C. Boscos

Prevalence of subclinical mastitis and influence of breed, parity, stage of lactation and mammary bacteriological status on Coulter Counter Counts and California Mastitis Test in the milk of Saanen and autochthonous Greek goats

Coulter Counter Counting (CCC), California Mastitis Test (CMT) and bacteriological examination were performed on milk samples from healthy primiparous Saanen, multiparous Saanen (SA) and multiparous autochthonous Greek (AG) goats, in six flocks in northern Greece. Mean CCC in bacteriologically negative milk samples were higher (P < 0.05) in AG goats than in multiparous SA, and in multiparous than in primiparous SA, throughout the lactation period. No differences were noticed with regard to the mean CCC in bacteriologically negative milk samples of primiparous SA, multiparous SA and multiparous AG goats from Day 50 up to Day 110 after kidding. Mean CCC in the milk of multiparous goats increased (P < 0.001) from Day 125 in AG goats and from Day 170 in SA goats up to Day 185 after kidding. Parity, breed and stage of lactation differences were not noticed with regard to the mean CMT scores. Prevalence of bacteria in the milk samples ranged between the flocks from 19.0% to 35.7% of the glands. The proportion of bacteriologically positive samples was higher (P < 0.05) in multiparous than in primiparous SA goats whereas no differences were noticed between multiparous SA and AG goats. Prevalence of coagulase-negative staphylococci, Staphylococcus aureus, Streptococcus spp. and other bacteria varied between the flocks. No breed or parity differences were observed with regard to the type of bacteria isolated. The presence of bacteria in caprine milk elevated both CCC and CMT scores in primiparous SA, multiparous SA and multiparous AG goats. This elevation was associated with the type of bacteria; S. aureus elevated CCC and CMT scores in milk more than coagulase-negative staphylococci did. The results of the present investigation lead to the conclusion that Coulter Counter and CMT could be appropriate only for the prediction of the presence of a major pathogen such as S. aureus in goats milk, even if we try to restrict their application to only one breed, to one parity or to a specific stage of lactation.

Effect of antioxidant supplementation in semen extenders on semen quality and reactive oxygen species of chilled canine spermatozoa.

The objective of this study was to evaluate quality of chilled dog semen processed with extenders containing various antioxidants. Single ejaculates from five dogs were always pooled and evaluated for concentration, sperm motility, progressive motility (RSF-movement), viability, acrosomal integrity and by the hypo-osmotic swelling (HOS)-test. Also, superoxide (O(2)(-)) production, hydroxyl radicals (OH) and total reactive oxygen species (tROS) were determined. Pooled semen was divided in seven aliquots (for control and test conditions), which were diluted to a final concentration of 67x10(6)spermatozoa/ml with TRIS-glucose-egg yolk extender with or without the following supplements: control (without antioxidants), vitamin C (0.5mM), N-acetyl-l-cysteine (NAC; 0.5mM), taurine (0.2mM), catalase (100u/ml), vitamin E (0.1mM) and 5-(4-dimethylamino-phenyl)-2-phenyl-penta-2,4-dienoic acid (B16; 0.1mM). The semen aliquots were chilled and preserved at 4 degrees C. Portions of chilled semen were removed at 24 and 72h, and semen quality was evaluated after rewarming. At 24h the mean (+/-S.E.M.) sperm motility was higher (p<0.001) when vitamin E, taurine and B16 were added in the extender, whereas more spermatozoa with RSF-movement were observed (p<0.001) in the vitamin E, catalase, B16 and taurine groups. Sperm viability was higher (p=0.040) in B16 and vitamin E groups and the percentage of swollen spermatozoa was higher (p=0.002) only in the B16 group. Acrosomal integrity and OH were not significantly influenced by any of the antioxidants tested. Superoxide production was significantly lower when vitamin C, B16 and vitamin E were added in semen extenders compared with the control (p=0.017). All antioxidant groups, except vitamin C and NAC, contained less tROS compared to the control group, but only the B16 group value differed significantly (p=0.05). At 72h sperm motility was higher (p<0.001) when vitamin E, catalase, B16, taurine and NAC were added in the extender. More spermatozoa with RSF-movement were observed (p<0.001) in the vitamin E, catalase, B16, taurine and NAC treatment groups. Sperm viability was higher (p=0.001) when vitamin E, B16, taurine and vitamin C were added in semen extenders. HOS-test percentages were higher (p=0.016) in the B16, vitamin E, catalase and NAC groups. Acrosomal integrity was not influenced in any case. Production of O(2)(-) was significantly higher using catalase compared to all the other groups (p=0.006), while OH was not significantly influenced by any of the antioxidants tested. The addition of vitamin E, catalase and B16 in semen extenders resulted in significantly lower tROS values compared with the controls (p<0.0005). The results suggest that vitamin E and B16 had the most pronounced effect in preserving semen quality of chilled dog spermatozoa.

