Ewe Milk Samples Research Articles

Exploring Listeria monocytogenes in Ewe Milk: ssrA Gene-based Real-time PCR Identification, Phylogenetic Analysis, and Antibacterial Assessment of Magnesium Oxide Nanoparticles Synthesized with Myrtus communis Leaf Extract

Background: Listeria monocytogenes, a zoonotic pathogen affecting humans and animals, exhibits a global distribution, including Iraq. This study focused on the rapid identification and phylogenetic analysis of L. monocytogenes in freshly collected ewe milk samples from various farms in Al-Qadisiyah province, Iraq.Methods: The study was conducted with care and precision, involving 150 milk samples. These samples were subjected to traditional bacterial isolation and identification using the enrichment culture method and biochemical tests, with the PCR technique confirming the results. The antibacterial activity of MgONPs was then assessed using the disc diffusion method, ensuring a comprehensive and reliable approach to the study.Result: The results show that 150 ewe milk samples underwent real-time PCR (RT-PCR) targeting the ssrA gene, followed by partial 16S rRNA gene sequencing (PSGS) of purified conventional PCR products. Furthermore, the study entails the biosynthesis of magnesium oxide nanoparticles using Myrtus communis leaf extract, followed by a comprehensive characterization utilizing UV-spectra, FTIR, SEM, and TEM techniques. The Agar well diffusion method assessed the antibacterial efficacy of these Biosynthesized nanoparticles against L. monocytogenes. The RT-PCR results revealed the presence of L. monocytogenes in 36 out of 150 samples (24%). Subsequent PCR analysis confirmed the presence of the pathogen in 30 out of these 36 positive samples (83.33%). Sequencing of two purified PCR products demonstrated 100% nucleotide identity with global isolates from Iraq and Turkey. Furthermore, the study demonstrated that L. monocytogenes exhibited substantial sensitivity (24.66 ± 0.3) to the biosynthesized magnesium oxide nanoparticles. These findings underscore the speed and precision of the RT-PCR method for detecting L. monocytogenes in fresh ewe milk samples.Conclusion: This comprehensive investigation enhances our understanding of L. monocytogenes prevalence in ewe milk and highlights the potential of Myrtus communis -derived nanoparticles for combating this pathogen.Keywords: Antibacterial nanoparticles; Listeria monocytogenes; Magnesium oxide; Myrtus communis sheep milk; ssrA gene

Effects of management strategies during early lactation and weaning on etiological agents of ovine subclinical mastitis and antimicrobial susceptibility of milk-derived bacterial isolates.

Subclinical mastitis is a common intramammary disease in sheep production systems. Expenses associated with compromised animal performance, therapeutic interventions, and decreased ewe longevity make efforts to minimize its prevalence worthwhile. The objectives of this study were to 1) quantify the prevalence of subclinical mastitis throughout lactation, 2) evaluate the impact of bedding treatments on subclinical mastitis during early lactation, 3) evaluate the efficacy of prophylaxis and feed restriction during weaning on subclinical mastitis cure rates, and 4) identify levels and types of antimicrobial resistance in milk-derived bacteria. Ewe milk samples were collected at days 1, 2, and 28 post-partum, weaning, and 3-d post-weaning for bacterial identification via culture-based methods. Staphylococcus spp. and Streptococcus spp. isolates were subjected to in vitro antimicrobial susceptibility testing. The overall prevalence of subclinical mastitis defined by culture growth ranged between 22% and 66% and differences were observed between post-weaning and days 1 and 28 milk samples. Commonly isolated bacteria include coagulase-negative staphylococci (CoNS; 59%), Bacillus spp. (35%), Mannheimia haemolytica (10%), Staphylococcus aureus (8%), Streptococcus spp. (5%), and Corynebacterium spp. (5%). Early milk samples (days 1 and 2) were compared between jug bedding treatment: jugs were recently vacated, cleaned, and dusted with barn lime before adding fresh straw (CLEAN) or jugs were previously vacated and fresh straw was added atop soiled bedding (SOILED). Jug bedding treatment did not affect the prevalence of subclinical mastitis, though CoNS had greater sulfadimethoxine resistance in SOILED isolates than CLEAN isolates (P = 0.03). Three different weaning treatments were used: ewes were injected with penicillin at weaning (PENN), ewes had restricted feed access 48 h prior to and 72 h post-weaning (FAST), or a combination of these treatments (COMBO). Weaning treatment did not affect the prevalence of subclinical mastitis or cure rate from weaning to 3-d post-weaning, though all PENN and no FAST milk S. aureus isolates were resistant against tetracycline (P = 0.08). Subclinical mastitis prevalence tended to decrease from weaning to post-weaning (P = 0.08). These data show that subclinical mastitis is common throughout lactation and the levels of antimicrobial resistance of bacteria isolated from ewe milk are generally low against commonly used antimicrobials.

