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Bacterial and fungal contaminants in caprine and ovine cheese: a Meta-analysis assessment
Abstract and Figures
The dairy industry is of great importance to the European economy contributing towards € 8.7 billion of the total trade surplus. Caprine and ovine milk amount to 3.1% of the 152 million tonnes of milk produced in Europe, 95% of which is transformed into dairy products such as cheese. These cheeses are mostly produced in small holdings from untreated milk, making them a high-risk dairy product for human consumption. A total of 49 foodborne disease outbreaks caused by dairy products were registered in 2017 in Europe. Therefore, these products remain a serious health risk. This meta-analysis examined 30 studies assessing bacterial or fungal contamination of caprine or ovine milk cheeses. The significantly contaminating microbes were found to be Acremonium spp. (19%), Aspergellus spp. (23%), Bacillus spp. (2%), Brucella spp. (34%), Enterobactericae spp. (36%), Enterococcus spp. (28%), Escherichia spp. (15%), Fusarium spp. (21%), Geotrichum spp. (22%), Listeria spp. (11%), Mucor spp. (15%), Penicillium spp. (25%), Phoma spp. (20%), Rhizopus spp. (15%), Salmonella spp. (3%), Scopulariopsis spp. (19%) and Staphylococcus spp. (25%) in caprine and ovine cheese, indicating a variety of food pathogens as well as spoilers. Raw milk is nutritious hence prone to contamination. However, since traditional cheese is often made from untreated milk, it is important to educate cheesemakers of key safety measures and goof manufacturing practice allowing for the safe production of these food items.
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... After receipt, solid foods such as poultry, minced meat, perishable products, cheese, bolón, and fruits were aseptically cut into 2-5 g pieces. Broiler specimens were collected from fur-bearing areas, as suggested elsewhere 18 . For samples containing a mixture of liquid and solid foods such as ceviche, Encebollado, fruit salads, and sauces. ...
The current study was carried out to estimate the risk of disease probability from the consumption of foods such as meats, poultry, unpasteurized cheeses, fruit-based drinks, ready-to-eat fruits, and typical preparations such as Encebollado, ceviche, and Bolón de Verde contaminated with Salmonella, Escherichia coli and Listeria monocytogenes in Ecuador using a quantitative microbiological risk assessment (QMRA). A first-order Monte Carlo simulation probabilistic distribution approach was adopted to assess the occurrence of pathogens in the tested foods. The scenario was simulated using the concentration levels concerning the contaminant and food consumption obtained through an online survey with a sample size of 202 people. A model (100,000 iterations) was run and created in an Excel spreadsheet using @Risk software. The results obtained are the risk of infection (possibilities of becoming infected by eating the food evaluated) and the contaminant dose per portion consumed. Additionally, an exponential model with a single dose was used for risk characterization to determine the probability of becoming ill from contaminated food. The QMRA model performed a prediction for the mean risk of Salmonella infection from ground beef consumption of 1.33 E - 04 log 10 cfu / serving, while the exponential model estimated a value of 1.0 log cfu - serving. In the case of Listeria monocytogenes, the QMRA estimated an average probability of infection in unpasteurized fresh cheese of 5.9E-05 compared to the average disease risk estimated in the risk characterization for L. monocytogenes of 9.50E-13. The QMRA estimated an average risk of infection by E. coli for Encebollado and ceviches of 5.6E -03 compared to the average risk of disease estimated in the risk characterization for Escherichia coli of 0.387 log cfu -ration. These results suggest the need to adopt effective mitigation strategies. Control parameters such as temperature during the supply chain and good hygiene practices during manufacturing can effectively control food-associated pathogens. More data is necessary toimprove the evaluation of the risk developed. Keywords: Microbiological risk análisis, Infectious biological agents (Salmonella, Listeria monocytogenes, Escherichia coli), QMRA (Quantitative Microbial Risk Assessment), Typical foods in Ecuador, Population food consumption in three main cities of Ecuador.
... This may be due to the fact that Enterobacteriaceae spp is also strongly associated with spoilage and coercion of human health. Enterobacteriaceae spp were major bacteria in the spoilage of caprine and ovine cheese (Griffin et al., 2020), It is also the main microorganism responsible for the spoilage of dandelion (Dermesonluoglu et al., 2016). Enterobacteriaceae spp and so on were the main genera on the surface of the kitchens of food service facilities, posing a certain risk to food safety (Lim et al., 2021), and is also an important cause of healthcare-associated bloodstream infections (BSIs) in children (Boguniewicz et al., 2021). ...
Poly (lactic acid) (PLA) composite films with the addition of mesoporous silica nanoparticles MSN (0, 2, 4 and 6 wt%) loaded with 10 wt% citral (CIT) were prepared for application in Chanterelles packaging. Composite films with added MSN/CIT showed good mechanical properties, especially 4MSN/CIT/PLA. Changes in physicochemical properties and bacterial flora of Chanterelles during packaging and storage were tested. Compared with CIT/PLA, Chanterelles packed with 4MSN/CIT/PLA showed about 1.62-times lower browning value, 1.53-times lower electrolyte permeability, and 1.83- and 1.78-times lower PPO and POD, respectively, at 12 day. Better physicochemical properties of Chanterelles can be maintained. For bacterial flora changes, Chanterelles packaged with 4MSN/CIT/PLA had more stable flora (p < 0.05) and lower species diversity during storage (p < 0.05), effectively controlling the growth and reproduction of their dominant spoilage bacteria (Enterobacteriaceae spp). In conclusion, the composite membranes obtained by the addition of MSN/CIT to PLA have great potential in the storage of Chanterelles.
... Therefore, raw caprine and ovine milk can be potential carriers of foodborne pathogens that could be linked to serious health risks. A recent meta-analysis study showed that the most prevalent genera are Listeria spp., Penicillium spp., Salmonella spp., Staphylococcus spp., and Escherichia spp., but data on the frequency of microbial contamination in cheese from small ruminants' milk (goat and sheep) are few [9]. The main sources of milk contamination are poor livestock salubriousness and herd health conditions, mastitis ubiquity, and improper milking and conservation practices [10]. ...
Milk is an excellent medium for the growth of several bacteria and other microorganisms and thus, it has been extensively studied. An always current issue in the dairy industry is mastitis, which causes losses in milk volume and profits. In many cases, milk is used raw or treated at low temperatures for further dairy processes while there are quite a few cases in which foodborne-related outbreaks have occurred. Both culture-based methods and PCR were used to assess the presence of certain pathogens related to both contagious and environmental pathogens, especially the emerging pathogenic bacterium Klebsiella pneumoniae, as well as Staphyloccocus aureus and Enterobacter spp., which are associated with mastitis in milk samples from different lactating ruminant species (cows, goats, and sheep) and to further evaluate the significance of the isolated pathogens to public health. Even though significant mastitis contagious pathogens such as Staphylococcus aureus and Staphylococcus epidermidis were not detected, environmental pathogens related to poor hygiene conditions at the farm level (K. pneumoniae, Staphylococcus saprophyticus, and Enterococcus spp.) were detected. In particular, K. pneumoniae and Staphylococcus saprophyticus were present in ovine milk samples while bovine and caprine milk samples were contaminated with Enterococcus spp. The presence of these bacteria underlines the significant role of environmental hygiene especially since Staphylococcus saprophyticus and Enterococcus spp. are related to urinary tract infections and all of the tested pathogens may carry antibiotic resistance genes. More specifically, 20% of the isolated Klebsiella pneumoniae strains were found resistant to carbapenem antibiotics. The presence of emerging K. pneumoniae in ovine milk samples also indicates the need for new policies in terms of safety testing. Suggestions of monitoring processes carried out by the relevant authorities are discussed.
... The presence of foodborne pathogens and the inherent variability in artisanal food products characteristics are clearly interconnected. Some of them can be associated with contaminated raw materials, deficient hygienic-sanitary conditions, inappropriate fermentations, recontamination events, incorrect production settings or unexpected situations during the manufacturing process (Griffin et al., 2020). These situations have been frequently found in artisanal productions, which also highlight the need of considering raw milk cheeses as products of high concern for public health. ...
