Ewes' milk samples with low (<500000 ml−1), medium (1000000–1500000 ml−1) and high (>2500000 ml−1) somatic cell counts (SCC) were used to manufacture hard ewes' cheese using the Zamorano cheese manufacturing protocol. Cheeses that had been ripened for 1, 2 and 3 months were used to obtain isoelectric ovine casein that was analysed by capillary electrophoresis. The texture of the cheeses during ripening was determined instrumentally using the Warner-Bratzler maximum shear force and assessed for sensory qualities by consumers using hedonic tests. The study revealed that the pH value and the lactose content of the milk were affected by high SCC and that the coagulation properties were dependent on the somatic cell content. The protein and moisture contents of the cheeses were unaffected by SCC but a significant increase of pH with ripening time were observed in high-SCC cheeses. The results also pointed to a significant increase in proteolysis related to SCC levels, showing that intact casein, both αs1 and β-casein, decreased as the SCC of milk increased, and that the proteolytic fragments, mainly I-αs1, increased with SCC levels. Significant differences in texture were found among the samples, the cheeses made with high levels of SCC being significantly less compact at each ripening time. The differences in texture were detected by the consumers, who reported defects in cheeses made with high levels of SCC. Indeed, high SCC cheeses were significantly less well accepted.

PCR primers were developed for the specific detection of Clavispora lusitaniae, Debaryomyces hansenii var hansenii, Geotrichum candidum, Kluyveromyces lactis and K. marxianus and Yarrowia lipolytica, yeast species commonly found on the surface of smear cheese. Forty eight representative strains frequently found in smear cheeses or taxonomically related to the target yeasts were used as templates, to validate the designed primers. The specific and selective detection of these yeasts was effective in situ, in Livarot smear, without yeast isolation and culture and was comparable with data obtained with a conventional method. The primers described here have thus potential for PCR studies applied to cheese. It should also be possible to use some of these primers with other substrates.

To determine the effect of feeding propionibacteria on metabolic indicators during lactation, multiparous and primiparous Holstein cows were fed one of three dietary treatments in a 2×3 factorial design from 2 weeks prepartum to 30 weeks post partum: (1) Control (primiparous n=5, multiparous n=8) fed a total mixed ration (TMR); (2) high-dose group (primiparous n=6, multiparous n=5) fed TMR plus 6×1011cfu/head daily (high-dose P169) of propionibacterium strain P169; or (3) low-dose group (primiparous n=8, multiparous n=6) fed TMR plus 6×1010 cfu/head daily (low-dose P169) of P169. Blood samples were collected weekly and analysed for plasma concentrations of glucose, insulin, insulin-like growth factor-I (IGF-I), leptin, nonesterified fatty acids (NEFA) and cholesterol. Between weeks 25 and 30, all groups received bovine somatotropin (bST) every 2 weeks. Low-dose P169 multiparous cows had lower (P<0·05) plasma insulin and glucose concentrations than high-dose P169 multiparous cows, whereas high-dose P169 primiparous cows had lower glucose but greater insulin concentartions than low-dose P169 primiparous cows (P<0·05). Plasma insulin[ratio ]glucose molar ratios were 13–18% lower (P<0·05) in low-dose P169 cows than in control or high-dose P169 cows. Plasma IGF-I, NEFA and leptin levels did not differ among diet groups between weeks 1 and 25. Low-dose P169 multiparous cows had 25% greater plasma cholesterol levels than high-dose P169 and control multiparous cows, but cholesterol levels in primiparous cows did not differ. During bST treatment, high-dose P169 multiparous cows and low-dose P169 primiparous cows had lower IGF-I levels than their respective controls and, regardless of parity, high-dose P169 cows had greater NEFA than control cows. Although supplemental feeding of P169 altered plasma hormones and metabolites, the particular effects were dependent on dose of P169 and parity of cows.

Milk fermented with mixed dahi cultures NCDC167, Lactococcus lactis ssp diacetylactis NCDC60 and two probiotic strains; Lactobacillus acidophilus NCDC14 and Lb. casei NCDC19 were evaluated after fermentation (14 h) and during 8 d storage at 7 °C. The β-galactosidase activity was found to increase after fermentation leading to the hydrolysis of lactose and production of glucose, galactose and oligosaccharides; that subsequently decreased during storage. The viable counts of lactococci and lactobacilli decreased during storage yet remained >106 cfu/ml after storage. The results of present study indicate that all the selected cultures have ability to produce oligosaccharides (prebiotics) due to transgalactosidal and lactose hydrolysis activities of β-galactosidase. The cultures developed an active synbiotic formula by maintaining sufficient probiotic viable counts to exert health benefits to the consumers.

