Kinetics of lung lesion development and pro-inflammatory cytokine response in pigs with vaccine-associated enhanced respiratory disease induced by challenge with pandemic (2009) A/H1N1 influenza virus.

The objective of this report was to characterize the enhanced clinical disease and lung lesions observed in pigs vaccinated with inactivated H1N2 swine δ-cluster influenza A virus and challenged with pandemic 2009 A/H1N1 human influenza virus. Eighty-four, 6-week-old, cross-bred pigs were randomly allocated into 3 groups of 28 pigs to represent vaccinated/challenged (V/C), non-vaccinated/challenged (NV/C), and non-vaccinated/non-challenged (NV/NC) control groups. Pigs were intratracheally inoculated with pH1N1 and euthanized at 1, 2, 5, and 21 days post inoculation (dpi). Macroscopically, V/C pigs demonstrated greater percentages of pneumonia compared to NV/C pigs. Histologically, V/C pigs demonstrated severe bronchointerstitial pneumonia with necrotizing bronchiolitis accompanied by interlobular and alveolar edema and hemorrhage at 1 and 2 dpi. The magnitude of peribronchiolar lymphocytic cuffing was greater in V/C pigs by 5 dpi. Microscopic lung lesion scores were significantly higher in the V/C pigs at 2 and 5 dpi compared to NV/C and NV/NC pigs. Elevated TNF-α, IL-1ß, IL-6, and IL-8 were detected in bronchoalveolar lavage fluid at all time points in V/C pigs compared to NV/C pigs. These data suggest H1 inactivated vaccines followed by heterologous challenge resulted in potentiated clinical signs and enhanced pulmonary lesions and correlated with an elevated proinflammatory cytokine response in the lung. The lung alterations and host immune response are consistent with the vaccine-associated enhanced respiratory disease (VAERD) clinical outcome observed reproducibly in this swine model.

Short communication: Allele, genotype, and haplotype data for bovine spongiform encephalopathy-resistance polymorphisms from healthy US Holstein cattle.

Bovine spongiform encephalopathy (BSE) is a neurodegenerative disease of cattle caused by abnormally folded prion proteins. Two regulatory region polymorphisms in the bovine prion gene are associated with resistance to classical BSE disease: a 23-bp region in the promoter that contains a binding site for the repressor protein RP58, and a 12-bp region in intron 1 that has a binding site for the transcription factor SP1. The presence of these binding sites enhances BSE resistance in cattle, whereas cattle that lack these regions are more susceptible to the disease. The present study examined the allele, genotype, and haplotype frequencies for the 23-bp and 12-bp polymorphisms in Holstein cattle from 9 different US states, and these frequencies were compared with data previously established for Holstein cattle from the United Kingdom, Germany, and Japan. Additionally, the coding region of the prion gene was sequenced from the US samples. Finally, archival samples from US Holstein sires born between 1953 and 1957 were analyzed. We found that the resistant allele and genotype frequencies for the US Holstein cattle were as high, or higher, relative to that observed in other countries. Furthermore, the current US frequencies were comparable to those determined in the archival samples from the 1950s. Based on the frequencies of these regulatory region polymorphisms, the US Holstein population is not at a greater risk for BSE than Holsteins worldwide.

Cumulative physiological events influence the inflammatory response of the bovine udder to Escherichia coli infections during the transition period.

A high proportion of intramammary coliform infections present at parturition develop disease characterized by severe inflammatory signs and sepsis during the first 60 to 70 d of lactation. In the lactating bovine mammary gland, the innate immune system plays a critical role in determining the outcome of these infections. Since the beginning of the 1990s, research has increased significantly on bovine mammary innate defense mechanisms in connection with the pathogenesis of coliform mastitis. Neutrophils are key effector cells of the innate immune response to intramammary infection, and their function is influenced by many physiological events that occur during the transition period. Opportunistic infections occur when the integrity of the host immune system is compromised by physical and physiological conditions that make the host more susceptible. The innate immune system of many periparturient cows is immunocompromised. It is unlikely that periparturient immunosuppression is the result of a single physiological factor; more likely, several entities act in concert, with profound effects on the function of many organ systems of the periparturient dairy cow. Their defense system is unable to modulate the complex network of innate immune responses, leading to incomplete resolution of the pathogen and the inflammatory reaction. During the last 30 yr, most efforts have been focused on neutrophil diapedesis, phagocytosis, and bacterial killing. How these functions modulate the clinical outcome of coliform mastitis, and how they can be influenced by hormones and metabolism has been the subject of intensive research and is the focus of this review. The afferent (sensing) arm of innate immunity, which enables host recognition of a diverse array of pathogens, is the subject of intense research interest and may contribute to the variable inflammatory response to intramammary infections during different stages of lactation. The development of novel interventions that modulate the inflammatory response or contribute to the elimination of the pathogen or both may offer therapeutic promise in the treatment of mastitis in periparturient cows.

