Impact of subclinical and clinical mastitis on sensitivity to pain of dairy cows

A total of 90 cows from three commercial farms were used to evaluate the relationship between subclinical mastitis and clinical mastitis and thermal nociceptive threshold. Milk strips from all udder quarters were tested for clinical mastitis with visual inspection of milk and udder alterations and for subclinical mastitis using California Mastitis Test. Milk yield was recorded, milk was sampled and further analyzed for somatic cells count (SCC). Cows were considered healthy when SCC<200 000 cells/ml and no visual alterations in milk and/or udder, with mild subclinical mastitis when SCC>200 000 cells/ml and no visual alterations in milk and/or udder, with moderate subclinical mastitis when SCC>500 000 cells/ml and no visual alterations in milk and/or udder and with clinical mastitis when visual alterations in milk and/or udder were detected. Nociceptive threshold was evaluated with the thermal threshold meter apparatus applied to the rear legs. Thermal threshold (TT) decreased when we compared healthy cows with cows presenting clinical mastitis and tended to decrease when we compare healthy cows with those with moderate subclinical mastitis. TT was lower at the ipsilateral rear leg compared with the contralateral leg to the infected mammary gland. TT linearly decreases as _log10SCC increased and it showed sharp decrease as _log10SCC exceed the value of 6.4. Increase in one unit of _log10SCC increased the odds of low thermal threshold (lower than 55.8°C). Subclinical mastitis might be a welfare issue as it tended to decrease nociceptive thermal threshold.     

Evaluation of quality changes in udder quarter milk from cows with low-to-moderate somatic cell counts

Much emphasis has been put on evaluating alterations in milk composition caused by clinical and subclinical mastitis. However, little is known about changes in milk composition during subclinical mastitis in individual udder quarters with a low-to-moderate increase in milk somatic cell count (SCC). This information is needed to decide whether milk from individual udder quarters with a moderate-to-high increase in milk SCC should be separated or not. The aim of this study was to determine how milk composition in separate udder quarters is affected when cow composite milk has low or moderately increased SCC levels. Udder quarter and cow composite milk samples were collected from 17 cows on one occasion. Milk yield was registered and samples were analyzed for SCC, fat, total protein, whey proteins, lactose, citric acid, non-protein nitrogen (NPN), lactoferrin, protein profile, free fatty acids (FFAs), lactate dehydrogenase (LDH), proteolysis, sodium and potassium. Bacteriological samples were collected twice from all four quarters of all cows. The cows were divided into three groups depending on their SCC at udder quarter level. The first group comprised healthy cows with four udder quarters with low SCC, <50 000 cells/ml; composition was equal when opposite rear and front quarters were compared. In the second and the third groups, cows had one udder quarter with 101 000 cells/ml < SCC < 600 000 cells/ml and SCC > 700 000 cells/ml, respectively. The remaining udder quarters of these cows had low SCC (<100 000 cells/ml). Despite the relatively low average cow composite SCC = 100 000 cells/ml of Group 2, milk from affected udder quarters exhibited lower casein number, content of lactose and β-casein (β-CN), while the content of whey protein, sodium, LDH and α-lactoalbumin (α-la) were higher compared to healthy opposite quarters. In addition to these changes, milk from affected udder quarters in Group 3 also exhibited lower values of potassium and αs1-casein (αs1-CN) and higher values of lactoferrin when compared to milk from opposite healthy quarters. This indicates that even when the SCC in cow composite milk is low, there might exist individual quarters for which milk composition is changed and milk quality impaired.     

Effects of bovine mammary gland biopsy and increased milking frequency on post-procedure udder health,histology, and milk yield

Sixteen cows in early lactation were randomly distributed into two groups in order to evaluate the effects of mammary biopsies and increased milking frequency on tissue characteristics, post-biopsy udder health and histology. One group was milked twice a day (2×) starting on the 2nd day after calving, until 28 days in milk (DIM). The other group was milked four times a day (4×) from two to 21 DIM, and twice a day (2×) from 22 to 28 DIM. On days 2, 7, 14, 21, and 28 postpartum, one fragment of secretory tissue was collected from one mammary quarter at a time. Collections were alternated between the four mammary quarters per collection day. A total of 80 mammary tissue samples were collected. Qualitative and quantitative analyses of the tissues were conducted by histologic examination. Animal health was assessed by observation of feed intake behavior immediately after biopsy, and weight and body condition score before and one week after biopsy. Udder health was assessed daily from calving to 60 DIM with California Mastitis Test (CMT) and by noting alterations in the milk such as blood, milk clots, blood clots, clinical signs of mastitis. Milk composition and somatic cell count (SCC) were analyzed before and after the biopsies. Milk production was evaluated before biopsy, on the day of biopsy, and after the biopsy. An average of 10 fields at 40× magnification was obtained from each sample. There were no evident changes in mammary morphology as a result of milking two or four times/day at any of the evaluated time points. Biopsy wounds healed rapidly without infection. Intramammary bleeding and CMT alterations were observed in 96% and 75% of the biopsied mammary quarters, respectively. Clinical mastitis was diagnosed in 12% of the biopsied quarters. Different milking frequencies had no effect on the frequency and duration of post-biopsy alterations. Milk production decreased after biopsies done on days 2 for 2× and 4× groups, but it returned to pre-biopsy values within 1 day. Milk composition and SCC were affected transiently. Increased milking frequency did not influence udder health. Post-biopsy recovery was rapid and the procedure proved effective without damaging the cows’ health.     

