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The utilization of animal donors of rumen fluid for laboratory experiments can raise ethical concerns, and alternatives to the collection of rumen fluids from live animals are urgently requested. The aim of this study was to compare the fresh rumen fluid (collected at slaughter, W) with that obtained from a continuous fermenter (RCF) and three methods of rumen fluid preservation (refrigeration, R, chilling, C, and freeze-drying, FD). The fermentability of different inoculum was evaluated by three in vitro tests (neutral detergent fiber (NDF) and crude protein (CP) degradability and gas production, NDFd, RDP and GP, respectively) using six feeds as substrates. Despite the two types of inoculum differed in terms of metabolites and microbiota concentration, the differences in vitro fermentability between the two liquids were less pronounced than expected (-15 and 20% for NDFd and GP when the liquid of fermenter was used and no differences for RDP). Within each in vitro test, the data obtained from rumen and from fermenter liquids were highly correlated for the six feeds, as well as between W and R (r: 0.837-0.985; p < 0.01). The low fermentative capacity was found for C and, particularly, FD for liquids. RCF could be used to generate inoculum for in vitro purposes and short-term refrigeration is a valuable practice to manage inoculum.

The aim of the study was to evaluate the use of three by-products as growing substrates for Hermetia illucens (Black Soldier Fly (BSF)) larvae: okara, maize distiller, brewer's grains, and a control hen diet. The study focused on larval growth and bioconversion performance, production of methane by larvae and environmental burden of larvae production, using Life Cycle Assessment (LCA) on a lab scale. Chemical composition of substrates differed: okara had the highest crude protein and ether extract contents, while brewer's grains showed the highest fiber content. Larvae fed on a hen diet and maize distiller exhibited the highest final weights (2.29 and 1.97 g, respectively). Larvae grown on okara showed the highest indexes for waste reduction and efficiency of conversion of the ingested feed. The BSF larvae did not produce any detectable traces of CH 4 . LCA evaluation showed that larvae production on a hen diet resulted in the most impact for most of environmental categories, for the inclusion of soybean meal in the diet (for climate change, 5.79 kg CO 2 eq/kg dry larvae). Feed production activities resulted in the main contributions to environmental impact. In order to compare the larvae production obtained on all substrates, an environmental impact was attributed to okara and brewer's grain through a substitution method, and, by this approach, the best sustainable product resulted from the larvae grown on the maize distiller.

Clostridia in the milk can lead to late blowing, a cheese defect. Clostridia are ubiquitous, deriving from both the farm environment and the feed ingested by the cows, and are transferred into the milk through faecal contamination. Our aim was to investigate the effect of different in-parlour practices on the content of anaerobic spore-forming bacteria in milk, and to monitor the variation in spore content in the feed and environment. The experiment, conducted in an experimental dairy during autumn, was repeated in exactly the same way for two consecutive years. The experimental design applied three different milking routines in three consecutive 7-d periods: forestripping alone (F); forestripping and post-dipping (F+Post); pre-dipping, wiping, forestripping and post-dipping (Pre+F+Post). Teat skin swabs and samples of feed, faeces, bedding materials and milk were collected for microbiological analyses. The dietary forage of the lactating cows included maize silage, which, in both years, was found to have the highest level of clostridial spore contamination. Pre-dipping with a detergent/emollient solution, and drying with a disposable paper towel, proved much more efficient in reducing spore contamination than forestripping alone, both on the teats (1·30 vs. 2·20 log10 MPN/swab; P < 0·001) and in the milk (1·82 vs. 2·47 log10 MPN/L, P < 0·02), while post-dipping had little influence on spore count. The standard plate count in milk was significantly lower with Pre+F+Post treatment than with F (3·80 vs. 4·51 log10 CFU/mL, P < 0·01). The teat preparation procedure did not influence the lactic acid bacterial levels in the milk, which is very positive in that decreased lactic acid bacterial content can lessen raw milk cheese quality.

