Depletion of CD4 T lymphocytes at the time of infection with M. avium subsp. paratuberculosis does not accelerate disease progression.

A calf model was used to determine if the depletion of CD4 T cells prior to inoculation of Mycobacterium avium subsp. paratuberculosis (Map) would delay development of an immune response to Map and accelerate disease progression. Ileal cannulas were surgically implanted in 5 bull calves at 2 months of age. Two calves were depleted of CD4 T cells by intravenous injection of anti-bovine CD4 antibody administered 24h prior to inoculation with Map. The two CD4-depleted calves and one non-depleted calf were inoculated via ileal cannula with 1 × 10(8)cfu live Map every 3 days for a total of 4 inoculations. Two additional calves served as non-depleted and uninfected controls. Injection with the anti-CD4 mAb reduced the frequency of CD4 T cells from a pre-depletion average of 15% to less than 1% in PBMC at 24h. However, a consistent proliferative response dominated by CD4 T cells, developed in both treated and untreated calves over the course of the 6-month study period. Recovery of Map from serial biopsies obtained from the CD4-depleted and non-depleted calves after Map infection did not differ. In addition, CD4 depletion did not increase the level of Map shed in the feces over the non-depleted animal.

Dynamics of bovine spleen cell populations during the acute response to Babesia bovis infection: an immunohistological study.

The spleen is a critical organ in defence against haemoparasitic diseases like babesiosis. Many in vitro and ex vivo studies have identified splenic cells working in concert to activate mechanisms required for successful resolution of infection. The techniques used in those studies, however, remove cells from the anatomical context in which cell interaction and trafficking take place. In this study, an immunohistological approach was used to monitor the splenic distribution of defined cells during the acute response of naïve calves to Babesia bovis infection. Splenomegaly was characterized by disproportionate hyperplasia of large versus small leucocytes and altered distribution of several cell types thought to be important in mounting an effective immune response. In particular, the results suggest that the initial crosstalk between NK cells and immature dendritic cells occurs within the marginal zone and that immature dendritic cells are first redirected to encounter pathogens as they enter the spleen and then mature as they process antigen and migrate to T-cell-rich areas. The results of this study are remarkably similar to those observed in a mouse model of malarial infection, suggesting these dynamic events may be central to the acute response of naïve animals to haemoparasitic infection.

Escherichia coli isolates' serotypes, genotypes, and virulence genes and clinical coliform mastitis severity.

Dairy cattle with clinical mastitis caused by Escherichia coli exhibit a wide range of disease severity, from mild, with only local inflammatory changes of the mammary gland, to severe, with significant systemic derangement. The present study was designed to examine the relationship between serotype and virulence genes of E. coli mastitis isolates, different levels of systemic disease severity, and farm from which the E. coli strain was obtained. One hundred twenty-three E. coli milk isolates were obtained from cows with clinical mastitis of varying systemic disease severity from 6 different farms. No predominant serotype was identified by farm or by systemic disease severity; however, the most frequent serotype, O158:NM (n = 3), was isolated from cows in the moderate severity group. Virulence genes evaluated were identified infrequently and were not associated with systemic disease severity. Evaluation of genetic similarity showed no clustering assigned by farm or mastitis severity based on systemic disease signs. We concluded that a high degree of genotypic variability is characteristic of E. coli strains causing clinical mastitis within and between different farms and systemic severity groups, and that specific cow factors probably play a more important role in determining systemic disease severity.

Biosecurity and biocontainment in alpaca operations.