Quantitative milk proteomics – Host responses to lipopolysaccharide‐mediated inflammation of bovine mammary gland

Intramammary infusion of lipopolysaccharide (LPS) in cows induces udder inflammation that partly simulates mastitis caused by infection with Gram‐negative bacteria. We have used this animal model to characterize the quantitiative response in the milk proteome during the time course before and immediately after the LPS challenge. Milk samples from three healthy cows collected 3 h before the LPS challenge were compared with milk samples collected 4 and 7 h after the LPS challenge, making it possible to describe the inflammatory response of individual cows. Quantitative protein profiles were obtained for 80 milk proteins, of which 49 profiles changed significantly for the three cows during LPS challenge. New information obtained in this study includes the quantified increase of apolipoproteins and other anti‐inflammatory proteins in milk, which are important for the cow's ability to balance the immune response, and the upregulation of both complement C3 and C4 indicates that more than one complement pathway could be activated during LPS‐induced mastitis. In the future, this analytical approach may provide valuable information about the differences in the ability of individual cows to resist and recover from mastitis.     

Bacteriostatic activity in cow's milk from udders infected with Streptococcus agalactiae and Staphylococcus aureus.

Milk samples from infected udders contained more lactoperoxidase and more thiocyanate than before infection. Irritation of 10 quarters of 5 cows caused the increase in the bacteriostatic activity of milk. Bacteriostatic activity of milk from the udders infected with staphylococci dropped after several weeks of chronic mastitis.     

Temporal changes in metabolites of prostanoids in milk of heifers after intramammary infusion of Escherichia coli organisms

A study was conducted to characterize the changes in the concentrations of three metabolites of prostanoids in the milk of a) heifers (n = 14; control) inoculated with Escherichia coli (E. coli) organisms into the udder and b) in heifers (n = 10; treatment) vaccinated with E. coli bacterin and treated similar to control heifers. Milk samples were obtained from the challenged quarter and analyzed for the concentrations of stable metabolites of thromboxane A2 (TXB2), prostacyclin (PCM) and prostaglandin E2 (PGEM) using radioimmunoassays. In control heifers milk TXB2 concentrations were significantly higher (P = 0.03) compared to treated heifers. Milk PCM concentrations increased significantly (P = 0.02) in control and treated heifers after the respective treatments, however, differences between the two groups were not significant. Milk PGEM concentrations also increased significantly (P = 0.02) in control and treated heifers after the respective treatments, and there were no differences between the two groups. Results of the present study suggest that, the prostanoids have a role in the pathophysiologic process of coliform mastitis.     

The Electrical Conductivity of Bovine Milk in Mastitis Diagnosis

The electrical conductivity (EC) of milk is mainly a function of the electrolyte concentration in the milk and therefore raised in mastitis. The present investigation was aimed at elaborating, if possible, a diagnostic model for screening purposes based on EC determinations and consistent with the diagnostic procedures and interpretations commonly used in laboratory milk diagnosis in the Nordic countries (Klastrup 1975). According to this diagnosis (here called reference diagnosis) cell numbers above 300,000/ml (cell count or the corresponding CMT-score) in foremilk quarter samples during the main part of the lactation period and significantly above the lowest value on within-udder comparison during late lactation are considered indicative of mastitis and bacteriological examinations are made when called for.     