Effects of dexamethasone on bovine circulating T lymphocyte populations.

In cattle, gamma delta T cells represent a higher proportion of circulating T cells than in humans. Bovine gamma delta T cells can be recognized by expression of gamma delta cell receptor (gamma delta TCR) determinants or by a 215/230-kDa surface antigen (WC1). WC1 is expressed on 90% or more of circulating bovine gamma delta T cells. The effects of dexamethasone on this and other subsets (CD3, CD2, CD4, CD8) of peripheral blood T lymphocytes were determined by flow cytometric analysis. Twelve 15-month old bulls were injected with dexamethasone (0.04 mg/kg/day) for 3 consecutive days and four bulls were untreated controls. Blood samples were collected daily for 3 days before dexamethasone injections and for an additional 7 days starting on the third day. Data were recorded as percent positive cells and as mean fluorescent intensity (MFI) of positive cells. Initially, CD3+ cells represented 65-73% of all peripheral blood clear cells (PBMC). Dexamethasone reduced CD3+/- cells (PBMC). Dexamethasone reduced CD3+ cells to 30% and these recovered to 50% positive cells by 9 days after the last dexamethasone injection. Loss of CD3+ cells was not due to reductions in alpha beta T cells because dexamethasone did not influence the percent CD2+, CD4+, or CD8+ cells. However, percent WC1+ cells rapidly declined from a baseline of 26.4% of PBMC to < 6% by the final injection. During injections, the MFI of WC1 increased. The MFI of WC1 returned to control values 7 days after the last injection of dexamethasone, but the percent gamma delta T cells recovered to only 14% WC1+ PBMC by the final day of the study. During its maximum effects on WC1, dexamethasone also caused a profound decrease of L-selectin MFI on remaining PBMC (mostly alpha beta T cells and monocyte/macrophages). In a second trial, two-color analyses determined that dexamethasone did not increase apoptosis in WC1+ cells and did not reduce L-selectin MFI on either CD2+ of WC1+ cells. The cumulative results suggested that dexamethasone promoted gamma delta T cell migration out of peripheral blood via an L-selectin-independent mechanism and that dexamethasone did not alter alpha beta T cell migration kinetics.

Regulation of L-selectin and CD18 on bovine neutrophils by glucocorticoids: effects of cortisol and dexamethasone.

The responsiveness of bovine neutrophil L-selectin and CD18 to in vivo glucocorticoid administration was characterized by flow cytometric analysis. Blood was sampled intensively from dairy cows treated for 3 days with placebo, cortisol, or dexamethasone. Immunostaining was performed on whole blood (100 microliters) that was left unstimulated or was stimulated with platelet-activating factor (PAF; 1 microgram/ml blood) prior to incubation with fluorescein isothiocyanate-conjugated monoclonal antibodies against L-selectin and CD18. Results were expressed as the percentage of positive-staining cells and as mean fluorescence intensity (MFI) of those cells. Total leukocyte count and leukocyte differentials were also performed on all blood samples. Dexamethasone caused nearly complete down-regulation of L-selectin (P < .01) on the surface of gated cells and reduced to half the MFI of CD18 (P < .01). Compared with values for the placebo group, dexamethasone began to cause L-selectin down-regulation within 8 h after the first injection and these effects persisted until 48 h after the third injection. This was correlated in time with an acute reduction in the proportion of cells that stained positive for L-selectin (from 98% before dexamethasone injections to a low of 17% by 40 h after the first injection). Dexamethasone also caused leukocytosis and neutrophilia during this time interval. In contrast, CD18 down-regulation was delayed until 16 h after the second dexamethasone injection and persisted for roughly 8 days. However, at no time during the experiment did dexamethasone influence the proportion of gated cells staining positive for CD18 (always 100%). Effects of cortisol were generally similar in pattern to those of dexamethasone but were more subtle and more readily detected when PAF was added to blood prior to immunostaining. These results strongly suggest that one mechanism of the anti-inflammatory action of glucocorticoids is to induce dramatic down-regulation of L-selectin and CD18 adhesion molecules on blood neutrophils.