To test the effects of year and processing plant on the nutritional value of canola meal (CM), 3 CM samples/yr were collected from each of 12 Canadian production plants over 4yr (total=144). Samples of CM were analyzed for differences in chemical composition and for in vitro ruminal protein degradability using the Michaelis-Menten inhibitor in vitro (MMIIV) method. In the MMIIV method, protein degradation rate (kd) was estimated by 2 methods: from net release (i.e., blank corrected) of (1) ammonia plus AA determined by o-phthaldialdehyde fluorescence (OPAF) assay or (2) ammonia, AA, plus oligopeptides determined by o-phthaldialdehyde absorbance (OPAA) assay; rumen-undegradable protein (RUP) was computed assuming passage rates of 0.16 and 0.06/h for, respectively, soluble and insoluble protein. Casein, solvent soybean meal (SSBM), and expeller soybean meal (ESBM) were included in all incubations as standard proteins. Differences among years and plants were assessed using the mixed procedures of SAS. Small but significant differences were found in CM among years for chemical composition, including N solubility; some of these differences may have been related to changes in our analytical methods over time. However, adjustment of degradation activity of individual in vitro incubations based on the mean degradation activity over all incubations yielded kd and RUP that did not differ by year using either assay. Simultaneously incubating CM samples from 2yr in the same in vitro runs confirmed that no year effects existed for kd or RUP. Differences existed in chemical composition of CM among the 12 processing plants over the 4yr of sample collection. Moreover, consistent differences in kd and RUP were observed among plants: kd ranged from 0.069 to 0.113/h (OPAA assay) and 0.075 to 0.120/h (OPAF assay), and RUP estimates ranged from 51 to 43% (OPAA assay) and 49 to 41% (OPAF assay). Regression of kd on insoluble N content of CM yielded correlation coefficients (R(2))=0.40 (OPAA assay) and 0.42 (OPAF assay), and regressions of kd on NDIN and N-fraction B3 yielded R(2)<0.02. Mean estimates from both OPAA and OPAF assays for casein, SSBM, ESBM, and CM were, respectively, kd=0.764, 0.161, 0.050, and 0.093/h and RUP=18, 33, 56, and 45%. A range of 8 percentage units from lowest to highest RUP suggests that substantial differences exist in metabolizable protein content of CM produced by different processing plants., (Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Background: Anaerobic spore-forming bacteria (ASFB) in milk derive from the farm environment, and the use of silages and management practices are the main responsible of milk ASFB contamination. The aim of this study was to evaluate the relationships between feeding, milking routine and cow hygiene and milk and Grana Padano cheese (produced with and without lysozyme) ASFB contamination., Results: The study involved 23 dairy farms. ASFB in corn silage were on average 2.34 ± 0.87 log10 MPN g(-1). For grass, Italian ryegrass and alfalfa, ASFB (log10 MPN g(-1)) were numerically higher for silages (3.22) than hays (2.85). The use of corn silages of high quality (high lactic and acetic acids concentrations) decreased the milk ASFB contamination, whilst the use of herbage silages did not affect it. The presence (>40%) of cows with dirty udders increased the ASFB contamination of milk, while forestripping had a positive effect (-9% ASFB). Ripened Grana Padano had an ASFB count below the analytical limit; Clostridium tyrobutyricum DNA was found only in wheels produced without lysozyme, which also showed late blowing., Conclusion: The factors increasing milk spore contamination were corn silage quality, cow udder hygiene and inadequate milking routine. Late blowing was present only in cheeses without lysozyme., (© 2014 Society of Chemical Industry.)

The effects of 18 essential oils, yeast, Quebracho tannin and Quillaja saponin on ruminal methane and gas production (GP) were studied in vitro. A lactating cow diet was incubated with rumen inoculum. Doses of the additives (mg/L) were: essential oils = 500; tannin = 67; yeast = 8.35 and 16.7; and saponin = 300. Lemongrass, estragole, eugenol, geraniol, limonen, thyme oil and thymol produced less gas (overall mean 33.8 mL/200 mg dry matter (DM)) than control (43.6 mL/200 mg DM; P < 0.001). Methane produced (mL/200 mg DM) by guaiacol (10.7), lemongrass (9.6), limonene (11.4), thyme oil (10.9) and thymol (2.1) was lower than control (12.5) (P < 0.001). Methane percentage on total GP was lower (P < 0.001) for cinnamic alcohol (25.4), guaiacol (24.5), thymol (19.7) and vanillin (26.3) than control (28.8). In a second experiment, thymol, guaiacol and yeast were added to the diet of dry fistulated cows to determine in situ neutral detergent fiber digestibility (NDFD) of six forages and in vivo dietary NDFD. Thymol and yeast decreased in situ NDFD after 24 h (33.9% and 33.5% vs. 38.1%; P = 0.008). Thymol decreased in vivo total tract NDFD (40.8% vs. 51.4%; P = 0.02). Differences in GP and methane levels were registered within classes of additives. A careful selection of additives may allow for the manipulation of ruminal fermentation., (© 2014 Japanese Society of Animal Science.)