A harmonised microbiological survey was performed in two artisanal factories of raw goat milk cheeses (A and B) located in the Andalusian region (Spain). A total of 165 different control points or samples (raw materials, final products, food-contact surfaces [FCS], and air) were examined as microbial and pathogen sources of contamination of artisanal goat raw milk cheeses. For raw milk samples analysed from both producers, the concentrations of aerobic-mesophilic bacteria (AMB), total coliforms, coagulase-positive Staphylococcus spp. (CPS), lactic-acid bacteria (LAB) and moulds and yeasts ranged between 3.48 and 8.59, 2.45–5.48, 3.42–4.81, 4.99–8.59 and 3.35–6.85 log cfu/mL respectively. For the same microbial groups, the analysis of raw milk cheeses revealed concentrations ranging from 7.82 to 8.88, 2.00–6.82, 2.00–5.28, 8.11–9.57 and 2.00–5.76 log cfu/g, respectively. Although the raw material analysed from producer A presented higher microbial loads and between-batch variability, it was B the producer with the most loaded final products. Regarding the microbial air quality, the fermentation area, storage room, milk reception and packaging room were the most AMB loaded places, while the ripening chamber was the area with higher fungal loads in bioareosol from both producers. Conveyor belts, cutting machine, storage boxes and brine tank were highlighted as the most contaminated FCS evaluated. Staphylococcus aureus was the only pathogen detected within the set of 51 isolates from samples by MALDI-TOF and molecular PCR, with a prevalence of 12.5% for samples from the producer B. The public health risk attributed to the consumption of artisanal goat cheese should not be neglected, and may consider the whole cheesemaking processing chain, from microbiological quality of raw milk to the ready-to-eat final product, especially concerning the presence of S. aureus.
... FBOs associated with Brucella, Campylobacter, S. aureus and Flavivirus in cheese were also reported in the EU and a multicountry outbreak of STEC O26 was linked to Romanian cheese (in 2016). 23 The occurrence of Brucella has been mostly linked to artisan cheeses made with raw milk (Griffin et al., 2020). Similarly, originating in the raw milk, ...
High-pressure processing (HPP) is a non-thermal treatment in which, for microbial inactivation, foods are subjected to isostatic pressures (P) of 400–600 MPa with common holding times (t) from 1.5 to 6 min. The main factors that influence the efficacy (log10 reduction of vegetative microorganisms) of HPP when applied to foodstuffs are intrinsic (e.g. water activity and pH), extrinsic (P and t) and microorganism-related (type, taxonomic unit, strain and physiological state). It was concluded that HPP of food will not present any additional microbial or chemical food safety concerns when compared
to other routinely applied treatments (e.g. pasteurisation). Pathogen reductions in milk/colostrum caused by the current HPP conditions applied by the industry are lower than those achieved by the legal requirements for thermal pasteurisation. However, HPP minimum requirements (P/t combinations) could be identified to achieve specific log10 reductions of relevant hazards based on performance
criteria (PC) proposed by international standard agencies (5–8 log10 reductions). The most stringent HPP conditions used industrially (600 MPa, 6 min) would achieve the above-mentioned PC, except for
Staphylococcus aureus. Alkaline phosphatase (ALP), the endogenous milk enzyme that is widely used to verify adequate thermal pasteurisation of cows’ milk, is relatively pressure resistant and its use
would be limited to that of an overprocessing indicator. Current data are not robust enough to support the proposal of an appropriate indicator to verify the efficacy of HPP under the current HPP conditions applied by the industry. Minimum HPP requirements to reduce Listeria monocytogenes levels by specific log10 reductions could be identified when HPP is applied to ready-to-eat (RTE) cooked meat products, but not for other types of RTE foods. These identified minimum requirements would result in the inactivation of other relevant pathogens (Salmonella and Escherichia coli) in these RTE foods to a similar or higher extent.
... Cheese is one of the utmost important fermented milk products produced and consumed by humans. In the European Union, 156.8 million tons of whole milk were processed in 2018, and 10.3 million tons of cheese was produced (Griffin et al., 2020). Of all the milk produced in the EU in 2018,37.7% was used for cheese production, being the main food obtained from milk. ...
Enterococcus spp. were isolated from PDO-cheese of Azeitão and Nisa at six cheesemaking units (Azeitão: A1, A2, A3, A4; and Nisa: N9, N10), over four years (2016–2019). Genomic typing was performed using RAPD and distinct enterococci (n = 145) were identified at the species level by multiplex-PCR and evaluated regarding antimicrobial drug resistance (AMR). Antibiotics from nine distinct classes (aminoglycosides, macrolides, oxazolidinones, chloramphenicol, streptogramins, tetracyclines, glycopeptides, β-lactams, and quinolones) were selected for AMR surveillance and breakpoint criteria defined by EUCAST and CLSI were considered and compared. Regarding species allocation, 78 enterococci were identified as E. faecium, 37 confirmed as E. faecalis and 30 as E. durans. High levels of resistance to quinupristin-dalfopristin, tetracycline and teicoplanin were observed. Some resistances to clinically relevant antimicrobials were also detected, including β-lactams, aminoglycosides, and glycopeptides. Two isolates were considered multidrug-resistant, one according to EUCAST and the other to CLSI breakpoint criteria. Overall, considering the absence of reports regarding enterococcal-related toxinfections or infections resulting from the consumption of PDO-cheeses, traditional foods harbouring these bacteria should be considered safe. However, the possibility of horizontal gene transfer events associated with antibiotic resistance determinants further highlights the importance for AMR surveillance along the food chain.
The fungal component of the microbiota, the mycobiota, has been neglected for a long time due to its poor richness compared to bacteria. Limitations in fungal detection and taxonomic identification arise from using metagenomic approaches, often borrowed from bacteriome analyses. However, the relatively recent discoveries of the ability of fungi to modulate the host immune response and their involvement in human diseases have made mycobiota a fundamental component of the microbial communities inhabiting the human host, deserving some consideration in host–microbe interaction studies and in metagenomics. Here, we reviewed recent data on the identification of yeasts of the Ascomycota phylum across human body districts, focusing on the most representative genera, that is, Saccharomyces and Candida . Then, we explored the key factors involved in shaping the human mycobiota across the lifespan, ranging from host genetics to environment, diet, and lifestyle habits. Finally, we discussed the strengths and weaknesses of culture‐dependent and independent methods for mycobiota characterization. Overall, there is still room for some improvements, especially regarding fungal‐specific methodological approaches and bioinformatics challenges, which are still critical steps in mycobiota analysis, and to advance our knowledge on the role of the gut mycobiota in human health and disease.
This article is categorized under: Immune System Diseases > Genetics/Genomics/Epigenetics
Immune System Diseases > Environmental Factors
Infectious Diseases > Environmental Factors
The mycobiome is the fungal component of the human microbial ecosystem that represents only a small part of this environment but plays an essential role in maintaining homeostasis. Colonization by fungi begins immediately after birth. The initial mycobiome is influenced by the gestational age of a newborn, birth weight, delivery method and feeding method. During a human’s life, the composition of the mycobiome is further influenced by a large number of endogenous and exogenous factors. The most important factors are diet, body weight, age, sex and antibiotic and antifungal therapy. The human mycobiome inhabits the oral cavity, gastrointestinal tract, respiratory tract, urogenital tract and skin. Its composition can influence the gut–brain axis through immune and non-immune mediated crosstalk systems. It also interacts with other commensals of the ecosystem through synergistic and antagonistic relationships. Moreover, colonization of the gut by opportunistic fungal pathogens in immunocompromised individuals can lead to clinically relevant disease states. Thus, the mycobiome represents an essential part of the microbiome associated with a variety of physiological and pathological processes. This review summarizes the current knowledge on the composition of the mycobiome in specific sites of the human body and its role in health and disease.
The massive food wastes pose a growing health concern for spreading of antibiotic resistance and pathogens due to food spoilage. However, little is known about these microbial hazards during collection, classification, and transportation before eventual treatment. Here, we profiled the temporal variations of antibiotic resistance genes (ARGs), pathogens, bacterial and fungal communities across four typical food wastes (vegetable, fish, meat, and rice) during storage at room temperature in summer (maximum 28-29 °C) of typical southeast city in China. A total of 171 ARGs and 32 mobile genetic elements were detected, and the absolute abundance of ARGs significantly increased by up to 126-fold with the storage time. Additionally, five bacterial pathogens containing virulence factor genes were detected, and Klebsiella pneumoniae was persistently detected throughout the storage time in all food types except rice. Moreover, fungal pathogens (e.g., Aspergillus, Penicillium, and Fusarium) were also frequently detected. Notably, animal food wastes were demonstrated to harbor higher abundance of ARGs and more types of pathogens, indicating a higher level of hazard. Mobile genetic elements and food types were demonstrated to mainly impact ARG profiles and pathogens, respectively. This work provides a comprehensive understanding of the microbial hazards associated with food waste recycling, and will contribute to optimize the food waste management to ensure biosecurity and benefit human health.