Biosecurity on South American camelid operations involves both external and internal measures to prevent the introduction and spread of disease. External biosecurity involves practices and techniques directed at the prevention of entry of new diseases into a group of animals. Internal biosecurity or biocontainment, involves practices and techniques that are directed at the prevention or spread of disease within an existing group of animals. External biosecurity is particularly important in North America camelid operations due to the extensive movement of animals for breeding or show purposes. Internal biosecurity typically involves this the prevention and treatment of failure of passive transfer, maintenance of proper nutrition and housing, and the implementation of an appropriate vaccination program for endemic or relevant diseases. Attention to appropriate cleaning and disinfection procedures related to housing, feeding, and treatment equipment is important for the maintenance of both internal and external biosecurity practices. This paper discusses various risk factors associated with the control of infectious disease in the context of external and internal biosecurity measures in camelids operations.

The innate immune response in calves to Boophilus microplus tick transmitted Babesia bovis involves type-1 cytokine induction and NK-like cells in the spleen.

The innate immune response to Babesia bovis infection in cattle is age-related, spleen-dependent and, in stabilate inoculated calves, has type-1 characteristics, including the early induction of IL-12 and IFN-gamma. In this study with three calves, parameters of innate immunity were followed for 2 weeks after tick transmission of B. bovis. Each calf survived the acute disease episode without drug intervention, and responded with increased levels of plasma interferon-gamma and type-1 cytokine expression, monocyte/macrophage activation, and CD8+ cellular proliferation in the spleen. The proliferating CD8+ population consisted primarily of NK-like cells, and the expansion occurred in parallel with an increase in IL-15 mRNA expression in the spleen.

Age-related innate immune response in calves to Babesia bovis involves IL-12 induction and IL-10 modulation.

There is a strong innate immunity in calves to infection with Babesia bovis. Interleukin (IL)-12 and IL-10 have been shown in vitro to be important immunoregulatory cytokines. Here we demonstrate in vivo that the protective innate response in young calves to infection with virulent B. bovis involves the early appearance of IL-12 and interferon-gamma (IFN-gamma) transcripts in the spleen. In contrast, IL-12 and IFN-gamma mRNA expression in the spleens of adult cattle that succumbed to the infection was delayed and depressed and occurred within the context of IL-10 expression. Also in contrast with calves, there was no detectable antibody response before death in adults. A vigorous CD8+ T-cell expansion occurred in the spleens of both calves and adults.

IL-4 and IL-10 inhibition of IFN-gamma- and TNF-alpha-dependent nitric oxide production from bovine mononuclear phagocytes exposed to Babesia bovis merozoites.

The requirement for IFN-gamma and/or TNF-alpha as co-stimulants with Babesia bovis merozoites for nitric oxide (NO) production was examined, as well as the regulatory role of IL-4 and IL-10. Purified B. bovis merozoites did not induce the production of NO in undifferentiated monocytes without addition of exogenous IFN-gamma and TNF-alpha unless the monocytes taken ex vivo were producing TNF-alpha endogenously. Under the latter condition, the NO production resulting from merozoite stimulation remained IFN-gamma-dependent. There was no evidence for endogenous synthesis of TNF-alpha in monocyte-derived macrophages (MDM), and merozoites alone were incapable of inducing TNF-alpha mRNA in MDM. However, while merozoites plus IFN-gamma induced TNF-alpha mRNA expression in MDM, NO was not produced. Both IL-4 and IL-10 inhibited expression of iNOS and production of NO in merozoite-stimulated monocytes.

Bovine neonatal immunology.

The majority of early, in utero immune development occurs independent of antigen exposure. Only later during development can a fetus respond to antigens, and even then the response depends on the stage of fetal development and the nature of the antigen. At birth, the neonate is rapidly exposed to large numbers of potential pathogens. Although immunocompetent, the neonate is immunonaive and dependent on passively acquired maternal immunoglobulins, immune cells, and other substances from colostrum for protection. Neonates that suffer failure of passive transfer of maternal immunoglobulins may be at increased risk for disease; however, many other factors interact in conjunction with the level of passively acquired immunoglobulin to determine the occurrence of disease. These include, but are not limited to, management, environment, hygiene, infection pressure, virulence of organisms, and antibody specificity. In addition to immunoglobulins, colostrum contains large numbers of immune cells and cytokines. It is thought that the primary role for the cellular component of colostrum is to interact with the development of local immunity and to modulate active immunization of the neonatal intestine. In particular, T lymphocytes are thought to transfer immune functions and secrete cytokines. Although most of the major cytokines have been identified in colostrum and milk, their biologic effects on the neonate have yet to be determined.

