Effect of microbial inoculation and storage length on the fermentation profile and nutritive value of high-moisture corn ensiled at 2 different dry matter concentrations.

The objective of this experiment was to evaluate the effect of microbial inoculation and storage length on the fermentation profile and nutrient composition of high-moisture corn (HMC) ensiled at 2 different dry matter (DM) concentrations. High-moisture corn was harvested when kernel DM concentrations were approaching 65% as-fed, and either left undried (HMC65; 67.6% DM) or dried at 40 °C to approximately 70% DM (HMC70; 71.0% DM), and then ensiled in quadruplicate vacuum pouches untreated (CON) or after one of the following inoculant treatments: 6.36 × 105 cfu of Lentilactobacillus buchneri DSM 12856, Lactiplantibacillus plantarum DSM 12836, and Pediococcus acidilactici DSM 16243 per g of HMC (LBLP); or 3.0 × 105 cfu of Lentilactobacillus buchneri DSM 12856, Lentilactobacillus diolivorans DSM 32074, and P. acidilactici DSM 16243 per g of HMC (LBLD). Vacuum pouches were allowed to ferment for 7, 14, 28, or 56 d. A three-way interaction was observed (P = 0.01) for the pH of HMC, where CON for HMC70 was greatest across storage lengths and HMC65 treatments generally had a lower pH than other treatments. Concentrations of total acids were greater (P = 0.001) in HMC65 than HMC70 and greater (P = 0.001) in HMC treated with LBLP and LBLD than CON. An interaction between DM concentration, microbial inoculation, and storage length was observed (P = 0.05) for concentrations of acetic acid. At 14 d, acetic acid concentrations were greater in HMC65 treated with LBLD than other treatments. Likewise, at 56 d, concentrations of acetic acid were greatest in HMC65 treated with LBLD, followed by HMC70 treated with LBLD. An interaction between DM concentration, microbial inoculation, and storage length was observed (P = 0.05) for 7-h starch disappearance (starchD). Across all DM concentration and inoculant treatment combinations, starchD increased with increasing storage length. StarchD was also generally greater for HMC65 treatments compared to HMC70, with small differences among inoculants. Results suggest that microbial inoculation can improve fermentation of HMC by increasing the production of antifungal acetic acid, but that DM concentration at ensiling remains a primary determinant of HMC fermentability.

High-moisture corn (HMC) short-term fermentation is affected by dry matter (DM) concentration. Thus, producers try to influence HMC fermentation by using microbial inoculants and by harvesting HMC at different DM concentrations. This study aimed to evaluate the effects of different DM concentrations, heterofermentative microbial inoculants, and storage length on the fermentation and nutritive value of HMC. Total acid production was greater in HMC with a lower DM and treated with microbial inoculants. Lactic acid concentrations were generally greater in lower DM HMC. Microbial inoculants increased the production of acetic acid, an antifungal acid. Because acetic acid can improve aerobic stability, these results demonstrate microbial inoculation and lower DM can improve HMC fermentation. However, DM concentration seems to influence fermentation to a greater extent than the use of microbial inoculants. Although starch concentration was not affected by microbial inoculants or DM concentration, starch digestibility was greater in lower DM HMC. This demonstrates lower DM may improve nutritive value in addition to improving fermentation by increasing the production of total acids and lactic acid in HMC.

Effect of cutting height, microbial inoculation, and storage length on fermentation profile and nutrient composition of whole-plant corn silage.

