奶牛隐性乳腺炎病原菌的分离鉴定和体细胞数及酶的相关性研究

对330个乳样分别用加州乳腺炎检验(California Mastitis Test,CMT),体细胞计数(Somatic Cell Count,SCC)和细菌的分离培养(Isolation of bacteria)法进行隐性乳腺炎的检测,结果表明3种方法检测的结果差异不显著(P>0.05);对检测患有隐性乳腺炎的241乳区进行了细菌的分离培养,结果表明引起隐性乳腺炎的最主要致病菌为金黄色葡萄球菌、凝固酶阴性葡萄球菌、链球菌、酵母菌、假单胞菌和棒状杆菌、其它细菌的检出率差异根显著(P<0.05)。对分离出优势细菌的部分乳样进行了SCC与LDH,NAGase,ALP和ACP几种酶的测定和相关性分析,结果表明:LDH与SCC有显著的相关性(r=0.7936),并且LDH与NA-Gase,ALP,ACP都有明显的相关性;NAGase与ACP,LDH有明显的相关性,但与ALP没有相关性;ALP只与ACP和LDH有显著的相关性外,与SCC,NAGase没有相关性;ACP与SCC,LDH,NAGase,ALP都有极显著的相关性。     

Prevalence of coagulase-negative staphylococci in bovine mastitis in Zimbabwe

This study was carried out to determine the prevalence of coagulase-negative staphylococci in clinical and subclinical mastitis in commercial and small-scale farms in Zimbabwe. Thirty five quarter milk samples from clinical mastitis cases and 371 quarter milk samples from cows with subclinical mastitis were cultured for bacterial pathogens. The most frequent pathogens isolated in clinical mastitis were the enteric bacteria (31.4%), followed by coagulase negative staphylococci (22.9%) and then Staphylococcus aureus (17.1%), whereas in subclinical mastitis S. aureus (34.2%) and coagulase-negative staphylococci were (33.2%) the most common. Bacillus species were only isolated in milk samples from subclinical mastitis. Coagulase-negative staphylococci were observed in mixed infections with other bacteria in only 2.2 of the 406 milk samples from clinical and subclinical mastitis where they were isolated together with Bacillus species in 6 of the 9 mixed infection cases. About 95% of the milk samples from which 131 coagulase-negative staphylococci were isolated had correspondingly high somatic cell counts. The coagulase-negative staphylococci isolated most frequently were S. chromogenes (7.9%), S. epidermidis (7.4%) and S. hominis (5.9%). They were all associated with high somatic cell counts. All the coagulase-negative staphylococci isolates were susceptible to cloxacillin and erythromycin, and more than 90% of the isolates were susceptible to neomycin, penicillin and streptomycin. The highest resistance was to tetracycline (17.6%), followed by lincomycin (13.7%). About 8% of the isolates were resistant to both penicillin and streptomycin.     

Antitrypsin and N-acetyl-beta-D-glucosaminidase as markers of mastitis in a herd of Ayshire cows

Microplate analyzers for N-acetyl-beta-D-glucosaminidase (NAGase) and antitrypsin in milk were high-capacity systems for determination of subclinical mastitis on a mammary-quarter basis in a herd of 80 Ayshire cows. Infected quarters could be better differentiated from non-infected quarters by interquarter evaluation of milk NAGase and antitrypsin, than by adaptation of threshold values based on measured NAGase and antitrypsin values. This differentiation between healthy quarters and mastitic quarters was further improved by using absolute values and interquarter evaluations in combination. Therefore, quarter-based sample collection in combination with microplate technology and computerization is proposed as the method of choice for monitoring udder health in herds.     

The prevalence of subclinical mastitis in dairy goats in Kenya

California mastitis test (CMT), direct leukocytes counts and bacteriological examination were performed on 630 milk samples from apparently healthy mammary glands of dairy goats comprising a mixed population of German Alpine, Toggenberg, Saanen and Galla crosses to find the prevalence of subclinical mastitis. The prevalence of subclinical mastitis was 9.8% according to CMT, 9.7% according to direct leukocyte counts and 28.7% by bacterial isolation during a 3-month period. The proportion of the bacteriologically positive milk samples was significantly (P <0.01) higher than that positive for CMT and direct leukocyte counts. There was a significant (P < 0.01) correlation between CMT and direct leukocyte counts. There was no significant direct relationship between bacterial isolation and CMT Bacterial organisms were isolated in 22.5% of the 568 CMT-negative milk samples. The results suggest that bacterial organisms isolated from the CMT-negative milksamples were either latent infections or did not stimulate any significant increase in somatic cell counts that could be detected by either the CMT or direct leukocyte counts. The observations of this study indicate that the mere presence of bacteria in goat's milk does not mean that the udder is infected and so does not warrant antibiotic therapy.     

