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.     

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.     

Prevalence and effects of intramammary infection in beef cows

Prevalence and effects of intramammary infection in 322 beef cows was determined during three calving intervals. Intramammary infection was confirmed in 37% of cows and 18.1% of quarters. Coagulase-positive staphylococci accounted for 17.9% of infections with Staphylococcus aureus isolated from 7.1% of cows. Coagulase-negative staphylococci and micrococci accounted for the remainder of infectious organisms. Butterfat and total protein levels were reduced 27.3 (P less than .05) and 25.5% (P less than .01), respectively, in milk from quarters infected with S. aureus. Somatic cell counts were elevated (P less than .001) with 3,827 X 10(3) cells/ml for S. aureus-infected quarters as compared with 555 X 10(3) cells/ml for uninfected quarters. Somatic cell counts were negatively correlated with 210-d calf weaning weights. Staphylococcus aureus-infected cows weaned calves weighing 19.1 kg less (P less than .01) than uninfected cows. At a present market value of $1.65/kg, economic losses were placed at $31.43/calf from cows infected with S. aureus in one or more quarters.     

Vaccination of cows with rough Escherichia coli mutants fails to protect against experimental intramammary bacterial challenge

Vaccination of cows with rough Escherichia coli mutants fails to protect against experimental intramammary bacterial challenge. Vaccine A, a heat-killed Re mutant of strain K12, (UB 1574), was administered as a single parenteral and local dose to 5 cows with 3 control animals and Vaccine B, a heat-killed mutant of O111:B4 (J5) was administered as three parenteral doses into 5 cows with 5 control animals. Following intramammary challenge with a smooth wild-type strain (P4), an acute, severe clinical mastitis developed in all 14 quarters (9 vaccine A and 5 vaccine B) of the vaccinated animals which was indistinguishable from that in the 11 quarters of the control animals. Following vaccine B there was an elevation in serum IgG1 and IgG2 antibody to the common core antigen of endotoxin which, in contrast to the control animals, showed a further increase after intramammary infection.     

The effect of polyethylene intramammary devices on the somatic cell count and milk production of dairy cows

SUMMARY Polyethylene intramammary devices (IMO) were inserted into all 4 quarters of 15 multiparous dairy cows. Fifteen cows, matched for parity and production, were controls. The insertion of IMD's was easily achieved and produced no adverse effects. Throughout the 150-day test period, the mean somatic cell count of cows with IMD's in situ was 216,000 cells per ml, compared with 119,000 cells per ml in controls (P<0.01). Total production of milk, butterfat and protein was not significantly different between the 2 groups. The new infection rate was too low to allow assessment of the value of the IMD for mastitis prevention.     

Somatic cell count in milk of selenium-supplemented dairy cows after an intramammary challenge with Staphylococcus aureus

The objective of this study was to evaluate the effect of selenium (Se) supplementation on milk somatic cell count (SCC) in dairy cows. Twelve multiparous Holstein-Friesian cows were fed a diet containing a suboptimal Se concentration (<0.05 ppm, dry basis) starting 2 months before calving. Supplemented cows (n=6) received a single s.c. injection of barium selenate (1 ml/50 kg BW) 45 days prior to calving, whereas control group was kept unsupplemented. Twenty weeks after calving, two mammary quarters (right side) of each cow were challenged with 205,000 cfu/ml of Staphylococcus aureus (strain Newbould 305). Blood was collected bi-weekly until day 150 of lactation for the analysis of blood glutathione peroxidase (GPx1; EC 1.11.1.9) activity. To re-isolate the challenging pathogen and to evaluate SCC, aseptic milk samples were collected daily starting on the day of challenge, and finishing 7 days after inoculation. Unsupplemented cows had a lower activity of GPx1 through the experiment (P<0.001). Natural log SCC (lnSCC) was higher in unsupplemented than Se-supplemented cows (P=0.04), showing evidence of significance after 5 days. Selenium supplementation of dairy cows fed a diet containing a suboptimal Se concentration, resulted in higher blood activity of GPx1, and lower mean lnSCC after an intramammary challenge with Staph. aureus.     