This study aimed to evaluate the effects of cutting height, heterofermentative microbial inoculants, and storage length on the fermentation profile and nutrient composition of whole-plant corn silage. The experiment was a completely randomized design with a 2 (cutting height) × 3 (microbial inoculation) × 5 (storage length) factorial arrangement of treatments. Corn forage was harvested at two cutting heights: either 25 cm (REG) or 65 cm (HI). Then, forage was inoculated with one of three microbial inoculants: (1) 300,000 CFU/g of fresh forage of Pediococcus acidilactici DSM 16243, Lentilactobacillus buchneri DSM 12856, and L. diolivorans DSM 32074 (LBLD; Bonsilage Speed inoculant, Provita Supplements Inc., Mendota Heights, MN), (2) 500,000 CFU/g of fresh forage of Lactiplantibacillus plantarum DSM 12837 and L. buchneri DSM 16774 (LPLB; Bonsilage Corn + WS inoculant, Provita Supplements Inc., Mendota Heights, MN), or (3) distilled water (CON). Last, forage was randomly assigned to ferment for 5, 7, 14, 28, or 56 d of storage in vacuum-sealed bags. Silage pH was affected by a three-way interaction (P = 0.01), where CON treatments decreased continually over time while LPLB and LBLD began to increase at later storage lengths. Acetic acid concentration was greater (P = 0.001) in LPLB and LBLD than CON silage after 56 d of storage. Silage treated with LBLD did not have detectable levels of propionic acid (P > 0.05), although 1-propanol concentration was greater (P = 0.001) in LBLD treatments after 56 d of storage. The concentrations of total acids and acetic acid were greater (P = 0.01 and P = 0.001, respectively) for REG silage compared to HI. Additionally, HI silage had greater (P = 0.001 and P = 0.001, respectively) concentrations of dry matter (DM) and starch, while neutral detergent fiber (aNDF) and lignin concentrations were lower (P = 0.001 and P = 0.001, respectively) in HI silage compared to REG silage. Last, HI silage had a greater (P = 0.001) NDF digestibility than REG silage. The results of this study demonstrate that increasing cutting height can improve nutrient composition of whole-plant corn silage. Additionally, results demonstrate that heterofermentative microbial inoculants can be used to shift silage fermentation to the production of lactic and acetic acids.

Effects of cultivar and harvest days after planting on dry matter yield and nutritive value of teff.

One of the most pressing issues facing the dairy industry is drought. In areas where annual precipitation is low, irrigation for growing feed presents the greatest water-utilization challenge for dairy producers. Here, we investigated the effects of cultivar and harvest days after planting (DAP) on dry matter (DM) yield and nutritive value of teff (Eragrostis tef), a warm-season annual grass native to Ethiopia that is well adapted to drought conditions. Eighty pots were blocked by location in a greenhouse and randomly assigned to four teff cultivars (Tiffany, Moxie, Corvallis, and Dessie) and to five harvest times (40, 45, 50, 55, or 60 DAP). Cultivars had no effect on DM yield and nutritive value. As harvest time increased from 40 to 60 DAP, DM yield and ash-free neutral detergent fiber (aNDFom) concentrations increased, while crude protein (CP) concentrations and in vitro NDF digestibility decreased. To assess carryover effects of time of harvest on yield and nutritive value, two additional cuttings were taken from each pot. Increasing first-cutting harvest time decreased CP concentrations in the second cutting and reduced DM yields in the second and third cutting. Harvesting teff between 45 and 50 DAP best optimized forage yield and nutritive value in the first and subsequent cuttings.

Normalization of cholesterol metabolism in spinal microglia alleviates neuropathic pain.

Neuroinflammation is a major component in the transition to and perpetuation of neuropathic pain states. Spinal neuroinflammation involves activation of TLR4, localized to enlarged, cholesterol-enriched lipid rafts, designated here as inflammarafts. Conditional deletion of cholesterol transporters ABCA1 and ABCG1 in microglia, leading to inflammaraft formation, induced tactile allodynia in naive mice. The apoA-I binding protein (AIBP) facilitated cholesterol depletion from inflammarafts and reversed neuropathic pain in a model of chemotherapy-induced peripheral neuropathy (CIPN) in wild-type mice, but AIBP failed to reverse allodynia in mice with ABCA1/ABCG1-deficient microglia, suggesting a cholesterol-dependent mechanism. An AIBP mutant lacking the TLR4-binding domain did not bind microglia or reverse CIPN allodynia. The long-lasting therapeutic effect of a single AIBP dose in CIPN was associated with anti-inflammatory and cholesterol metabolism reprogramming and reduced accumulation of lipid droplets in microglia. These results suggest a cholesterol-driven mechanism of regulation of neuropathic pain by controlling the TLR4 inflammarafts and gene expression program in microglia and blocking the perpetuation of neuroinflammation.

Prostate cancer management choices in patients undergoing multiparametric magnetic resonance imaging/ultrasound fusion biopsy compared to systematic biopsy.