Effect of N‐acetyl‐L‐cysteine Supplementation in Semen Extenders on Semen Quality and Reactive Oxygen Species of Chilled Canine Spermatozoa

The objective of this study was to evaluate the quality of chilled dog semen processed with extenders containing various concentrations of N-acetyl-L-cysteine (NAC). Ejaculates from five dogs were collected, pooled and evaluated for concentration, motility, rapid steady forward movement (RSF-movement), viability, acrosomal integrity and by the hypo-osmotic swelling test (HOST). In addition, superoxide anion (O(2)(-*)) production, hydroxyl radicals (OH(*)) and total reactive oxygen species (tROS) were determined. The pool was divided into five aliquots, which were diluted to a final concentration of 66.66 x 10(6) spermatozoa/ml with Tris-glucose-egg yolk extender containing one of the following concentrations of NAC (0, 0.5, 1, 2.5 or 5 mm). The semen aliquots were chilled and preserved at 4 degrees C. Semen quality was evaluated after rewarming at 72 h. Sperm motility was significantly higher with the 0.5 mm concentration compared with the control group (p = 0.001). Rapid steady forward movement was higher with the 0.5 and 1 mm concentrations compared with the control and 5 mm group (p < 0.001). Viability and HOST percentages were not significantly altered. Compared with the control, the 5 mm concentration showed significantly reduced percentages of spermatozoa with normal acrosomes (p = 0.049). None of the ROS values at 72 h were significantly affected by the presence of NAC in semen extenders, although all NAC concentrations showed lower O(2)(-*) and OH(*) values compared with the control. Only the concentrations of 1 and 5 mm inhibited the significant increase of tROS values after 72 h, compared with the fresh semen value. In conclusion, NAC supplementation of semen extenders is beneficial to semen motility of canine spermatozoa during chilling with the 0.5 mm concentration being the most effective, although no significant ROS inhibition was observed at 72 h.

Quality and reactive oxygen species of extended canine semen after vitamin C supplementation

The objective of this study was to evaluate the quality of extended dog semen processed with diluents containing various concentrations of vitamin C. Ejaculates from five dogs were collected, pooled and evaluated for concentration, sperm motility, rapid steady forward movement (RSF-movement), viability, acrosomal integrity and by the hypo-osmotic swelling test. Also, superoxide (O(2)(-)*) production, hydroxyl radicals (OH*) and total reactive oxygen species (tROS) were determined. The pool was divided in five aliquots, which were diluted to a final concentration of 66 x 10(6) spermatozoa/ml with a Tris-glucose-egg yolk extender containing one of the following concentrations of vitamin C (0, 0.1, 0.5, 1 or 2.5 mM). The semen aliquots were chilled and preserved at 4 degrees C. Portions of chilled semen were removed at 24 and 72 h, and semen quality was evaluated after rewarming. This process was repeated 10 times in pooled semen of the same origin and data were analysed by one-way analysis of variance. At both times, none of the semen quality parameters were positively influenced (p>0.05) by vitamin C supplementation. At 24 h, none of the reactive oxygen species (O(2)(-)*, OH*, tROS) were significantly altered. At 72 h, significant reductions of O(2)(-)* production were observed by the concentrations of 0.1, 0.5 and 2.5 mM compared with the 0 mM concentration (p=0.049). Also, at 72 h, the 2.5 mM concentration showed significantly lower OH* values in comparison with the control group (p=0.048). In conclusion, addition of vitamin C to semen extenders does not benefit the quality of canine extended spermatozoa.

Use of enzyme-immunoassay for oestradiol-17β and progesterone quantification in canine serum

The objective of this investigation was to develop and evaluate competitive inhibition-enzyme-immunoassays for canine serum oestradiol-17beta (E(2)) and progesterone (P(4)) quantification. Sera from 56 healthy bitches at various stages of oestrus cycle and pregnancy were tested. For E(2) measurement, each sample (0.4 ml) was extracted with diethyl ether and after solvent evaporation the resultant hormone was reconstituted to one-fifth of the original sample volume in aqueous buffer. Each reconstitute (30 microl) was assayed for E(2) to estimate respective serum concentration. For P(4), each sample (10 microl) was directly assayed without extraction. The classic cyclic hormonal pattern during the oestrus cycle of the bitch was observed. The brief, sharp dominance of E(2) during the follicular phase was followed by the long-lasting dominance of P(4) during the luteal phase (late oestrus, dioestrus or pregnancy). During the anoestrus phase both hormones were found at basal levels, with the exception of E(2) during late anoestrus which appeared to be rising. Both assays had acceptable specificity (cross-reactions < or =10%), precision (coefficient of variation (C.V.) < 7%) and accuracy (E(2) recovery: 97%; P(4) recovery: 104.7%). The sensitivity of E(2) and P(4) assay was 4 pgml(-1) and 0.28 ngml(-1), respectively.