Efficacy of extended pirlimycin therapy for treatment of experimentally induced Streptococcus uberis intramammary infections in lactating dairy cattle

Streptococcus uberis is an important cause of mastitis in dairy cows throughout the world, particularly during the dry period, around the time of calving, and during early lactation. Strategies for controlling S. uberis mastitis have not received adequate research attention and are therefore poorly defined and inadequate. Objectives of the present study were to evaluate the efficacy of extended therapy regimens with pirlimycin for treatment of experimentally induced S. uberis intramammary infections in lactating dairy cows during early lactation and to evaluate the usefulness of the S. uberis experimental infection model for evaluating antimicrobial efficacy in dairy cows. The efficacy of extended pirlimycin intramammary therapy regimens was investigated in 103 mammary glands of 68 dairy cows that became infected following experimental challenge with S. uberis during early lactation. Cows infected with S. uberis in one or both experimentally challenged mammary glands were randomly allocated to three groups, representing three different treatment regimens with pirlimycin, including 2-day (n = 21 cows, 31 mammary quarters), 5-day (n = 21 cows, 32 quarters), and 8-day (n = 26 cows, 40 quarters). For all groups, pirlimycin was administered at a rate of 50 mg of pirlimycin hydrochloride via intramammary infusion. A cure was defined as an experimentally infected mammary gland that was treated with pirlimycin and was bacteriologically negative for the presence of S. uberis at 7, 14, 21, and 28 days after treatment. Experimental S. uberis intramammary infections were eliminated in 58.1% of the infected quarters treated with the pirlimycin 2-day regimen, 68.8% for the 5-day regimen, and 80.0% for the 8-day regimen. Significant differences (P <.05) in efficacy were observed between the 2-day and 8-day treatment regimens. The number of somatic cells in milk decreased significantly following therapy in quarters for which treatment was successful in eliminating S. uberis. However, there was no evidence to suggest that extended therapy with pirlimycin resulted in a greater reduction in somatic cell counts in milk than the 2-day treatment. The S. uberis experimental infection model was a rapid and effective means of evaluating antimicrobial efficacy during early lactation at a time when mammary glands are highly susceptible to S. uberis intramammary infection.     

Efficacy of vaccination and antimicrobial treatment to eliminate chronic intramammary Staphylococcus aureus infections in dairy cattle

OBJECTIVE: To determine whether a combination of vaccination and extended intramammary antimicrobial treatment would eliminate chronic intramammary Staphylococcus aureus infections in lactating dairy cows. DESIGN: Randomized controlled clinical trial. ANIMALS: 50 dairy cows with chronic mastitis caused by S aureus. PROCEDURE: Cows were identified and paired within herd on the basis of days in milk, lactation number, milk production, and numbers of quarters infected. Treated cows (n=20) received 3 doses of a polyvalent S aureus bacterin on days 1, 15, and 21 of the study along with intramammary administration of pirlimycin in all 4 quarters once daily for 5 treatments (days 16 to 20). Control cows (n=23) received no treatment. Follow-up samples for bacteriologic culture were collected for at least 3 months after treatment to determine treatment success rates. RESULTS: Significantly more S aureus infections were eliminated from treated cows (8/20 [40%]), compared with control cows (2/23 [9%]). The proportion of infected quarters that yielded negative results throughout the follow-up period was also significantly higher in treated cows (13/28 [46%]) than in control cows (2/41 [5%]). CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that a combination of vaccination and antimicrobial treatment can be successful in eliminating some cases of chronic intramammary S aureus infections in dairy cattle. However, it is important to consider extended treatment protocols carefully because many cows are likely to remain infected with S aureus despite treatment and vaccination.     

Effects of intramammary infection and parity on calf weaning weight and milk quality in beef cows

The objectives of this study were to determine 1) the effect of intramammary infection on calf weaning weight, milk somatic cell count, and milk composition, and 2) the effect of parity on percentages of infected cows, infected quarters, and blind quarters. The number of infected quarters, milk somatic cell counts, milk components, and intramammary infection were studied at weaning in 164 beef cows. The percentage of infected cows ranged from 61.9% at first parity to 66.7% at fifth to ninth parities. Cows with three or four infected quarters had higher (P < .01) milk somatic cell counts than cows with zero, one, or two infected quarters. Among bacterial isolates, Staphylococcus aureus-infected quarters had the highest (P < .01) milk somatic cell count. Percentages of butterfat and lactose were lower (P < .01) in milk from infected quarters than from uninfected quarters. Infections by S. aureus and coagulase-negative staphylococci were the most common and accounted for 67 to 78% of the infections. Percentages of infected quarters and infections caused by S. aureus increased with parity (P < .01). Intramammary infections did not affect (P > .10) calf weaning weight. In conclusion, intramammary infection had no effect on calf weaning weight but increased milk somatic cell count and decreased the percentage of protein, lactose, solids-not-fat, and butterfat. The number of infected and blind mammary quarters increased with parity.     