Derivation of multivariate indices of milk composition, coagulation properties, and curd yield in Manchega dairy sheep

This study approaches the interrelation patterns between composition of milk and whey, curd yield, chromaticity, syneresis, and technological quality of Manchega sheep milk using multivariate factor analysis. In addition, the effect of the main husbandry components (flock, prolificacy, season of the year, stage of lactation, and parity) on the common latent factors that define the pattern of variation of Manchega milk was assessed. For this purpose, 1,200 individual Manchega ewe milk samples from 4 different flocks registered under the Protected Designation of Origin Queso Manchego were analyzed (50 ewes/flock). Samples were collected in 2 different seasons of the year (spring and autumn) and at 3 time points per season: early, mid-, and late lactation. The obtained results suggested that curd yield mainly depends on milk composition, and the retention of water in the curd is related to coagulation traits. Thus, composition and moisture content could be useful indicators to assess the efficiency and quality of milk intended for cheesemaking, regardless of the analysis of coagulation properties. Finally, in terms of husbandry, a direct effect of flock and stage of lactation was observed on all analyzed factors, with a lower influence of season and parity.

Quality of the ewes’ milk as a raw material in the dairy industry in the Republic of North Macedonia

This study is based on analyzing physico-chemical and microbiological characteristics of ewe milk from three different regions, which is used as a raw material in dairy industry of North Macedonia. All milk samples (n=123) were collected on dairy reception points of dairy industry in Pelagonia region, in a period of six (6) months. The average values for chemical composition of milk were in the range as follows: 6.83±0.22% to 7.68±0.36% for milk fat, 5.52±0.09% to 6.06±0.23% for proteins, 4.62±0.08% to 4.79±0.05% for lactose, 11.16±0.11% to 1.51±0.24% for non-fat dry matter (NFDM), -0.561 to -0.543oC for freezing point and 8.61±0.4 oSH to 8.88±0.47oSH for titrable acidity. The microbiological and hygienic status of examined ewe milk samples was of poor hygienic quality. The average value of the examined samples for somatic cell count ranged from 753.333 cfu/mL to 1.125.000 cfu/mL and the average number of total bacteria count was from 532.444 cfu/mL to 1.986.222 cfu/mL.

Effect of Molasses and Dried Orange Pulp as Sheep Dietary Supplementation on Physico-Chemical, Microbiological and Fatty Acid Profile of Comisana Ewe's Milk and Cheese.

The use of agro-industrial by-products for ruminant feed represents both an economical and environmental convenient way for reducing waste discharge and waste management costs for food industries. Large amounts of waste from citrus processing industries are available in Sicily, Italy. In the present study, the effect of dried citrus pulp as sheep dietary supplementation was evaluated on physico-chemical, microbiological and fatty acid composition of resulting milk and cheese. Pelleted feed integrated with molasses and blond orange pulp, replacing cane molasses, beet pulp and part of the maize and sunflower in ration, were administrated to ewes as an experimental treatment The experiment involved sixty Comisana breed sheep divided into two groups and two feeding trials (experimental and control). Ewe's milk and cheese samples were collected from January to April and analyzed for physico-chemical, microbiological and fatty acid profile composition. Results suggested that both the experimental milk and cheese were different from the controls. In particular, an increase of experimental milk yield and fat content were registered whilst the cheese samples exhibited a significant decrease of pH values and an increase in fat and protein contents. In addition, an increase of conjugated linoleic acids as well as of the oxidative stability were observed indicating the beneficial effect of dietary supplementation. Furthermore, among the main microbial groups, the experimental and control samples, no differences were detected. However, with the exception of streptococci, which was found higher in experimental cheeses, and staphylococci, which was significantly reduced by experimental feed. Moreover, the application of culture-independent methods highlighted the dominance of Lactobacillus rhamnosus/casei group in the experimental cheese, suggesting a driving role of the dietary supplementation in the cheese microbiota composition. The present study demonstrated that the inclusion of citrus by-products in the diet of small dairy ruminants is a promising feeding, which could positively affect milk composition and cheese manufacture.

