Scanning electron microscopy-aided observations on and therapy of teat canal infections

An examination of teat canal swabs established that 51 teat canals out of 68 quarters of machine-milked cows were colonized by Staphylococcus aureus. Only 31 of these quarters yielded milk from which S. aureus could be cultured, and 6 out of the 31 produced milk containing somatic cell counts in excess of 500 000/ml. No inhibitory substances could be detected in milk samples 12 h after 10 mg of sodium cloxacillin had been deposited in the test canal on 1-4 successive occasions. Teat canal swabs and milk sample cultures of the same quarters became and remained bacteriologically negative for at least a week after the last treatment. Six quarters, which according to the International Dairy Federation criteria were suffering from subclinical mastitis, became negative after local teat canal therapy. Scanning electron micrographs of one infected teat canal revealed the presence of cocci in depressions and crevices on the epithelial surface, suggesting that such cocci are not always flushed out into milk samples. Teat canal therapy should make a marked contribution to the control of bovine mastitis.     

Bovine serum albumin and cell counts in the diagnosis of subclinical udder infection

Summary

Puncture of the milk cisterns was performed in 120 bacteriologically positive quarters of forty‐seven lactating dairy cows on three farms. This method was used to determine whether the existing infection was an infection of the teat canal or one of the udder. The results were related to the concentration of bovine serum albumin (BSA) and the cell count in the milk. Of the bacteriologically negative quarters, both BSA levels (in 91 per cent of the quarters the BSA concentration was 0.20 mg. per ml. of milk or less) and cell counts (92 per cent contained less than 500,000 cells per ml. of milk) were low. In cases of udder infection with primary pathogenic bacteria there was a marked increase in cell count (90 per cent more than 500,000 cells per ml. of milk), whereas the increase in BSA was rather small (51 per cent still contained 0.20 mg. BSA per ml. of milk or less). While the difference in cell counts of milk from quarters with udder infections and teat canal infections with primary pathogenic bacteria was significant, the difference between the BSA levels of these two groups was not. Therefore, the cell count supplies more reliable information than does the BSA level of the milk. Of all infections, 23 per cent were found to be infections of the teat canal.     

Changes in the bovine udder quarters naturally infected by Corynebacterium bovis

Of 272 bovine udder quarters studied for mastitis, 19 of them naturally infected with Corynebacterium bovis alone, were compared with 16 others infected by C. bovis together with other bacteria and another 36 non-infected quarters. While there was no significant difference in milk somatic cell counts between the quarters infected by C. bovis alone and those affected by C. bovis together with other bacteria (33.37 +/- 20.28 X 10(3) and 33.86 +/- 23.18 X 10(3)/ml of milk, respectively), there was a significant difference between these and the non-infected quarters (5.60 +/- 3.23 X 10(3)/ml of milk). Microscopically, quarters infected by either C. bovis alone or C. bovis in combination with other bacteria had inflammatory changes in the teat cisterns, Furstenberg's rosettes and/or mammary parenchyma. The non-infected quarters had no changes. In all 82 quarters no pathological changes could be seen in the teat canals.     

Role of eicosanoids, histamine, and serotonin in the pathogenesis of Klebsiella pneumoniae-induced bovine mastitis

By inoculating Klebsiella pneumoniae into the teat canals of mammary glands, coliform mastitis was induced experimentally in 6 lactating cows. Release of eicosanoids, histamine, and serotonin in plasma and milk was studied in response to 2 doses of K pneumoniae. A low dose (mean, 5,000 organisms/ml) was inoculated into cows 1 through 4, and a high dose (mean, 200,000 organisms/ml) was inoculated into cows 5 and 6. Milk and blood samples were collected before inoculation (0 hours), and hourly, from 3 to 24 hours after inoculation. Concentrations of prostaglandin F2 alpha (PGF2 alpha), prostaglandin E (PGE), thromboxane B2 (TxB2), histamine, and serotonin were measured in plasma and milk obtained from control (NaCl solution-inoculated) and infected quarters. Fluorometric analysis of milk from infected quarters revealed significantly increased histamine and serotonin concentrations regardless of the dose of K pneumoniae. The mean (+/- SEM) peak concentrations of histamine were significantly (P less than 0.01) increased from the preinoculation value of 44 (+/- 12) ng/ml to 312 (+/- 104) ng/ml in milk from infected quarters and 72 (+/- 24) ng/ml in milk from control quarters. The mean peak concentration of serotonin increased significantly from the preinoculation concentration of 436 (+/- 37) ng/ml to 1,754 (+/- 662) ng/ml and 4,867 (+/- 1,248) ng/ml in milk from control (P less than 0.02) and infected (P less than 0.001) quarters, respectively. However, serotonin concentration in milk from infected quarters was approximately 2.8 times greater than that in milk from control quarters. Concentrations of PGF2 alpha, PGE, and TxB2 in milk and plasma were evaluated by radioimmunoassay.(ABSTRACT TRUNCATED AT 250 WORDS)     