Sequence of the bovine CD18-encoding cDNA: comparison with the human and murine glycoproteins.

The bovine cDNA (CD18) encoding CD18, a cell-surface glycoprotein involved in multiple leukocyte functions, was sequenced and compared with the human and murine sequences. Portions of the 5'- and 3'-untranslated regions of the nucleotide sequences are conserved among the three species, including a 3' A+T-rich region believed to regulate mRNA stability and translational efficiency. The 2833-bp bovine sequence coded for a protein of 769 amino acids (aa). Overall, the deduced aa sequences were greater than 80% identical among the three species. The aa 96-389 and those in the cytoplasmic domain were very highly conserved with approx. 95% aa identity. All Cys residues and potential Asn-glycosylation sites present in the bovine sequence were also present in the human and murine sequences. The aa identity was also found in those regions where mutations were found to cause the genetic disease, leukocyte adhesion deficiency. These data identify functionally important regions of the CD18 mRNA and protein.

Cloning and sequencing of a cDNA encoding bovine intercellular adhesion molecule 3 (ICAM-3).

A cDNA encoding a putative bovine intercellular adhesion molecule (ICAM)-3, a ligand of the leukocyte integrin LFA-1 (CD11a/CD18), was sequenced and compared with human ICAM sequences. The 1635-bp bovine sequence codes for a protein of 544 amino acids (aa). This putative bovine ICAM-3 has five immunoglobulin (Ig)-like domains similar to human ICAM-1 and ICAM-3, and belongs to the Ig gene superfamily. The overall identities of the deduced aa sequence with those of human ICAM-3 and ICAM-1 are 61% and 58%, respectively. The predicted number and positions of Cys residues are all conserved between the bovine and human ICAM 3 aa sequences.

Integrin Mac-1 and beta-amyloid in microglial release of nitric oxide.

The beta-amyloid protein associated with Alzheimer's disease (AD) has been well characterized biochemically; however, its primary biological function and mode of action in AD has not been determined. We have shown previously that beta-amyloid (beta25-35), in combination with interferon-gamma (IFN-gamma), can induce nitric oxide release from cultured hippocampal microglial cells. In the present study, binding of beta-amyloid with the leukocyte integrin Mac-1, a cell surface receptor on microglia, was studied by observing (1) inhibition of beta-amyloid (beta25-35)-mediated release of nitric oxide from cultured microglial cells following exposure to monoclonal antibodies against Mac-1 (anti-CD18 and anti-CD11b) and (2) competitive binding of fluorochrome-labeled beta25-35 with anti-CD18 or anti-CD11b using fluorescent flow cytometry. Wt.3 (anti-CD18 antibody) and OX42 (anti-CD11b antibody) were as effective as opsonized zymosan at inducing the release of nitric oxide from microglia. Furthermore, Wt.3 and OX42 acted synergistically to induce maximum nitric oxide release. An interaction between beta-amyloid and CD18 of Mac-1 was evidenced by the suppressive action of beta25-35 on Wt.3-mediated release of nitric oxide and the synergistic action between OX42 and beta25-35 in inducing nitric oxide release from microglia. The tissue culture study was supported by competitive binding assays of fluorochrome-labeled beta25-35 and Mac-1 antibodies (Wt.3 or OX42). The majority of microglial cells (71%) did bind biotinylated beta-amyloid in the presence of cytochalasin B, suggesting that beta-amyloid binding to microglia is a receptor-mediated event. Furthermore, pre-exposure to Wt.3, but not OX42, significantly decreased binding of biotinylated beta25-35 to microglia. These findings suggest that CD18 of Mac-1 may play a role in beta-amyloid-mediated release of nitric oxide.

Cloning, sequencing, and analysis of cDNA encoding bovine granulocyte-colony stimulating factor.