Acute phase proteins in milk in naturally acquired bovine mastitis caused by different pathogens

The concentrations of haptoglobin (Hp) and serum amyloid A (SAA) and the activity of N-acetyl-β-D-glucosaminidase (NAGase) in milk from 234 cows with spontaneous mastitis caused by different pathogens were measured to assess whether they corresponded with the clinical signs of mastitis and whether there were any differences between pathogens. Ninety-eight of the cows had clinical mastitis and 136 had subclinical mastitis. There were statistically significant positive correlations between the concentrations of SAA and Hp and the activity of NAGase. Significant differences in the concentrations of acute phase proteins and NAGase activity were found in milk from cows with mastitis caused by different pathogens. The highest concentrations of Hp and NAGase were found in cases of mastitis caused by Escherichia coli and Arcanobacterium pyogenes, and the lowest concentrations were from cases of mastitis caused by coagulase-negative staphylococci. Very low SAA concentrations were found in milk from the cases caused by A pyogenes, in contrast to cases caused by other major mastitis pathogens. The median concentration of SAA was over 10 times higher in cases of mastitis caused by E coli than in mastitis caused by other pathogens. There were significant differences in the mean Hp concentration and NAGase activity between clinical and subclinical mastitis. In approximately one-third of the samples, the Hp concentration was below the detection limit, potentially compromising the use of Hp as a mastitis marker.     

Direct and indirect measurement of somatic cell count as indicator of intramammary infection in dairy goats

Background

Mastitis is the most important and costly disease in dairy goat production. Subclinical mastitis is common in goats and is mainly caused by contagious bacteria. Several methods to diagnose subclinical mastitis are available. In this study indirect measurement of somatic cell count (SCC) by California Mastitis Test (CMT) and direct measurement of SCC using a portable deLaval cell counter (DCC) are evaluated. Swedish goat farmers would primarily benefit from diagnostic methods that can be used at the farm. The purpose of the study was to evaluate SCC measured by CMT and DCC as possible markers for intramammary infection (IMI) in goats without clinical symptoms of mastitis. Moreover to see how well indirect measurement of SCC (CMT) corresponded to direct measurement of SCC (DCC).

Method

Udder half milk samples were collected once from dairy goats (n = 111), in five different farms in Northern and Central Sweden. Only clinically healthy animals were included in the study. All goats were in mid to late lactation at sampling. Milk samples were analyzed for SCC by CMT and DCC at the farm, and for bacterial growth at the laboratory.

Results

Intramammary infection, defined as growth of udder pathogens, was found in 39 (18%) of the milk samples. No growth was found in 180 (81%) samples while 3 (1%) samples were contaminated. The most frequently isolated bacterial species was coagulase negative staphylococci (CNS) (72% of all isolates), followed by Staphylococcus aureus (23% of all isolates). Somatic cell count measured by DCC was strongly (p = 0.000) associated with bacterial growth. There was also a very strong association between CMT and bacterial growth. CMT 1 was associated with freedom of IMI while CMT ≥2 was associated with IMI. Indirect measurement of SCC by CMT was well correlated with SCC measured by DCC.

Conclusions

According to the results, SCC measured with CMT or DCC can predict udder infection in goats, and CMT can be used as a predictor of the SCC.     

Comparison of milk antitrypsin,albumin, n-acetyl-β-d-glucosaminidase,somatic cells and bacteriological analysis as indicators of bovine subclinical mastitis

Thirty-two quarters, five of which harbored subclinical mastitis, were examined daily for one month. The usefulness of milk antitrypsin, BSA (bovine serum albumin), NAGase, somatic cells and bacteriological analysis in differentiating the inflamed quarters from the healthy control quarters was analysed. Inter-quarter evaluation clearly improved each indirect mastitis parameter; NAGase and antitrypsin were better indicators of differences between infected and non-infected quarters than BSA or the somatic cell count.     

Comparison of milk antitrypsin, albumin, n-acetyl-beta-D-glucosaminidase, somatic cells and bacteriological analysis as indicators of bovine subclinical mastitis

Thirty-two quarters, five of which harbored subclinical mastitis, were examined daily for one month. The usefulness of milk antitrypsin, BSA (bovine serum albumin), NAGase, somatic cells and bacteriological analysis in differentiating the inflamed quarters from the healthy control quarters was analysed. Inter-quarter evaluation clearly improved each indirect mastitis parameter; NAGase and antitrypsin were better indicators of differences between infected and non-infected quarters than BSA or the somatic cell count.     

