Role of Calf-Adapted Escherichia coli in Maintenance of Antimicrobial Drug Resistance in Dairy Calves

The prevalence of antimicrobial drug-resistant bacteria is typically highest in younger animals, and prevalence is not necessarily related to recent use of antimicrobial drugs. In dairy cattle, we hypothesize that antimicrobial drug-resistant, neonate-adapted bacteria are responsible for the observed high frequencies of resistant Escherichia coli in calves. To explore this issue, we examined the age distribution of antimicrobial drug-resistant E. coli from Holstein cattle at a local dairy and conducted an experiment to determine if low doses of oxytetracycline affected the prevalence of antimicrobial drug-resistant E. coli. Isolates resistant to tetracycline (>4 μg/ml) were more prevalent in <3-month-old calves (79%) compared with lactating cows (14%). In an experimental trial where calves received diets supplemented with or without oxytetracycline, the prevalence of tetracycline-resistant E. coli was slightly higher for the latter group (P = 0.039), indicating that drug use was not required to maintain a high prevalence of resistant E. coli. The most common resistance pattern among calf E. coli isolates included resistance to streptomycin (>12 μg/ml), sulfadiazine (>512 μg/ml), and tetracycline (>4 μg/ml) (SSuT), and this resistance pattern was most prevalent during the period when calves were on milk diets. To determine if prevalence was a function of differential fitness, we orally inoculated animals with nalidixic acid-resistant strains of SSuT E. coli and susceptible E. coli. Shedding of SSuT E. coli was significantly greater than that of susceptible strains in neonatal calves (P < 0.001), whereas there was no difference in older animals (P = 0.5). These data support the hypothesis that active selection for traits linked to the SSuT phenotype are responsible for maintaining drug-resistant E. coli in this population of dairy calves.     

Assessment of a brown midrib (BMR) mutant gene on the nutritive value of sudangrass using in vitro and in vivo techniques

The brown midrib (BMR) gene has been reported to reduce the lignin concentration in plants, which contributed to increased fiber digestion in ruminants. Three studies were completed to compare the digestibility of a BMR mutant of sudangrass (sorghum bicolor subsp. Drummondii) versus a non-BMR (‘Piper’) variety when included in diets fed to sheep (Study 1), to complete a rumen in vitro assessment of sheep and lactating cow diets (Study 2), and to compare digestibility when included in the diet fed to lactating dairy cows (Study 3). Four wether sheep were used in a 2 × 2 Latin square experiment (Study 1) with total fecal collection to determine total tract apparent digestibility of pelleted Piper (P) and BMR (P-BMR) sudangrass hays. Forage pellets consisted of either P-BMR or P hay with added urea to meet the maintenance crude protein (CP) requirement of the sheep. Digestibility of organic matter (OM; P<0.01), dry matter (DM; P<0.01), acid detergent fiber (ADF; P<0.05), and neutral detergent fiber (aNDFom; P<0.07) was higher for P-BMR than P sudangrass. In vitro rumen digestibility of aNDFom using cattle rumen fluid was higher at 24 (P<0.01), 48 (P<0.01) and 72 h (P<0.01) of fermentation for P-BMR versus P (Study 2). Four lactating Holstein dairy cows (251 ± 30 days in milk) and fitted with ruminal and duodenal cannulae were used in a 4 × 4 Latin square experiment. Total mixed rations (TMR) contained 180 g/kg DM shredded sudangrass hay and 180 g/kg sliced alfalfa hay, but the proportion of P to P-BMR sudangrass varied as 100:0, 66:34, 34:66, or 0:100. Yields of milk and milk protein were highest at the 66:34 level (Quadratic: P=0.06 and 0.07, respectively), but composition of milk fat, protein and lactose, as well as DM intake, did not differ (Study 3), probably because forestomach and total tract apparent digestion of aNDFom and OM did not differ due to sudangrass source.     

Effects of a combination of fibrolytic and amylolytic enzymes on ruminal enzyme activities,bacterial diversity,blood profile and milk production in dairy cows