Mold spoilage of dairy products such as yogurt is a concern in dairy industry. Not only does it lead to substantial food waste, economic losses, and even brand image damage, but it may also cause public health concern due to the potential production of mycotoxin. Good hygiene practices are necessary to prevent contamination, but contamination may nevertheless occur at the production site and, not least, at the site of the consumer. In recent years, there has been a growing interest from consumers for "clean label" food products, which are natural, less-processed, and free of added, chemical preservatives, and a wish for shelf lives of considerable length in order to minimize food waste. This has sparked an interest in using lactic acid bacteria (LAB) or their metabolites as biopreservatives as a way to limit the growth of spoilage organisms in dairy products. A range of compounds produced by LAB with potential antifungal activity have been described as contributing factors to the inhibitory effect of LAB. More recently, growth inhibition effects caused by specific competitive exclusion have been elucidated. It has also become clear that the sensitivity toward both individual antifungal compounds and competition mechanisms differ among molds. In this review, the main spoilage molds encountered in dairy products are introduced, and an overview of the antifungal activity of LAB against different spoilage molds is presented including the main antifungal compounds derived from LAB cultures and the sensitivity of the spoilage molds observed toward these compounds. The recent findings of the role of competitive exclusion with emphasis on manganese depletion and the possible implications of this for biopreservation are described. Finally, some of the knowledge gaps, future challenges, and trends in the application of LAB biopreservation in dairy products are discussed.
Cheese has important nutritional properties as a result of their protein, lipid and essential minerals content. This study analyzed the bioavailability of Ca, Mg and Zn in Brazilian cheeses, using the in vitro INFOGEST digestion method coupled with assays in Caco-2 cells. Furosine content (marker of the early Maillard reaction) was also evaluated. Two self-produced fresh cheeses, cow and goat Minas frescal; and two commercial matured goat cheeses, Blue and Pyramid, were analyzed. Mineral bioaccessibility after in vitro digestion was in the range of 44.4 - 74.7%, 54.8 - 66.1% and 18.2 - 38.7% for Ca, Mg and Zn, respectively. Bioaccessibilty was significantly affected by the mineral and fat content level. The digested fresh cheeses (Minas frescal) showed the highest values of soluble Ca and Mg; and the Pyramid goat cheese presented the greatest Ca and Zn transport efficacy across Caco-2 cell monolayers. Furosine could be a useful indicator to evaluate cheese ripeness and its indirect effect on the mineral availability.
Krokmach is a traditional Bulgarian fermented dairy product. The microbiota composition was investigated for the first time by next-generation sequencing. The results revealed a quite specific constitution comprising several bacterial species never reported before for fermented dairy products. It resembles those of soft and smear cheeses rather than that of yoghurt. The dominant species was Lactococcus lactis subsp. lactis (75% of the operational taxonomic unit (OTU) counts). The overall composition was quite unusual with 8.5% of Exiguobacterium sp. OTU counts, as well as the presence of Apiactobacillus kunkeei, Megamonas sp. and Kluyvera georgiana among the dominant and subdominant species and genera.
Food can be spoiled by microbial contaminants rendering it unsafe for human consumption, and requires regular testing to ensure its safety. Current techniques involve the destruction of food matrices for microbial assessment. In contrast, emerging technologies such as hyperspectral imaging (HSI) can be integrated into the manufacturing chain as on-line or in-line systems for frequent microbial assessment. This study aims to develop a model cheese system with a high surface area to optimise and deploy HSI for microbial detection. Compatibility and stability studies were conducted on the model cheese system by measuring physiochemical parameters, such as pH moisture content, water activity, total protein content, and sugars. Microbial testing was also carried out to quantify the natural or background microflora of the model cheese after rennet and Lactobacillus spp. starter cultures were added. Penicillium chrysogenum was isolated from commercial cheese samples using 18S rRNA gene amplification and sequencing. Finally, the model cheese was challenge tested with P. chrysogenum isolated from commercial samples and imaged using a hyperspectral camera. As a proof of concept, an untrained principal component analysis model was developed to identify model cheese inoculated with P. chrysogenum at mycelial growth or after conidia formation, illustrating a versatile model cheese system that can be deployed for non-destructive assessment of fungal contaminants with HSI.
The application of non-destructive process analytical technologies in the area of food science got a lot of attention the past years. In this work we used hyperspectral imaging to detect mould on milk agar and cheese. Principal component analysis is applied to hyperspectral data to localise and visualise mycelia on the samples' surface. It is also shown that the PCA loadings obtained from a set of training samples can be applied to hyperspectral data from new test samples to detect the presence of mould on these. For both the agar and cheeselets, the first three principal components contained more than 99
%
of the total variance. The spatial projection of the second principal component highlights the presence of mould on cheeselets. The proposed analysis methods can be adopted in industry to detect mould on cheeselets at an early stage and with further testing this application may also be extended to other food products.
This report of EFSA and the European Centre for Disease Prevention and Control presents the results of the zoonoses monitoring activities carried out in 2015 in 32 European countries (28 Member States (MS) and four non-MS). Campylobacteriosis was the most commonly reported zoonosis and the increasing European Union (EU) trend for confirmed human cases since 2008 continued. In food, the occurrence of Campylobacter remained high in broiler meat. The decreasing EU trend for confirmed human salmonellosis cases since 2008 continued, but the proportion of human Salmonella Enteritidis cases increased. Most MS met their Salmonella reduction targets for poultry. More S. Enteritidis isolates were reported and S. Infantis was confirmed as the most frequent serovar isolated from domestic fowl. In foodstuffs, the EU level Salmonella non-compliance for minced meat and meat preparations from poultry was low. Despite the significant increasing trend since 2008, the number of human listeriosis cases stabilised in 2015. In ready-to-eat foods, Listeria monocytogenes seldom exceeded the EU food safety limit. The decreasing EU trend for confirmed yersiniosis cases since 2008 continued. Positive findings for Yersinia were mainly reported in pig meat and products thereof. The number of confirmed shiga toxin-producing Escherichia coli (STEC) infections in humans was similar to 2014. In food, STEC was most frequently reported in meat from ruminants. A total of 4,362 food-borne outbreaks, including waterborne outbreaks, were reported. Bacteria were the most commonly detected causative agents, followed by bacterial toxins, viruses, other causative agents and parasites. The causative agent remained unknown in 33.5% of all outbreaks. As in previous years, Salmonella in eggs continued to represent the highest risk agent/food combination. The report further summarises trends and sources for tuberculosis due to Mycobacterium bovis, Brucella, Trichinella, Echinococcus, Toxoplasma, rabies, Coxiella burnetii (Q fever), West Nile virus and tularaemia.
This study investigated the microbiological quality and the incidence of some pathogens in raw goat's and ewe's milk in Egypt. Microbiological analysis revealed that the mean aerobic plate count was 9.11±2.47×10⁶ and 2.04±0.91×10⁶ CFU/ml for goat's and ewe's milk, respectively. Enterobacteriaceae were detected in 24 (68.6%) goat's milk samples and 21 (60%) ewe's milk samples, with mean count values of 2.53±0.57×10⁶ and 1.67±0.87×10⁵ CFU/ml in goat's and ewe's milk samples, respectively. Coliforms were detected in 68.57% and 60%, with mean count values of 6.47±2.17×10⁵ and 1.66±0.85×10⁵ CFU/ml in goat's and ewe's milk samples, respectively. Escherichia coli was detected in 5 (14.3%) and 4 (11.4%), Staphylococcus aureus was detected in 11 (31.43%) and 13 (37.14%), with mean count values of 1.41×10⁴ and 6.67×10⁴ CFU/ml in goat's and ewe's milk, respectively. On the other hand, Salmonella and Listeria monocytogenes were not detected in the examined samples. Obtained results highlighted the poor microbiological and sanitary quality of goat's and ewe's milk produced in Egypt.
When we speak about heterogeneity in a meta-analysis, our intent is usually to understand the substantive implications of the heterogeneity. If an intervention yields a mean effect size of 50 points, we want to know if the effect size in different populations varies from 40 to 60, or from 10 to 90, because this speaks to the potential utility of the intervention. While there is a common belief that the I(2) statistic provides this information, it actually does not. In this example, if we are told that I(2) is 50%, we have no way of knowing if the effects range from 40 to 60, or from 10 to 90, or across some other range. Rather, if we want to communicate the predicted range of effects, then we should simply report this range. This gives readers the information they think is being captured by I(2) and does so in a way that is concise and unambiguous. Copyright © 2017 John Wiley & Sons, Ltd.