Detection of capsular polysaccharide in milk of cows with natural intramammary infection caused by Staphylococcus aureus

Detection of capsular polysaccharide (CP) in milk of cows with natural intramammary infection caused by Staphylococcus aureus was attempted. Five quarters of 5 cows harboring S aureus strains that produce type-8 CP were selected. Using an ELISA with a monoclonal antibody, type-8 CP was not detected in extracts prepared from fresh milk collected aseptically. By contrast, CP was easily detectable after incubation of infected milk at 38 C for 20 hours. Quantitation of CP in extracts from incubated milk samples by use of ELISA indicated a great variation of CP expression by strains. Although an incubation step was necessary to detect CP, results of the study indicate that CP may be expressed in vivo during intramammary infection caused by S aureus.     

Effect of a milk-derived factor on the inflammatory response to Staphylococcus aureus intramammary infection.

Daily injections of an anti-inflammatory milk-derived factor (MDF) into mice increased resistance to Staphylococcus aureus challenge, and reduced leukocyte infiltration. Intraperitoneal injection of MDF into lactating mice prior to S. aureus intramammary challenge resulted in greater milk secretory activity and less inflammation compared with untreated controls, but had little effect on the number of S. aureus recovered from mammary tissue. Infusion of MDF directly into mouse mammary glands prior to challenge reduced S. aureus recovered after challenge. Incubation of bovine mammary macrophages in medium supplemented with MDF enhanced phagocytosis of opsonized S. aureus. In addition, infusion of 5 mg MDF into uninfected bovine mammary glands 24 h prior to S. aureus challenge resulted in fewer infections (five of ten) than in control quarters (seven of nine). Repeated daily injections of 5 mg MDF into S. aureus-infected quarters increased the percent of mammary neutrophils and decreased the recovery of S. aureus, but did not eliminate infections. Intravenous injection of 8 g MDF into cows resulted in pronounced leukopenia while the accompanying effect on mammary leukocytes was less marked but followed a similar course. Results suggest that the use of MDF in mice enhanced resistance to experimental infection and was beneficial in maintaining mammary secretory activity and reducing inflammation after bacterial challenge. In the cow, MDF promoted phagocytosis in vitro and was effective against challenge when infused intramammarily.     

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.     

The ability of the enzyme-linked immunosorbent assay to detect antibody against Staphylococcus aureus in milk following experimental intramammary infection

Changes in the milk antibody levels against Staphylococcus aureus were measured at the start of an experimental intramammary instillation of either S. aureus (Study I) or Staphylococcus hyicus (Study II). A commercial enzyme-linked immunosorbent assay system was used. Twenty-one Holstein cows were enrolled in Study I and 15 Holstein cows were used in Study II. Pathogen instillation began 21 days before the start of the non-lactating period. Cows received intramammary antibiotic treatment in all quarters immediately after the last milking, the start of the non-lactating period. Lacteal secretions were collected before the start of the non-lactating period, and during the immediate postpartum period in both studies, and during the non-lactating period in Study I. Milk was cultured for mastitis pathogens and S. aureus antibody levels and somatic cell counts were determined from all samples. There was an approximate 2-week delay in the elevation in antibody levels in response to the instillation of S. aureus. Antibody levels remained elevated in cows with S. aureus intramammary infections postpartum, but were below threshold in cows where intramammary infections were cured during the non-lactating period. Antibody levels were elevated by S. hyicus intramammary infections, remained elevated for the first 12 days postpartum, but were below threshold by day 21 postpartum. Cows with incipient intramammary S. aureus infections might be misclassified as false negatives by the antibody test. However, results suggest that cows with S. hyicus intramammary infections that were not cured would not be misclassified if milk is withheld from test for the first 30 days postpartum, as recommended by the manufacturer of the test.     