Detection of ovine intramammary infection with the California mastitis test

The overall sensitivity of the California mastitis test (CMT) for detecting intramammary infection (the likelihood of a positive CMT score in the presence of intramammary infection) was 69.3%. The specificity of the CMT (likelihood of a negative test in the absence of intramammary infection) was 76.5%. When only infections by major ovine mastitis pathogens were considered, the sensitivity increased to 100% and the specificity decreased to 71.1%. Bacterial cultural examination and CMT scores were recorded for 526 samples of milk obtained from 106 brood ewes. Positive CMT scores were recorded for all samples from udder halves infected with major ovine mastitis pathogens (coagulase-positive staphylococci, Escherichia coli, and Pasteurella haemolytica). The CMT scores for samples from udder halves infected with coagulase-negative staphylococci were variable.     

Innate immune response in experimentally induced bovine intramammary infection with Staphylococcus simulans and S. epidermidis

ABSTRACT: Coagulase-negative staphylococci (CNS) are in several countries the most common bacteria isolated in subclinical mastitis. To investigate the innate immune response of cows to infections with two common mastitis-causing CNS species, Staphylococcus epidermidis and Staphylococcus simulans, experimental intramammary infection was induced in eight cows using a crossover design. The milk somatic cell count (SCC), N-acetyl-β-D-glucosaminidase (NAGase) activity, milk amyloid A (MAA), serum amyloid A (SAA) and proinflammatory cytokines interleukin (IL)-1β, IL-8, and tumor necrosis factor α (TNF-α) were determined at several time points before and after challenge. All cows became infected and showed mild to moderate clinical signs of mastitis. The spontaneous elimination rate of the 16 infections was 31.3%, with no difference between species. Infections triggered a local cytokine response in the experimental udder quarters, but cytokines were not detected in the uninfected control quarters or in systemic circulation. The innate local immune response for S. simulans was slightly stronger, with significantly higher concentrations of IL-1β and IL-8. The IL-8 response could be divided into early, delayed, or combined types of response. The CNS species or persistency of infection was not associated with the type of IL-8 response. No significant differences were seen between spontaneously eliminated or persistent infections.     

Normal somatic cell count and subclinical mastitis in Murrah buffaloes

This study was conducted to investigate the normal somatic cell count (SCC) and to define subclinical mastitis in Murrah buffaloes. Data were collected from 60 clinically normal buffaloes stationed at five farms of Chitwan Nepal and Buffalo Research Center, Hissar, India. Somatic cell count was measured using the Newman-Lampert staining technique. The upper limit of SCC was determined >or=200 000/ml of milk based on the mean +/- 2SD of a total SCC. Abnormal data of the SCC was repeatedly removed, which lie beyond the values of more than mean + 2SD until all the data come to lie within (mean + 2SD). Averages of SCC of right front and right hind quarters were significantly higher than left front and left hind quarters. Nearly 94% of California mastitis test (CMT) negative quarters were having somatic cells >or=200 000/ml. The mean SCC of CMT positive quarter was significantly higher (P < 0.01) than CMT negative quarters. Subclinical mastitis was diagnosed on the basis of samples with SCCs >or=200 000/ml with positive bacterial cultures. Subclinical mastitis was found in 21.7% buffaloes and 8% of the quarter foremilk samples. Neutrophil counts were significantly higher in subclinical mastitis milk.     

Identification of subclinical mastitis with a hand-held electrical conductivity meter

A hand-held, commercially available instrument for measuring the electrical conductivity of milk (the Milk Checker) has been examined for its usefulness and accuracy in detecting subclinical and clinical mastitis. Foremilk from uninfected quarters had an electrical conductivity of 5.4 to 5.6 millisiemens (mS)/cm. Milk from cows with subclinical Staphylococcus aureus infections had a higher milk conductivity (7.1 to 7.5 mS/cm) but milk from cows with subclinical S uberis infections showed no increase in conductivity (5.3 to 5.6 mS/cm). However, experimental S uberis infections could be detected by a 50 per cent increase in the electrical conductivity of foremilk two milkings before visible signs of mastitis were apparent. The equipment could be a useful advisory/veterinary tool but is unlikely to be used routinely in the milking parlour.     