We hypothesised that adding a combination of fibrolytic and amylolytic enzymes to the diet of early-lactation dairy cows would improve rumen enzyme activity and bacterial diversity, promote energy metabolism, and benefit milk production in cows. Twenty multiparous early-lactation (90 ± 5 d) Holstein cows with similar body conditions were randomly allocated to control (CON, n = 10) and experimental (EXP, n = 10) groups in a completely randomised single-factor design. The CON was fed only a basal total mixed ration diet, and the diet of the EXP was supplemented with a combination of fibrolytic and amylolytic enzymes at 70 g/cow/d (cellulase 3 500 CU/g, xylanase 2 000 XU/g, β-glucanase 17 500 GU/g, and amylase 37 000 AU/g). The experiment lasted 28 days, with 21 days for adaptation and 7 days for sampling. Enzyme addition increased the activity levels of α-amylase and xylanase, and the ammonia-N concentration (P < 0.05) tended to increase the activity of β-glucanase (P = 0.08) in rumen fluid. However, there was no significant difference in the rumen bacterial richness and diversity, phylum (richness > 0.1%) or genus (richness > 1%) composition between the CON and EXP groups (P > 0.05). A tendency of difference was found between CON and EXP (R = 0.22, P = 0.098) in principal component analysis. Ten genera showed different abundances across the CON and EXP groups (linear discriminant analysis effect size, linear discriminant analysis > 2). EXP increased the ratio of albumin to globulin and the concentrations of total cholesterol and low-density lipoprotein cholesterol (P < 0.05) and tended to increase triglycerides (P = 0.09) in blood. Milk yield, 3.5% fat-corrected milk yield and energy-corrected milk yield increased with enzyme supplementation (P < 0.05). The production levels of milk fat and lactose increased, but the percentage of solids, not fat and protein, decreased in EXP (P < 0.05). Although the DM intake was not affected, the feed efficiency tended to increase (P = 0.07) in EXP. In conclusion, dietary supplementation with a mixture of fibrolytic and amylolytic enzymes on multiparous early-lactation dairy cows increased α-amylase and xylanase activity levels in rumen fluid, enhanced milk performance and tended to improve the feed efficiency in cows.     

Feeding a High Concentrate Diet Down-Regulates Expression of ACACA,LPL and SCD and Modifies Milk Composition in Lactating Goats

High concentrate diets are fed to early and mid-lactation stages dairy ruminants to meet the energy demands for high milk production in modern milk industry. The present study evaluated the effects of a high concentrate diet on milk fat and milk composition, especially, cis-9, trans-11 CLA content in milk and gene expression of lactating goats. Eight mid-lactating goats with rumen fistula were randomly assigned into a high concentrate diet (HCD) group and low concentrate diet (LCD) group. High concentrate diet feeding significantly increased lipopolysaccharides (LPS) in plasma and decreased milk fat content, vaccenic acid (VA) and cis-9, trans-11 CLA in milk of the lactating goats. The mRNA expression levels of sterol regulatory element binding protein B 1c (SREBP1c), lipoprotein lipase (LPL), fatty acid synthetase (FASN) and acetyl-CoA carboxylase α (ACACA, ACCα) involving in lipid metabolism were analyzed, and ACACA and LPL all decreased in their expression level in the mammary glands of goats fed a high concentrate diet. DNA methylation rate of stearoyl-CoA desaturase (SCD) was elevated and decreased, and SCD mRNA and protein expression was reduced significantly in the mammary glands of goats fed a high concentrate diet. In conclusion, feeding a high concentrate diet to lactating goats decreases milk fat and reduced expression of SCD in the mammary gland, which finally induced cis-9, trans-11 CLA content in milk.     

Inclusion of cocoa by-product in the diet of dairy sheep: Effect on the fatty acid profile of ruminal content and on the composition of milk and cheese

In this study, we hypothesized that dietary cocoa bean shell (CBS) as a partial replacer of human edible cereal grains in the diet of lactating ewes may affect performance and milk and cheese composition. Twenty Comisana lactating ewes allotted into control (CTRL; n = 10) or cocoa (CBS; n = 10) group received alfalfa hay ad libitum and 800 g of conventional (CTRL) or experimental (CBS) concentrate containing 11.7% CBS to partially replace corn and barley of the CTRL concentrate. Milk yield and composition did not differ between groups, and only urea concentration was lower in CBS milk. Dietary CBS increased cheese fat and reduced protein percentage in CBS group. Fatty acid composition of rumen content partially reflected that of the ingested diet, with total saturated fatty acids (SFA), total monounsaturated fatty acids (MUFA), 16:0, 18:0 and 18:1c9 greater in the CBS group. Moreover, all the identified trans- and cis-18:1 isomers were greater in CBS rumen content. Milk and cheese showed a similar fatty acid composition. Total MUFAs were greater in milk and cheese of CBS, mainly due to the proportion of 18:1c9, and conversely, total polyunsaturated fatty acids (PUFA), PUFAn-6 and PUFAn-6-to-PUFAn-3 ratio was greater in CTRL group. Concluding, the inclusion of CBS in the diet of lactating ewes within the limit imposed by the current legislation did not cause detrimental effects on animal performance and milk composition. Interestingly, dietary CBS reduced milk urea concentration probably due to the phenols contained in CBS concentrate. However, our results support that biohydrogenation was weakly impaired by dietary CBS. Finally, CBS negatively affected cheese nutritional characteristics due to lower protein and greater fat content, but improved fat health indexes in milk and cheese.     

Does Dietary Mitigation of Enteric Methane Production Affect Rumen Function and Animal Productivity in Dairy Cows?

