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Neutrophils are the most important polymorphonuclear leukocytes (PMNL), representing the front-line defense involved in pathogen clearance upon invasion. As such, they play a pivotal role in immune and inflammatory responses. Isolated PMNL from 5 mid-lactating Holstein dairy cows were used to evaluate the in vitro effect of methionine (Met) and choline (Chol) supplementation on mRNA expression of genes related to the Met cycle and innate immunity. The target genes are associated with the Met cycle, cell signaling, inflammation, antimicrobial and killing mechanisms, and pathogen recognition. Treatments were allocated in a 3 × 3 factorial arrangement, including 3 Lys-to-Met ratios (L:M, 3.6:1, 2.9:1, or 2.4:1) and 3 levels of supplemental Chol (0, 400, or 800 μg/mL). Three replicates per treatment group were incubated for 2 h at 37°C and 5% atmospheric CO 2. Both betaine-homocysteine S-methyltransferase and choline dehydrogenase were undetectable, indicating that PMNL (at least in vitro) cannot generate Met from Chol through the betaine pathway. The PMNL incubated without Chol experienced a specific state of inflammatory mediation [greater interleukin-1β (IL1B), myeloperoxidase (MPO), IL10, and IL6] and oxidative stress [greater cysteine sulfinic acid decarboxylase (CSAD), cystathionine gamma-lyase (CTH), glutathione reductase (GSR), and glutathione synthase (GSS)]. However, data from the interaction L:M × Chol indicated that this negative state could be overcome by supplementing additional Met. This was reflected in the upregulation of methionine synthase (MTR) and toll-like receptor 2 (TLR2); that is, pathogen detection ability. At the lowest level of supplemental Chol, Met downregulated GSS, GSR, IL1B, and IL6, suggesting it could reduce cellular inflammation and enhance antioxidant status. At 400 μg/mL Chol, supplemental Met upregulated PMNL recognition capacity [higher TLR4 and L-selectin (SELL)]. Overall, enhancing the supply of methyl donors to isolated unstimulated PMNL from mid-lactating dairy cows leads to a low level of PMNL activation and upregulates a cytoprotective mechanism against oxidative stress. Enhancing the supply of Met coupled with adequate Chol levels enhances the gene expression of PMNL pathogen-recognition mechanism. These data suggest that Chol supply to PMNL exposed to low levels of Met effectively downregulated the entire repertoire of innate inflammatory-responsive genes. Thus, Met availability in PMNL during an inflammatory challenge may be sufficient for mounting an appropriate biologic response.

Genetic variability and population structure of Cercospora beticola, the causal agent of Cercospora leaf spot in sugarbeet, from four sugarbeet-growing regions of Greece were investigated using growth rate, pathogenicity, and mini-and microsatellite DNA fingerprinting. Mycelial growth and pathogenicity were very diverse within and between groups, and no correlation was found between these features and the geographic origin of the isolates. High diversity was found by micro-and minisatellite fingerprinting, with an average gene diversity of 0.21, and no significant differences among populations. Among the 46 isolates, 45 different genotypes were identified, showing a high degree of genotype diversity. Analysis of the genetic profiles provided no evidence for regional patterns of variation (ΦF ST =0.01, P = 0.261) and the analysis of molecular variation (AMOVA) revealed that genetic variability was due mainly to variations within (99%) rather than between (1%) populations. Such a low level of genetic differentiation is reflected by a migration rate value Nm of 4.7. The high migration rate cannot be referred to splash dispersed conidia. To justify the absence of a regional structure in these C. beticola populations, we must suppose the existence of a long-distance means of dispersal, such as seed transmission and/or man mediated transmission.

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