The detection of subclinical mastitis in the bactrian camel (Camelus bactrianus) by somatic cell count and California mastitis test

Milk samples (n=160) from 7 clinically healthy bactrian camels were cultured to detect subclinical udder infection. The samples were assessed by the Californian mastitis test (CMT) and somatic cell count (SCC). Bacteria were recovered from 36 (22.5%) of the milk samples. Staphylococcus aureus and coagulase-negative staphylococci (CNS) were the main organisms found.Infected quarters had significantly higher mean values for the SCC (p<0.01) and CMT (p<0.001) than non-infected quarters. All 7 camels were infected with CNS but only 4 with S. aureus. CMT values for S. aureus-infected camels were significantly higher than for those only infected with CNS. The values for SCC and CMT were significantly influenced by the stage of lactation (p<0.05). No significant difference was found from the effect of the quarters. Both SCC and CMT were of value in predicting the infection status of the udder.Abbreviations CMT California mastitis test - SCC somatic cell count - CNS coagulase-negative staphylococci     

Prevalence, causes and laboratory diagnosis of subclinical mastitis in the goat

Five goat herds were examined to determine the prevalence and causes of subclinical mastitis and to assess the value of some laboratory tests currently used on milk samples as aids in the diagnosis of caprine mastitis. In the 170 samples taken from the pairs of mammary glands of 85 goats, the prevalence of infection in the different herds ranged from 15 per cent to 79 per cent of halves. Just over one-third (36 per cent) of all halves were infected, the organisms isolated being coagulase-negative staphylococci (80 per cent), coagulase-positive staphylococci (16 per cent), alpha-haemolytic streptococci (2 per cent) and Pasteurella haemolytica (2 per cent). Neither anaerobic organisms nor mycoplasmas were found. Tests confirmed that the coagulase-positive staphylococci were pathogens but that the coagulase-negative staphylococci rarely caused detectable disease in the caprine udder. The large between-herd variation in the geometric mean cell counts of uninfected milk samples means that somatic cell counts, the Whiteside test and the California mastitis test, are unreliable as aids in the diagnosis of caprine mastitis.     

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.     

Causative organisms and somatic cell counts in subclinical intramammary infections in milking goats in the UK

The bacterial causes of subclinical mastitis were determined in samples of milk taken from one half of the udders of 159 goats in three different herds. The mean prevalence of subclinical infection was 33 percent, with prevalences of 26 percent, 39 percent and 42 percent in the three herds. Staphylococcus aureus was isolated from seven (13 percent) of the 53 infected halves, coagulase-negative staphylococci accounted for 47 percent, Corynebacterium species for 31 percent and alpha-haemolytic streptococci for 6 percent of the infected samples. The mean somatic cell count of the uninfected milk samples was 428,000 cells/ml, and 93 percent of uninfected samples had counts less than 1,000,000 cells/ml; the mean cell count of the infected samples was 2,785,000 cells/ml.     

Persistent Experimental Salmonella dublin Intramammary Infection in Dairy Cows

Experimental intramammary infections were induced in five post-parturient Holstein cows by inoculation of low numbers (5000 colony forming units) of virulent Salmonella dublin via the teat canal of mammary gland quarters. Rectal temperature, pulse and respiratory rates, milk yield, and milk quality as assessed by the California Mastitis Test (CMT) and somatic cell counts (SCC) were recorded every 12 hours at milking. Bacteriologic cultures of foremilk quarter samples and feces were obtained daily, as were complete blood counts. ELISA titers for IgG and IgM recognizing S. dublin lipopolysaccharide (LPS) were obtained weekly on serum and quarter milk samples. All cows excreted S. dublin intermittently from infected quarters, but no changes were detected in rectal temperature, appearance of the mammary gland or secretions, CBC, milk yield, and pulse and respiratory rates. Somatic cell counts were modestly increased in infected quarters as compared with uninfected quarters (P = .015, paired t test); however, CMT scores after infection remained low, and were not significantly different from pre-infection scores (P greater than .10, sign test). After infection, administration of dexamethasone resulted in signs of clinical mastitis and increased excretion of S. dublin from mammary quarters (P = .0004, paired t test). One cow had necrotizing mastitis and S. dublin septicemia and was euthanatized. In the four surviving cows, clinical improvement was observed after systemic gentamicin therapy and intramammary infusion with polymyxin B, but all cows continued to excrete S. dublin intermittently from one or more quarters and occasionally from feces for the remaining period of observation. All infected cows demonstrated a rise in IgG and IgM ELISA titers recognizing S. dublin LPS in serum and milk. At necropsy (13-25 weeks postinfection), S. dublin was recovered only from the mammary tissue or supramammary lymph nodes in three of four cows. In one cow, mammary gland and lymph-node samples were negative for S. dublin despite positive milk cultures. In all cows, histopathologic examination revealed multifocal areas of chronic active mastitis. These lesions were similar to histopathologic findings from mammary gland carriers with naturally acquired S. dublin infection.     