It has been suggested that the rumen microbiome and rumen function might be disrupted if methane production in the rumen is decreased. Furthermore concerns have been voiced that geography and management might influence the underlying microbial population and hence the response of the rumen to mitigation strategies. Here we report the effect of the dietary additives: linseed oil and nitrate on methane emissions, rumen fermentation, and the rumen microbiome in two experiments from New Zealand (Dairy 1) and the UK (Dairy 2). Dairy 1 was a randomized block design with 18 multiparous lactating cows. Dairy 2 was a complete replicated 3 x 3 Latin Square using 6 rumen cannulated, lactating dairy cows. Treatments consisted of a control total mixed ration (TMR), supplementation with linseed oil (4% of feed DM) and supplementation with nitrate (2% of feed DM) in both experiments. Methane emissions were measured in open circuit respiration chambers and rumen samples were analyzed for rumen fermentation parameters and microbial population structure using qPCR and next generation sequencing (NGS). Supplementation with nitrate, but not linseed oil, decreased methane yield (g/kg DMI; P<0.02) and increased hydrogen (P<0.03) emissions in both experiments. Furthermore, the effect of nitrate on gaseous emissions was accompanied by an increased rumen acetate to propionate ratio and consistent changes in the rumen microbial populations including a decreased abundance of the main genus Prevotella and a decrease in archaeal mcrA (log₁₀ copies/ g rumen DM content). These results demonstrate that methane emissions can be significantly decreased with nitrate supplementation with only minor, but consistent, effects on the rumen microbial population and its function, with no evidence that the response to dietary additives differed due to geography and different underlying microbial populations.     

Long-Term Effects of Subacute Ruminal Acidosis (SARA) on Milk Quality and Hepatic Gene Expression in Lactating Goats Fed a High-Concentrate Diet

Purpose

The mechanism underlying the decline in milk quality during periods of feeding high-concentrate diets to dairy ruminants is not well documented. The aim of this study was to investigate the metabolic changes in the liver that contribute to the input of substrate precursors to the mammary gland after feeding a high-concentrate diet to lactating goats for a long period.

Experimental Design

Eight mid-lactating goats with rumen cannulas were randomly assigned to two groups. For 9 weeks, the treatment group was fed a high-concentrate diet (60% concentrate of dry matter, HC) and the control group was fed a low-concentrate diet (40% concentrate of dry matter, LC). Ruminal fluid, plasma, and liver tissues were sampled, microarray techniques and real-time polymerase chain reaction were used to evaluate metabolic parameters and gene expression in liver.

Results

Feeding a 60%-concentrate diet for 9 weeks resulted in a significant decrease in rumen pH. Changes in fat and protein content also occurred, which negatively affected milk quality. Plasma levels of leptin (p = 0.058), non-esterified fatty acid (p = 0.071), and glucose (p = 0.014) increased markedly in HC group. Plasma cortisol concentration was significantly elevated in the treatment group (p<0.05). Expression of the glucocorticoid receptor protein gene was significantly down-regulated (p<0.05) in the liver. The expression of genes for interleukin 1β, serum amyloid A, C-reactive protein, and haptoglobin mRNA was significantly increased (p<0.05) in the HC group. GeneRelNet analysis showed that gene expression involved in inflammatory responses and the metabolism of lipids, protein, and carbohydrate were significantly altered by feeding a high-concentrate diet for 9 weeks.

Conclusions

Activation of the acute phase response and the inflammatory response may contribute to nutrient partitioning and re-distribution of energy in the liver, and ultimately lead to a decline in milk quality.     

Ethnic Variations in Central Corneal Thickness in a Rural Population in China: The Yunnan Minority Eye Studies

Purpose

To describe the ethnic differences in central corneal thickness (CCT) in population-based samples of ethnic Bai, Yi and Han people living in rural China.

Methods

6504 adults (2119 ethnic Bai, 2202 ethnic Yi and 2183 ethnic Han) aged 50 years or older participated in the study. Each subject underwent standardized ocular examinations and interviewer-administered questionnaires for risk factor assessment. CCT was measured for both eyes using an ultrasound pachymeter. Regression and principal component analysis were performed to examine the relationship of ethnicity and other factors with CCT.

Results

The mean CCT readings were 536.4 ± 34.2 μm in ethnic Bai, 532.1 ± 32.1 μm in ethnic Yi and 529.6 ± 32.7 μm in ethnic Han adults (P<0.001), respectively. There was a decreasing trend of mean CCT with increasing age across all ethnic groups. In multivariate linear regression models, increasing CCT was associated with younger age (P<0.001), male gender (P<0.001), Bai (P<0.001) or Yi (P<0.001) ethnicity, greater body mass index (P<0.001), higher systolic blood pressure (P<0.001), greater corneal curvature (P<0.001), deeper anterior chamber (P < 0.001), and thicker lens (P<0.001). Ethnicity contributed significantly to presence of thin cornea (60%; P< 0.001) compared with other factors. CCT had similar impact on intraocular pressure readings across all ethnic groups.

Conclusions

This study of more than 6500 multiethnic participants demonstrates significant ethnic variations in CCT, with Han ethnicity having the thinnest cornea compared with ethnic minorities. These data are essential to guide future multiethnic clinical trials on CCT-related ocular conditions such as glaucoma.     