Multivariate analysis of the milk coagulation process in ovine breeds from Spain

In Spain, ewe milk is mainly used for cheesemaking, and farming systems have traditionally been based on the use of autochthonous breeds. However, in recent years, the progressive introduction of highly productive foreign breeds in Spanish farms has led to an increasing interest in the characterization of dairy sheep breeds to evaluate whether genetic selection schemes should focus on productivity or milk technological aptitude. The purpose of this work was to explore milk composition and coagulation to classify 4 of the main dairy sheep breeds used in Spain. This study included 832 individual ewe milk samples from the breeds Manchega, Assaf, Merino de Grazalema, and Merino de Los Pedroches. Samples were analyzed for native pH, composition (fat, protein, lactose, and total solids), coagulation properties, and individual laboratory curd yield. An indicator of coagulation efficiency was also determined. Canonical discriminant analysis was performed to establish differences and similarities among breeds based on the measured variables. In addition, cluster analysis was performed to study and quantify the concrete relationships among the discriminated groups. Discriminant analysis proved to be a powerful tool to accurately draw distinctions between breeds. In all cases, discrimination among breeds was evident and the 4 breeds could be easily differentiated. Cluster analysis showed greater similarity between Merino de Grazalema and Assaf compared with the other breeds, and F-statistics indicated a higher discriminating ability for the variables related to milk composition. However, Merino de Grazalema and Manchega were difficult to separate according to milk composition, but the coagulation process differenced them clearly. Coagulation also evidenced similarities between Manchega and Merino de Los Pedroches, although the latter was revealed to be the most different breed of all 4, which could lay the ground for its differentiation as an independent breed in the Official Catalogue of Spanish Livestock Breeds.

An investigation of oral moxidectin carryover to nursing lambs via milk

The purpose of this study was to investigate the concentrations of moxidectin in the plasma and milk of lactating ewes and in the plasma of their nursing lamb. Four, single lamb bearing Border Leister x Dorset ewes were administered a single oral dose of moxidectin (0.2mg/kg). Plasma and milk samples were collected nine times within the first 72h and at 30 and 60 d post anthelmintic administration. Samples were measured using high-performance liquid chromatography. A pharmacokinetic analysis of moxidectin was conducted on ewe plasma and resulted in a CMAX of 13.1ng/mL at 8.2h (TMAX). Ewe milk and lamb plasma samples were reported descriptively. Three of the four ewes on trial demonstrated quantifiable levels of moxidectin in milk at baseline sampling. Ewe milk samples reached an average peak concentration of 151.1ng/mL 12h post administration. Moxidectin levels were detected up to 60 d in all milk samples. Lamb plasma samples reached an average peak concentration of 2.8ng/mL 24h post administration to the ewe. Results from the current study demonstrate that orally administrated moxidectin is carried over into the milk and evident in the plasma of nursing lambs. Further research is needed in order to understand how moxidectin concentrations and exposure time influences the fetus and nursing lamb in regards to the development of anthelmintic resistance.