OBJECTIVES: To assess management choices in patients who undergo magnetic resonance imaging (MRI)/ultrasound (MRI/US) fusion-guided prostate biopsy compared to patients who undergo systematic biopsy. METHODS: We compared men who underwent MRI/US fusion-guided prostate biopsy to those who underwent systematic 12-core biopsy from 2014 to 2016. Patient demographics and pathologic findings were reviewed. The highest grade group per case was considered for analysis. RESULTS: Follow-up was available on 133 patients who underwent MRI/US targeted biopsy and 215 patients who underwent systematic biopsy. There was no difference in prebiopsy prostate-specific antigen (PSA) (10.1 ± 10.0 vs. 12.9 ± 20.5, P = 0.11) between the 2 cohorts. Patients in the MRI cohort were more likely to have had a previous prostate biopsy (P<0.0001). Overall, more patients in the MRI cohort choose active surveillance compared to the standard cohort (49.6% vs. 24.2%, P<0.0001), confirmed on multivariate logistic regression model adjusting for age, PSA density, prior biopsy history, race, grade group, and provider (P = 0.013). This finding held true independently for patients with grade groups 1 and 2 tumors (P = 0.02 and P = 0.005, respectively) and in a multivariate logistic regression model adjusting for grade group 1 and 2 tumors (P = 0.0051). In the standard cohort, more patients chose radiation over prostatectomy (47.2% vs. 24.4%, P<0.0001). On multivariate analysis, race was an independent predictor of active surveillance, with African Americans less likely to undergo active surveillance. CONCLUSIONS: Patients who undergo MRI/US targeted biopsy are more likely to choose active surveillance over early definitive treatment compared to men diagnosed on systematic biopsy when adjusting for tumor grade, PSA density, prior biopsy history, race, and provider.

Apocynin-treatment reverses hyperoxaluria induced changes in NADPH oxidase system expression in rat kidneys: a transcriptional study.

PURPOSE: We have previously shown that production of reactive oxygen species (ROS) is an important contributor to renal injury and inflammation following exposure to oxalate (Ox) or calcium-oxalate (CaOx) crystals. The present study was conducted, utilizing global transcriptome analyses, to determine the effect of Apocynin on changes in the NADPH oxidase system activated in kidneys of rats fed a diet leading to hyperoxaluria and CaOx crystal deposition. APPROACH: Age-, sex- and weight-matched rats were either fed regular rat chow or regular rat chow supplemented with 5% w/w hydroxy-L-proline (HLP). Half of the rats on the HLP diet were also placed on Apocynin-supplemented H(2)O. After 28 days, each rat was euthanized, their kidneys freshly explanted and dissected to obtain both cortex and medulla tissues. Total RNA was extracted from each tissue and subjected to genomic microarrays to obtain global transcriptome data. KEGG was used to identify gene clusters with differentially expressed genes. Immunohistochemistry was used to confirm protein expressions of selected genes. RESULTS: Genes encoding both membrane- and cytosolic-NADPH oxidase complex-associated proteins, together with p21rac and Rap1a, were coordinately up-regulated significantly in both renal medulla and cortex tissues in the HLP-fed rats compared to normal healthy untreated controls. Activation of NADPH oxidase appears to occur via the angiotensin-II/angiotensin-II receptor-2 pathway, although the DAG-PKC pathway of neutrophils may also contribute. Immuno histochemical staining confirmed up-regulated gene expressions. Simultaneously, genes encoding ROS scavenger proteins were down-regulated. HLP-fed rats receiving Apocynin had a complete reversal in the differential-expression of the NADPH oxidase system genes, despite showing similar levels of hyperoxaluria. CONCLUSIONS: A strong up-regulation of an oxidative/respiratory burst involving the NADPH oxidase system, activated via the angiotensin-II and most likely the DAG-PKC pathways, occurs in kidneys of hyperoxaluric rats. Apocynin treatment reversed this activation without affecting the levels of hyperoxaluria.

A structural element that facilitates proton-coupled electron transfer in oxalate decarboxylase.