Bacteremia associated with naturally occuring acute coliform mastitis in dairy cows.

OBJECTIVE: To determine the incidence of bacteremia in dairy cows with naturally occurring acute coliform mastitis (ACM) with a wide range of disease severity. DESIGN: Cohort study. ANIMALS: 144 dairy cows with ACM from 6 herds. PROCEDURE: Cows were examined at time of identification of ACM (time 0) and classified as having mild, moderate, or severe mastitis on the basis of rectal temperature, hydration status, rumen contraction rate, and attitude. Cows were reexamined at 24 or 48 hours. Bacteriologic culturing of milk and blood (30 ml), CBC, and serum biochemical analysis were performed at each time point. Appropriate samples were obtained at a single point from herdmates without mastitis (controls) that were closely matched for lactation number and days since parturition. Blood culture results were compared among severity groups and controls by use of chi2 tests, as was outcome of an ACM episode for cows grouped by blood bacterial isolates. RESULTS: Bacteria were isolated from 52 blood samples from 46 of 144 (32%) cows with ACM, which was significantly more than control cows (11/156; 7.1%). Group-1 isolates (Escherichia coli, Pasteurella multocida, Mannheimia haemolytica, Klebsiella pneumoniae, Enterobacter agglomerans, and Salmonella enterica serotype Typhimurium) were identified in 20 of 144 (14%) cows with ACM and 0 of 156 control cows. Group-1 isolates were identified in 4.3, 9.1, and 42% of cows classified as having mild, moderate, and severe ACM, respectively. Escherichia coli and K pneumoniae milk and blood isolates obtained from the same cow were of the same genotype. Bacillus spp were identified in 21 of 144 (15%) cows with ACM, which was significantly more than control cows (3/156; 1.9%). Thirty-five percent of cows with a group-1 isolate died during the mastitis episode. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that bacteremia develops in a substantial proportion of cows with ACM. Classification of severity of disease is important for establishment of effective treatment protocols; parenteral antimicrobial treatment may be indicated in cows with ACM.

The age-related immunity in cattle to Babesia bovis infection involves the rapid induction of interleukin-12, interferon-gamma and inducible nitric oxide synthase mRNA expression in the spleen.

Young calves possess a strong innate immunity against Babesia bovis infection that lasts for approximately 6 months after birth and is abrogated with the removal of the spleen. This immunity is characterized as cellular involving a soluble mediator. Nitric oxide has been implicated by virtue of its babesiacidal affects in vitro, but questioned to be as effective in vivo, due to its ability to downregulate type-1 immunity. Spleen cells were obtained from 4-month-old calves and adult steers and processed for monitoring cytokine and inducible nitric oxide synthase (iNOS) mRNA expression during the response to initial B. bovis infection. The data provided evidence of a transient role for nitric oxide in innate immunity, characterized by brief iNOS induction in the spleen of calves that was not detectable in the spleens of adults. The iNOS message followed the early induction of interleukin (IL)-12 and interferon (IFN)-gamma message in calves. The induction of IL-12 and IFN-gamma message in adults was delayed until IL-10 message was induced. Transformation growth factor-beta mRNA expression levels were greater in spleen cells from adults early in infection and then declined, whereas expression levels increased in spleen cells from calves later in the infection process. Together, the data support the concept of 'first come, first serve' cytokine influence over cellular activities, the importance of a type-1 response in the control of an initial infection and the need for tight regulation in order to prevent pathology associated with over production of nitric oxide and inflammatory cytokines.