Effects of a ruminally protected B-vitamin supplement on milk yield and composition of lactating dairy cows

It is not clear if B vitamins supplied to the small intestine of dairy cows from dietary and rumen microbial sources are provided in sufficient quantity to maximize animal performance. Our objective was to determine effects of adding a ruminally protected B vitamin blend supplement, containing biotin, folic acid, pantothenic acid and pyridoxine, to the diet of high producing dairy cows on their productivity. Two dairy facilities located in California (USA) were used, one with mid lactation Holstein cows (Experiment 1) and the other with early lactation Holstein cows (Experiment 2). In each Experiment, cows were randomly assigned to treatment in a 2 × 2 crossover design with 28 d (Experiment 1) or 35 d (Experiment 2) experimental periods. In Experiment 1, milk and milk fat yield were unaffected by treatment, although milk fat proportion was lower (37.1 versus 36.3 g/kg; P<0.01), but milk protein yield was higher (1.21 versus 1.24 kg/d; P=0.02) in cows fed B vitamins. In Experiment 2, milk (39.60 versus 40.46 kg/d; P=0.02), milk fat (1.40 versus 1.47 kg/d; P<0.01) and milk protein yield (1.10 versus 1.16 kg/d; P<0.01), as well as milk energy output (113.2 versus 117.8 MJ/d; P<0.01) were all higher with B vitamin feeding. Body condition score (BCS) increased more with B vitamin feeding in Experiment 2, but was unaffected in Experiment 1. Body locomotion score (BLS) increased with B vitamin feeding in both experiments (P=0.01 and < 0.01, respectively), possibly an indication of reduced locomotory ability. Overall, productivity of high producing lactating dairy cows responded positively to feeding a mixture of ruminally protected B vitamins, although differences in the extent of the positive responses between experiments perhaps suggests that early lactation cows, with lower DM intake to milk yield ratios, may be more responsive to ruminally protected B vitamins than mid lactation cows, with higher DM intake to milk yield ratios.     

Pten Regulates Development and Lactation in the Mammary Glands of Dairy Cows

Pten is a tumor suppressor gene regulating many cellular processes, including growth, adhesion, and apoptosis. In the aim of investigating the role of Pten during mammary gland development and lactation of dairy cows, we analyzed Pten expression levels in the mammary glands of dairy cows by using western blotting, immunohistochemistry, and quantitative polymerase chain reaction (qPCR) assays. Dairy cow mammary epithelial cells (DCMECs) were used to study the function of Pten in vitro. We determined concentrations of β-casein, triglyceride, and lactose in the culture medium following Pten overexpression and siRNA inhibition. To determine whether Pten affected DCMEC viability and proliferation, cells were analyzed by CASY-TT and flow cytometry. Genes involved in lactation-related signaling pathways were detected. Pten expression was also assessed by adding prolactin and glucose to cell cultures. When Pten was overexpressed, proliferation of DCMECs and concentrations for β-casein, triglyceride, and lactose were significantly decreased. Overexpression of Pten down-regulated expression of MAPK, CYCLIN D1, AKT, MTOR, S6K1, STAT5, SREBP1, PPARγ, PRLR, and GLUT1, but up-regulated 4EBP1 in DCMECs. The Pten siRNA inhibition experiments revealed results that opposed those from the gene overexpression experiments. Introduction of prolactin (PRL) increased secretion of β-casein, triglyceride, and lactose, but decreased Pten expression levels. Introduction of glucose also increased β-casein and triglyceride concentrations, but did not significantly alter Pten expression levels. The Pten mRNA and protein expression levels were decreased 0.3- and 0.4-fold in mammary glands of lactating cows producing high quality milk (milk protein >3.0%, milk fat >3.5%), compared with those cows producing low quality milk (milk protein <3.0%, milk fat <3.5%). In conclusion, Pten functions as an inhibitor during mammary gland development and lactation in dairy cows. It can down-regulate DCMECs secretion of β-casein, triglyceride, and lactose, and plays a critical role in lactation related signaling pathways.     

Influence of exogenous enzymes on nutrient digestibility, extent of ruminal fermentation as well as milk production and composition in dairy cows