The conformational properties of an active-site loop segment, defined by residues Ser(161)-Glu(162)-Asn(163)-Ser(164), have been shown to be important for modulating the intrinsic reactivity of Mn(II) in the active site of Bacillus subtilis oxalate decarboxylase. We now detail the functional and structural consequences of removing a conserved Arg/Thr hydrogen-bonding interaction by site-specific mutagenesis. Hence, substitution of Thr-165 by a valine residue gives an OxDC variant (T165V) that exhibits impaired catalytic activity. Heavy-atom isotope effect measurements, in combination with the X-ray crystal structure of the T165V OxDC variant, demonstrate that the conserved Arg/Thr hydrogen bond is important for correctly locating the side chain of Glu-162, which mediates a proton-coupled electron transfer (PCET) step prior to decarboxylation in the catalytically competent form of OxDC. In addition, we show that the T165V OxDC variant exhibits a lower level of oxalate consumption per dioxygen molecule, consistent with the predictions of recent spin-trapping experiments [Imaram et al. (2011) Free Radicals Biol. Med. 50, 1009-1015]. This finding implies that dioxygen might participate as a reversible electron sink in two putative PCET steps and is not merely used to generate a protein-based radical or oxidized metal center.

Gene expression profiling of early-phase Sjögren's syndrome in C57BL/6.NOD-Aec1Aec2 mice identifies focal adhesion maturation associated with infiltrating leukocytes.

PURPOSE: Despite considerable efforts, the molecular and cellular events in lacrimal gland tissues initiating inflammatory responses leading to keratoconjunctivitis sicca (KCS), autoimmunity, and Sjögren's syndrome (SjS) have yet to be defined. To determine whether altered glandular homeostasis occurs before the onset of autoimmunity, a temporal transcriptome study was carried out in an animal model of primary SjS. METHODS: Using oligonucleotide microarrays, gene expression profiles were generated for lacrimal glands of C57BL/6.NOD-Aec1Aec2 mice 4 to 20 weeks of age. Pairwise analyses identified genes differentially expressed, relative to their 4-week expression, during the development of SjS-like disease. Statistical analyses defined differentially and coordinately expressed gene sets. The PANTHER (Protein ANalysis THrough Evolutionary Relationships) classification system was used to define annotated biological processes or functions. RESULTS: Temporal transcript expression profiles of C57BL/6.NOD-Aec1Aec2 lacrimal glands before, or concomitant with, the first appearance of inflammatory cells revealed a highly restricted subset of differentially expressed genes encoding interactive extracellular matrix proteins, fibronectin, integrins, and syndecans. In contrast, genes encoding interepithelial junctional complex proteins defined alterations in tight junctions (TJ), adherens, desmosomes, and gap junctions, suggesting perturbations in the permeability of the paracellular spaces between epithelial barriers. Correlating with this were gene sets defining focal adhesion (FA) maturation and Ras/Raf-Mek/Erk signal transduction. Immunohistochemically, FAs were associated with infiltrating leukocytes and not with lacrimal epithelial cells. CONCLUSIONS: For the first time, FA maturations are implicated as initial biomarkers of impending autoimmunity in lacrimal glands of SjS-prone mice. Changes in TJ complex genes support an increased movement of cells through paracellular spaces.

EPR spin trapping of an oxalate-derived free radical in the oxalate decarboxylase reaction.

EPR spin trapping experiments on bacterial oxalate decarboxylase from Bacillus subtilis under turn-over conditions are described. The use of doubly (13)C-labeled oxalate leads to a characteristic splitting of the observed radical adducts using the spin trap N-tert-butyl-α-phenylnitrone linking them directly to the substrate. The radical was identified as the carbon dioxide radical anion which is a key intermediate in the hypothetical reaction mechanism of both decarboxylase and oxidase activities. X-ray crystallography had identified a flexible loop, SENS161-4, which acts as a lid to the putative active site. Site directed mutagenesis of the hinge amino acids, S161 and T165 was explored and showed increased radical trapping yields compared to the wild type. In particular, T165V shows approximately ten times higher radical yields while at the same time its decarboxylase activity was reduced by about a factor of ten. This mutant lacks a critical H-bond between T165 and R92 resulting in compromised control over its radical chemistry allowing the radical intermediate to leak into the surrounding solution.