Neutrophils play a critical role in defending against bacterial infections. Hematopoietic growth factors are a class of regulatory cytokines that are required for stimulation, proliferation, and differentiation of blood cells. Granulocyte colony stimulating factor (G-CSF) is a cytokine that induces proliferation and maturation of precursor myeloid cells in the bone marrow into fully differentiated neutrophils. G-CSF also modulates the functional activity of mature neutrophils. Treatment with G-CSF significantly enhances neutrophil phagocytic activity and killing of bacteria and fungi. We have isolated and sequenced a cDNA clone encoding bovine G-CSF (bG-CSF) from an endothelial cell cDNA library using primers designed from ovine G-CSF. The full length cDNA is 1460 nucleotides with 585 nucleotides comprising the open reading frame. Sequence analysis shows 95% identity with ovine, 89% with porcine, 85% with human, and 76% with murine G-CSF. The deduced G-CSF protein consists of 174 amino acids with 95% identity to ovine, 86% to porcine, 81% to human, and 71% to murine. The signal peptide of G-CSF is 21 amino acids long which is nine amino acids shorter than that of human and murine G-CSF. RT-PCR analysis shows that neither freshly isolated nor ConA stimulated neutrophils express G-CSF mRNA. Mononuclear cells, however, expressed G-CSF mRNA after 48 h incubation with or without ConA stimulation.

Cytokine gene expression in ileal tissues of cattle infected with Mycobacterium paratuberculosis.

Cytokine gene expression in ileal tissues of cattle infected with Mycobacterium paratuberculosis was evaluated. The effects of infection with M. paratuberculosis on cytokine production may influence immune regulation at the site of colonization, resulting in the chronic inflammatory state associated with the latter stages of this disease. Ileal samples were obtained at necropsy from noninfected control cows (n=8) and clinically infected cows (n=7) and processed for immunohistochemistry and in situ hybridization. Cows infected with M. paratuberculosis were in the latter stages of disease with clinical signs such as weight loss, watery diarrhea, and inappetence. Among cytokines we studied, interleukin-1alpha (IL-1alpha), IL-1beta, IL-6, and interferon-gamma (IFN-gamma) were expressed significantly more in infected animals than in noninfected control animals. The expression of tumor necrosis factor-alpha (TNF-alpha), however, was not different between the two groups of cattle. In addition, immunohistochemical staining demonstrated that the number of resident macrophages in the ileum of infected animals was three times greater than that of noninfected cows. In contrast to this, ileal tissues from noninfected control animals contained 1.5 times more neutrophils than the ileal tissues from cows infected with M. paratuberculosis. These data demonstrate that localized ileal cytokine production is different between cows chronically infected with M. paratuberculosis and noninfected control cows.

Expression, purification, and in vitro biological activities of recombinant bovine granulocyte-colony stimulating factor.

Neutrophils are essential components of the innate immune system and they play a critical role in the defense of host against bacterial and fungal infections. The colony stimulating factors are a class of glycoproteins that are required for proliferation, differentiation, and functional activation of hematopoietic progenitor cells. Granulocyte-colony stimulating factor (G-CSF) is a member of this regulatory family of cytokines that specifically stimulates proliferation and maturation of precursor cells in the bone marrow into fully differentiated and functional neutrophils. G-CSF also modulates the biological activities of mature neutrophils in circulation. A bovine G-CSF (bG-CSF) cDNA clone (previously isolated and sequenced in our laboratory) was expressed in Escherichia coli and the biological activities of the solubilized protein from purified inclusion bodies were examined. Flow cytometric analysis of membrane antigen density of neutrophils activated with bG-CSF revealed an upregulation in the expression of CD11a (>114%), CD11b (>148%), CD11c (>87%), and CD18 (>109%). Expression of L-selectin was decreased by more than 43%. There was no change, however, in the expression of CD14. These findings indicate that recombinant bG-CSF (rbG-CSF) expressed in E. coli is biologically active and exerts the same type of effects on neutrophils in vitro as those of human G-CSF (hG-CSF).

Cloning, sequencing and analysis of cDNA encoding bovine intercellular adhesion molecule-1 (ICAM-1).

Intercellular adhesion molecule-1 (ICAM-1) is an inducible glycoprotein that interacts with the leukocyte beta 2-integrins, LFA-1 and Mac-1. We have isolated and analyzed a cDNA clone coding for the putative bovine ICAM-1 gene and compared it with known comparative sequences from other species as well as bovine ICAM-3. The 3398-bp bovine ICAM-1 cDNA sequence codes for 535 amino acids and shows 57% homology with human ICAM-1 and 47% homology with bovine ICAM-3 at the amino acid levels. The predicted number and positions of cysteine residues in bovine ICAM-1 are all conserved among species including bovine ICAM-3. It has two arginine-glycine-aspartate (RGD) sites in the extracellular region and a serine residue in the cytoplasmic tail. Northern blot results show that the bovine ICAM-1 gene is expressed in stimulated leukocytes whereas bovine ICAM-3 is expressed predominantly in resting neutrophils.

A colorimetric assay for quantitating bovine neutrophil bactericidal activity.

