西杂牛♀）的肉用性能。随机选取同期出生断奶的杂交一代和西牛与西杂牛各9头（公牛）,在同一饲养管理条件下持续育肥至20月龄,屠宰后取股二头肌研究肉质特性,并利用气相色谱仪研究肌内脂肪酸的组成及含量。结果显示,和西牛股二头肌的剪切力、失水率和蒸煮损失显著低于西杂牛（p<0.01）,熟肉率、肉色L*、a*、b*、C值均显著高于西杂牛（p<0.05）;和西牛股二头肌中饱和脂肪酸（SFA）显著低于西杂牛（p<0.01）,不饱和脂肪酸（UFA）显著高于西杂牛（p<0.05）,单不饱和脂肪酸（MUFA）与多不饱和脂肪酸（PUFA）均显著高于西杂牛（p<0.05）,功能性脂肪酸γ-亚麻酸、花生四烯酸和EPA显著高于西杂牛（p<0.01）;n-6/n-3PUFA显著低于西杂牛（p<0.05）,P/S值显著高于西杂牛（p<0.01）。结果表明和西牛肉质较好,牛肉中功能脂肪酸的营养价值得到了改善和提高。      

Escherichia coli and enterococci at beaches in the Grand Traverse Bay,Lake Michigan: sources,characteristics, and environmental pathways

This study quantified Escherichia coli (EC) and enterococci (ENT) in beach waters and dominant source materials, correlated these with ambient conditions, and determined selected EC genotypes and ENT phenotypes. Bathing-water ENT criteria were exceeded more frequently than EC criteria, providing conflicting interpretations of water quality. Dominant sources of EC and ENT were bird feces (10(8)/d/bird), storm drains (10(7)/d), and river water (10(11)/d); beach sands, shallow groundwater and detritus were additional sources. Beach-water EC genotypes and ENT phenotypes formed clusters with those from all source types, reflecting diffuse inputs. Some ENT isolates had phenotypes similar to those of human pathogens and/or exhibited high-level resistance to human-use antibiotics. EC and ENT concentrations were influenced by collection time and wind direction. There was a 48-72-h lag between rainfall and elevated EC concentrations at three southern shoreline beaches, but no such lag at western and eastern shoreline beaches, reflecting the influence of beach orientation with respect to cyclic (3-5 d) summer weather patterns. In addition to local contamination sources and processes, conceptual or predictive models of Great Lakes beach water quality should consider regional weather patterns, lake hydrodynamics, and the influence of monitoring method variables (time of day, frequency).     

Diversity,distribution and antibiotic resistance of Enterococcus spp. recovered from tomatoes,leaves, water and soil on U.S. Mid-Atlantic farms

Antibiotic-resistant enterococci are important opportunistic pathogens and have been recovered from retail tomatoes. However, it is unclear where and how tomatoes are contaminated along the farm-to-fork continuum. Specifically, the degree of pre-harvest contamination with enterococci is unknown. We evaluated the prevalence, diversity and antimicrobial susceptibilities of enterococci collected from tomato farms in the Mid-Atlantic United States. Tomatoes, leaves, groundwater, pond water, irrigation ditch water, and soil were sampled and tested for enterococci using standard methods. Antimicrobial susceptibility testing was performed using the Sensititre microbroth dilution system. Enterococcus faecalis isolates were characterized using amplified fragment length polymorphism to assess dispersal potential. Enterococci (n = 307) occurred in all habitats and colonization of tomatoes was common. Seven species were identified: Enterococcus casseliflavus, E. faecalis, Enterococcus gallinarum, Enterococcus faecium, Enterococcus avis, Enterococcus hirae and Enterococcus raffinosus. E. casseliflavus predominated in soil and on tomatoes and leaves, and E. faecalis predominated in pond water. On plants, distance from the ground influenced presence of enterococci. E. faecalis from samples within a farm were more closely related than those from samples between farms. Resistance to rifampicin, quinupristin/dalfopristin, ciprofloxacin and levofloxacin was prevalent. Consumption of raw tomatoes as a potential exposure risk for antibiotic-resistant Enterococcus spp. deserves further attention.     