Background: Camelina sativa (CS) is an oilseed crop used for biofuel production. By-products from oil extraction are high in protein and can be used in ruminant rations; more information about their nutritive value is required also considering the antinutrional factor content of the by-products. The aim of this study was to evaluate the nutritive value of CS meal genotypes in comparison with canola., Results: Ten CS genotypes and one canola cultivar were evaluated. Meals were obtained from seeds after solvent oil extraction. CS average crude protein (CP) content (g kg⁻¹ dry matter) was 457. Numerical differences in lysine and sulfur amino acid content were observed among CS genotypes. Glucosinolate (mmol kg⁻¹) content was higher for CS (23.1) than canola (7.2). Sinapine content (g kg⁻¹) was lower for CS (2.79) than for canola (4.32). Differences were observed among CS genotypes for rumen undegraded protein (RUP). Average RUP (g kg⁻¹ CP) was 316 for CS and 275 for canola., Conclusions: CS meal has potential for use in ruminant rations as a high-quality protein source. In vivo studies are needed to compare CS with other protein sources used in cattle rations. Implementation of breeding programs for improved meal quality is recommend., (© 2013 Society of Chemical Industry.)

Background: To investigate the incidence, risk factors and clinical implications of venous thromboembolism (VTE) in advanced cancer patients treated in phase I studies., Patients and Methods: Patients enrolled and treated in phase I studies conducted by SENDO (Southern Europe New Drugs Organization) Foundation between 2000 and 2010 in 15 experimental centers were considered for the study. Clinical data, including adverse events, were prospectively collected during the studies and retrospectively pooled for VTE analysis., Results: Data of 1415 patients were considered for analysis. Five hundred and twenty-six (37.2%) patients were males, and median age was 57.3 years (range: 13-85). Eighty-five percent of patients had metastatic disease, while the remaining had locally advanced irresectable disease. For 706 (49.9%) of the patients, the study treatment was with cytotoxic agent(s) only, for 314 with target therapy(ies) only, while the remaining patients received a target therapy in combination with a cytotoxic drug. Fifty-six (3.96%) patients who developed a VTE, almost all (89.3%) during the course of treatment, the remaining during the follow-up. At univariate analysis, the Khorana score, the combination of an antiangiogenic agent with a cytotoxic drug, and the time from first cancer diagnosis to study entry (as continuous variable) were associated with a statistically significant increase of VTE occurrence. The multivariate analysis confirmed only a statistically significant association for the Khorana score. The hazard ratio of VTE occurrence was 7.88 [95% confidence interval (CI) 2.86-21.70) and 2.74 (95% CI 1.27-5.92) times higher for the highest (≥3) and intermediate (1-2) scores as compared with score = 0., Conclusions: VTE is a relatively common complication among patients treated in the context of phase I studies. The Khorana score predicts VTE development and can be used to identify patients at high of VTE.

Zinc is important in a multitude of biological functions, including regulation of the immune response, modulation of keratogenesis and wound healing, maintenance of normal reproductive function, and acuity of taste and smell. Zinc-responsive dermatosis is an uncommon disease of dogs resulting from either an absolute or relative deficiency in zinc. Dermatological lesions are characterized by erythema, alopecia, scales, and crusts that primarily affect the head. Two forms of the disease exist: a familial form affecting Alaskan Malamutes and Siberian Huskies and a form that affects growing puppies fed zinc-deficient or oversupplemented diets. The history, clinical signs, and skin biopsy results are typically diagnostic. Life-long zinc supplementation is usually necessary in the familial form of the disease, although dietary correction alone may be curative in the second form. Lethal acrodermatitis is a rare inherited disorder of Bull Terriers that does not respond to zinc supplementation and is invariably fatal.