Determination of insecticide Deltamethrin residues in local and imported raw milk samples collected from different animal's species and the effect of processing heat treatment on its content in milk

A total of 163 milk samples (500 ml each) of cows, ewe, goats, buffaloes, camels were collected randomly at weekly intervals (10 samples/ week) from both Abu-Ghraib and Al-Fudhailiya villages and also for different local retail markets inside the Baghdad province and these samples were including milk of different animal species, milk cans, bulk milk tanks and imported Ultra-high temperature processing milk. Among the total milk samples only 138 milk samples were examined during two climatic periods where the first period was in summer that extended from the beginning of September to the end of October 2015 while the second period was in winter that extended from the beginning of January to the end of February 2016. Besides that, some of the selected positive samples for Deltametrin residues were subjected to one of the commercial heat treatments such as 63°C/30 min., 80°C/5 min. and 100°C/5 min. to evaluate the efficiency of heat exposure on the degradation of Deltametrin residues in milk. The results pointed out that milk samples containing the higher fat percentage exhibited significantly (P<0.05) the highest concentration of Deltametrin in summer (0.08ppm) than in winter (0.008 ppm) seasons. It was clearly obvious that the detectable concentrations of the Deltametrin were higher in buffalo's and ewe milk samples than those found in cows, goats and camels and such results could be attributed to the higher fat content of buffalos and ewes milk than the other animals as well as the lipophilic nature of the Deltamethrin. In other word, increased the fat percentages of milk was being associated with an increased level of Deltametrin residues due to the lipophilic nature of the Deltametrin pesticide. The current results revealed that milk samples that were collected from buffaloes, ewes and cows recorded significantly (P<0.05) the highest Deltametrin residues in summer season where their mean levels that exceeded the accepted MRLs of 0.05 ppm to milk samples of goats and camels that had significantly (P<0.05) the lowest mean levels of Deltametrin residues where their means levels were 0.038 and 0.032 ppm respectively. There was a significant (P<0.05) seasonal variation of the Deltametrin concentrations in milk samples for each animal species where all the milk samples that were collected from buffaloes, ewe, cows, goats and camels had significantly (P<0.05) higher mean levels of Deltametrin residues in summer season than in winter season, milk samples that were collected from milk cans (5, 25 and 50 kg) recorded significantly (P<0.05). The highest Deltametrin residues during the summer season in comparison to 10 tons bulk milk tank samples.

Caprine lentivirus in sheep milk and semen

ABSTRACT With the objective of detecting the presence of caprine lentivirus (CLV) in ewe milk and in ram semen, ten matrixes and four reproducers experimentally infected with CLV were used. Samples of ewe milk were collected during the four months of lactation, five collections per animal, totaling 50 samples. Regarding the rams, eight semen collections were made per animal, during one year of experimentation, totaling 32 samples. The milk and semen samples were submitted to DNA extraction and the nested polymerase chain reaction test (nPCR) to detect CLV proviral DNA. Eight (16%) of the milk samples were positive in nPCR originating from two ewes. Only one (3.12%) semen sample was positive. The amplification products were sequenced, and were confirmed to be a CLV genomic sequence. Thus, the presence of CLV proviral DNA in sheep milk and semen was demonstrated, confirming the feasibility of infection between species, and alerting to the risk of spreading infections.

Biodiversity and origin of the microbial populations isolated from Masske, a traditional Iranian dairy product made from fermented Ewe's milk

Masske is a traditional Iranian dairy product containing 50% butterfat made from fermented ewe's milk. Overall, 672 bacterial isolates were collected from ewe's milk, fermented milk (FM) and Masske samples that were produced in households located in southern regions of the Khorasan Province in Iran. To identify lactic acid bacteria in these samples, a total of 79 Gram‐positive and catalase‐negative isolates were analysed. The identification of isolates was achieved by phenotypic and sequential analysis of the 16S rRNA gene. Enterococcus faecium and Aerococcus viridans were the most frequently isolated species in the samples, but the most commonly present bacteria in Masske were Streptococcus thermophilus.

Lead and cadmium in raw buffalo, cow and ewe milk from west Azerbaijan, Iran

In this study, 300 raw buffalo, cow and ewe milk samples from five townships in west Azerbaijan, Iran, were analysed. Lead and cadmium were determined using atomic absorption spectrophotometry. Mean concentration of lead and cadmium in buffalo milk samples was 0.018 ± 0.001 and 0.003 ± 0.001 mg/kg, respectively. Mean concentration of lead and cadmium in cow milk samples was 0.007 ± 0.001 and 0.001 ± 0.001 mg/kg, respectively, and in ewe milk, these mean values were 0.010 ± 0.001 and 0.002 ± 0.001 mg/kg, respectively. Statistical analyses showed that lead and cadmium concentrations in buffalo milk were significantly higher than those in cow and ewe milk. Moreover, the concentration of these heavy metals in ewe milk was significantly higher than that in cow milk. It was also found that concentration of these selected toxic metals in milk increased with increasing age of the animals.