Conjugated linoleic acid (CLA) has a range of biological properties, including effects on lipid metabolism, milk and body composition in animals. This study investigated the effects of dietary CLA on lactating rats and development of the suckling pups. Dams were fed either a control diet or the same diet supplemented with 25 g/kg of a fat supplement containing 540 g CLA/kg (final concentration of 13·5 g CLA/kg diet) from parturition to the 15th day post-partum. The CLA mixture used in this study contained the following isomers (per 100 g): cis-9, trans-11 (24 g); cis-10, trans-12 (35 g); cis-8, trans-10 (15 g); cis-11, trans-13 (17 g) and others (9 g). On d 15 post partum, CLA supplementation reduced milk fat content by 33% and pup growth by 21%. The milk fatty acid profile, with decreased content of short and medium chain acids, suggests CLA inhibition was more pronounced for de novo lipid synthesis. Consistent with these results, activities of fatty acid synthase, glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase were reduced by CLA treatment in the mammary gland and liver. In contrast, the activity of NADP-malate dehydrogenase was unchanged.

The effects of minerals on casein micelle stability of individual cows' milk, throughout a complete lactation, were investigated. Calcium and calcium ions, magnesium, phosphorus, sodium, potassium and citrate contents were analysed, together with the following physical properties of milk; pH, ethanol stability, rennet clotting time and coagulum firmness. There was an inverse non-linear relationship between free calcium ion concentration and ethanol stability (ES; r=0·84). Rennet coagulation time showed a weaker relationship with free calcium ion concentration (r=0·44) but a stronger relationship with pH (r=0·66). In addition, samples containing higher amounts of free calcium ions produced a firmer gel. Citrate in natural samples acts as a stabilizing factor, as it slightly improves milk stability. Potassium, on the other hand, exhibited a negative correlation, but only with rennet clotting time (r=−0·52). Throughout lactation the average values were; free Ca2+ concentration 1·88 mM, pH 6·63, ES 83·2% and clotting time 13·6 min. The equilibrium relationship between pH and free Ca2+ concentration was investigated by adjusting milk pH from 5·9 to 7·1, using acid and alkali. There was a good inverse linear relationship between pH and log (free Ca2+) for individual milk samples, with a gradient of −0·62 and a standard deviation of 0·042.

Udder defence mechanisms are not completely explained by current mastitis research. The anatomical construction of the udder implies that infection of one udder quarter does not influence the immune status of neighbouring quarters. To test this hypothesis, we compared the immune reactions of individual udder quarters in response to microbial attacks. In the course of immune reactions, polymorphonuclear leucocytes (PMN) release oxygen radicals, which can be determined by chemiluminescence (CL). Milk from 140 udder quarters of 36 cows was analysed for somatic cell count (SCC), differential cell count, viability and CL activity. Quarters with an SCC <100000 cells/ml and free of pathogens were defined as uninfected, all other quarters were categorized as infected. Three groups of cows were classified cytologically: group A (healthy, 11 animals, SCC limit <100000 cells/ml); group B (moderate mastitis, 8 cows, SCC [ges ]100000 and <400000 cells/ml in at least one quarter); and group C (severe mastitis, 17 cows, SCC [ges ]400000 cells/ml in at least one quarter). Infected and uninfected quarters in groups B and C were analysed separately. Viability of PMN leucocytes was significantly (P=0·0012) lower in group A (72·6%) than in healthy quarters of group C (84·0%). Lowering the SCC limit of healthy quarters to <50000 cells/ml (group A: all quarters within the udder) revealed striking differences between samples of groups B and C: in addition to varying differential cell counts and viabilities, CL activity of group B<50 (2929 CL units/million PMN) was markedly lower than that of the other groups (5616 in group A<50 and 6445 CL units/million PMN in group C<50). These results allow the conclusion that the infection of one udder quarter influences the cell activity of neighbouring quarters. When the SCC threshold for healthy quarters was reduced to 50000 cells/ml, greater differences in cell activities were detected between healthy udders and healthy quarters of infected udders.

The occurrence of styrene in food may be an important aroma defect (celluloid odour), even at very low concentrations (Miltz et al. 1980) causing consumer rejection and is therefore a problem for the food industry. We examined the biosynthetic pathway leading to styrene formation by Penicillium camemberti using labelled compounds. As styrene is strongly hydrophobic and volatile, we first had to develop a continuous extraction process. Using resins XAD2 it was reasonable to suspect phenylalanine (Phe) as the precursor. The addition of Phe marked with 13C on the ring provokes the accumulation of labelled styrene. The enzyme activities involved were also tentatively measured. Styrene appears to be synthesized from phenylalanine by phenylalanine ammonia-lyase activity followed by a decarboxylation catalysed by a cinnamic acid decarboxylase.