Experimental colonization of the bovine teat duct with Corynebacterium bovis and the effect on milk somatic cell counts.

Colonization with Corynebacterium bovis was established in 59 of 64 (92%), 58 of 59 (98%) and 19 of 34 (56%) of uninfected bovine mammary quarters following inoculation of 83.3 X 10(4) colony-forming units (CFU) of the organism into the teat cistern, 4.7 X 10(3) CFU 5 mm into the teat duct or by exposure of the teat orifice to a milk culture containing 1.6 X 10(7) CFU/mL respectively. Mean somatic cell counts for foremilk samples from 122 quarters were significantly higher after colonization with C. bovis (145,900/mL) compared to before exposure (130,900/mL).     

Bacterial cure and somatic cell count response of dairy cows with a positive California Mastitis Test at calving to therapy with cephapirin sodium

Cows without signs of clinical mastitis were evaluated by the California Mastitis Test at calving (Day 0). Milk samples from 117 of 184 quarters (64 cows) were positive by this test for mastitis and were submitted for bacterial culture and determination of somatic cell counts. Cows with infected quarters were randomly allocated to treatment with cephapirin sodium by intramammary infusion or to be untreated as controls. Two and 4 weeks following calving, milk was again sampled from the infected quarters and tested. By the 4-week evaluation, the quarters treated with cephapirin sodium had significantly (P < or = .05) fewer positive bacterial cultures and somatic cell counts were significantly (P < or =.05) reduced compared with untreated control quarters.     

Concentrations of triiodothyronine (T3), tumour necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) in milk from healthy and naturally infected quarters of cows

The effect of naturally acquired bacterial infection of the bovine udder on the activity of 5'-thyroxine monodeiodinase (5'-MD), and on the concentrations of the pro-inflammatory cytokines interleukin (IL)-6 and tumour necrosis factor (TNF)-alpha in milk, from healthy (control) and inflamed quarters, was determined. The diagnostic procedure included history and clinical examination of the udder, macroscopic evaluation of secretions, the Californian Mastitis Test, determination of somatic cell counts and bacteriological examination of milk. It has been found that the milk triiodothyronine (T3) content and the 5'-MD activity from inflamed quarters were decreased when compared with controls. The decrease in the milk T3 from subclinical mastitic quarters was manifested when somatic cell counts were > 10(6) ml(-1). TNF-alpha was on average 2-fold higher in infected milk, and the concentration of IL-6 was unchanged. These results suggest that the decreased T3 content in mammary secretions during naturally occurring mastitis is associated with the severity of inflammation, increased TNF-alpha concentration and impaired enzymatic activity of 5'-MD.     

Effect of intramammary devices on the outcome of induced Escherichia coli infection of bovine mammary quarters

Eighteen Holstein cows, free of intramammary infection, were fitted with smooth (n = 9) or abraded (n = 9) intramammary devices (IMD) in 2 diagonally opposed quarters within 4 weeks after calving. The 2 other quarters of each cow were used as controls. Three to 6 weeks after IMD insertion, depending on when milk somatic cell counts returned to a base-line value of less than 4 X 10(5)/ml, all cows were subjected to bacterial challenge exposure in the front or rear quarters by intracisternal injection of about 30 colony-forming units of Escherichia coli/quarter. Challenge exposure was done immediately after milking. Three weeks after the initial bacterial exposure, the other quarter pairs were similarly challenge exposed. Quarter bacteriologic status, concentration of milk somatic cells, and clinical observations (rectal temperature, milk appearance, udder palpation, and general condition of the cow) were monitored. Infection developed in 14 of 16 (88%) quarters with smooth IMD vs 16 of 16 (100%) control quarters and in 7 of 17 (41%) quarters with abraded IMD vs 17 of 17 (100%) control quarters. The difference in infection frequency between quarters with smooth IMD and quarters with abraded IMD was significant (P less than 0.05). Protection against establishment of infection was associated with somatic cell counts greater than 8.0 X 10(5)/ml in milk collected immediately after milking (7 of 12 quarters) or 4 hours later (11 of 12 quarters). In 10 quarters (59%) of cows fitted with abraded IMD, secretory abnormalities appeared before bacterial challenge inoculation. Abnormal milk or visible blood was observed over periods varying from 2 weeks after insertion through the entire lactation.     