Experimental intramammary inoculation with Mycoplasma bovis in vaccinated and unvaccinated cows: effect on the mycoplasmal infection and cellular inflammatory response

The effect of vaccination on mycoplasmal infection and the cellular inflammatory response was evaluated in 4 vaccinated and 4 control cows experimentally challenged in 2 of 4 quarters with live Mycoplasma bovis. In unchallenged quarters during the first three weeks after experimental challenge exposure, 6 of 8 quarters on control cows, and 7 of 8 quarters on vaccinated cows became infected with low numbers (10(2)-10(4) cfu/ml) of M bovis. During the same period all challenge-infused quarters on both control and vaccinated animals became infected with high numbers (10(9) cfu/ml) of M bovis. Thereafter, all quarters on vaccinated cows became culture-negative for M bovis, while 2 of 8 unchallenged quarters, and 4 of 8 challenged quarters on 3 of 4 control cows remained infected. A cellular inflammatory response as measured by the California Mastitis Test accompanied the experimental infection in proportion to the infection level except in challenged quarters on vaccinated cows after the first three weeks post challenge in which the cellular inflammatory response remained high despite the advent of negative M bovis culture results. This study indicates that the course of experimental M bovis mastitis can be affected by vaccination, and that vaccination results in an adverse cellular inflammatory response in challenged quarters.     

Milk leucocyte population patterns in bovine udder infection of different aetiology

This study compared the different leucocyte populations in milk from udders infected with different mastitic pathogens and in different stages of infection. Milk samples were collected from quarters free of intramammary infection, acutely infected with Escherichia coli or Staphylococcus aureus and chronically infected with S. aureus, coagulase-negative staphylococci (CNS) or Streptococcus dysgalactiae. Udder bacteriological status was confirmed after three consecutive bacteriological examinations from weekly quarter milk samples. At the time of the trial, milk samples were tested for somatic cell count (SCC) and differential cell count by both light microscopy (LM) and flow cytometry. Monoclonal antibody (mAb) CD11a/CD18 was used in order to differentiate between leucocytes and epithelial cells when tested by flow cytometry. Udder quarters free of intramammary infection had a mean SCC lower than 107 x 10(3) cells/ml in which the epithelial cells were the main cell type followed by polymorphonuclear cells (PMNs), while macrophages and lymphocytes had a lower concentration. Only 56% of the cells were labelled with the mAb anti-CD11a/CD18. In either acute E. coli- or S. aureus-infected quarters, SCC were significantly higher (P < 0.0001) than in samples from the time of inoculation, with over 90% of the cells labelled with the mAb anti-CD11a/CD18. The main cell type was neutrophils. In chronically infected cows, differences in SCC and in leucocyte patterns were found between infecting pathogens as well as between quarters harbouring the same pathogen. In all the chronically infected quarters, SCC was significantly higher (P < 0.05) than in uninfected ones. The distribution of the leucocyte patterns in the quarters infected with S. dysgalactiae did not differ from that in quarters with acute infection with both E. coli and S. aureus. In the cows chronically infected with S. aureus or CNS, the proportion of PMN was higher but not significantly different from quarters free of intramammary infection, while epithelial cells were significantly lower (P < 0.05). The T lymphocytes bearing CD4+ or CD8+ were significantly higher in quarters chronically infected with S. aureus than in quarters free of intramammary infection and in quarters acutely infected with either E. coli or S. aureus. In all samples B cells were negligible.     

Effect of intramammary infusion of bacterial lipopolysaccharide on experimentally induced Staphylococcus aureus intramammary infection

Mastitis due to Staphylococcus aureus is a significant problem in the dairy industry and is refractory to antibiotic treatment and/or vaccine prevention. Relative to other mastitis-causing pathogens, S. aureus elicits a diminutive host inflammatory response during intramammary infection. To determine whether induction of a heightened inflammatory response could influence outcome of infection, the highly pro-inflammatory molecule bacterial lipopolysaccharide (LPS) was infused into udder quarters experimentally infected with S. aureus. Relative to S. aureus-infected udder quarters receiving saline, quarters infused with LPS demonstrated a heightened inflammatory response as demonstrated by the induction of TNF-alpha and higher milk somatic cell counts and albumin levels. Although there was no overall effect on bacterial clearance, a trend toward reduced bacterial numbers during the immediate pro-inflammatory response following LPS infusion was observed suggesting that this novel approach to treating S. aureus intramammary infection may warrant further investigation.     