This experiment studied effects of a mixture of exogenous enzymes (ZADO^®) from anaerobic bacteria on ruminal fermentation, feed intake, digestibility, as well as milk production and composition in cows fed total mixed rations (TMRs; 0.7 corn silage and 0.3 of a concentrate mixture). Twenty lactating multiparous Brown Swiss cows (500 ± 12.4 kg live weight) were randomly assigned into two experimental groups of 10 immediately after calving and fed a TMR with or without (CTRL) addition of 40 g/cow/d of enzymes for 12 weeks. Addition of enzymes increased (P<0.05) rumen microbial N synthesis. Intake of dry matter (DM) and organic matter (OM) was positively influenced (P<0.05) by supplementation, and digestibility of all nutrients was higher (P<0.05) in the total tract of supplemented cows, although the magnitude of the improvement varied among nutrients, with the highest improvement in aNDFom and ADFom (418–584 and 401–532 g/kg respectively; P<0.05) than the other nutrients. Supplementation of enzymes also increased (P<0.05) rumen ammonia N and total short chain fatty acid (SCFA) concentrations, and individual SCFA proportions were also altered with an increase in acetate (61.0–64.8 mol/100 mol; P=0.05) before feeding, and acetate and propionate increased 3 h post-feeding (60.0–64.0 and 18.3–20.8 mol/100 mol respectively; P<0.05). Milk and milk protein production was higher (12.8–15.7 and 0.45–0.57 kg/d respectively; P<0.05) for cows fed the ZADO^® supplemented diet. This exogenous enzyme product, supplemented daily to the TMR of cows in early lactation, increased milk production due to positive effects on nutrient intake and digestibility, extent of ruminal fermentation and microbial protein synthesis.     

Effects of feeding cows in early lactation with soy hulls as partial forage replacement on heat production,retained energy and performance

This study measured the effects of replacing corn silage and vetch hay by soy hulls in total mixed rations (TMRs) fed to 25 pairs of cows through 90 d in milk, on dry matter (DM) intake, in vivo digestibility, milk yield and composition, onset of normal estrous activity, body condition score (BCS), health and the energy balance of lactating cows. The partitioning of metabolizable energy (ME) intake between heat production (HP) and retained energy (RE) in milk and body change of each cow was measured. The two TMRs differed in the content of forage and forage aNDFom [235 g/kg versus 350 g/kg; and 128 g/kg versus 187 g/kg DM, in the experimental (EXP) and control (CON) diets, respectively]. This was reflected by an increase in voluntary DM intake by 7.2% (P=0.02) in the EXP group as compared with the CON. In vivo DM and aNDFom digestibility were 4.9% (P=0.03) and 22.7% higher (P=0.01), respectively, in the EXP group than in the CON. The higher DM intake and digestibility of the EXP TMR were reflected by a concomitant increase of 7.4% in milk yield and 10.8% in RE (P=0.01) of the EXP cows as compared with the CON. The two dietary groups expressed similar somatic cell counts, and metabolic disorders (i.e., ketosis and/or lameness), as well as pedometer activity (steps/h) suggesting similar udder health, behavior and animal welfare. A trend to an earlier return to normal ovarian activity occurred in the EXP cows as reflected by fewer days to 1st ovulation and advanced outset of estrous cycle. Despite the higher RE of the EXP cows, the HP of both groups was maintained at an upper level of 141–136 MJ/cow/d during the 90 d of the experiment.     

Cellular and Physiological Effects of Dietary Supplementation with β-Hydroxy-β-Methylbutyrate (HMB) and β-Alanine in Late Middle-Aged Mice

There is growing evidence that severe decline of skeletal muscle mass and function with age may be mitigated by exercise and dietary supplementation with protein and amino acid ingredient technologies. The purposes of this study were to examine the effects of the leucine catabolite, beta-hydroxy-beta-methylbutyrate (HMB), in C₂C₁₂ myoblasts and myotubes, and to investigate the effects of dietary supplementation with HMB, the amino acid β-alanine and the combination thereof, on muscle contractility in a preclinical model of pre-sarcopenia. In C₂C₁₂ myotubes, HMB enhanced sarcoplasmic reticulum (SR) calcium release beyond vehicle control in the presence of all SR agonists tested (KCl, P<0.01; caffeine, P = 0.03; ionomycin, P = 0.03). HMB also improved C₂C₁₂ myoblast viability (25 μM HMB, P = 0.03) and increased proliferation (25 μM HMB, P = 0.04; 125 μM HMB, P<0.01). Furthermore, an ex vivo muscle contractility study was performed on EDL and soleus muscle from 19 month old, male C57BL/6nTac mice. For 8 weeks, mice were fed control AIN-93M diet, diet with HMB, diet with β-alanine, or diet with HMB and β-alanine. In β-alanine fed mice, EDL muscle showed a 7% increase in maximum absolute force compared to the control diet (202 ± 3vs. 188± 5 mN, P = 0.02). At submaximal frequency of stimulation (20 Hz), EDL from mice fed HMB plus β-alanine showed an 11% increase in absolute force (88.6 ± 2.2 vs. 79.8 ± 2.4 mN, P = 0.025) and a 13% increase in specific force (12.2 ± 0.4 vs. 10.8 ± 0.4 N/cm², P = 0.021). Also in EDL muscle, β-alanine increased the rate of force development at all frequencies tested (P<0.025), while HMB reduced the time to reach peak contractile force (TTP), with a significant effect at 80 Hz (P = 0.0156). In soleus muscle, all experimental diets were associated with a decrease in TTP, compared to control diet. Our findings highlight beneficial effects of HMB and β-alanine supplementation on skeletal muscle function in aging mice.     