Patients with Kaposi's sarcoma (KS) have a human herpesvirus-8 (HHV-8) load higher than patients without KS and present a CD8(+) T-cell activation with production of Th1-type cytokines both in tissues and peripheral blood mononuclear cells (PBMC). Because in tissues of KS patients detection of inflammatory cytokines (IC) can precede detection of HHV-8 DNA and because signs of immunoactivation and/or dysregulation can precede KS development, we investigated the effect of IC on HHV-8 infection. To achieve this goal, PBMC and purified cell populations from 45 patients with KS and 45 patients at risk of KS were analyzed for HHV-8 DNA and/or gene expression and for cell survival, growth, and phenotype before or after culture with or without the IC increased in KS. The results indicate that PBMC that are polymerase chain reaction (PCR)-positive at day 0 generally loose the virus upon culture. However, the presence of IC maintains HHV-8 DNA load in cultured cells. In addition, IC increase viral load to detectable levels in PBMC from serologically positive patients that were PCR-negative before culture. gamma Interferon is sufficient for these effects, whereas tumor necrosis factor and interleukin-6 have little or no activity. The increase of HHV-8 DNA by IC is observed after short-term (7 days) or long-term (28 days) culture of the cells and occurs in one or both of the two circulating cell types that are infected in vivo: B cells and monocytes. In both cases it is associated with lytic gene expression, suggesting that virus reactivation is one of the most likely mechanisms for the effect of IC on virus load. However, IC have also effects on the cells target of HHV-8 infection, because they increase B-cell survival and induce the growth and differentiation of monocytes into KS-like spindle cells with markers of endothelial macrophages. Because cells with markers of endothelial macrophages are present in blood and lesions from KS patients and are infected by HHV-8, these data may explain the high HHV-8 load associated with KS development and suggest that infected monocytes may carry the virus to tissues, transmit the infection, or differentiate in loco in spindle cells with endothelial macrophage markers.

Kaposi's sarcoma (KS) is an angioproliferative disease associated with infection by the human herpesvirus-8 (HHV-8). HHV-8 possesses genes including homologs of interleukin-8 (IL-8) receptor, Bcl-2, and cyclin D, which can potentially transform the host cell. However, the expression of these genes in KS tissues is very low or undetectable and HHV-8 does not seem to transform human cells in vitro. In addition, KS may not be a true cancer at least in the early stage. This indicated that besides its transforming potential, HHV-8 may act in KS pathogenesis also through indirect mechanisms. Evidence suggests that KS may start as an inflammatory-angiogenic lesion mediated by cytokines. However, little is known on the nature of the inflammatory cell infiltration present in KS, on the type of cytokines produced and on their role in KS, and whether this correlates with the presence of HHV-8. Here we show that both acquired immunodeficiency syndrome (AIDS)-KS and classical KS (C-KS) lesions are infiltrated by CD8+ T cells and CD14+/CD68+ monocytes-macrophages producing high levels of gamma-interferon (gamma IFN) which, in turn, promotes the formation of KS spindle cells with angiogenic phenotype. gamma IFN, in fact, induces endothelial cells to acquire the same features of KS cells, including the spindle morphology and the pattern of cell marker expression. In addition, endothelial cells activated by gamma IFN induce angiogenic lesions in nude mice closely resembling early KS. These KS-like lesions are accompanied by production of basic fibroblast growth factor, an angiogenic factor highly expressed in primary lesions that mediates angiogenesis and spindle cell growth. The formation of KS-like lesions is upregulated by the human immunodeficiency virus Tat protein demonstrating its role as a progression factor in AIDS-KS. Finally, gamma IFN and HLA-DR expression correlate with the presence of HHV-8 in lesional and uninvolved tissues from the same patients. As HHV-8 infects both mononuclear cells infiltrating KS lesions and KS spindle cells, these results suggest that HHV-8 may elicit or participate in a local immune response characterized by infiltration of CD8+ T cells and intense production of gamma IFN which, in turn, plays a key role in KS development.

Objective: To determine the response rate of zinc-responsive dermatosis to zinc supplementation, the optimal dosage of zinc required for resolution of lesions, the rate of recurrence of lesions, and to develop recommendations for maintenance dosages of zinc to be administered to dogs with this type of zinc-responsive dermatosis., Design: Retrospective case series., Animals: 17 northern-breed dogs with a diagnosis of zinc-responsive dermatosis., Procedure: Histologic evaluation of skin biopsy specimens and review of medical records. Additional information was obtained from veterinarians and owners via a telephone questionnaire., Results: In 12 of 17 dogs, lesions were unilateral initially, then became symmetrical as the disease progressed. Pyoderma was evident in 5 of 17 dogs, whereas 10 were pruritic. Most lesions initially developed between September and April, and 12 of 17 dogs developed lesions in February, October, and November. Initial dosages of zinc supplement ranged from 0.8 to 4.6 mg/kg of body weight/d (0.36 to 2.09 mg/lb/d). Effective/ maintenance dosages ranged from 0.5 mg/kg (0.23 mg/lb), twice weekly, to 8.0 mg/kg/d (3.6 mg/lb/d). Fifteen of 17 dogs had complete resolution of lesions after zinc supplementation. Lesions recurred in 9 of 16 dogs. Approximately half of the recurrent lesions were a result of a missed dose or a decrease in dosage or frequency of zinc supplementation., Clinical Implications: An initial dosage of zinc supplement of 1.0 mg of elemental zinc/kg (0.45 mg of elemental zinc/lb), PO, every 24 hours is recommended. Treatment should be continued for 1 month to determine response to treatment, and the daily dosage should be increased by 50% if the initial dosage is not effective. Dogs are prone to recurrence of lesions if a dose of zinc is missed or the dosage or frequency is decreased.