Cutaneous metastases of a mammary carcinoma in a llama.

An 8-year-old, female llama was evaluated for nonhealing, ulcerative, cutaneous lesions, which also involved the mammary gland. Biopsies of the lesions distant from and within the mammary gland area revealed an aggressive carcinoma. The tumor was confirmed at necropsy to be a mammary gland adenocarcinoma with cutaneous metastasis.

Use of systemic disease signs to assess disease severity in dairy cows with acute coliform mastitis.

OBJECTIVE: To evaluate the use of systemic disease signs for classifying severity of acute coliform mastitis in dairy cows. DESIGN: Prospective cohort study. ANIMALS: 144 dairy cows. PROCEDURE: Cows were examined at the time of initial identification of disease (time 0) and classified as having mild, moderate, or severe disease on the basis of rectal temperature, hydration status, rumen contraction rate, and attitude. A CBC and serum biochemical analyses were performed, and milk samples were submitted for bacterial culture at time 0 and 48 hours later. RESULTS: 69 cows were classified as having mild disease, 44 as having moderate disease, and 31 as having severe disease. Median WBC and neutrophil counts were significantly lower in cows with moderate or severe disease at time 0 than in cows with mild disease. Band neutrophil count was significantly higher at 48 hours and serum calcium concentration was significantly lower at time 0 and at 48 hours in cows with severe or moderate disease, compared with cows with mild disease. Twenty-eight, 51, and 77% of cows with mild, moderate, and severe disease, respectively, had > 100,000 colony-forming units/ml of milk at time 0. The odds that a cow with severe disease would die or be culled were 3.6 times the odds for a cow with moderate disease and 11.2 times the odds for a cow with mild disease. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that a classification scheme based on readily observable systemic disease signs can be used to classify disease severity in cows with acute coliform mastitis.

Passive transfer of colostral immunoglobulins in calves.

Passive transfer of colostral immunoglobulins has long been accepted as imperative to optimal calf health. Many factors, including timing of colostrum ingestion, the method and volume of colostrum administration, the immunoglobulin concentration of the colostrum ingested, and the age of the dam have been implicated in affecting the optimization of absorption. The practice of colostrum pooling, the breed and presence of the dam, and the presence of respiratory acidosis in the calf also may affect passive transfer. Various tests have been reported to accurately measure passive transfer status in neonatal calves. The radial immunodiffusion and the enzyme-linked immunosorbent assay (ELISA) are the only tests that directly measure serum IgG concentration. All other available tests including serum total solids by refractometry, sodium sulfite turbidity test, zinc sulfate turbidity test, serum gamma-glutamyl transferase activity, and whole blood glutaraldehyde gelation estimate serum IgG concentration based on concentration of total globulins or other proteins whose passive transfer is statistically associated with that of IgG. This paper presents a comprehensive review of the literature of passive transfer in calves including factors that affect passive transfer status, testing modalities, effects of failure of passive transfer on baseline mortality, consequences of failure of passive transfer, and some treatment options. Many previously accepted truisms regarding passive transfer in calves should be rejected based on the results of recent research.

Septic orchitis in an alpaca.

An adult, intact male alpaca was presented with an acute onset of unilateral scrotal swelling. Following complete physical and ultrasonographic examination, the most likely differential diagnoses were orchitis, hematoma, and testicular torsion. The animal was castrated and histopathologic evaluation revealed unilateral orchitis. Streptococcus equi subsp. zooepidemicus was cultured.

Flow cytometric analysis of an immunodeficiency disorder affecting juvenile llamas.