Efficacy of flunixin meglumine for the treatment of endotoxin-induced bovine mastitis

The clinical effect of flunixin meglumine administration was determined in cows with acute mastitis induced by intramammary administration of endotoxin. In 12 lactating cows, 10 micrograms of Escherichia coli 026:B6 endotoxin were administered via a teat cannula into the teat cistern of single randomly selected rear quarters. Cows were challenge exposed as pairs. One cow in each pair was administered parenteral flunixin meglumine (6 cows) and 1 cow per pair was administered saline solution (6 cows). Multiple doses (7) of 1.1 mg of flunixin meglumine/kg of body weight or saline solution were administered at 8-hour intervals beginning 2 hours after endotoxin. Cow and quarter clinical signs as well as milk somatic cell concentrations, bovine serum albumin, electrical conductivity, and milk production were determined before and for 14 days after endotoxin inoculation. Intramammary endotoxin produced signs characteristic of acute coliform mastitis. Quarter and systemic abnormalities occurred and milk production was reduced by approximately 50% at 12 hours after endotoxin. Flunixin meglumine therapy significantly (P less than or equal to 0.05) reduced rectal temperatures and quarter signs of inflammation and improved clinically graded depression when compared with these signs in saline solution-treated controls. Milk production and laboratory indicators of inflammation in milk were not significantly (P greater than 0.05) different for flunixin meglumine vs saline solution controls. The clinical response observed was consistent with the antipyretic, analgesic, and anti-inflammatory properties of flunixin meglumine.     

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.     

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.     

Effect of the intramammary device on milk infection status, yield, and somatic cell count and on the morphological features of the lactiferous sinus of the bovine udder

Twenty primiparous heifers were fitted intramammarily with polyethylene coils in both quarters of one random right or left udder half at 5 days after parturition. Foremilk samples were collected and udder-half milk yields were measured at the afternoon milking on days - 1, 3, 7, and 14 and on every 14th day for 8 months after the device was inserted. Three weeks after the heifers were fitted with the intramammary device, 6 were euthanatized for gross observation of devices and tissues and cytologic evaluation of the gland cistern epithelium. There were significantly fewer bacterial isolations (P less than 0.01) and less clinical mastitis (P less than 0.05) in treated quarters than in the control quarters. Coagulase-negative staphylococci were isolated at frequencies of 0 and 15.2% for treated and control quarters. The reduction in isolation frequency for treated, compared with control, quarters was less marked with other organisms. Intramammary devices in no way interfered with the milking process. Milk yields per milking were 4.2 kg for treated udder halves compared with 4.4 kg for control halves; however, this 0.2 kg difference was not significant. Mean milk somatic cell counts, as determined by electronic counter, were 34 X 10(3) and 81 X 10(3) cells/ml for control and treated quarters (P less than 0.05). Mean bovine serum albumin values were 0.160 and 0.175 mg/ml for control and treated udder halves (P less than 0.05), indicating an increased capillary permeability due to the device. Quantitative morphologic analysis of gland cisterns showed a significant (P less than 0.05) change toward a single layer of epithelial cells in treated quarters compared with a double layer in control quarters.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intensification of milk somatic cell response to intramammary device

Treatments consisting of copper-impregnated polyethylene intramammary device (PIMD-Cu), PIMD-Cu which had been abraded to ensure exposure of surface copper (APIMD-Cu), PIMD which had been abraded in an identical manner (APIMD), and an untreated control were established in mammary quarters of 2 cows. Quarters selected were bacteria free and had milk somatic cell counts (MSCC) in strippings of less than 240 X 10(3) ml. Milk somatic cell counts were determined from strippings immediately after milking and 6 hours later. Cows were monitored for 4 weeks after devices were inserted. Milk samples (250 ml) were collected and analyzed for free fatty acids, a measure of hydrolytic rancidity, at 14 days. The devices were removed from mammary quarters after 4 weeks and examined with a scanning electron microscope. Surfaces of the devices were assessed for plaque buildup and presence of leukocytes. Immediately after and 6 hours later, geometric mean MSCC for APIMD, APIMD-Cu, PIMD-Cu, and control quarters average 946/1,556, 1,479/2,882, 512/1,148, and 161/190 X 10(3) ml, respectively. There was no difference in free fatty acid values of samples from treated and control quarters. Plaque formation was observed over the entire surface of APIMD. Numerous leukocytes were found associated with plaque and appeared to initiate development of plaque. Smaller amounts of plaque were found on APIMD-Cu, and minimal amounts were found on PIMD-Cu. Results indicate that modification of PIMD by abrading or addition of copper will increase MSCC to concentrations (greater than 900 X 10(3) ml) that should be protective against establishment of infection by mastitis pathogens.     