The effects of colchicine on milk secretion, mammary metabolism and blood flow in the goat

Mammary function in the conscious goat was studied during colchicine-induced depression of milk secretion in one mammary gland. Milk yield of the treated gland was reduced to approximately a quarter of previous, while there were significant increases in afternoon milk yield from the untreated glands on the 2nd and 3rd days after treatment in goats in late lactation. Milk composition in the untreated glands was not significantly affected. In the treated gland, milk [Na+], [Cl-], [citrate] and [protein] increased while [K+] and [lactose] decreased, although the time course of these changes differed; milk [fat] was unaffected. Mammary extractions ((A-V)/A) of glucose, acetate and most amino acids were significantly decreased during the period of maximal inhibition of secretion. There were no significant changes in arterial plasma concentrations of glucose, acetate or any essential amino acids. In another series of experiments, mammary blood flow increased and then returned to normal after colchicine treatment even though milk yield and mammary glucose uptake decreased markedly; oxygen uptake was not significantly affected. The results are discussed in relation to the actions of colchicine on the mammary secretory cell, to the normal control of mammary blood flow and to the mechanism of compensation by the untreated gland.     

Temporal changes in metabolites of prostanoids in milk of heifers after intramammary infusion of organisms

A study was conducted to characterize the changes in the concentrations of three metabolites of prostanoids in the milk of a) heifers (n=14; control) inoculated with ( ) organisms into the udder and b) in heifers (n=10; treatment) vaccinated heifers. Milk samples were obtained from the challenged quarter and analyzed for the concentrations of stable metabolites of thromboxane A₂ (TXB₂), concentrations were significantly higher (P=0.03)bar compared to treated heifers. Milk PCM concentrations increased significantly (P=0.02) in control and treated heifers after the respective treatments, however, differences between the two groups were not significant. Milk PGEM concentrations also increased significantly (P=0.02) in control and treated heifers after the respective treatments, and there were no differences between the two groups. Results of the present study suggest that, the prostanoids have a role in the pathophysiologic process of coliform mastitis     

Survey of yeast mastitis in dairy herds of small-type farms in the Lublin region,Poland

The purpose of this study was to isolate fungi from the quarter milk of cow udders in 9 dairy farms. The samples were collected between May 1996 and April 2000 in the Lublin region. Six hundred and four milk samples collected from the quarters of 172 cows with clinical and subclinical mastitis were examined. Milk samples were plated as soon as possible on blood agar (BA), Mac Conkey agar, aesculin-tallium acetate-crystal violet blood agar, and Sabouraud agar with chloramphenicol and gentamicin. Fifty eight (9.6%) of the samples were positive for fungi. All of the fungal isolates were yeasts of the genera Candida, Rhodotorula and Trichosporon. We also isolated Streptococcus agalactiae (4.9%), Str. spp.(6.4%), Staphylococcus aureus (10.4%), coagulase-negative staphylococci-CNS (36.6%), Escherichia coli (3.5%), other microorganisms (2.6%) and no growth (25.8%). This revised version was published online in June 2006 with corrections to the Cover Date.     

Chronic ethanol intake in lactating rats: milk analysis

Milk was analyzed at 5 and 15 days of lactation in rats fed an ethanol liquid diet or appropriate control diet. Alcoholic rats showed blood ethanol levels as high as 43 mM at the end of lactation. Milk from ethanol-fed rats showed an increase in pH, protein and lipids and a decrease in lactose, compared with controls. Chronic ethanol consumption seems to reduce the yield of milk. The nutritional status of sucklings from alcoholic mothers seems to be related more to the quantity of milk than to its quality.     

Relationship between the bovine major histocompatibility complex (BoLA), erythrocyte markers and susceptibility to mastitis in Icelandic cattle

Summary. Milk and blood samples were obtained from three Icelandic dairy herds. The herds were monitored regularly for mastitis incidence. Cell counts, adenosine triphosphate (ATP) and antitrypsin levels of the milk samples were recorded. In addition, red cell and BoLA typing were performed on the blood. Although cell counts and ATP levels showed significant associations with mastitis, antitrypsin levels did not. Red blood cell antigens N'₂ and S₁ and the lymphocyte antigen detected by the monoclonal antibody M7 were associated with low cell counts, whilst BoLA w6 and w6.1 were associated with high cell counts. BoLA w6.2 and w11 showed significant association with high antitrypsin levels. Only ED116 showed a significant association with mastitis.     