Human herpesvirus 8 (HHV-8) is a recently discovered, virus that is highly associated with Kaposi's sarcoma (KS) and AIDS-associated body cavity lymphomas, although it is also found in some normal individuals. HHV-8 is related by nucleotide sequence homology to herpesvirus saimiri (HVS), which causes T cell lymphomas in some New World monkeys, and to Epstein-Barr virus (EBV), a human herpesvirus linked etiologically with Burkitt's lymphoma and nasopharyngeal carcinoma. We report that, like HVS but unlike EBV, HHV-8 contains a gene (ORF74) with significant sequence homology to the high-affinity IL-8 receptor, a member of the alpha (CXC) chemokine receptor family of transmembrane G protein-coupled receptors. We also show by reverse transcription PCR that the chemokine receptor-related HHV-8 gene is detectable in some RNA samples from KS tissue, and that its expression varies independently from that of ORF26, a minor capsid protein. The presence of a potential chemokine receptor in HHV-8 and its expression in KS tissue suggests that it may be important in the regulation of viral gene expression and may play a role in the etiology of KS and AIDS-related body cavity lymphomas.

Basic fibroblast growth factor (bFGF) and human immunodeficiency virus type 1 (HIV-1) Tat protein synergize in inducing angiogenic Kaposi's sarcoma-like lesions in mice. Synergy is due to Tat, which enhances endothelial cell growth and type-IV collagenase expression in response to bFGF mimicking extracellular matrix proteins. The bFGF, extracellular Tat and Tat receptors are present in HIV-1-associated KS, which may explain the higher frequency and aggressiveness of this form compared to classical Kaposi's sarcoma where only bFGF is present.

In the present study the requirements for in vitro infection of antigen-specific CD8+ cytotoxic T lymphocytes (CTL) with human immunodeficiency virus -1(HIV-1) were investigated. CD3+CD8+CD4- HIV-1 nef-specific CTL become infected with HIV-1 after short-term co-culture with HLA-matched HIV-1-infected CD20+ B lymphoblastoid cells (B-LCL) which are specifically killed. Similar results were observed with an allospecific CD8+ CTL population. In addition, co-culture experiments showed that once infected with HIV-1, these CD8+ CTL could spread the infection further to uninfected CD4+ lymphocytes. In contrast, CD8+ CTL did not become infected with HIV-1 when co-cultured with HLA-mismatched HIV-1-infected B-LCL which are not killed. These observations in vitro could have relevance in peripheral lymphoid organs contributing to the progressive decrease of HIV-specific CD8+ CTL activity that is associated with the progression to AIDS.

Human T-cell leukaemia-lymphoma virus (HTLV) is an exogenous human retrovirus distinct from all known animal retroviruses. HTLV is closely linked to a subtype of adult T-cell malignancies and except for isolated cases, has not been found associated with any other form of leukaemia, lymphoma or other cancers (see refs 1, 2 for review). HTLV can be transmitted to cord blood T lymphocytes in vitro and the infected cells exhibit characteristics of transformed neoplastic T cells. We have recently cloned DNA sequences derived from approximately 1 kilobase (kb) of the 5' and 3' termini of the HTLV genome, as well as a 4-5-kb defective HTLV provirus flanked by cellular sequences. The availability of these probes has enabled us to carry out a limited survey of different fresh or cultured cells from patients of different lymphoid and myeloid malignancies for HTLV-related DNA sequences. The results presented here show that cells from all Japanese patients with adult T-cell leukaemia and several patients with various mature T-cell malignancies from elsewhere contained one or more copies of a highly conserved HTLV genome. The infected cells are of clonal origin. Fresh cells from 1 of the 10 myeloid leukaemic patients contained exogenous DNA sequences distantly related to HTLV.