Fungi isolated from bovine udders , and their possible sources

AIM: To identify fungi isolated from infections of the bovine mammary gland, and establish their possible sources. METHODS: From a herd of 420 cows, milk samples were collected from all quarters at calving and cultured to detect causative organisms. Quarters identified as infected with fungi were further sampled during early lactation. Samples from feedstuffs, the feed pad and ends of teats were also collected and analysed for the presence of fungi. RESULTS: Eleven of 420 cows were diagnosed with intramammary infections (IMI) caused by yeasts (nine cows, 10 quarters) and moulds (two cows, three quarters). Six of the yeast species had previously been reported as being responsible for mastitis. Elevated somatic cell counts (SCC) were observed in many quarters, but most infections were eliminated spontaneously. Two of the fungi isolated from milk samples were also isolated from feedstuffs and teat swabs, and seven other fungi isolated from milk samples were not isolated from feed, the feed pad or cows' teats. CONCLUSIONS: Isolation of fungi from the udder is rarely reported in dairy cows in New Zealand. In this herd, contamination of the end of the teat originating from feedstuffs and possibly exacerbated by the use of a feed pad may have led to the establishment of IMI caused by fungi. CLINICAL RELEVENCE: Fungi are infrequently if ever reported in mastitis trial data or surveys in New Zealand and are probably of little clinical significance. KEY WORDS: Fungi, yeast, mould, bovine, intramammary infection, somatic cell count, mastitis.     

The use of polyethylene intramammary device in protection of the lactating bovine udder against experimental infection with Staphylococcus aureus or Streptococcus agalactiae.

The susceptibility of lactating bovine udder quarters fitted with a polyethylene intramammary device to infection was investigated. Following experimental challenge with Streptococcus agalactiae or Staphylococcus aureus, the incidence of infection was significantly (p less than 0.05) lower in intramammary device-fitted quarters compared to control quarters. In general, total foremilk and strippings milk somatic cell counts for intramammary device-fitted and control quarters were not significantly (p less than 0.05) different. Differential foremilk and strippings milk somatic cell counts were significantly (p less than 0.05) higher in samples from intramammary device-fitted quarters compared to control quarters.     

Physiological and morphological changes in bovine mammary glands following intramammary infusion of recombinant interferon-gamma.

Eleven lactating dairy cows were used to evaluate the response of bovine mammary glands to increasing doses of recombinant bovine interferon (rBoIFN)-gamma. Right front and rear quarters were intramammarily infused with eight different doses (10(2) U to 2 x 10(8) U/quarter) of rBoIFN-gamma; each dose was tested in at least two quarters. Left udder halves served as within animal controls in which quarters were injected with a saline placebo or were not infused at all. Milk secretion samples for compositional analysis were collected from each quarter prior to infusion and at 6, 24, 36 and 48 h following infusion. Animals were slaughtered immediately following the 48 h sampling period and mammary tissue was obtained for morphometric analyses. Milk composition was similar between control quarters and those quarters infused with up to 10(5) U of rBo-IFN-gamma during the entire sampling period. Quarters infused with 10(6) U and 10(7) U of rBoIFN-gamma had higher milk somatic cell counts (SCC) following treatment compared with preinfusion values. Changes in the composition of mammary secretion were most dramatic in quarters infused with greater than or equal to 10(8) U of rBoIFN-gamma as indicated by the significant increase in SCC and milk pH with a concomitant decrease in lactose concentration when compared with pre-infusion values or with control quarters. Morphometric analysis of tissue demonstrated an increase in stroma, a decrease in luminal area, and a marked increase in the number of infiltrating leukocytes in those quarters infused with the higher doses of rBoIFN-gamma.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characteristics of an unusual mycoplasma isolated from a case of caprine mastitis and arthritis with possible systemic manifestations