Sources of enterococci in Idiazábal-type cheese

Enterococci are a ubiquitous group and are natural constituents of the intestinal flora of nearly all animals and humans and can reach high levels in a variety of farmhouse cheeses. The purpose of this study was to determine the origin of the different enterococcal strains present in cheeses at different stages of ripening by typing the enterococci isolated from the raw milk, the cheeses, the cheesemaking environment, and from the faecal matter of the ewes and humans associated with cheese production. The potential presence of vancomycin-resistant enterococci (VRE) at all stages of the process and in the cheeses was also considered. The study was carried out at two separate cheesemaking dairy plants, and samples of the ewes' faeces, the shepherds' and cheesemakers' stools, teat cups, vat, brine, holding tank milk, vat milk, and the cheeses after brining and after 1, 15, and 60 days of ripening were collected. Cheesemaking procedures at the two plants were similar, yet the enterococcal levels and species observed differed at all the sample collection points, though E. faecalis predominated in all the milk and cheese samples. The traceability study performed for the species E. faecalis present at all the sample collection points suggested that the cheesemaker and the cheesemaking equipment were the source of the enterococci in the cheeses, though other possible non-faecal sources remain to be determined. VanC1 and vanC2/C3 enterococcal strains were isolated from the ovine faeces, teat cup, brine, and vat samples at cheesemaking dairy plant A, while only two vanC2/C3 strains were isolated from ovine faeces samples at dairy plant B. No VRE strains were detected in any of the milk or cheese samples.     

BACTERIOLOGICAL ANALYSIS OF FROZEN SHRIMPS. 1. Total Plate Count-Coliforms and Enterococci in Precooked Frozen Chilean Shrimp

SUMMARY– 392 samples of precooked frozen shrimps from 2 Chilean industries (A and B) were analyzed for total bacterial count, conforms and enterococci throughout a period of 8 months. 1-lb samples of breaded shrimp were received directly from the manufacturers after a freezing period of 10 days at —18δC. Total bacterial count ranged from 10⁴ to 10⁵ organisms perg. Conforms were absent in 65% of the samples from A, and in 40% of those from 8; 89.6% of the samples from A and 50.1% of those from B were bacteriologically acceptable considering a limit of not more than 50 coliforms per g. 98% of the frozen shrimp samples belonging to A contained enterococci, as did 66% of the samples from B. Smaller percentages (17% for A and 54% for B) of acceptable samples are obtained from both industries when 100 enterococci per g is considered as the limit. During the period of observation some sanitary measures were adopted and subsequent coliform counts improved. In plant A working conditions are better and the understanding of bacteriological grounds for the proper handling of food materials has led to the elaboration of a product of consistently better quality. Enterococci counts are in contradiction with coliform counts, since the low-level coliform samples are rejected on the basis of their enterococcal content. In plant B there is a better correlation between coliform and enterococcal counts. Though not investigated, this may be related to the precooking system employed: steaming in an enclosed conveyor in A versus immersion in boiling sea-water in B.     

Determination of Milk Fat Adulteration with Vegetable Oils and Animal Fats by Gas Chromatographic Analysis

This study assessed the potential application of gas chromatography (GC) in detecting milk fat (MF) adulteration with vegetable oils and animal fats and of characterizing samples by fat source. One hundred percent pure MF was adulterated with different vegetable oils and animal fats at various concentrations (0%, 10%, 30%, 50%, 70%, and 90%). GC was used to obtain the fatty acid (FA) profiles, triacylglycerol (TG) contents, and cholesterol contents. The pure MF and the adulterated MF samples were discriminated based on the total concentrations of saturated FAs and on the 2 major FAs (oleic acid [C18:1n9c] and linoleic acid [C18:2n6c], TGs [C52 and C54], and cholesterol contents using statistical analysis to compared difference. These bio‐markers enabled the detection of as low as 10% adulteration of non‐MF into 100% pure MF. The study demonstrated the high potential of GC to rapidly detect MF adulteration with vegetable and animal fats, and discriminate among commercial butter and milk products according to the fat source. These data can be potentially useful in detecting foreign fats in these butter products. Furthermore, it is important to consider that several individual samples should be analyzed before coming to a conclusion about MF authenticity.