Although there has been little study of the origin of intramammary infection (IMI) in goats, a common view is that most bacterial infection in goats occurs during milking. In the present study, the dynamics of occurrence of udder infection during and between lactations in three Anglo-Nubian goat farms in Israel was monitored. Coagulase-negative staphylococci were the predominant bacteria in the IMIs. We found that about 15% of the yearling does were already infected with bacteria when they joined the flock, whereas about 8% of the goats that dried-off returned with new IMIs. Moreover, virtually none of the goats acquired infection during lactation. Thus, our study showed that the aetiology of IMI in goats is very similar to that in dairy cows. A preventive treatment during the dry period should, therefore, be considered as an effective means of reducing the current rate of bacterial infections in goats.

In the study presented in this article, the influence of added α-lactalbumin and β-lactoglobulin on the changes that occur in casein micelles at 250 and 300 MPa were investigated by in-situ measurement of light transmission. Light transmission of a serum protein-free casein micelle suspension initially increased with increasing treatment time, indicating disruption of micelles, but prolonged holding of micelles at high pressure partially reversed HP-induced increases in light transmission, suggesting reformation of micellar particles of colloidal dimensions. The presence of α-la and/or β-lg did not influence the rate and extent of micellar disruption and the rate and extent of reformation of casein particles. These data indicate that reformation of casein particles during prolonged HP treatment occurs as a result of a solvent-mediated association of the micellar fragments. During the final stages of reformation, κ-casein, with or without denatured whey proteins attached, associates on the surface of the reformed particle to provide steric stabilisation.

Milk somatic cell count (SCC) is the gold standard in diagnosis of subclinical mastitis, and is also an important parameter in quality programmes of dairy cooperatives. As routine SCC analysis is usually restricted to central laboratories, much effort has been invested in the search for alternative biomarkers of mastitis and milk quality, including the presence in the milk of the acute phase proteins (APP), haptoglobin (Hp) and serum amyloid A (SAA). The aim of this study was to investigate relationships between Hp, SAA and SCC in quarter, cow composite, and bulk tank milk samples. Cows (n=165), without any clinical signs of disease or abnormalities in the milk or udder, from three different dairy farms, were used. Cow composite milk samples from all cows delivering milk at the sampling occasion were taken once in each herd. In one of the farms, representative quarter milk samples (n=103) from 26 cows were also collected. In addition, bulk tank milk samples from 96 dairy farms were included in the study. Samples were analysed for Hp, SAA and SCC, and relationships between the parameters were evaluated at quarter, cow and tank milk levels using Chi-square analysis. Milk samples were categorized according to their SCC, and the presence, or no presence, of SAA and Hp, based on the detection limits of the screening methods (0·3 mg/l and 1·0 mg/l for SAA and Hp, respectively). Hp and SAA were found in milk at quarter, cow composite and bulk tank levels. A large proportion (53%) of the animals had detectable milk concentrations of APP, and SAA was detected more frequently, and at higher concentrations than Hp, regardless of sample type. SAA was detected in as many as 82% of the bulk tank milk samples. Significant relationships were found between Hp, SAA and SCC at quarter and cow composite milk levels, but only between SAA and SCC at bulk tank milk level. Detectable levels of APP were more common at high SCC.

We investigated effects of subclinical intramammary infection (IMI) on milk somatic cell count (SCC) and milk composition in udder halves of dairy goats. A total of 35 mixed-age Alpine does (70 udder halves; approximately 55 kg body weight) were rotationally grazed on a mixture of vegetative forages (wheat/berseem clover, sudan grass and cowpeas). Milk samples for bacterial analysis and SCC were collected monthly from both halves from April to September, 2001. Across stages of lactation, 19–31% of udder halves became infected. The prevalence of IMI exhibited quadratic patterns through multi-peaked responses within each stage of lactation. Higher rates of IMI were observed during the early stage of lactation (19% in May) and in the late stage of lactation (31% in September). Coagulase negative Staphylococcus (CNS, 43·7%), Staph. aureus (35·4%), and Pseudomonas aeruginosa (12·4%) were the most prevalent pathogens. Within single-strain IMI, log SCC (6·24) was lower (P<0·01) for CNS than those derived from IMI by Staph. aureus (6·49), Ps. aeruginosa (6·53) or Serratia spp. (6·90). Infected udder halves had a higher average SCC (4761 v. 2259×103 cells/ml; P<0·01) than uninfected halves, but uninfected halves often had similar levels of SCC to infected halves. Daily average milk production was not significantly different between infected and non-infected goats and the relationship between IMI and SCC was not always correlated. Effective mastitis screening requires bacteriological culture since SCC was not highly correlated.