Plasma progesterone concentration in relation to ovulation rate and embryo yield in Chios ewes superovulated with PMSG

Abstract The main purpose of this work was to investigate the relationship between plasma progesterone concentration and the number of ovulations and/or the number of embryos collected from Chios ewes induced to superovulate with various doses of PMSG. The oestrous cycles of the animals were synchronized by means of MAP intravaginal sponges for 14 days and PMSG was injected i.m. (1500 IU, Group 1; 1000 IU, Group 2; 750 IU, Group 3; 500 IU, Group 4; 0 IU, Group 5) at the time of sponge withdrawal. Seven days after sponge removal and 5 days after mating, mid-ventral laparotomy was performed and the uterine horns and/or oviducts were flushed. The number and diameter of corpora lutea (CL), the number of large (diameter > 0.5 cm) anovulated follicles and the total ovarian response (TOR = CL + large anovulated follicles) were recorded. The embryos were examined under a dissecting microscope and evaluated according to morphological criteria. Blood samples were collected once daily for 4 days starting on the day of sponge withdrawal. One more sample was taken on the day of embryo collection. Progesterone concentration was determined using a conventional ELISA. A significant positive correlation was found between plasma progesterone concentration and number of corpora lutea (r = 0.61, P

Effects of long-term recombinant bovine somatotropin (bST) administration on milk yield, milk composition and mammary gland health of dairy ewes

Abstract The present investigation was carried out to test the effects of recombinant bovine somatotropin (bST) on galactopoiesis and mammary gland health of the ewe. Twenty-two polytocous lactating Chios ewes were assigned into two groups (11 in control group and 11 in treatment group). Treated ewes were injected every second week with 160 mg bST in a prolonged release formulation, from the fifth day after delivery until the end of lactation. All animals were fed the same ration. Supplementation with bST significantly ( P P P P >0.05). Mean Somatic Cell Count (SCC) from animals without subclinical mastitis were progressively increased ( P P P >0.05). Coagulase-negative staphylococci were the predominant organisms isolated from milk samples in both groups. Prevalence of subclinical mastitis was unaffected after bST administration ( P >0.05) although treated animals tended to show higher rates.

Teat lesions predispose to invasion of the ovine mammary gland by Mannheimia haemolytica.

Teat lesions, produced in ewes by an experimental chapping procedure, were found to facilitate experimental infection with Mannheimia haemolytica, as assessed by observations on infection of the teat skin, teat duct and mammary gland, and on the production of mastitis. The origin of the M. haemolytica strain used (ovine tonsillar or mammary infection) did not appear to influence the results. In a second experiment, in which ewes continued to suckle their lambs but were not deliberately infected, chapping was shown to favour infection by Staphylococcus epidermidis, Staphylococcus aureus and M. haemolytica.

The effect of bovine somatotropin (bST) administration on reproduction, progesterone concentration during lactation and LH secretion during estrus, in dairy ewes

Twenty-two polytocous lactating Chios ewes were used to test the effects of bovine somatotropin (bST) on reproduction, progesterone concentration and LH secretion during estrus. Half of the ewes were injected every second week with 160 mg bST in a prolonged release vehicle, from the fifth day post partum until the end of lactation, while the remaining ones were used as controls. All animals were fed the same amount of ration. Supplementation with bST resulted in an increase of milk production (P<0.05) and an insignificant trend for delayed resumption of normal estrous cycles. Although there were no differences between groups, there was also a tendency for the bST group to display lower progesterone concentrations during the first three fortnights after the onset of normal estrous cycles and higher ones during the last three fortnights of the experiment, compared with the control group. Duration of the first normal luteal phase after delivery of the bST group was found to be shorter compared with the control group (P<0.05). After estrous synchronization the bST group showed a shorter estrus compared with the control group (P<0.05). Average and baseline LH concentrations during synchronized estrous in the bST group was lower (P<0.001) compared with the control group. Additionally, the conception rate did not differ between the two groups. This study supports the concept that the beneficial effects of bST treatment on milk production outweigh the potential deleterious effects on reproduction.

The induction of lymphoid follicle-like structures in the ovine teat duct following experimental infection with Mannheimia haemolytica.

The objectives of this study were to investigate the early stages of experimental infection of the ovine mammary gland with Mannheimia haemolytica and to identify the lymphocyte subsets accumulating at the teat duct. M. haemolytica was inoculated into one teat of each of 25 ewes and clinical, bacteriological, cytological, haematological, physicochemical, gross pathological, histopathological and immunohistochemical examinations were carried out. Clinical signs of inflammation were evident by 8 h but had subsided 2 days after challenge. Polymorphonuclear neutrophils (PMN) predominated in milk films up to 1 day following challenge, but the proportion of lymphocytes and macrophages progressively increased thereafter. Total blood leucocyte counts decreased immediately after challenge and then rose until 1 day after challenge with immature PMNs comprising >3% of the total. The pH of the mammary secretions from the challenged side was increased (>7.0). Focal lymphoid accumulations were observed in the lamina propria at the junction of the teat duct and cistern, including CD79(+), CD3(+) and gammadelta T cells, CD68(+) and MHC-II(+) cells with a particular increase in the numbers of CD8(+) T cells from days 3 to 5 after challenge. The findings suggest that these organised lymphoid structures are inducible and contribute to the defence of the infected teat when the PMN-macrophage response is overwhelmed.