Ricotta fresca cheese is the product of Sardinian dairy industry most exposed to microbial post-process contamination. Due to its technological characteristics, intrinsic parameters, pH (6.10-6.80) and water activity (0.974-0.991), it represents an excellent substrate for the growth of spoilage and pathogenic microorganisms, which are usually resident in cheese-making plants environments. Generally, ricotta fresca has a shelf life of 5-7 days. For this reason, at industrial level, modified atmosphere packaging (MAP) is used to extend the durability of the product. However, few investigations have been conducted to validate the use of MAP in ricotta fresca. The aim of this work is to evaluate the shelf lifeNoof ricotta fresca under MAP. A total of 108 samples were collected from three Sardinian industrial cheese-making plants and analysed within 24 h after packaging and after 7, 14 and 21 days of refrigerated storage. Aerobic mesophilic bacteria, mesophilic and thermophilic cocci and lac-tobacilli, Enterobacteriaceae and E. coli, L. monocytogenes, Pseudomonas spp., Bacillus cereus, yeasts and moulds, and the chemical-physical parameters and composition of the product were determined. At the end of the shelf life, Pseudomonas spp. and Enterobacteriaceae reached high concentrations, 5 to 7 and 3 to 6 log10 colony forming unit g?1, respectively. The presence of environmental contaminants indicates that the use of MAP without the appropriate implementation of prerequisite programmes is not sufficient to extend the durability of ricotta fresca. Gas mixture and packaging material should be selected only on the basis of scientific evidence of their effectiveness.
Ricotta salata cheese is a salted variety of ricotta traditionally made in Sardinia (Italy) from the whey remaining after the production of Pecorino Romano PDO or other sheep’s milk cheeses. Ricotta salata cheese is very critical for the possible growth of pathogenic and spoilage microorganisms. Sporadic cases of listeriosis associated with ricotta salata cheese have been reported over recent years. The objective of the present study was to assess the evolution of spoilage and pathogen microorganism of vacuum packed ricotta salata cheese during the entire product shelf-life. The durability study was conducted on 18 vacuum packed ricotta salata cheese samples analyzed at the beginning of the shelf-life and after 60 and 90 days of refrigerated storage. Pathogen as Listeria monocytogenes and Bacillus cereus were never detected. During shelf-life total bacterial counts ranged between 7.90±0.64 CFU g-1 and 9.19±0.58 CFU g-1 on the rind and between 2.95±0.68 CFU g-1 and 4.27±1.10 CFU g-1 in the inner paste, while Enterobacteriaceae ranged between 4.22±0.66 CFU g-1 and 5.30±0.73 CFU g-1 on the rind and 3.13±1.80 CFU g-1 and 2.80±0.88 CFU g-1 in the inner paste. Considered the technology, the intrinsic properties and the almost absence of competing microflora, ricotta salata cheese can support the growth of spoilage and pathogen microorganism originating from the processing environment. The high level of total bacterial counts and Enterobacteriaceae observed both on the rind and the inner paste suggest contamination of the product from the processing environment. Therefore it is essential a strict implementation of hygiene during processing in order to reduce the load of environmental contaminants that may grow during refrigerated storage.
In recent years, the incidence of foodborne diseases caused by shiga toxin-producing
Escherichia coli (STEC) has increased globally. For this reason, within the specific regional
control plan for the detection of STEC in food products in Italy, the presence of STEC in
unpasteurized milk cheeses was investigated. In total 203 samples obtained from March 2011
to December 2013 were analysed, with two standard methods (ISO 16654:2001 and ISO
13136:2012). Two strains of E. coli O157 were isolated (2/161, 1.2%) but did not carry any
virulence-associated genes and 22 stx-positive samples (22/146, 15.1%) were detected in
enrichment cultures, mostly from ovine cheeses. Only two strains isolated from different ovine
cheeses carried stx gene and none of these was eae-positive. This study confirms the presence
of stx-positive E. coli and suggests that this type of food cannot be excluded as a potential
vehicle of STEC.
The objective of this study was to determine the occurrence of antimicrobial resistance among Staphylococcus aureus and to
estimate the presence of methicillin-resistance in S. aureus (MRSA) isolates obtained by culture and polymerase chain reaction
(PCR) methods. For this purposes, 100 Iranian white and feta cheese samples collected from different suppliers were initially
evaluated for the occurrence of S. aureus by culturing methods. The obtained isolates were subjected to disc diffusion antimicrobial
susceptibility tests and a PCR method to detect the mecA gene. Out of the 100 cheese samples examined, 25 (25%) samples were
contaminated with S. aureus with a mean of 5.74 ± 5.67 log cfu/g. Out of the 25 isolates, 23 (92%) were found to be resistant to at
least one antibiotic or more, tested by a disk diffusion method. The highest rate of antibiotic resistance was observed to penicillin G
(92%) followed by ampicillin (73%) and cloxacillin (68%). None of the isolates was resistant to gentamycin and vancomycin. Eight
(34.78%) of the 23 S. aureus isolates were genotypically confirmed as MRSA. The results indicate that the presence of antimicrobial
resistant strains of S. aureus in Iranian cheese samples constitute a potential risk for human health. This calls for better control of the
spread of antimicrobial resistant strains as well as cheese contamination sources.
This study aimed to evaluate Arcobacter species contamination of industrial sheep ricotta cheese purchased at retail and to establish if the dairy plant environment
may represent a source of contamination. A total of 32 sheep ricotta cheeses (1.5 kg/pack) packed in a modified atmosphere
were purchased at retail, and 30 samples were collected in two sampling sessions performed in the cheese factory from surfaces
in contact with food and from surfaces not in contact with food. Seven out of 32 samples (21.9%) of ricotta cheese collected
at retail tested positive for Arcobacter butzleri at cultural examination; all positive samples were collected during the same sampling and belonged to the same batch. Ten
surface samples (33.3%) collected in the dairy plant were positive for A. butzleri. Cluster analysis identified 32 pulsed-field gel electrophoresis (PFGE) patterns. The same PFGE pattern was isolated from
more than one ricotta cheese sample, indicating a common source of contamination, while more PFGE patterns could be isolated
in single samples, indicating different sources of contamination. The results of the environmental sampling showed that A. butzleri may be commonly isolated from the dairy processing plant investigated and may survive over time, as confirmed by the isolation
of the same PFGE pattern in different industrial plant surface samples. Floor contamination may represent a source of A. butzleri spread to different areas of the dairy plant, as demonstrated by isolation of the same PFGE pattern in different production
areas. Isolation of the same PFGE pattern from surface samples in the dairy plant and from ricotta cheese purchased at retail
showed that plant surfaces may represent a source of A. butzleri postprocessing contamination in cheeses produced in industrial dairy plants.
Contamination by fungi of the genus Aspergillus with special reference to the possible detection of aflatoxin-producing strains of Aspergillus flavus was studied in 52 samples of commercial cheeses made with different types of milk (8 of cow's milk, 12 of ewe's milk, 13 of goat's milk, and 19 of milk mixtures of various species: cow, ewe, and goat) produced in southern Spain. The frequency of appearance of various species of Aspergillus, A. glaucus, A. niger, A. nidulans, A. sulphureus, A. Terreus, and A. flavus, in the different types of cheese was determined. In 4 (2 of goat's milk cheese and 2 of cheeses made with milk from various species) out of 52 samples (7.69%), aflatoxin-producing strains of A. flavus were detected.
Pecorino Siciliano (PS) "primosale" is a traditional Sicilian fresh soft cheese made from sheep's milk. Short-ripening time and production from unpasteurized or raw milk can facilitate bacterial contamination of PS "primosale". The microbiological quality of "primosale" on retail sale in the street markets of Palermo, Italy was studied by detecting the common food pathogens Listeria monocytogenes and Staphylococcus aureus and indicator microorganisms, such as Escherichia coli, Enterobacteriaceae and Staphylococcaceae. In our study, 4% and 44% of the samples, respectively, did not comply with the acceptability levels fixed by European regulations for S. aureus and E. coli. A high contamination of bacteria belonging to Enterobacteriaceae and Staphylococcaceae was found in 42% and 50% of the cheeses analyzed, respectively. Such results indicate poor husbandry and poor hygiene practices during milk collection or preservation or during cheese production processes and handling. In addition, the retail sale conditions may have played a role in cheese contamination since a correlation was found between poor microbiological quality and some selling parameters. This study emphasizes the need to improve production hygiene throughout the PS food chain in line with the traditional cheese-making procedures. Labelling of PS with clear information on whether the cheese was prepared from raw milk also requires improvement.