Genetic correlations between production and disease traits during first lactation in Holstein cows

The aim of this study was to estimate genetic correlations between milk yield, somatic cell score (SCS), mastitis, and claw and leg disorders (CLDs) during first lactation in Holstein cows by using a threshold–linear random regression test-day model. We used daily records of milk, fat and protein yields; somatic cell count (SCC); and mastitis and CLD incidences from 46 771 first-lactation Holstein cows in Hokkaido, Japan, that calved between 2000 and 2009. A threshold animal model for binary records (mastitis and CLDs) and linear animal model for yield traits were applied in our multiple trait analysis. For both liabilities and yield traits, additive genetic effects were used as random regression on cubic Legendre polynomials of days on milk. The highest positive genetic correlations between yields and disease incidences (0.36 for milk and mastitis, 0.56 for fat and mastitis, 0.24 for protein and mastitis, 0.32 for milk and CLD, 0.44 for fat and CLD and 0.31 for protein and CLD) were estimated at about the time of peak milk yield (36 to 65 days in milk). Selection focused on early lactation yield may therefore increase the risk of mastitis and CLDs. The positive genetic correlations of SCS with mastitis or CLD incidence imply that selection to reduce SCS in the early stages of lactation would decrease the incidence of both mastitis and CLD.     

Relation of Milk Production Loss to Milk Somatic Cell Count

Milk production loss was studied in relation to increased somatic cell count (SCC). Available data were weekly test-day milk yields and SCC (in 1,000 cells/ml), and mastitis incidences. In total, 18,131 records from 274 cows were used. Production loss was determined for test-day kg milk, kg protein, and kg energy-corrected milk. Least-squares analysis of variance was used to estimate the direct effect of Log10(SCC) on production. The recorded measures of production were first corrected for fixed effects, with adjustment factors estimated from a healthy data-set. The average daily milk yield was 19.7 kg/day in first lactation and 22.0 in later lactations. The geometric mean of SCC was 63.1 in first lactation and 107.2 in later lactations. The incidence of clinical mastitis treated by a veterinarian was 19.8% of the lactations-at-risk. Linear relationships were found between the production parameters and Log10(SCC). Quadratic and cubic effects were evaluated, but were found to contribute little to the overall fit of the models. The individual milk yield loss was 1.29 kg/day for each unit increase in Log10(SCC) for cows in first lactation. Milk yield decreased by 2.04 kg/day per unit Log10(SCC) for older cows. Corresponding values for protein yield were 0.042 and 0.067 kg/day for first and later lactations, respectively.     

Enhanced levels of immunoreactive β-casomorphin-8 in milk of breastfeeding women with mastitis

An incorrect, superficial suckling technique in breastfeeding frequently leads to milk congestion and sometimes mastitis. In the present study we have examined whether milk congestion may affect levels of the atypical opioid β-casomorphin-8 in milk and in plasma. We also investigated whether the rate of acute psychosis during the first half year after parturition has declined in Sweden over the years. Milk and plasma samples were collected for peptide analysis from 14 women with mastitis and 10 controls. We found that in a group of 14 late cases of mastitis (median 48 days post partum) the detected mean level of β-casomorphin-8 in milk was significantly higher and somewhat higher in plasma at the acute stage compared with 2–3 weeks later, after recovery when the symptoms had disappeared, as well as compared to the control subjects. Swedish official statistics show that the incidence of acute psychosis in the first month and in the first half year after birth has declined by a half during the last 30 years. A relationship between postpartum psychosis and elevated β-casomorphin-8 levels in CSF has been suggested from earlier studies. In this study, milk congestion led to enhanced levels of β-casomorphin-8 in milk, which may be related to postpartum psychosis and probably also to ‘the postnatal blues’.     

The release of bradykinin in bovine mastitis.

The kinin peptides are released during inflammation and are amongst the most potent known mediators of vasodilatation, pain and oedema. Despite early reports of the presence of kinins in milk, no previous study has investigated the role of the kinin system in bovine mastitis. The present study indicated that mastitis was accompanied by raised levels of bradykinin (BK) in milk and the increased levels of BK correlated with the severity of mastitis. Raised BK levels in mastitic milk were not dependent on the presence of inflammatory cells, nor were they secondary to changes in blood levels of BK. In milk from sub-clinically inflamed quarters, BK was raised in those milks where Staphylococcus aureus (S. aureus) was isolated but not in those milks where no pathogen was isolated. Increasing S. aureus artificially, also caused an increase in the milk BK. Increases in milk BK were not restricted only to the mastitic quarters of the udder. In udders in which mastitis was detected in one or more quarters, BK increases were also detected in the apparently uninvolved quarters.     