A colorimetric assay was developed for quantitating bovine neutrophil bactericidal activity against Staphylococcus aureus. The procedure used the tetrazolium compound, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT). The assay was conducted by incubating antibody-opsonized S. aureus with neutrophils in microtiter plates for 1 h at a ratio of 10 bacteria per neutrophil. Neutrophils were then lysed with saponin. The MTT was added and samples were incubated for 10 min. Live S. aureus reduced MTT to purple formazan. Dead bacteria and lysed neutrophils did not react with MTT. Bacterially-reduced formazan was solubilized by adding isopropanol and formazan production was quantitated by measuring absorption at 560 nm. Absorption of formazan was directly related to viable bacteria cell number and was used to determine the number of S. aureus not killed by neutrophils. The percentage of bacteria killed by neutrophils was determined by extrapolation from a standard formazan curve that was derived by incubating MTT with known numbers of S. aureus. The colorimetric MTT assay detected suppressed bactericidal activity after in vitro treatment of bovine neutrophils with colchicine, cytochalasin B, or phorbol 12-myristate 13-acetate. In vitro treatment of neutrophils with low levels of recombinant bovine interferon gamma (rBoIFN-gamma) enhanced bactericidal activity, whereas high levels decreased activity. These results suggest the colorimetric MTT bactericidal assay is efficacious in detecting modulation of bovine neutrophil bactericidal activity. Furthermore, the MTT assay has many advantages over traditional bactericidal assays in that it is sensitive, inexpensive, requires less than 3 h to complete, and can analyze many neutrophil samples in a single day.

Local and systemic immune response in the cow after intramammary vaccination during lactation.

The local and systemic immune response of the lactating cow during the 10 week period after intramammary (IMM) vaccination with killed Mycobacterium bovis (M. bovis) was evaluated. Antigen (tuberculin)-reactive lymphocytes were present in the milk as early as 2 weeks post-vaccination, and in the blood at 6 weeks after vaccination. The milk lymphocytes, compared to the blood lymphocytes were consistently more responsive to tuberculin. Both blood and milk lymphocytes responded in vitro to the lectins, phytohemagglutinin-A (PHA-P) and concanavalin A (Con A), although the milk lymphocytes were consistently less responsive than the blood lymphocytes during the period. Anti-tuberculin antibody was significantly elevated in the milk and blood of the vaccinated animals at 10 weeks post-vaccination. Infusion of tuberculin into the mammary glands of the cows 10 weeks after vaccination resulted in a marked increase in the number of milk leukocytes. The influx of leukocytes initially consisted of polymorphonuclear leukocytes (PMNL), and later, mononuclear leukocytes. Intramammary vaccination also resulted in antigen recognition at sites distant from the mammary gland.

Detection of conglutinin in bovine serum by complement-dependent agglutination of Escherichia coli.

Agglutination of Escherichia coli (ECA) by normal bovine serum was shown to be prevented by heating serum to 56 degrees C for 30 min, but restored by normal horse, swine, rabbit or guinea pig sera. Further investigation of the ECA reaction using techniques to distinguish between conglutination and immunoconglutination indicated ECA to be a conglutination reaction. Testing of 264 sera obtained from 22 normal cattle over a period of 5 months did not show individual or seasonal variation in ECA. Changes in ECA and conglutination were detected in sera of periparturient cows. The ECA reaction is a simple technique for detecting conglutinin in bovine serum.

Cloning and sequencing of cDNA encoding bovine tumor necrosis factor (TNF)-receptor I.

Tumor necrosis factor (TNF) is a multipotent cytokine produced by activated macrophages and lymphocytes and its activity is mediated by specific cell surface receptors, TNF-RI and TNF-RII. We have isolated and analyzed a cDNA encoding bovine TNF-RI gene and compared it with known TNF-RI sequences from other species. The cDNA sequence for the coding region of bovine TNF-RI shows 80% homology with porcine TNF-RI and 77% with human TNF-RI. The cDNA sequence of bovine TNF-RI codes for 471 amino acids and shows 75% and 67% identity with the amino acid sequences of porcine TNF-RI and human TNF-RI, respectively. The predicted bovine TNF-RI amino acid sequence consists of a signal peptide, an extracellular domain, a transmembrane region and a cytoplasmic tail. The extracellular region contains four repeated cysteine rich domains, which are conserved in all species. Northern blot results show that bovine TNF-RI gene is expressed in neutrophils and mononuclear leukocytes.