Food safety is a critical factor in the production of farmhouse cheese. In Ireland the varieties of farmhouse cheese produced reflect a much broader range than those produced commercially and some of these cheese varieties are associated with greater microbiological risk. These include cheese produced from unpasteurised milk and soft ripened cheese such as mould or smear-ripened cheeses which have high pH and relatively short ripening times. The aim of this study was to determine the microbiological quality of farmhouse cheeses in Ireland. Three hundred and fifty one cheese samples, from 15 cheese producers, were analysed for microbiological quality on a monthly basis throughout the year. The analyses included enumeration of Escherichia coli, Staphylococcus aureus and Listeria monocytogenes (using the relevant agars) and enrichment for L. monocytogenes. The cheeses selected were produced from ovine, caprine and bovine milk. Both unpasteurised and pasteurised milk cheeses were sampled and these included hard, semi-hard and soft cheeses, internal/external mould-ripened and smear-ripened cheeses and the cheeses represented different geographic regions. Of the cheeses tested, 94% were free of L. monocytogenes, all were within the EU limits for E. coli and only one cheese variety had S. aureus levels above the recommended numbers for the first 6 months of the year. Due to a modified production process the numbers were within the guidelines for the second six months. The results indicate that Irish farmhouse cheeses are of a high microbiological quality.
The microbiological quality, safety, and composition of mixtures of ewe's and goat's milk (90:10) used for cheesemaking were evaluated before and after thermization at 60 and 67 degrees C for 30 s. Such mild thermal treatments are commonly applied to reduce natural contaminants of raw milk before processing for traditional hard Greek cheeses. Raw milk samples had an average total bacterial count of 7.3 log CFU/ml; most of these bacteria were lactic acid bacteria (LAB) and pseudomonads. The LAB flora of raw milk was dominated by enterococci (40.8%), followed by lactococci (20.4%), leuconostocs (18.4%), and mesophilic lactobacilli (10.2%). Enterococcus faecalis (30.1%) and Enterococcus faecium (13.7%) were the most common LAB isolates, followed by Enterococcus durans, Lactococcus lactis subsp. lactis, Lactobacillus plantarum, and Leuconostoc lactis. Thermization at 60 degrees C for 30 s was effective for reducing raw milk contamination by enterobacteria (5.1 log CFU/ml), coagulase-positive staphylococci (3.3 log CFU/ml), and Listeria (present in 25-ml samples) to safe levels, but it also reduced mesophilic lactococci, leuconostocs, lactobacilli, and selected enterococci (72.0%) in thermized milk. Thermization at 67 degrees C for 30 s had a major inactivation effect on all bacterial groups. Two nisin-producing L. lactis subsp. lactis strains (M78 and M104) were isolated from raw milk, but neither nisin-producing nor other bacteriocin-producing LAB strains were isolated from thermized milk. Thus, thermization treatments control harmful bacteria but also may have a negative impact on milk quality by reducing desirable LAB and the biodiversity of raw milk bacteria overall, inactivating potentially protective LAB strains and enhancing the ability of potentially pathogenic enterococci to grow in fresh cheese curds.
One hundred and twenty-two samples of cheeses made from goat and sheep milk and a mixture of the two types, produced in Southern Italy by industrial establishments or artisans, were analysed for the detection of fungal contamination and the presence of potentially toxigenic moulds. Only organoleptically acceptable cheeses without evident fungal contamination were studied. Among these, 40 were unripened, 30 medium and 52 long ripened cheeses. Moulds were found in 54 of the 122 analysed samples (44.3%). The most contaminated cheeses were the medium and long ripened ones (46.3% and 35.2%), and the industrial cheeses (59.1%). The artisan cheeses were the least contaminated (26.8%). Among the cheeses that tested positive, Penicillium species were the most frequently isolated (72.9%), followed by Geotrichum spp. (7.3%), Aspergillus spp. (4.2%) and Mucor spp. (4.2%). The potentially toxigenic species within the genera Penicillium, Aspergillus and Fusarium were mainly detected in sheep cheeses.
Seventy raw milk cheeses made in different regions of Portugal, both hard and soft varieties, made with cow's, ewe's, or goat's milk or combinations of these, were sampled within their quoted shelf lives for microbiological safety. On the basis of the presence or numbers of Escherichia coli, E. coli O157, Staphylococcus aureus, Salmonella, and Listeria monocytogenes, cheeses were categorized as satisfactory, acceptable, unsatisfactory, or unacceptable and potentially hazardous. Twenty-two of the 70 cheeses were classified as satisfactory or acceptable. Thirty-seven of the cheeses were considered unsatisfactory because of the presence of E. coli, S. aureus, or both, while 11 of the cheeses were graded as unacceptable and potentially hazardous because of the presence of excessive numbers of S. aureus, E. coli, or L. monocytogenes and the presence of Salmonella in three of these. All cheeses graded as unacceptable and potentially hazardous were soft or semisoft cheeses made with ewe's and goat's milk, with the exception of two hard cheeses made with cow's milk. E. coli O157 was not detected in any of the cheeses. According to the present results, it seems that the presence or counts of pathogenic or indicator organisms in raw milk cheeses cannot be related to the processing conditions, milk type, or region of production.
Travel and migration are the major drivers of human brucellosis in Western Europe. The infection is usually transmitted through the consumption of unpasteurized milk or dairy products in or from endemic regions. Although eradicated from livestock in Germany and most Member States of the European Union, considerable numbers of domestic human brucellosis cases have been reported annually. The actual source of these autochthonous cases in non-endemic countries remains to be elucidated. We therefore evaluated the presence of Brucella spp. in 200 cheese samples originating from endemic countries which were sold at weekly markets, in supermarkets and by delis in Berlin (Germany) as well as online. The cheese samples included loose, non-labelled and pre-packed, labelled cheese of five types (brine, cream, soft, semi-hard and hard cheese), made from bovine, ovine and caprine milk. The cheese was mainly declared as raw milk cheese by the retailers. We screened for and confirmed the presence of Brucella-DNA in cheese using genus-specific quantitative real-time PCRs targeting IS711 and bcsp31, respectively. The molecular prevalence of Brucella was 20.5% (n = 41), but viable Brucellae could not be isolated from the positively tested samples using classical culture methods. The logistic regression model indicated that Brucella was significantly more often detected in late summer purchases (p = 0.036) as well as in cheese from Bulgaria, France, Greece and Turkey (p = 0.017). In contrast to the vendor information, essentially only three positive cheese samples were made from raw milk. Moreover, positive samples clustered at certain vendors which indicates large-scale illegal imports. In summary, Brucella in imported raw milk cheese seems to be still a challenge for food safety standards in the European Union. Uncontrolled import of dairy products from endemic regions might explain human Brucella infections acquired in non-endemic EU countries.
This review compiles published information concerning the incidence of pathogenic microorganisms — Listeria monocytogenes, Salmonella spp., Staphylococcus aureus and shiga-toxin producing Escherichia coli (STEC) — in goat and sheep raw milk and cheese. Meta-analytical data were extracted from 37 primary studies undertaken in Australia, Brazil, China, Colombia, Costa Rica, Czech Republic, Egypt, Germany, Greece, Iran, Italy, Malaysia, Mexico, Norway, Poland, Portugal, Spain, Sweden, Switzerland, Turkey, UK and USA. Pooled frequencies of detection of pathogens were found to be similar for sheep and goat raw milk: Salmonella (1.4–2.4%), L. monocytogenes (2.9–3.6%), STEC (4.3–4.8%) and S. aureus (35–39%). Likewise, in goat cheeses, regardless of being made of raw or heat-treated milk, S. aureus has been the most frequent contaminant (16.0%), whereas in raw milk cheeses, regardless of origin, the pooled prevalence of S. aureus is equally high in hard (34.6%) and soft cheeses (25.7%). L. monocytogenes is another important pathogen in sheep and goat milk cheeses (3.6–12.8%) while E. coli O157 strains with virulence genes (4.3%) also appear to persist during cheese manufacture. As expected, STEC has a higher pooled incidence in raw milk cheeses (10.0%) than in pasteurised milk cheeses (4.7%). Thus, the moderate contamination in raw milk and cheese of sheep and goat origin, revealed by this meta-analysis, advocates the reinforcement of general prevention measures such as close monitoring of hygiene on farms and eradication of disease by sheep and goat dairy farmers. Moreover, for the production of traditional cheeses made of raw milk, preventive measures during processing, namely, regular sterilisation of dairy equipment, process monitoring and hygiene of operators, should be even more stringent.
Caciofiore della Sibilla is a specialty ewe's milk cheese traditionally manufactured in a foothill area of the Marche region (Central Italy) with a crude extract of fresh young leaves of Carlina acanthifolia All. subsp. acanthifolia as a coagulating agent. The fungal dynamics and diversity of this specialty cheese were investigated throughout the manufacturing and 20-day ripening process, using a combined PCR-DGGE approach. The fungal biota of a control ewe's milk cheese manufactured with the same batch of milk coagulated with a commercial animal rennet was also monitored by PCR-DGGE, in order to investigate the contribution of the peculiar vegetable coagulant to the fungal diversity and dynamics of the cheese. Based on the overall results collected, the raw milk and the dairy environment represented the main sources of fungal contamination, with a marginal or null contribution of thistle rennet to the fungal diversity and dynamics of Caciofiore della Sibilla cheese.