Relations Between Udder Infection and Somatic Cells in Camel (Camelus dromedarius) Milk

Quarter milk samples (n = 391) from 101 camels were examined to study the occurrence and causes of mastitis in traditionally managed camels in eastern Sudan and to evaluate the value of the California Mastitis Test (CMT), somatic cell count (SCC) and adenosine triphosphate (ATP) in the detection of subclinical mastitis in the camel. One hundred and seventy (43.5%) of the quarter milk samples yielded pathogenic bacteria. Streptococcus agalactiae, other Streptococcus spp., Staphylococcus aureus, coag–ulase–negative staphylococci, and Escherichia coli were isolated from milk. Thirty–two (8.2%) quarter milk samples yielded mixed cultures, and 189 (48.3%) yielded no growth. Mean values for CMT, SCC and ATP were higher for quarters infected with major pathogens. However, a significant number of quarter milk samples had elevated values in these tests but were from quarters from which no bacteria were isolated. The ability of the tests to predict a positive bacteriology increased slightly when 2 or 3 tests were combined. kw|Keywords|k]inflammation; k]diagnostic tests; k]Mastitis; k]CMT; k]ATP; k]bacteriology; k]Sudan     

Variations During Lactation in Total and Differential Leukocyte Counts,N-acetyl-ß-D-glucosaminidase,Antitrypsin and Serum Albumin in Foremilk and Residual Milk from Non-infected Quarters in the Bovine

Quarter samples of foremilk and residual milk were taken approximately every second week from 2 days post partum (pp) throughout lactation month 9, from 5 dairy cows in their second lactation period. Bacteriologically positive milk samples were excluded. The aim was to study the variation in total and differential leukocyte counts, N-acetyl-ß-D-glucosaminidase (NAGase), antitrypsin (ATR) and serum albumin (BSA) in milk during the lactation period and different stages of oestrous cycle. Also the between milkings variation was studied from lactation month 4 to 9. At 2 days pp, each fraction of milk contained significantly higher numbers of leukocytes and had a higher activity of NAGase and ATR than later in the lactation period. In foremilk the highest content of BSA was also recorded at 2 days pp. From lactation month 2 to 9, stage of lactation had, in general, a slight effect on the variation in the variables measured. The total leukocyte count in residual milk tended to increase as lactation proceeded. The proportion of monocyte-macrophages in foremilk was significantly decreased during the last 4 months. NAGase and BSA in both fractions and ATR in residual milk increased significantly towards the end of the lactation period. From lactation month 4 to 9 the highest recorded ranges of variation between milkings, within quarter and stage of lactation, in the total leukocyte count, proportions of neutrophils, lymphocytes, monocyte-macrophages, NAGase, ATR and BSA in foremilk were 215 × 103ml, 42 %, 34 %, 54 %, 6.68 units, 0.36 units and 0.14 mg/ml respectively. The corresponding figures in residual milk were higher except for the variation in BSA which was slightly lower in residual milk than in foremilk. In residual milk there was a positive correlation between the proportion of neutrophils and the total leukocyte count, when calculated on data from all cows and the entire experimental period. During the oestrous periods, the proportion of neutrophils in residual milk was higher than during the dioestrous periods. Foremilk and residual milk differed in the total as well as the differential leukocyte counts in all the various stages of lactation, whereas the contents of NAGase, ATR and BSA were equal in both fractions. The exception was 2 days pp when the proportions of lymphocytes were equal in both fractions and BSA-significantly higher in foremilk than in residual milk.     

Isolation,Biochemical and Molecular Identification,and In-Vitro Antimicrobial Resistance Patterns of Bacteria Isolated from Bubaline Subclinical Mastitis in South India

Buffaloes are the second largest source of milk. Mastitis is a major impediment for milk production, but not much information is available about bubaline mastitis, especially subclinical mastitis. The aim of this study was to (a) investigate the application of various tests for the diagnosis of bubaline subclinical mastitis, (b) identify the major bacteria associated with it, and (c) evaluate the antibiotic resistance pattern of the bacteria. To this end, 190 quarter milk samples were collected from 57 domesticated dairy buffaloes from organized (64 samples) and unorganized (126 samples) sectors. Of these, 48.4%, 40.0%, 45.8%, 61.1%, and 61.6% were positive for subclinical mastitis by somatic cell count, electrical conductivity, California mastitis test, bromothymol blue test, and N-acetyl glucosaminidase test, respectively. As compared to the gold standard of somatic cell count, California mastitis test performed the best. However, a combination of the two methods was found to be the best option. Microbiological evaluation, both by biochemical methods as well as by monoplex and multiplex polymerase chain reaction, revealed that coagulase-negative staphylococci were the most predominant (64.8%) bacteria, followed by streptococci (18.1%), Escherichia coli (9.8%) and Staphylococcus aureus (7.3%). Most of the pathogens were resistant to multiple antibiotics, especially to β-lactam antibiotics. We propose that California mastitis test be combined with somatic cell count for diagnosis of subclinical mastitis in domestic dairy buffaloes. Further, our results reveal high resistance of the associated bacteria to the β-lactam class of antibiotics, and a possible major role of coagulase-negative staphylococci in causing the disease in India.