Prawn Shell Chitosan Has Anti-Obesogenic Properties,Influencing Both Nutrient Digestibility and Microbial Populations in a Pig Model

The potential of natural products to prevent obesity have been investigated, with evidence to suggest that chitosan has anti-obesity effects. The current experiment investigated the anti-obesity potential of prawn shell derived chitosan on a range of variables relevant to obesity in a pig model. The two dietary treatment groups included in this 63 day study were: T1) basal diet and T2) basal diet plus 1000 ppm chitosan (n = 20 gilts per group (70 ± 0.90 kg). The parameter categories which were assessed included: performance, nutrient digestibility, serum leptin concentrations, nutrient transporter and digestive enzyme gene expression and gut microbial populations. Pigs offered chitosan had reduced feed intake and final body weight (P< 0.001), lower ileal digestibility of dry matter (DM), gross energy (GE) (P< 0.05) and reduced coefficient of apparent total tract digestibility (CATTD) of gross energy and nitrogen (P<0.05) when compared to the basal group. Fatty acid binding protein 2 (FABP2) gene expression was down-regulated in pigs offered chitosan (P = 0.05) relative to the basal diet. Serum leptin concentrations increased (P< 0.05) in animals offered the chitosan diet compared to pigs offered the basal diet. Fatness traits, back-fat depth (mm), fat content (kg), were significantly reduced while lean meat (%) was increased (P<0.05) in chitosan supplemented pigs. Pigs offered chitosan had decreased numbers of Firmicutes in the colon (P <0.05), and Lactobacillus spp. in both the caecum (P <0.05) and colon (P <0.001). Bifidobacteria populations were increased in the caecum of animals offered the chitosan diet (P <0.05). In conclusion, these findings suggest that prawn shell chitosan has potent anti-obesity/body weight control effects which are mediated through multiple biological systems in vivo.     

Wet brewers' grains or bean curd pomance as partial replacement of soybean meal for lactating cows

The purpose of this study was to evaluate the effects of supplementation of the by-product, brewers' grains and the bean curd pomance, on the performance of lactating cows and their ruminal characteristics. Through this, we wanted to increase resource utilization and to eliminate pollution from these by-products. Thirty-two Holstein lactating cows were allocated randomly into four dietary treatment groups. The experimental diets were formulated to be isoenergetic and isonitrogenous according to National Research Council (1989), and containing 35% corn silage, 20% alfalfa hay and 45% concentrates on DM basis. The dietary treatments consisted of the inclusion of different by-products sources. These included soybean bean (as the control), brewers' grains, bean curd pomance, and the mixed by-products (containing the same amount of brewers' grains and bean curd pomance). The experimental feeding lasted for eight weeks after one week of adaptation. In addition, four rumen cannulated Holstein cows were used in a 4×4 Latin square with 10-day period for collecting rumen samples. Results showed that cows that were fed the bean curd pomance diet produced significantly more milk than those that were fed the brewers' grains diet or the mixed by-product diet. The cows on the control diet produced significantly less milk than the other treatment groups (P<0.05). Both the control and brewers' grains groups consumed more feed than those in the bean curd pomance and mixed by-products diet groups (P<0.05). The cows that were fed the mixed by-products diet produced a significantly lower percentage of milk fat, total solids, milk protein and milk lactose than the others (P<0.05). The milk lactose percentage was significantly higher for cows that were fed the bean curd pomance than those fed the brewers' grains diet (P<0.05). Cows that were fed the bean curd pomance produced significantly more milk fat, total solids, milk protein and milk lactose than the others (P<0.05). Cows that were fed the mixed by-products diet produced significantly lower amount of milk fat, total solids and milk protein than the control and brewers' grains group (P<0.05). These diets did not significantly influence the body weight of the cows. It, however, significantly influenced the ruminal characteristics (P<0.05).     

Studies of Cream Seeded Carioca Beans (Phaseolus vulgaris L.) from a Rwandan Efficacy Trial: In Vitro and In Vivo Screening Tools Reflect Human Studies and Predict Beneficial Results from Iron Biofortified Beans

