IMMUNE RESPONSE IN DAIRY COW FORESTOMACHS

Production diseases of dairy cows include several pathologies. These are considered man-made problems, mostly reported in the post-calving period of negative energy balance (NEB), when high-yielding dairy cows are unable to achieve a feed energy intake matching their high production requirements (Mulligan et al., 2008). A correct management of production diseases demands an early diagnostic approach and prognostic parameters. In ruminants, metabolic stress greatly influences the forestomachs physiology, including their immune response ability. Therefore a proper evaluation of production diseases in ruminants should also include markers of the innate immune response to metabolic stress (Amadori, 2016). Since forestomach walls express immune receptors and cytokines, and the rumen liquor is infiltrated by leukocytes, it is reasonable that they play an important role in the local immune response to metabolic stress (Ingvartsen et al., 2003; Trevisi et al., 2014a). My PhD hypothesis implies that ruminal fluids could be an important source of diagnostic information for the identification of herds at risk for production diseases, in addition to the traditional analyses. Accordingly, we further characterized the leukocytes subpopulations in the rumen liquor, highlighting the presence of B cells (the most frequent leukocyte population in the rumen liquor), T cells, and myeloid cells. We also compared the leukocyte composition in rumenocentesis versus esophageal probe samples, and we did not observe any significant statistical difference between the two sample collection techniques. We investigated the origin of the leukocytes of the rumen fluid and demonstrated that they partly derive from the oral cavity and reach the rumen through the saliva. On the basis of these findings, we carried out a field survey of innate immune parameters in rumen fluids of 128 animals from 12 farms, along with clinical inspections, assessment of milk yield, rumen pH and volatile fatty acids (VFA), and evaluation of major metabolic and hematologic parameters. Significant statistical correlations were found between immune markers in rumen fluids and biochemical parameters of dairy cows. In particular, a significant negative correlation was found between CD45 gene expression in rumen fluids (leukocyte infiltration) and ruminal pH. The infiltration of B cells was negatively affected by ruminal pH and high concentrations of volatile fatty acids (VFA). The same type of regulation was also exerted on the concentrations of IgM and total Ig. Interestingly, total Ig and IgM in rumen fluids showed a strong positive correlation with urea levels in blood, to some extent correlated with feed intake. We also investigated how the alteration of the ruminal microbiota could influence the immunologic profile of the rumen liquor. The use of a low-dose antibiotic (Monensin) led to significant differences in metabolic and immunological parameters in the rumen, and this resulted in a reduction of leukocyte colonization and immunoglobulin concentration in the ruminal fluid. Our procedures were included into a kit and tested in a group of 10 animals, half of them suffering from ruminal stasis; we could detect differences between the two groups in terms of leukocytes infiltration and Ig concentration. At a molecular level, the group with overt pathology showed a marked reduction of IGLC and KRT5 gene expression. Our data suggest that forestomach immune responses could be directed to “dangers” arising within the forestomach environment (diet unbalance, abnormal fermentations), but also arise as reporter system of disease conditions elsewhere in the body. The immune markers could integrate consolidated diagnostic parameters (e.g. rumen pH and VFA, milk cell counts, blood, fecal analytes) and contribute to robust, early diagnosis of production diseases of dairy cattle.