Expression and characterization of a recombinant soluble form of bovine tumor necrosis factor receptor type I.

A recombinant soluble bovine tumor necrosis factor receptor type I (sboTNF-RI) was expressed in the methylotrophic yeast Pichia pastoris and evaluated for its ability to inhibit bovine tumor necrosis factor alpha (TNF-alpha) cytotoxicity. A cDNA encoding the extracellular domain of bovine TNF-RI was placed under the control of the powerful and tightly regulated alcohol oxidase1 (AOX1) gene promoter of the pPICZa A vector and the resulting construct integrated into the 5' region of the alcohol oxidase genes of GS115 and KM71 strains of Pichia. Soluble bovine TNF-RI was secreted into the medium following induction of the AOX1 gene promoter with methanol, and purified to greater than 95% purity by ion-exchange chromatography. In in vitro assays, the purified recombinant sboTNF-RI will block the cytolytic activity of bovine TNF-alpha on WEHI 164 cells clone 13 by 50% when used at a concentration of 170 microg/ml, and by nearly 90% when used at a concentration of 310 microg/ml. Results of this study suggest that recombinant sboTNF-RI may have therapeutic value as a TNF inhibitor in cattle with coliform mastitis.

Bovine sire effects on daughters' in vitro blood neutrophil functions, lymphocyte blastogenesis, serum complement and conglutinin levels.

Blood neutrophil functions, lymphocyte blastogenic responses, serum complement, and serum conglutinin activity of 98 lactating Holstein cows from two genetic lines were evaluated. The genetic lines were produced in a selection experiment that created and perpetuated genetic differences in milk production for up to seven generations. No significant differences between the two genetic lines of cows were found for neutrophil function, lymphocyte blastogenic responses, serum complement levels, or serum conglutinin levels. Significant differences between sire progeny groups within lines were found for unstimulated and mitogen-stimulated lymphocyte blastogenesis (P less than 0.0001), and almost all neutrophil functions (antibody independent neutrophil cytotoxicity, antibody dependent neutrophil cytotoxicity, ingestion of bacteria, iodination, chemiluminescence, chemokinesis, and chemotaxis (P less than or equal to 0.05)). Sire progeny group differences (P less than or equal to 0.0001) within lines for serum complement and conglutinin activity were also found. Neutrophil chemiluminescence activity (positive relationship; P less than or equal to 0.001), concanavalin A-stimulated lymphocyte blastogenesis (positive relationship; P less than or equal to 0.004), and serum conglutinin activity levels (negative relationship; P less than or equal to 0.01) each had small but significant associations with the total milk somatic cell count. Cows seropositive for bovine leukosis virus had increased resting and mitogen-stimulated lymphocyte blastogenic activity and were associated with increased in vitro neutrophil random migration and production of superoxide anion. Estimates of genetic parameters of various immune cell functions, of serum complement and of conglutinin levels for daughters of 11 sires with 4-6 daughters in the data set were determined. In this report, genetic variation was demonstrated for nonspecific humoral and cellular immunity.

Study of immunological dysfunction in periparturient Holstein cattle selected for high and average milk production.

Data from twenty assays of traits associated with innate and adaptive immunity were evaluated from 137 periparturient Holstein cows. These cows had been selected through planned matings for four different levels of milk production (high and average pounds of milk, and high and average pounds of milk fat plus protein). For up to seven generations, the genetic lines were produced by mating females of each line to sires of corresponding merit. With the exceptions of neutrophil ingestion of Staphylococcus aureus and directed migration, all assays measuring neutrophil functions were depressed beginning 2 to 3 weeks before calving through 3 weeks after calving. Serum concentrations of immunoglobulin G1 decreased while those of immunoglobulin G2 increased around calving time. Serum complement and conglutinin concentrations decreased before calving and reached a minimum around calving time. Cows selected for high milk production (pounds of milk and pounds of milk fat plus proteins) had significantly higher (P < 0.10) numbers of circulating neutrophils and mononuclear cells, had higher (P < 0.10) neutrophil resting chemiluminescence and higher (P < 0.10) neutrophil directed migration than cows with average production potentials. There were significant (P < 0.001) sire progeny group differences for most traits associated with the immune system that we tested. These results can be considered encouraging, in that selection for high milk yield did not produce unfavorable correlated responses in the functional capacity of immune function traits, and that there is sufficient genetic variation in these immunological traits among sires of high genetic merit for milk production to potentially improve the immunocompetence of periparturient cows through planned mating experiments.