The aim of this study was to investigate the prevalence and virulence factors of Staphylococcus aureus isolates from sheep and goat dairy products using end-point PCR. A total of 90 samples were analyzed and 37 (41.1%) resulted positive for S. aureus. S. aureus genes were found in 7 (18.9%) out of 37 isolates: sec was the most frequently recovered gene (42.8%), followed by seh (28.6%), sea (14.3%) and see (14.3%). The mecA gene was detected in one isolate (2.7%), and this strain was also positive for the presence of seh. PCR revealed 35 isolates (94.6%) as positive for the icaA gene. No S. aureus isolate carried lukF-PV and lukS-PV genes. The presence of these potentially harmful bacteria is a potential public health risk, therefore processing innovation in the ovine and goat dairy industry is required in order to reduce the microbiological contamination of these foods.
Ricotta Salata is a traditional ripened and salted whey cheese made in Sardinia (Italy) from sheep's milk. This product is catalogued as ready-to-eat food (RTE) since it is not submitted to any further treatment before consumption. Thus, foodborne pathogens, such as Listeria monocytogenes, can represent a health risk for consumers. In September 2012, the FDA ordered the recall of several batches of Ricotta Salata imported from Italy linked to 22 cases of Listeriosis in the United States. This study was aimed at evaluating the presence and virulence properties of L. monocytogenes in 87 samples of Ricotta Salata produced in Sardinia. The ability of this product to support its growth under foreseen packing and storing conditions was also evaluated in 252 samples. Of the 87 samples 17.2% were positive for the presence of L. monocytogenes with an average concentration of 2.2 log10 cfu/g. All virulence-associated genes (prfA, rrn, hlyA, actA, inlA, inlB, iap, plcA, and plcB) were detected in only one isolated strain. The Ricotta Salata samples were artificially inoculated and growth potential (δ) was assessed over a period of 3 mo. The value of the growth potential was always >0.5 log10 cfu/g under foreseen packing and storing conditions. This study indicates that Ricotta Salata supports the L. monocytogenes growth to levels that may present a serious risk to public health, even while stored at refrigeration temperatures.
This review concentrates on information concerning microbiological hazards possibly present in raw milk dairy products, in particular cheese, butter, cream and buttermilk. The main microbiological hazards of raw milk cheeses (especially soft and fresh cheeses) are linked to Listeria monocytogenes, verocytotoxin-producing Escherichia coli (VTEC), Staphylococcus aureus, Salmonella and Campylobacter. L.monocytogenes, VTEC and S.aureus have been identified as microbiological hazards in raw milk butter and cream albeit to a lesser extent because of a reduced growth potential compared with cheese. In endemic areas, raw milk dairy products may also be contaminated with Brucella spp., Mycobacterium bovis and the tick-borne encephalitis virus (TBEV). Potential risks due to Coxiella burnetii and Mycobacterium avium subsp. paratuberculosis (MAP) are discussed. Pasteurisation ensures inactivation of vegetative pathogenic microorganisms, which increases the safety of products made thereof compared with dairy products made from raw milk. Several control measures from farm to fork are discussed.
The distribution of mould species was examined at several points of the processing chain in a Manchego cheese plant and associated dairy farms. Geotrichum and Fusarium were the genera most frequently isolated from milk samples as well as in cheeses ripened for one month, evidencing a direct transfer from raw milk. Conversely, the mycobiota of long-ripened cheeses consisted mainly of Penicillium species, which gained entry to the cheese through the air of ripening rooms. This study contributes to the understanding of the dynamics of fungal populations in semihard and hard cheeses, highlighting that airborne transfer from the stables could have a direct impact on their quality.
IntroductionIndividual studiesThe summary effectHeterogeneity of effect sizesSummary points
Animal products are one of the niches of bifidobacteria, a fact probably attributable to secondary contamination. In this study, 2 species of the genus Bifidobacterium were isolated by culture-dependent methods from ovine cheeses that were made from unpasteurized milk without addition of starter cultures. The isolates were identified as Bifidobacterium crudilactis and Bifidobacterium animalis subsp. lactis using matrix-assisted laser desorption/ionization time-of-flight analysis and sequencing of phylogenetic markers (16S rRNA, hsp60, and fusA).
Meat and meat products are the main vehicles of foodborne diseases in humans caused by pathogens such as Salmonella spp., Campylobacter spp., Listeria monocytogenes, Yersinia enterocolitica, verotoxigenic Escherichia coli (VTEC) and Staphylococcus aureus. In order to prioritise research on those microbial hazards, a meta-analysis study was conducted to summarise available information on the presence of such pathogens in meats produced in Portugal. By using a logit-transformed proportion as effect size parameterisation, a number of multilevel random-effect meta-analysis models were fitted to estimate mean occurrence rates of pathogens, and to compare them among meat categories (i.e., bovine meat, broiler meat, pork, minced beef and minced pork), and among meat product categories (i.e., intended to be eaten cooked, to be eaten raw and cured meats). The mean occurrence rate of Campylobacter in Portuguese broiler meat (40%; 95% CI: 22.0–61.4%) was about ten times higher than that of Salmonella (4.0%; 95% CI: 1.4–10.8%); although these levels were comparable to current EU ranges. Nevertheless, in the other meat categories, the meta-analysed incidences of Salmonella were slightly to moderately higher than EU averages. A semi-quantitative risk ranking of pathogens in Portuguese-produced pork pointed Salmonella spp. as critical (with a mean occurrence of 12.6%; 95% CI: 8.0–19.3%), and Y. enterocolitica as high (6.8%; 95% CI: 2.2–19.3%). In the case of the Portuguese meat products, the non-compliance to EU microbiological criteria for L. monocytogenes (8.8%; 95% CI: 6.5–11.8%) and Salmonella spp. (9.7%; 95% CI: 7.0–13.4%) at sample units level, in the categories ‘intended to be eaten cooked’ and ‘to be eaten raw’, were considerably higher than EU levels for ready-to-eat products in comparable categories. S. aureus was the pathogen of greatest concern given its high occurrence (22.6%; 95% CI: 15.4–31.8%) in meat products. These results emphasised the necessity of Portuguese food safety agencies to take monitoring, and training actions for the maintenance of good hygiene practices during the production of the great variety of traditional meat products. This meta-analysis study also highlighted important gaps of knowledge, and may assist food safety authorities in the prioritisation of microbiological hazards, and the implementation of essential food safety assurance systems at primary production.
Listeria monocytogenes is a ubiquitous bacterium widely distributed in the environment that can cause a severe disease in humans when contaminated foods are ingested. Cheese has been implicated in sporadic cases and in outbreaks of listeriosis worldwide. Environmental contamination, in several occasions by persistent strains, has been considered an important source of finished product contamination. The objectives of this research were to (i) evaluate the presence of L. monocytogenes within the factory environments and cheeses of three processing plants, artisanal producer of raw ewe's milk cheeses (APC), small-scale industrial cheese producer (SSI) and industrial cheese producer (ICP) each producing a distinct style of cheese, all with history of contamination by L. monocytogenes (ii) and identify possible sources of contamination using different typing methods (arsenic and cadmium susceptibility, geno-serotyping, PFGE). The presence of markers specific for 3 epidemic clones (ECI-ECIII) of L. monocytogenes was also investigated. Samples were collected from raw milk (n=179), whey (n=3), cheese brining solution (n=7), cheese brine sludge (n=505), finished product (n=3016), and environment (n=2560) during, at least, a four-year period. Listeria monocytogenes was detected in environmental, raw milk and cheese samples, respectively, at 15.4%, 1.1% and 13.6% in APC; at 8.9%, 2.9% and 3.4% in SSI; and at 0%, 21.1% and 0.2% in ICP. Typing of isolates revealed that raw ewe's milk and the dairy plant environment are important sources of contamination, and that some strains persisted for at least four years in the environment. Although cheeses produced in the three plants investigated were never associated with any case or outbreak of listeriosis, some L. monocytogenes belonging to specific PFGE types that caused disease (including putative epidemic clone strains isolated from final products) were found in this study.