Efficacy of extended pirlimycin hydrochloride therapy for treatment of environmental Streptococcus spp and Staphylococcus aureus intramammary infections in lactating dairy cows

Fifty-one chronically infected lactating dairy cows were used to evaluate the efficacy of extended pirlimycin therapy regimens for treatment of intramammary infections by environmental Streptococcus spp and Staphylococcus aureus. Cows (n = 47) with one or more infected mammary quarters were blocked by parity and randomly allocated to one of three groups for treatment with pirlimycin (50 mg/mammary quarter) as follows: one treatment per day for 2 days (n = 36 infected mammary quarters); one treatment per day for 5 days (n = 36 infected mammary quarters); and one treatment per day for 8 days (n = 20 infected mammary quarters). Four cows with nine infected mammary quarters were included as untreated controls. Milk samples from each mammary quarter were collected 7 days before treatment, immediately before treatment, and weekly for 4 weeks after the final treatment for microbiological evaluation. A bacteriologic cure was defined as a treated, infected quarter that was bacteriologically negative for the presence of previously identified bacteria at weekly intervals after treatment. Efficacy of pirlimycin therapy against intramammary infections caused by environmental Streptococcus spp and S. aureus was 44.4%, 61.1%, and 95.0% for the 2-, 5-, and 8-day treatment regimens, respectively. None of the infections in the untreated control quarters was cured. Significant differences in efficacy were detected between all pirlimycin groups and the untreated control group, between the 8- and 2-day treatment regimens, and between the 8-day and 5-day treatment regimens (P < or = .05). Results of this study indicate that extended pirlimycin therapy was effective in eliminating intramammary infections caused by environmental streptococci and S. aureus in lactating dairy cows.     

Relationships between teat-end bacteria and intramammary infections

Teat-end populations of staphylococci and streptococci were studied on teat apices of an experimental herd. Individual cows were found to have higher numbers of some of these microorganisms than other cows and certain teats had consistently higher populations than other teats in the same individual. A positive relationship was found between the numbers of staphylococci on teat-ends and presence of intramammary infections of the quarters. High teat-end bacterial numbers were present prior to the laboratory recognition of intramammary infections.     

The effect of intramuscular and intramammary vaccination of cows on antibody levels and resistance to intramammary infection by Staphylococcus aureus.

Cows were vaccinated simultaneously by the intramuscular and intramammary route with formolised Staphylococcus aureus cells of strains BB, Mexi and 3528. Vaccination resulted in slight increases in serum agglutinin titres, but the levels of the agglutinins in milk and colostrum were not higher in vaccinated cows than in the unvaccinated controls. Vaccination did not result in higher levels of IgM, IgG1, IgG2 or IgA immunoglobulins in serum, colostrum or milk as compared with controls. Vaccinated cows, particularly those in which strain 3528 was used, showed some resistance to infection following challenge with low numbers of viable S aureus Mexi, but there was no resistance to infection when similar numbers of the virulent BB strain were used for challenge. It is concluded that vaccination is unlikely to prevent infection of the bovine udder by S aureus.     

An intramammary implant in the prevention of infectious mastitis in cows

A sterile polyethylene device was introduced in the milk cistern of two mammary quarters of forty dairy cows. The cows were divided into three groups according to the length of exposure of the device in the cistern (3 days, 14 days, 365 days). The somatic cell counts were studied for a year in the first 10 ml fraction of milk and an increase in the somatic cell counts was found in the quarters having the intramammary device, as compared with the control quarters (having no devices). This difference was marked during lactation and prior to the onset of drying off. For the reduction in the frequency of occurrence of new natural intramammary infections, the activity of the device against S. aureus was limited and no activity against S. agalactiae was proved. The proportions of polymorphonuclears, round-cell elements and macrophages were histologically studied and compared for the udder quarters with and without the intramammary body in correlation with the time of exposure of the device to the milk cistern milieu. The most marked differences in favour of the udder quarters with the intramammary device were recorded in the alveoli containing more cells with a significant proportion of polymorphonuclear leucocytes. Small differences were found in the interstitial and subepithelial zone of the milk cistern. The activity of acid and alkaline phosphatase and adenosine triphosphatase was histochemically determined in the tissue structures of the udder and was found not to change under the influence of the device. Leucocytes and macrophages adhering to the surface of the body were observed under scanning electron microscope.