A study was conducted to determine whether intramammary antibiotic treatment of heifer mammary glands following the first milking after calving was effective for reducing the percentage of mammary quarters infected during early lactation. Jersey and Holstein heifers from two research herds were assigned to one of three treatment groups: (1) no intramammary infusion following the first milking after parturition, (2) intramammary infusion of all quarters with pirlimycin hydrochloride following the first milking after parturition and (3) intramammary infusion of all quarters with novobiocin sodium plus penicillin G procaine following the first milking after parturition. Almost 93% of Jersey heifers (40/43) and 73·1% of quarters (125/171) were infected at the first milking. Almost 77% of quarters (33/43) were cured following treatment with pirlimycin, 61·8% (21/34) were cured following treatment with penicillin-novobiocin and 39·6% (19/48) of infections were eliminated spontaneously in the untreated control group. Significantly fewer infections were observed in pirlimycin or penicillin-novobiocin treated mammary glands of Jersey heifers during early lactation than in untreated control mammary glands. Almost 89% of Holstein heifers (32/36) and 52·8% of quarters (76/144) were infected at the first milking. About 57% (12/21) of quarters were cured following treatment with pirlimycin, 41·4% (12/29) were cured following treatment with penicillin-novobiocin and 23·1% (6/26) of infections were eliminated spontaneously in the untreated negative control group. Significantly fewer infections were observed in pirlimycin treated mammary glands of Holstein heifers during early lactation than in untreated control mammary glands. However, no significant differences were observed following penicillin-novobiocin treatment of Holstein heifers after the first milking of lactation compared with untreated control quarters. Coagulase-negative staphylococci, Streptococcus uberis and Streptococcus dysgalactiae subsp dysgalactiae were isolated most frequently in heifers from both herds.

Phenomena generating FFAs, important flavour precursors, are significant in cheese ripening. In Camembert-like cheeses, it was intended to establish the relationships between the dynamics of FFA concentrations changes and the succession of ripening microflora during ripening. Experimental Camembert-type cheeses were prepared in duplicate from pasteurised milk inoculated with Kluyveromyces lactis, Geotrichum candidum, Penicillium camemberti, and Brevibacterium aurantiacum under aseptic conditions. For each cheese and each cheesy medium, concentrations of FFAs with odd-numbered carbons, except for 9[ratio ]0 and 13[ratio ]0, did not change over time. For long-chain FFAs, concentrations varied with the given cheese part (rind or core). K. lactis produced only short or medium-chain FFAs during its growth and had a minor influence on caproic, caprylic, capric, and lauric acids in comparison with G. candidum, the most lipolytic of the strains used here. It generated all short or medium-chain FFAs (4[ratio ]0–12[ratio ]0) during its exponential and slowdown growth periods and only long-chain ones (14[ratio ]0–18[ratio ]0) during its stationary phase. Pen. camemberti produced more long-chain FFAs (14[ratio ]0–18[ratio ]0) during its sporulation. Brev. aurantiacum did not generate any FFAs. The evidence of links between specific FFAs and the growth of a given microorganism is shown.

Several milk components related to immune defences (lysozyme, lactoferrin and γ-globulins, γ-G) and to inflammation (somatic cell counts, SCC; N-acetyl-β-glucosaminidase, NAGase; albumin) were considered. Forty-one quarters and 685 samples of 24 cows were included in the study; among them 534 samples were defined as negative (78·0%), 93 as diseased (13·5%) and 58 (8·5%) as subclinical. The pattern of each milk component in quarters always negative during the follow-up period was evaluated by a mixed model. Statistical analysis showed that days in milk (DIM), age (primiparous, pluriparous), herd and the interaction between herd and days in milk significantly influenced all the markers, with very few exceptions. A subset of samples including the negative quarters before the first outcome of an infection or a subclinical mastitis and the samples from quarters always negative was also selected. The analysis showed that herd, DIM and health status had a significant influence on most markers. Overall, primiparous cows were confirmed to have higher levels of most of the markers than pluriparous cows. The presence of a herd effect on non specific immune defences in fully negative quarters implies that when the mechanisms behind their release are fully elucidated, it might be possible to modulate them. Udder tissues were confirmed as an important source of some immune components, as supported by the inconsistency between SCC mean values and NAGase, lysozyme and lactoferrin values. Overall, quarters with high levels of NAGase, lysozyme and γ-G, exposed to bacteria, did not develop subclinical mastitis. Hence, invading pathogens could induce the development of subclinical IMI when these components and γG are in low concentration.