In the context of the prevailing trend toward more natural products, there seems to be an increasing preference for raw milk consumption as raw milk is associated with several perceived health benefits that are believed to be destroyed upon heating. However, many human pathogens can be isolated from raw cow milk. The prevalence of foodborne pathogens in raw cow milk varies, but their presence has been demonstrated in many surveys and foodborne infections have been repeatedly reported for Campylobacter, Salmonella spp. and human pathogenic verocytotoxin-producing Escherichia coli. In industrialized countries, milk-borne and milk product-borne outbreaks represent 2–6% of the bacterial foodborne outbreaks.The aim of this review is to present scientifically sound data regarding the risks and benefits related to the consumption of raw and heated cow milk. Both microbiological aspects (e.g., the prevalence of milk-borne pathogens, pathogen growth inhibition by antimicrobial systems and by lactic acid producing bacteria, probiotic bacteria, etc.) and nutritional or health aspects (nutritional value, immunity, allergies, lactose intolerance, diabetes, milk digestibility, etc.) are considered.As such, it is demonstrated that consumption of raw milk poses a realistic health threat due to a possible contamination with human pathogens. It is therefore strongly recommended that milk should be heated before consumption. With the exception of an altered organoleptic profile, heating (in particularly ultra high temperature and similar treatments) will not substantially change the nutritional value of raw milk or other benefits associated with raw milk consumption.
Traditional and novel approaches for the calculation of the heat treatment efficiency are compared in this work. The Mild Heat value (MH-value), an alternative approach to the commonly used sterilisation, pasteurisation and cook value (F, P, C-value), is calculated to estimate the efficiency of a mild heat process. MH-value is the time needed to achieve a predefined microbial reduction at a reference temperature and a known thermal resistant constant, z, for log-linear or specific types of non-log-linear microbial inactivation kinetics. An illustrative example is given in which microbial inactivation data of Listeria innocua CLIP 20-595 are used for estimating the inactivation parameters under isothermal conditions of 58, 60, 63 and 66 °C by the use of the log-linear and the Geeraerd et al. (2000) model. Thereafter, dynamic temperature profiles (targeting at 54 and 57 °C) representing milk thermisation are exploited for illustrating the application of MH-value. Finally, the equivalent holding times of different temperatures are calculated taking into account the observed non-linearity.
The aim of this study was to survey the presence of Staphylococcus aureus and Listeria monocytogenes during the cheese making process in small-scale raw milk cheese production in Norway. The prevalence of S. aureus in bovine and caprine raw milk samples was 47.3% and 98.8%, respectively. An increase in contamination during the first 2-3 h resulted in a 73.6% prevalence of contamination in the bovine curd, and 23 out of 38 S. aureus-negative bovine milk samples gave rise to S. aureus-positive curds. The highest contamination levels of S. aureus were reached in both caprine and bovine cheese after 5-6 h (after the first pressing). There was no contamination of L. monocytogenes in caprine cheeses and only one (1.4%) contaminated bovine cheese. This work has increased our knowledge about S. aureus and L. monocytogenes contamination during the process of raw milk cheese production and gives an account of the hygiene status during the manufacture of Norwegian raw milk cheeses.
The objective of this study was to address knowledge gaps identified in an earlier risk assessment of Staphylococcus aureus and raw milk cheese. A survey of fresh and short-time ripened cheeses produced on farm-dairies in Sweden was conducted to investigate the occurrence and levels of S. aureus, Listeria monocytogenes and Escherichia coli, to characterize S. aureus isolates with special emphasis on enterotoxin genes, antibiotic resistance, bio-typing and genetic variation, and to collect information related to production practices. In general, the hygienic quality of farm-dairy cheeses appeared to be of an acceptable microbiological quality, e.g. L. monocytogenes and staphylococcal enterotoxin were not detected in cheese samples. However, E. coli and enterotoxigenic S. aureus were frequently found in raw milk cheeses and sometimes at levels that are of concern, especially in fresh cheese. Interestingly, levels in raw milk fresh cheese were significantly lower when starter cultures were used. Up to five S. aureus colonies per cheese, if possible, were characterized and about 70% of isolates carried one or more enterotoxin genes, most common were sec and sea. The Ovine biotype (73%) was most common among isolates from goat milk cheese and the Human biotype (60%) from cow milk cheese. Of all isolates, 39% showed decreased susceptibility to penicillin, but the proportion of isolates from cows' cheese (66%) compared to isolates from goats' cheese (27%) was significantly higher. S. aureus isolates with different properties were detected in cheese from the same farm and, sometimes even the same cheese. Isolates with the same pulsed-field gel electrophoresis (PFGE)-pattern were detected on geographically distant dairies. This indicates that multiple sources and routes of contamination are important. To improve the safety of these products efforts to raise awareness of the importance of hygiene barriers and raw milk quality as well as improved process control can be suggested, e.g. use of starter cultures and monitoring of fermentation with a pH-meter. For future safety assessments, a better understanding of factors determining toxin production in these cheeses is needed.
The aim of this study was to investigate the presence of Staphylococcus aureus (S. aureus) and staphylococcal enterotoxins (SEs) genes in sheep cheese and dairy dessert samples by multiplex PCR (mPCR) technique. A total of 150 samples were analyzed consisting of 50 dairy dessert samples and 100 sheep cheese. Coagulase positive staphylococci (CPS) were found in 86 (57.3%) out of 150 analyzed samples. S. aureus were isolated from 60 (60%), 26 (52%) of sheep cheese and from of dairy desserts, respectively. Five suspected colonies were tested from each sheep cheese and dairy dessert samples for phenotypic and genotypic characterizations. A total of 430 isolates from the 86 positive samples were investigated in this study. Eighty (18.6%) isolates were characterized as S. aureus. The enterotoxin genes (sea, seb, sec, sed) were found in 13 (3.02%) out of 80 isolates. From cheese isolates, sea, seb and sed were detected in 5 (1.6%), 2 (0.6%), 1 (0.3%), respectively. From dairy dessert isolates, sea, sec and sed were detected in 3 (2.3%), 1 (0.76%), 1 (0.76%), respectively. The presence of SEs was identified in 12 (2.8%) out of 80 isolates by using ELISA technique. It was determined that these SEs had a distribution of 7 (1.6%) SEA, 2 (0.46%) SEB, 1 (0.23%) SEC, and 2 (0.46%) SED. SEs were found in 7 (2.3%) cheese and 5 (3.8%) dairy dessert isolates. In conclusion, S.aureus and their SEs were found to be present in sheep cheese and dairy desserts in this study. It is emphasized that the presence of S. aureus and their SEs genes in sheep cheese and dairy desserts may be regarded as a potential risk for human health.
Food is an important vehicle for transmission of Shiga toxin-producing Escherichia coli (STEC). To assess the potential public health impact of STEC in Swiss raw milk cheese produced from cow's, goat's, and ewe's milk, 1,422 samples from semihard or hard cheese and 80 samples from soft cheese were examined for STEC, and isolated strains were further characterized. By PCR, STEC was detected after enrichment in 5.7% of the 1,502 raw milk cheese samples collected at the producer level. STEC-positive samples comprised 76 semihard, 8 soft, and 1 hard cheese. By colony hybridization, 29 STEC strains were isolated from 24 semihard and 5 soft cheeses. Thirteen of the 24 strains typeable with O antisera belonged to the serogroups O2, O22, and O91. More than half (58.6%) of the 29 strains belonged to O:H serotypes previously isolated from humans, and STEC O22:H8, O91:H10, O91:H21, and O174:H21 have also been identified as agents of hemolytic uremic syndrome. Typing of Shiga toxin genes showed that stx(1) was only found in 2 strains, whereas 27 strains carried genes encoding for the Stx(2) group, mainly stx(2) and stx(2vh-a/b). Production of Stx(2) and Stx(2vh-a/b) subtypes might be an indicator for a severe outcome in patients. Nine strains harbored hlyA (enterohemorrhagic E. coli hemolysin), whereas none tested positive for eae (intimin). Consequently, semihard and hard raw milk cheese may be a potential source of STEC, and a notable proportion of the isolated non-O157 STEC strains belonged to serotypes or harbored Shiga toxin gene variants associated with human infections.
For 5 months, the udders of milking ewes, raw ewe's milk, cheese, and the plant and environment of a cheese manufacturer in Portugal were investigated using standard methods for the presence of Listeria spp. An association between subclinical mastitis and Listeria monocytogenes in a single lactating sheep was investigated by visual inspection of udders for signs of inflammation, application of somatic cell counts, the California mastitis test, pH measurement to milk, and culture of L. monocytogenes and Staphylococcus spp. To track the routes of contamination by L. monocytogenes, 103 isolates were characterized by molecular serotyping and amplified fragment length polymorphism, and a selection was further tested by pulsed-field gel electrophoresis. This study provides molecular and epidemiological evidence tracking the persistence of a single L. monocytogenes strain causing a subclinical udder infection without obvious inflammation in a single ewe. This infection was the likely source of contamination of raw milk that was subsequently used to produce unpasteurised milk cheese and resulted in a single strain of this bacterium colonizing the processing environment and the final cheese product.