Determination of IgG levels in bulk ewe's milk

Colostrum addition to milk results in a reduction of the yield during cheese manufacturing. Determination of IgG levels is accepted as an evidence of this troublesome habit due to the high level of this protein in colostrum. This study shows for the first time the IgG levels in bulk ewe's milk samples from Spain throughout the year. IgG quantification was carried out by a sandwich ELISA that recognizes specifically ovine IgG. The mean value of IgG concentration obtained from 481 samples was 0.271±0.253mg/ml. In general, this value is within the range considered acceptable for mature milk according to quality payment systems. Surprisingly, a high mean concentration was obtained in samples collected in winter, 0.742±0.378mg/ml, suggesting that births were grouped in that period. These results point up that standardization of ewe's milk production has not been achieved yet. For this reason, determination of immunoglobulin levels periodically would be a good parameter to control milk quality.

Prediction of fatty acid profiles in cow, ewe, and goat milk by mid-infrared spectrometry

Mid-infrared (MIR) spectrometry was used to estimate the fatty acid (FA) composition in cow, ewe, and goat milk. The objectives were to compare different statistical approaches with wavelength selection to predict the milk FA composition from MIR spectra, and to develop equations for FA in cow, goat, and ewe milk. In total, a set of 349 cow milk samples, 200 ewe milk samples, and 332 goat milk samples were both analyzed by MIR and by gas chromatography, the reference method. A broad FA variability was ensured by using milk from different breeds and feeding systems. The methods studied were partial least squares regression (PLS), first-derivative pretreatment + PLS, genetic algorithm + PLS, wavelets + PLS, least absolute shrinkage and selection operator method (LASSO), and elastic net. The best results were obtained with PLS, genetic algorithm + PLS and first derivative + PLS. The residual standard deviation and the coefficient of determination in external validation were used to characterize the equations and to retain the best for each FA in each species. In all cases, the predictions were of better quality for FA found at medium to high concentrations (i.e., for saturated FA and some monounsaturated FA with a coefficient of determination in external validation >0.90). The conversion of the FA expressed in grams per 100mL of milk to grams per 100g of FA was possible with a small loss of accuracy for some FA.

Antibacterial activity and cholesterol assimilation of lactic acid bacteria isolated from traditional Iranian dairy products

In this study, lactic acid bacteria were isolated from ewe milk, traditional yoghurt and sour buttermilk samples collected from different areas of Azarbayjan-e-sharqi in Iran. All the isolates were screened for their ability to produce bacteriocin like inhibitory substances (BLIS) by studying their inhibitory action against pathogens like Listeria monocytogenes, Salmonella enteritidis and Staphylococcus aureus, after eliminating the effect of organic acids and hydrogen peroxide. According to results, four of the isolates identified as Lactobacillus brevis, Lactobacillus pentosus, Pedoicoccus acidilactici and Lactobacillus paracasei were unaffected by the action of pH neutralization and hydrogen peroxide and showed inhibitory action against the tested pathogens. The inhibitory activities demonstrated by these isolates were completely inhibited in the presence of proteolytic enzymes.The isolates in study were further characterized for their cholesterol reduction ability. Cholesterol assimilation by both viable and dead cells of these strains was determined in MRS broth containing 0.3g/100 mL bile salt. According to results, highest level of cholesterol removal was recorded in L. brevis, while all the other isolates in study were also able to reduce cholesterol to lesser extent. To conclude, the Lactic Acid Bacteria isolated from these traditional products might be exploited for their probiotic potential for future studies.