Recombinant bovine lactoferrin N-terminal polypeptide (rbLF-N) Escherichia coli expression system was constructed and the rbLF-N antimicrobial activity was displayed by enzymatic proteolysis in this study. A 162 bp 5′-terminal fragment of bovine lactoferrin (bLF) gene from bovine liver gDNA was amplified by PCR. The DNA fragment containing exon-2 of the bLF gene was cloned into the expression vector pGEX-4T1 and the glutathione-S-transferase–rbLF-N (GST-rbLF-N) fusion protein was obtained by over-expression in Esch. coli BL21(DE3). After thrombin/pepsin digestion, the rbLF-N was released from the fusion protein. The recombinant peptide was separated and identified by SDS-PAGE, HPLC and LC-MS/MS analysis. A very strong anti-food-born microbial pathogen activity of the rbLF-N peptides was displayed through bio- and kinetic-assays in vitro. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the rbLF-N peptide for bacterial pathogens Staphylococcus aureus, Streptococcus mutans, Esch. coli and Klebsiella pneumoniae were 11·7, 11·7, 11·7, 23·4 μg and 23·4, 11·7, 11·7, 46·4 μg, respectively. This study created a new route for exploring lactoferrin peptide application in food science.

The objective was to characterize the antibody isotype responses to vaccination with a commercial Staphylococcus aureus bacterin in dairy heifers. Twenty-five Holstein-Friesian dairy heifers were assigned at random to one of two groups, vaccinates (n=14) or controls (n=11). Vaccinates received two 5-ml doses of Lysigin 28 d apart in late gestation. Both groups were challenged with a heterologous serotype 5 strain of Staph. aureus by aseptic intramammary infusion on days 6, 7 and 8 of lactation. Samples for serum antibody isotype (IgG, IgG1, IgG2 and IgM) determinations were taken before each vaccination, immediately prior to challenge (6 d post-calving) and at the end of the study (28 d post-calving). Samples for milk antibody isotype determinations were taken immediately prior to challenge and at the end of the study. Antibody isotype sample-to-positive ratios (S[ratio ]P ratio=(mean test sample OD−mean within-plate negative control OD)/(mean within-plate positive control OD−mean within-plate negative control OD)) were determined in milk and serum using a series of ELISAs coated with different strains of Staph. aureus belonging to capsular polysaccharide (CP) serotype 5 or 8 or surface polysaccharide (SP) serotype 336. Vaccinates had higher mean serum IgG1 and IgG2 S[ratio ]P ratios than controls against all three strains of Staph. aureus (P[les ]0·023). Vaccinates had higher mean milk IgG S[ratio ]P ratios than controls against CP8 and SP336 strains of Staph. aureus (P[les ]0·030). Hence, a humoural immune response to the vaccine was detected in serum and milk, but responses varied according to strain and antibody isotype tested.

The bovine's long generation interval results in a delay of several years when evaluating mammary specific transgenes in genetically engineered animals. This experiment was conducted to evaluate the feasibility of reducing that waiting period. Lactation was induced in prepubertal bull and heifer calves as a means of predicting transgene behaviour during subsequent post-parturient lactations in the heifers themselves, and in daughters sired by the bulls. The animals carry a lactation-specific transgene encoding lysostaphin, an antimicrobial protein that kills Staphlococcus aureus, a mastitis-causing pathogen. Oestrogen, progesterone and dexamethasone were administered as previously described (Ball et al. 2000) to nine heifers (five transgenics) ranging in weight from 80 to 145 kg. Eight bull calves (seven transgenics) weighing 81–178 kg received additional oestrogen and progesterone injection prior to dexamethasone treatment. All nine heifers responded to the milk induction scheme yielding between 19 ml and 4·5 l over 5 d. Milk volume from the four responding males (30 μl to 2·5 ml) was significantly less than that harvested from females (P=0·025). Only bull calves >117 kg had a positive response. Lysostaphin was detected in all transgenic prepubertal heifers and in two transgenic prepubertal bull calves induced. A positive relationship was observed between lysostaphin's stapholytic activity in the two types of lactations (r2=0·907, P<0·001) thus providing a useful means of predicting subsequent lysostaphin production in post-partum milk.