Prevalence of subclinical udder infections and individual somatic cell counts in three dairy goat herds during a full lactation

For dairy goats, both the determination of the somatic cell counts (SCC) and the interpretation of these values may be a problem. Several investigations have shown that SCC for goat's milk, even from not infected mammary halves, are often higher than for cows milk. In the three herds examined about 40% of mammary halves and 30% of the goats were infected. However large differences between the three herds could be observed. In most cases, infections were caused by coagulase negative staphylococci (CNS) or corynebacteria. The SCC of individual milk samples from goats without any udder infection hardly differed from those of goats with at least one udder half infected with CNS. In 20% and 30% of the cases the SCC was higher than 750'000 cells/ml, respectively. The relation between California Mastitis Test (CMT) reactions and udder infections was not very close. Over 20% of mammary halves infected with CNS showed negative CMT reactions. On the other hand, 25% of samples from mammary halves without a proven infection reacted positively. The large differences in individual cell counts on herd and animal level indicate that production and breeding systems might be important reasons for the higher SCC. As a consequence, the most common methods for or the control of udder health and udder infections (SCC, California Mastitis Test) are of limited value for goats. Since there was only a weak relation between milk quality properties and SCC, any arguments for the introduction of legal limits below 1 million cells per ml can hardly be found.     

Bayesian latent class evaluation of three tests for the screening of subclinical caprine mastitis in Bangladesh

Routine monitoring for subclinical infection is one of the key mastitis control approaches. However, the accuracy of the most commonly used screening tests has not yet been established. The aim of the present study was therefore to evaluate the accuracy of three screening tests, namely California mastitis test (CMT), white side test (WST), and surf field mastitis test (SFMT) for the screening of subclinical caprine mastitis. A cross-sectional study based on 484 randomly collected milk (242 goats) samples from three districts of Bangladesh was conducted for the screening of subclinical mastitis by the aforementioned tests. The Bayesian latent class model was implemented in WinBUGS to estimate the tests’ characteristics and true prevalence of subclinical mastitis. The Bayesian posterior estimates of sensitivities with a 95% credible intervals (CrIs) were 98.60% (95.18–99.95%), 98.28% (94.56–99.92%), and 89.98% (83.39–95.03%), and specificities with 95% CrIs were 99.19% (98.11–99.96%), 99.27% (97.34–99.98%), and 99.28% (97.35–99.98%), respectively for CMT, WST, and SFMT. The true prevalence of subclinical caprine mastitis was estimated to be 43.49% (95% CrI 37.46–48.98%). The positive predictive values (PPV) of the three tests were similar. The serial and parallel interpretation of any test pairs increased the PPV and negative predictive value respectively close to 100%. Based on the simplicity, cost and performance as well WST and SFMT simultaneously could be recommended for the screening of caprine subclinical mastitis in Bangladesh.

     

Observations on intramammary infections in first calf heifers in early lactation

Intramammary infections and mastitis were monitored on four occasions at three-week intervals in 61 first calf heifers in five dairy herds during the first ten weeks of lactation. Of 940 quarter milk samples examined 65% were classed as negative, 10.4% as positive for mastitis (mainly subclinical), 1.8% as group 3 (infection present but no elevation in somatic cell count) and 22.8% as group 4 (elevated somatic cell count). Seventy-seven percent of the infections detected and identified were those due to coagulase-negative staphylococci, the main species being S. hyicus, S. epidermidis, S. simulans and S. hominis. Other infections detected with Corynebacterium pyogenes (three samples), Escherichia coli (one sample), Micrococcus spp. (one sample), S. aureus (two samples) and Streptococci (non-agalactiae) (seven samples).The geometric mean somatic cell count for 23 quarters infected with coagulase-negative staphylococci was 311 x 10(3) cells / mL compared to 134 x 10(3) cells / mL in noninfected adjacent contralateral quarters. The respective figures for% cell volume in Channel 8 (mainly neutrophils) were 10.6% and 3.5%. There was a highly significant association between herd and the proportion of quarter milk samples in the four mastitis categories.