Sheep and goat edible products (mainly meat and dairy products) have interesting characteristics in their levels of flavour, taste, aromas and leanness as well as the specific composition of fats, proteins, amino and fatty acids. Their quality is very much linked to historical and cultural uniqueness right through the production, marketing and consumption chains. This refers, at least in the Mediterranean region, to farming systems with dominant extensive grazing situations, specific technologies and conditions for slaughtering as well as for the transformation processes of cheese-making and its maturing; they are also characterised by traditional nutritional habits of the consumers. While the organoleptic properties of the dairy products are very important, the sanitary aspects become more and more influential and tend to modify the accepted definitions of these products and their quality (e.g. non-pasteurised milk, production chains, hygiene, transformation methodologies, etc.). Today there are major research efforts to ameliorate the production aspects but also the quality of the products; better cheese yield with the work undertaken on alpha-s1-casein, flavour and taste of cheeses is improved by the work on lipolysis and the study of the molecules responsible for the taste of the cheeses, studies on the fattening level of animals including genetic variability, and a better understanding of the fat distribution on carcasses. The future evolution of these products is difficult to foresee. Will quality be the criterion which will give the market direction in the developed countries (standardised products and top quality special products)? In the developing regions will we be able to maintain the actual production which is well adapted to the local demand or will this not be handicapped by the fragility and marginalisation of the existing livestock farming systems and untenable rises in costs?
In this study, a total of 105 samples including 35 raw milk, 35 cows' milk cheese, and 35 ewes' milk cheese samples were obtained from Kirikkale city and investigated for the presence and contamination level of Brucella species. For this reason, milk and cheese samples were collected in aseptic conditions and brought to the laboratory under cold conditions. The method suggested from FARREL was used for the isolation and identification of Brucella spp. In addition, Most Probable Number Technique was used to determine the contamination level in Brucella spp. positive samples. According to analysis findings, B. melitensis was isolated from 5 (14.2%) of 35 ewes' milk cheese samples at the level of 3.6 x 10(1)-9.3 x 10(3) MPN/g. Brucella spp. were not detected in any of raw milk and cows' milk cheese samples. In conclusion this study reinforced the previous reports that ewes' milk cheese is an important source of Brucella spp. and they have been risk for public health.
The aim of this study was to determinate the prevalence, serotypes and virulence genes of Shiga toxin-producing Escherichia coli (STEC) strains isolated from different dairy products (DP) in Spain with the purpose of determining whether DP represent a potential source of STEC pathogenic for humans. A total of 502 DP were examined from 64 different ovine and caprine flocks and 6 dairy plants in Extremadura (Western Spain). Samples were collected monthly between March 2003 and June 2004 and included 360 unpasteurised milk obtained from the bulk tank, 103 fresh cheese curds and 39 cheeses. Samples obtained were examined for STEC using genotypic (PCR) methods. STEC strains were detected from 39 (10.8%) bulk tank, 4 (3.9%) fresh cheese curds and 2 (5%) cheese, whereas O157:H7 serotype were isolated from one (0.3%) bulk tank. A total of 9 STEC strains (O27:H18, O45:H38, O76:H19, O91:H28, O157:H7, ONT:H7, ONT:H9 and ONT:H21) were identified in this study. One of them, the serotype O27:H18, has not been reported previously as STEC. PCR showed that 3 strains carried stx1 genes, 5 possessed stx2 genes and 1 both stx1 and stx2. Whereas all STEC caprine isolates showed ehxA genes, only O157:H7 serotype showed eae virulence genes. The strain O157:H7 isolated possessed intimin type gamma1 and belonged to phage type 31. This study confirms that dairy product is an important reservoir of STEC pathogenic for humans.
The aim of the present study was to investigate the occurrence of Escherichia coli O157:H7 and other Shiga toxin-producing E. coli (STEC) in 'Castellano' cheese, a non-cooked and hard or semi-hard Spanish cheese made from ewe's milk. A total of 83 raw milk cheese samples with different ripening times (2.5, 6 and 12 months) were taken at 30 cheese factories. Samples were examined for the presence of STEC using in the first stage the Association of Official Analytical Chemists (AOAC) official method number 997.11, and then, in the second stage, isolates were tested for virulence genes using genotypic (PCR) methods. Three STEC strains were detected in two samples (2.4%) of 'Castellano' cheese, one with 2.5 and the other one with 12 month-ripening period. From those STEC isolates, two were identified as E. coli O14 and the third presented an O-specific polysaccharide not-groupable serologically (ONG). PCR showed that all isolates were characterized by harbouring the Shiga toxin (stx) stx1 gene and by the absence of the genes for stx2, eaeA, and ehxA virulence factors. This study revealed the potential of STEC to survive in long-ripened-hard cheeses.
Two studies of retail fresh, ripened and semi-hard cheeses made from raw, thermized or pasteurized milk were undertaken in the UK during 2004 and 2005 to determine the microbiological quality of these products. Using microbiological criteria in European Commission Recommendations 2004/24/EC and 2005/175/EC, 2% of both raw, thermized (37/1819 samples) and pasteurized (51/2618 samples) milk cheeses were of unsatisfactory quality. Raw or thermized milk cheeses were of unsatisfactory quality due to levels of Staphylococcus aureus at 10(4)cfu g(-1), Escherichia coli at 10(5)cfu g(-1), and/or Listeria monocytogenes at 10(2)cfu g(-1), whereas pasteurized milk cheeses were of unsatisfactory quality due to S. aureus at 10(3)cfu g(-1) and/or E. coli at 10(3)cfu g(-1). Salmonella was not detected in any samples. Cheeses were of unsatisfactory quality more frequently when sampled from premises rated as having little or no confidence in management and control systems, and stored/displayed at above 8 degrees C. Raw or thermized milk cheeses were also more likely to be of unsatisfactory quality when they were unripened types, and pasteurized milk cheeses when they were: semi-hard types; from specialist cheese shops or delicatessens; cut to order. These results emphasize the need for applying and maintaining good hygiene practices throughout the food chain to prevent contamination and/or bacterial growth. Labelling of cheeses with clear information on whether the cheese was prepared from raw milk also requires improvement.
Goats' milk cheeses (n=181) from the Hessian market (retail shops, weekly markets, farm markets) were quantitatively analysed for Staphylococcus (S.) aureus, and 14 were found positive. From these samples, 64 isolates of S. aureus were characterized biochemically and genetically, including their potential to produce staphylococcal enterotoxins (SE). SE genes sea to selo was studied by PCR and gene expression was evaluated by reverse transcriptase (RT)-PCR. SEA-SEE production in culture was determined by enzyme immunoassay (EIA). One isolate produced SEA, 18 isolates (from 4 samples) produced SEC, while SEB, SED, and SEE were not found. Toxin production was in agreement with PCR and RT-PCR results for the presence and expression, respectively, of the corresponding toxin genes. Trans-SE genes seg, sei, selm, seln, and selo were detected in 14 isolates from 4 cheese samples, exclusively as clusters. These samples were all from small-scale producers which directly or indirectly market their products regionally. No isolate was positive for seh or sej. RT-PCR detected the presence of the corresponding mRNA for all genes except selo, further indicating the possibility that respective proteins indeed have been produced in culture. These results suggest that S. aureus in goats' milk cheese potentially produces SE like proteins, besides SEA and SEC.
The aim of this study was to describe the prevalence, serotypes and virulence genes of Shiga toxin-producing Escherichia coli (STEC) isolated from raw milk cheese samples collected at producer level with the purpose of determining whether raw milk cheeses in Switzerland represent a potential source of STEC pathogenic for humans.
Raw milk cheese samples (soft cheese n=52; semi-hard and hard cheese n=744; all produced from Swiss cows’, goats’ and sheep milk) collected at producer level throughout Switzerland within the national sampling plan during the period of March 2006 to December 2007 were analyzed.
Of the 432 cheese samples obtained in the year 2006 and the 364 samples obtained in the year 2007, 16 (3.7%) and 23 (6.3%), respectively were found to be stx positive. By colony dot-blot hybridization, non-O157 STEC strains were isolated from 16 samples. Of the 16 strains 11 were typed into seven E. coli O groups (O2, O15, O22, O91, O109, O113, O174), whereas five strains were non-typeable (ONT). Among the 16 STEC strains analyzed, stx1 and stx2 were detected in one and 15 strains. Out of the 15 strains with genes encoding for the Stx2 group, four strains were positive for stx2, six strains for stx2d2, two strains for stx2-O118, one strain for stx2-06, one strain for stx2g, one strain for stx2 and stx2d2, and one strain for stx2 and stx2g. Furthermore, three STEC strains harbored E-hlyA as a further putative virulence factor. None of the strains tested positive for eae (intimin).
Results obtained in this work reinforce the suggestion that semi-hard raw milk cheese may be a potential vehicle for transmission of pathogenic STEC to humans.
Milk and milk product statistics - Statistics Explained
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