The present study was undertaken to characterize the immune system of llamas and alpacas and establish the basis for an immunodeficiency disorder affecting juvenile llamas. Flow cytometric (FC) analysis of the immune system with a panel of monoclonal antibodies (mAbs) revealed the immune system of llamas and alpacas is similar in leukocyte subset composition to that in ruminants. Peripheral blood mononuclear cells in adults are comprised of surface immunoglobulin (sIg(+)) B-cells (31%+/-8 S.D.), alphabeta T-cells (27%+/-12 S.D.), WC1(+) gammadelta T-cells (16%+/-11 S.D.), and 5-16% monocytes. In contrast to cattle, goats, and sheep, however, the frequency of WC1(+) gammadelta T-cells is not high in juveniles but similar to the frequency in adults. Also, sIg(+) B-cells are present in high concentration in juveniles (43%+/-11 S.D. ). Expression of major histocompatibility class II molecules on resting T-cells was low or absent. Comparative analysis of peripheral blood lymphocyte composition in normal juvenile llamas and llamas presenting with the signs of the juvenile llama immunodeficiency syndrome (JLIDS) revealed the concentration of B-cells is extremely low (1-5%) in affected animals. The findings suggest JLIDS is attributable to an autosomal recessive genetic defect in the development of B-cells.

Regulation of the immunoglobulin G1 receptor: effect of prolactin on in vivo expression of the bovine mammary immunoglobulin G1 receptor.

Induction of colostrogenesis in non-pregnant cows was used to evaluate the relationship between prolactin (PRL) and mammary immunoglobulin G1 (IgG1) receptor expression. Six of eleven non-pregnant, non-lactating Holstein cattle responded to a standard lactation induction protocol by development of elevated IgG1 concentrations in mammary secretions. In order to increase the diversity in PRL concentrations, two of the six cattle were treated with bromocriptine, and two others were treated with recombinant bovine PRL. Serum alpha-lactalbumin, serum PRL and mammary secretion IgG1 concentrations were measured throughout the experiment. Biopsies of mammary tissue were collected after induction of lactation, and after treatments to alter serum PRL. Immunohistochemistry was used to evaluate IgG1 receptor expression. Administration of recombinant bovine (rbPRL) was associated with increased lactogenic activity, decreased secretion IgG1 concentrations, and decreased IgG1 receptor expression. Decreased serum PRL, due to bromocriptine, was associated with decreased lactogenic activity and maintenance of IgG1 receptor expression. Results of this experiment are consistent with an effect of PRL in decreasing the expression of the bovine mammary IgG1 receptor at the onset of lactogenesis.

Detection of low serum immunoglobulin concentrations in clinically ill calves.

The sensitivity, specificity, and accuracy of classification of 4 tests for failure of passive transfer (FPT) were examined in clinically ill neonatal calves. Comparisons were made with serum IgG1 concentrations determined by radial immunodiffusion. Serum samples were obtained from 27 clinically ill calves < or = 21 days of age. The results of 4 commonly used assays, the sodium sulfite turbidity test, the zinc sulfate turbidity test, refractometry, and the serum gamma-glutamyl transferase (GGT) activity test, were compared with radial immunodiffusion determinations of serum IgG1 concentration. Serum GGT activity using a 50 IU/L threshold resulted in correct classification of the highest percentage of calves (93%) with regard to their passive transfer status. The sodium sulfite test with a 1+ end point and refractometry using a 5.5 g/dL end point resulted in correct classification of 85% of the calves studied. When using the sodium sulfite test, the 2+ and 3+ test end points had lower specificity, 0.58 and 0.00, respectively, than the 1+ end point. This loss in specificity resulted in misclassification of calves with adequate serum immunoglobulin concentrations as having FPT. The zinc sulfate turbidity test was inadequately specific (0.33) and resulted in misclassification of 33% of calves.

Squamous cell carcinoma originating from a cutaneous scar in a llama.

A nonhealing wound associated with a laceration in a 12-year-old llama was evaluated. Initial attempts at closure were unsuccessful and biopsy revealed scar tissue. Subsequent biopsies, 18 mo later, revealed squamous cell carcinoma with regional metastasis. This report describes squamous cell carcinoma, secondary to a traumatic wound in a llama.