A mycoplasma designated strain GM790A was isolated from milk and internal organs of 2 lactating goats showing mastitis and arthritis. The isolate was not related serologically to any of the currently known ovine-caprine mycoplasmas, except an isolate designated Mycoplasma sp. G, first recorded from the external ear canal of clinically normal goats in Australia. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and restriction enzyme DNA studies of strain GM790A and Mycoplasma sp. G revealed similar but not identical patterns. The inoculation of strain GM790A into the teat canal of 2 lactating goats resulted in an abrupt diminution of lactation leading to mastitis and agalactia in about 3 days. A maximum of 1 x 10(7) colony-forming units (CFU) of the mycoplasma were shed per ml of mammary secretion. Milk production partially resumed at a low level 3 weeks postinoculation, the longest period tested, but the milk still contained 1 x 10(2) CFU of the agent. The results of this study indicate that strain GM790A possesses pathogenic potential for the goat and most probably represents a new species of the genus Mycoplasma.     

Effect of abraded intramammary device on outcome in lactating cows after challenge exposure with Streptococcus uberis

Intramammary devices (IMD) were abraded with medium-grade emery cloth or were left smooth. One IMD of each type was inserted into a mammary quarter of each of 5 lactating cows. The remaining 2 quarters served as controls. Quarter foremilk, bucket milk, and stripping milk samples were collected for 3 consecutive days at 2 weeks after IMD insertion, and milk somatic cell counts (SCC) were determined. Milk samples also were collected immediately after and 0.5, 1, 2, 4, 6, 8, and 11 hours after milking. All quarters were challenge exposed with 250 colony-forming units of Streptococcus uberis at 2 months after IMD insertion. Foremilk and stripping milk samples were collected for bacteriologic culture and SCC at the next 10 milkings. Mean foremilk, bucket milk, and stripping milk SCC (X 10(6) cells/ml) were 0.18, 0.07, and 0.91, respectively, for quarters with abraded IMD; 0.06, 0.05, and 0.43, respectively, for quarters with smooth IMD; and 0.03, 0.03, and 0.15, respectively, for control quarters. Mean SCC after milking (X 10(6) cells/ml) for the various intervals were 0.70, 1.29, 0.70, 0.97, 1.15, 1.17, 0.77, and 0.85 for quarters with abraded IMD; 0.43, 0.62, 0.61, 0.45, 0.64, 0.60, 0.31, and 0.26 for quarters with smooth IMD; and 0.15, 0.24, 0.15, 0.19, 0.15, 0.15, 0.14 and 0.06 for control quarters. After challenge exposure, 2 of 5 of the quarters with abraded IMD, 4 of 5 of the quarters with smooth IMD, and 8 of 9 control quarters became infected. Results indicated that abraded IMD increased SCC in stripping milk to concentrations that provided 60% protection against challenge exposure with S uberis.     

A comparative field trial of cephalonium and cloxacillin for dry cow therapy for mastitis in Australian dairy cows

OBJECTIVE: To investigate and compare the therapeutic efficacy of dry cow agents containing either cephalonium or cloxacillin within Australian dairy herds. DESIGN: A treatment-control trial. METHODS: Milk from infected quarters of cows with high somatic cell counts in milk on eight Australian dairy farms was cultured to identify bacterial pathogens. Cows were randomly assigned to treatment groups and one group was treated with cephalonium at drying off and the other group was treated with cloxacillin at drying off. Milk samples from infected quarters were collected immediately after calving and were cultured for pathogens. The effect of treatment on bacteriological cure was examined and somatic cell counts from infected cows from the first two herd tests after calving were examined for a treatment effect. On four farms, milk samples were collected for culture from all cases of clinical mastitis identified within the first 7 days after calving. The effect of treatment upon incidence of clinical mastitis after calving was examined. RESULTS: There was no significant difference between treatments on quarter cure rates for new infections, for chronic infections and for infections with Staphylococcus aureus, Streptococcus agalactiae and Streptococcus uberis. Infected quarters treated with cephalonium had a significantly higher cure rate than quarters treated with cloxacillin when Corynebacterium bovis and Staphylococcus epidermids were included as pathogens combined (80.3% versus 70.7%). There was no significant difference between the treatments on somatic cell counts of infected cows at the first two herd tests after calving. There was no difference between treatments on the incidence of clinical mastitis in the first 7 days after calving.