Use of Front-Face Fluorescence Spectroscopy to Differentiate Sheep Milks from Different Genotypes and Feeding Systems

The objective of the present study was to assess the potential of front-face fluorescence spectroscopy coupled with chemometric tools for the evaluation of the quality of milk samples according to the feeding system and genotype. Fifty (n = 50) ewe's milk samples were scanned after excitation set at 250, 290, 322, and 380 nm and emission set at 410 nm. Thirty out of the 50 samples composed the first trial and were obtained from two different genotypes (i.e., Comisana versus Sicilo-Sarde); the second trial was composed of 20 samples obtained from the Sicilo-Sarde genotype with two different feeding systems in pen (soybean versus scotch bean). Milk samples were divided into four groups named Sicilo-Sarde with pasture feeding (Spas), Comisana with pasture feeding (Cpas), Sicilo-Sarde feeding on scotch bean (Ssco), and Sicilo-Sarde feeding on soybean (Ssoy). The factorial discriminant analysis was applied to the: (i) four groups (i.e., Spas, Ssco, Ssoy, and Cpas) and (ii) three groups composed only of Sicilo-Sarde genotype (i.e., Spas, Ssco, and Ssoy). Considering the four groups, the best result was obtained with the excitation vitamin A spectra since correct classification amounting to 76% was observed. When the factorial discriminant analysis was performed with the three groups belonging to the Sicilo-Sarde genotype, the best result was obtained again with vitamin A spectra (i.e., emission and excitation spectra) since 88.6% of correct classification was observed. Concatenation technique applied to the five fluorescence spectra improved the rate of classification between the four groups since 44 out of 50 samples were correctly classified. No misclassification was observed between milk samples collected from ewes with pasture feeding from the pen feeding. It was concluded from the obtained results that fluorescence spectroscopy could be considered as a powerful tool for differentiating between raw milks according to both genotype and feeding system.

Short communication: Jenny milk as an inhibitor of late blowing in cheese: A preliminary report

Late blowing on semihard and hard cheese may have an important economic effect on dairy production. Many studies have attempted to prevent this defect by physical treatment, the use of additives, and the use of bacteriocins. In this paper, we look at the effect of jenny milk as an inhibitor of blowing caused by clostridia and coliforms in ewe cheese making. Bulk ewe and jenny milk samples were collected in the morning by mechanical milking and were refrigerated at 4°C. On the collected samples, the count of somatic cells, coliforms, Clostridium butyricum, and Escherichia coli were determined. The bulk raw milk was divided in two 45-L vats: vat 1 was used as a control, whereas 0.5L of jenny milk was added to vat 2. Four semihard cheeses, weighing about 2kg each, were made from each vat. Cheese making was replicated twice. After a ripening period of 60 d, the count of coliforms and of C. butyricum was determined. In the treated group, a significant inhibition of coliform bacteria was observed. The addition of jenny milk in cheese making may prove to be a useful and innovative approach for the inhibition of spore-forming clostridia strains.

Heavy metals in raw cow and ewe milk from north-east Iran

The presence of toxic metals in milk may create significant health problems for the population. In this study, 1440 raw cow and ewe milk samples from 18 townships in north-east Iran were analysed in four different seasons. Lead, cadmium and mercury levels were determined using atomic absorption spectrophotometry. Mean concentration of lead, cadmium and mercury in cow milk samples was 12.9 ± 6.0, 0.3 ± 0.3 and 3.1 ± 0.3 ng g−1, respectively, and in ewe milk samples, these mean values were 14.9 ± 7.8, 1.6 ± 1.2 and 3.1 ± 0.3 ng g−1, respectively. Statistical analyses showed that lead and cadmium concentrations in ewe milk were significantly higher than in cow milk. Concentrations of these metals in ewe milk varied significantly with different seasons. As the concentrations did not exceed the safety limits, they could not pose a serious danger to public health.