Iron (Fe) deficiency is a highly prevalent micronutrient insufficiency predominantly caused by a lack of bioavailable Fe from the diet. The consumption of beans as a major food crop in some populations suffering from Fe deficiency is relatively high. Therefore, our objective was to determine whether a biofortified variety of cream seeded carioca bean (Phaseolus vulgaris L.) could provide more bioavailable-Fe than a standard variety using in-vivo (broiler chicken, Gallus gallus) and in-vitro (Caco-2 cell) models. Studies were conducted under conditions designed to mimic the actual human feeding protocol. Two carioca-beans, a standard (G4825; 58μg Fe/g) and a biofortified (SMC; 106μg Fe/g), were utilized. Diets were formulated to meet the nutrient requirements of Gallus gallus except for Fe (33.7 and 48.7μg Fe/g, standard and biofortified diets, respectively). In-vitro observations indicated that more bioavailable-Fe was present in the biofortified beans and diet (P<0.05). In-vivo, improvements in Fe-status were observed in the biofortified bean treatment, as indicated by the increased total-body-Hemoglobin-Fe, and hepatic Fe-concentration (P<0.05). Also, DMT-1 mRNA-expression was increased in the standard bean treatment (P<0.05), indicating an upregulation of absorption to compensate for less bioavailable-Fe. These results demonstrate that the biofortified beans provided more bioavailable Fe; however, the in vitro results revealed that ferritin formation values were relatively low. Such observations are indicative of the presence of high levels of polyphenols and phytate that inhibit Fe absorption. Indeed, we identified higher levels of phytate and quercetin 3–glucoside in the Fe biofortified bean variety. Our results indicate that the biofortified bean line was able to moderately improve Fe-status, and that concurrent increase in the concentration of phytate and polyphenols in beans may limit the benefit of increased Fe-concentration. Therefore, specific targeting of such compounds during the breeding process may yield improved dietary Fe-bioavailability. Our findings are in agreement with the human efficacy trial that demonstrated that the biofortified carioca beans improved the Fe-status of Rwandan women. We suggest the utilization of these in vitro and in vivo screening tools to guide studies aimed to develop and evaluate biofortified staple food crops. This approach has the potential to more effectively utilize research funds and provides a means to monitor the nutritional quality of the Fe-biofortified crops once released to farmers.     

TNFα Altered Inflammatory Responses,Impaired Health and Productivity,but Did Not Affect Glucose or Lipid Metabolism in Early-Lactation Dairy Cows

Inflammation may be a major contributing factor to peripartum metabolic disorders in dairy cattle. We tested whether administering an inflammatory cytokine, recombinant bovine tumor necrosis factor-α (rbTNFα), affects milk production, metabolism, and health during this period. Thirty-three Holstein cows (9 primiparous and 24 multiparous) were randomly assigned to 1 of 3 treatments at parturition. Treatments were 0 (Control), 1.5, or 3.0 µg/kg body weight rbTNFα, which were administered once daily by subcutaneous injection for the first 7 days of lactation. Statistical contrasts were used to evaluate the treatment and dose effects of rbTNFα administration. Plasma TNFα concentrations at 16 h post-administration tended to be increased (P<0.10) by rbTNFα administration, but no dose effect (P>0.10) was detected; rbTNFα treatments increased (P<0.01) concentrations of plasma haptoglobin. Most plasma eicosanoids were not affected (P>0.10) by rbTNFα administration, but 6 out of 16 measured eicosanoids changed (P<0.05) over the first week of lactation, reflecting elevated inflammatory mediators in the days immediately following parturition. Dry matter and water intake, milk yield, and milk fat and protein yields were all decreased (P<0.05) by rbTNFα treatments by 15 to 18%. Concentrations of plasma glucose, insulin, β-hydroxybutyrate, non-esterified fatty acids, triglyceride, 3-methylhistidine, and liver triglyceride were unaffected (P>0.10) by rbTNFα treatment. Glucose turnover rate was unaffected (P = 0.18) by rbTNFα administration. The higher dose of rbTNFα tended to increase the risk of cows developing one or more health disorders (P = 0.08). Taken together, these results indicate that administration of rbTNFα daily for the first 7 days of lactation altered inflammatory responses, impaired milk production and health, but did not significantly affect liver triglyceride accumulation or nutrient metabolism in dairy cows.     

Effect of Monensin and Lasalocid-Sodium on the Growth of Methanogenic and Rumen Saccharolytic Bacteria

It is thought that monensin increases the efficiency of feed utilization by cattle by altering the rumen fermentation. We studied the effect of monensin and the related ionophore antibiotic lasalocid-sodium (Hoffman-LaRoche) on the growth of methanogenic and rumen saccharolytic bacteria in a complex medium containing rumen fluid. Ruminococcus albus, Ruminococcus flavefaciens, and Butyrivibrio fibrisolvens were inhibited by 2.5 μg of monensin or lasalocid per ml. Growth of Bacteroides succinogenes and Bacteroides ruminicola was delayed by 2.5 μg of monensin or lasalocid per ml. Populations of B. succinogenes and B. ruminicola that were resistant to 20 μg of either drug per ml were rapidly selected by growth in the presence of each drug at 5.0 μg/ml. Selenomonas ruminantium was insensitive to 40 μg of monensin or lasalocid per ml. Either antibiotic (10 μg/ml) inhibited Methanobacterium MOH, Methanobacterium formicicum, and Methanosarcina barkeri MS. Methanobacterium ruminantium PS was insensitive to 40 μg of monensin or 20 μg of lasalocid per ml. The methanogenic strain 442 was insensitive to 40 μg of monensin but sensitive to 10 μg of lasalocid per ml. The results suggest that monensin or lasalocid acts in the rumen by selecting for succinate-forming Bacteroides and for S. ruminantium, a propionate producer that decarboxylates succinate to propionate. The selection could lead to an increase in rumen propionate formation. Selection against H₂ and formate producers, e.g. R. albus, R. flavefaciens, and B. fibrisolvens, could lead to a depression of methane production in the rumen.     