Immune competence of the mammary gland as affected by somatic cell and pathogenic bacteria in ewes with subclinical mastitis

Immune competence of the ewe mammary gland was investigated by monitoring the leukocyte differential count, cytokine pattern, and endogenous proteolytic enzymes in milk samples with different somatic cell counts (SCC) and pathogenic bacteria. Furthermore, the leukocyte differential count and T-lymphocyte populations were evaluated in ewe blood. A total of 1,500 individual milk samples were randomly selected from the pool of the samples collected during sampling and grouped into 5 classes of 300 samples each, on the basis of SCC. Classes were <300,000cells/mL, from 300,000 to 500,000cells/mL, from 501,000 to 1,000,000cells/mL, from 1,001,000 to 2,000,000cells/mL, and >2,000,000cells/mL. Microbiological analyses of ewe milk were conducted to detect mastitis-related pathogens. Sheep whose udders were without clinical abnormalities, and whose milk was apparently normal but with at least 103cfu/mL of the same pathogen were considered to have subclinical mastitis and therefore defined as infected. Polymorphonuclear neutrophilic leukocytes (PMNL) and macrophages increased with SCC, whereas lymphocytes decreased. Milk samples with SCC >1,000,000cells/mL showed differences in leukocyte populations between uninfected and infected ewes, with higher percentages of PMNL and macrophages and lower percentages of lymphocytes in infected animals. Nonviable PMNL levels were the highest in ewe milk samples with SCC <300,000cells/mL; starting from SCC >500,000cells/mL, nonviable PMNL were higher in uninfected ewes than in infected ones. In infected animals giving milk with SCC >1,000,000cells/mL, a higher CD4+/CD8+ ratio was observed, suggesting that the presence of pathogens induced an activation of both CD4+ and CD8+. The levels of tumor necrosis factor-α and IL-12 were higher in infected than uninfected ewes, irrespective of SCC. Plasmin activity increased along with SCC and was always higher in infected than uninfected animals; cathepsin D increased starting from 1,001,000cells/mL in milk samples from noninfected ewes and starting from 301,000cells/mL in milk samples from infected animals. The associations between somatic cells, cytokines, endogenous proteolytic enzymes, and pathogenic bacteria can be used to better understand the pathogenesis of subclinical mastitis in ewes and the effect on the immune response of ewe mammary gland.

Characterization of coagulase-positive staphylococci isolated from tank and silo ewe milk

The prevalence of coagulase-positive staphylococci (CPS) was studied among 390 samples of ewe milk. Fifty-seven (14.85%) samples of tank milk and all samples (6) of silo milk gave a positive result on Baird-Parker agar base supplemented with rabbit plasma fibrinogen, whereas amplification of the coagulase (coa) gene was successful in 6 (1.56%) samples of tank milk and in all silo samples. Phenotypic and genetic analysis of 153 isolates identified 151 (98.69%) as Staphylococcus aureus. Amplification of the coa gene was positive for 149 isolates (97.39%). The staphylococcal enterotoxin (SE) C gene was detected in 116 strains (75.82%), whereas more than one SE gene was carried in 5 strains (3.26%). None of the isolates harbored the genes for SEE or for methicillin resistance. A high prevalence of CPS carrying enterotoxin genes should be of concern because ewe milk is mainly processed into raw milk cheeses. The detection of the coa gene from milk samples could help to assess the microbiological safety of raw milk intended for direct use in the dairy industry.

Fungal diversity in cow, goat and ewe milk

Knowledge of fungal diversity in the environment is poor compared with bacterial biodiversity. In this study, we applied the denaturing high-performance liquid chromatography (D-HPLC) technique, combined with the amplification of the ITS1 region from fungal rDNA, for the rapid identification of major fungal species in 9 raw milk samples from cow, ewe and goat, collected at different periods of the year. A total of 27 fungal species were identified. Yeast species belonged to Candida, Cryptococcus, Debaryomyces, Geotrichum, Kluyveromyces, Malassezia, Pichia, Rhodotorula and Trichosporon genera; and mold species belonged to Aspergillus, Chrysosporium, Cladosporium, Engyodontium, Fusarium, Penicillium and Torrubiella genera. Cow milk samples harbored the highest fungal diversity with a maximum of 15 species in a single sample, whereas a maximum of 4 and 6 different species were recovered in goat and ewe milk respectively. Commonly encountered genera in cow and goat milk were Geotrichum candidum, Kluyveromyces marxianus and Candida spp. (C. catenulata and C. inconspicua); whereas Candida parapsilosis was frequently found in ewe milk samples. Most of detected species were previously described in literature data. A few species were uncultured fungi and others (Torrubiella and Malassezia) were described for the first time in milk.