Polymorphic Variants of SCN1A and EPHX1 Influence Plasma Carbamazepine Concentration,Metabolism and Pharmacoresistance in a Population of Kosovar Albanian Epileptic Patients

Aim

The present study aimed to evaluate the effects of gene variants in key genes influencing pharmacokinetic and pharmacodynamic of carbamazepine (CBZ) on the response in patients with epilepsy.

Materials & Methods

Five SNPs in two candidate genes influencing CBZ transport and metabolism, namely ABCB1 or EPHX1, and CBZ response SCN1A (sodium channel) were genotyped in 145 epileptic patients treated with CBZ as monotherapy and 100 age and sex matched healthy controls. Plasma concentrations of CBZ, carbamazepine-10,11-epoxide (CBZE) and carbamazepine-10,11-trans dihydrodiol (CBZD) were determined by HPLC-UV-DAD and adjusted for CBZ dosage/kg of body weight.

Results

The presence of the SCN1A IVS5-91G>A variant allele is associated with increased epilepsy susceptibility. Furthermore, carriers of the SCN1A IVS5-91G>A variant or of EPHX1 c.337T>C variant presented significantly lower levels of plasma CBZ compared to carriers of the common alleles (0.71±0.28 vs 1.11±0.69 μg/mL per mg/Kg for SCN1A IVS5-91 AA vs GG and 0.76±0.16 vs 0.94±0.49 μg/mL per mg/Kg for EPHX1 c.337 CC vs TT; P<0.05 for both). Carriers of the EPHX1 c.416A>G showed a reduced microsomal epoxide hydrolase activity as reflected by a significantly decreased ratio of CBZD to CBZ (0.13±0.08 to 0.26±0.17, p<0.05) also of CBZD to CBZE (1.74±1.06 to 3.08±2.90; P<0.05) and CDR_CBZD (0.13±0.08 vs 0.24±0.19 μg/mL per mg/Kg; P<0.05). ABCB1 3455C>T SNP and SCN1A 3148A>G variants were not associated with significant changes in CBZ pharmacokinetic. Patients resistant to CBZ treatment showed increased dosage of CBZ (657±285 vs 489±231 mg/day; P<0.001) but also increased plasma levels of CBZ (9.84±4.37 vs 7.41±3.43 μg/mL; P<0.001) compared to patients responsive to CBZ treatment. CBZ resistance was not related to any of the SNPs investigated.

Conclusions

The SCN1A IVS5-91G>A SNP is associated with susceptibility to epilepsy. SNPs in EPHX1 gene are influencing CBZ metabolism and disposition. CBZ plasma levels are not an indicator of resistance to the therapy.     

Nevirapine,Sodium Concentration and HIV-1 RNA in Breast Milk and Plasma among HIV-Infected Women Receiving Short-Course Antiretroviral Prophylaxis

Introduction

Risk factors for breast milk transmission of HIV-1 from mother to child include high plasma and breast milk viral load, low maternal CD4 count and breast pathology such as mastitis.

Objective

To determine the impact of nevirapine and subclinical mastitis on HIV-1 RNA in maternal plasma and breast milk after intrapartum single-dose nevirapine combined with either 1-week tail of Combivir (zidovudine/lamivudine) or single-dose Truvada (tenofovir/emtricitabine).

Methods

Maternal plasma and bilateral breast milk samples were collected between April 2008 and April 2011 at 1, 4 and 6 weeks postpartum from HIV-infected Tanzanian women. Moreover, plasma samples were collected at delivery from mother and infant.

Results

HIV-1 RNA was quantified in 1,212 breast milk samples from 273 women. At delivery, 96% of the women and 99% of the infants had detectable nevirapine in plasma with a median (interquartile range, IQR) of 1.5 μg/mL (0.75–2.20 μg/mL) and 1.04 μg/mL (0.39–1.71 μg/mL), respectively (P < 0.001). At 1 week postpartum, 93% and 98% of the women had detectable nevirapine in plasma and breast milk, with a median (IQR) of 0.13 μg/mL (0.13–0.39 μg/mL) and 0.22 μg/mL (0.13–0.34 μg/mL), respectively. Maternal plasma and breast milk HIV-1 RNA correlated at all visits (R = 0.48, R = 0.7, R = 0.59; all P = 0.01). Subclinical mastitis was detected in 67% of the women at some time during 6 weeks, and in 38% of the breast milk samples. Breast milk samples with subclinical mastitis had significantly higher HIV-1 RNA at 1, 4 and 6 weeks (all P < 0.05).

Conclusion

After short-course antiretroviral prophylaxis, nevirapine was detectable in most infant cord blood samples and the concentration in maternal plasma and breast milk was high through week 1 accompanied by suppressed HIV-1 RNA in plasma and breast milk.