Potential subchronic food safety of the stacked trait transgenic maize GH5112E-117C in Sprague-Dawley rats

The food safety of stacked trait genetically modified (GM) maize GH5112E-117C containing insect-resistance gene Cry1Ah and glyphosate-resistant gene G2-aroA was evaluated in comparison to non-GM Hi-II maize fed to Sprague-Dawley rats during a 90-day subchronic feeding study. Three different dietary concentrations (12.5, 25 and 50 %, w/w) of the GM maize were used or its corresponding non-GM maize. No biologically significant differences in the animals’ clinical signs, body weights, food consumption, hematology, clinical chemistry, organ weights and histopathology were found between the stacked trait GM maize groups, and the non-GM maize groups. The results of the 90-day subchronic feeding study demonstrated that the stacked trait GM maize GH5112E-117C is as safe as the conventional non-GM maize Hi-II.     

Nutritional assessment and fate of DNA of soybean meal from roundup ready or conventional soybeans using rats

This study was conducted to compare the safety of soybean meal prepared from genetically modified (GM) glyphosate-tolerant (Roundup Ready; RR) soybeans and conventional soybeans. Eighty Sprague-Dawley rats (40 males and 40 females) were randomly allotted to one of four groups according to sex and body weight for a 13-week feeding experiment. The rats were fed corn-based diets containing 60% conventional soybean meal, a mixture of 30% conventional and 30% RR soybean meal, 60% or 90% RR soybean meal. All diets were adjusted to an identical nutrient level except the 90% RR diet. The two soybean meals were similar in chemical analysis and amino acid composition. During the 13-week growth trial, body weight (P < 0.05) and feed intake (P < 0.05) decreased only in rats fed with 90% RR soybean meal at the first week. No treatment-related deaths occurred during the experiment. Gross necropsy findings, haematological or urinalysis values and clinical serum parameters showed no meaningful differences between rats fed the control and RR soybean meals. A 145 bp of cp4 epsps gene specific for the GM constructs from RR soybean meal or a 407 bp of lec gene from endogenous soybean DNA could not be detected in investigated masseter muscle samples. No adverse effects of glyphosate-tolerant soybean meal on rats were seen even at levels as high as 90% of the diet.     

Safety assessment of transgenic Bacillus thuringiensis rice T1c-19 in Sprague-Dawley rats from metabonomics and bacterial profile perspectives

Bacillus thuringiensis rice is facing commercialization as the main food source in the near future. The unintended effects of genetically modified (GM) organisms are the most important barriers to their promotion. We aimed to establish a new in vivo evaluation model for genetically modified foods by using metabonomics and bacterial profile approaches. T1c-19 rice flour or its transgenic parent MH63 was used at 70% wt/wt to produce diets that were fed to rats for ～ 90 days. Urine metabolite changes were detected using (1)H NMR. Denaturing gradient gel electrophoresis and real-time polymerase chain reaction (RT-PCR) were used to detect the bacterial profiles between the two groups. The metabonomics was analyzed for metabolite changes in rat urine, when compared with the non-GM rice group, where rats were fed a GM rice diet. Several metabolites correlated with rat age and sex but not with GM rice diet. Significant biological differences were not identified between the GM rice diet and the non-GM rice diet. The bacteria related to rat urine metabolites were also discussed. The results from metabonomics and bacterial profile analyses were comparable with the results attained using the traditional method. Because metabonomics and bacterial profiling offer noninvasive, dynamic approaches for monitoring food safety, they provide a novel process for assessing the safety of GM foods.     

Nutritional assessment and fate of dna of soybean meal from roundup ready or conventional soybeans using rats

This study was conducted to compare the safety of soybean meal prepared from genetically modified (GM) glyphosate-tolerant (Roundup Ready; RR) soybeans and conventional soybeans. Eighty Sprague-Dawley rats (40 males and 40 females) were randomly allotted to one of four groups according to sex and body weight for a 13-week feeding experiment. The rats were fed corn-based diets containing 60% conventional soybean meal, a mixture of 30% conventional and 30% RR soybean meal, 60% or 90% RR soybean meal. All diets were adjusted to an identical nutrient level except the 90% RR diet. The two soybean meals were similar in chemical analysis and amino acid composition. During the 13-week growth trial, body weight (P?P?cp4 epsps gene specific for the GM constructs from RR soybean meal or a 407?bp of lec gene from endogenous soybean DNA could not be detected in investigated masseter muscle samples. No adverse effects of glyphosate-tolerant soybean meal on rats were seen even at levels as high as 90% of the diet.     

Overexpression of Toll-like receptor 4 enhances LPS-induced inflammatory response and inhibits Salmonella Typhimurium growth in ovine macrophages

The Toll-like receptor 4 (TLR4) plays a crucial role in innate inflammatory responses, as it recognizes gram-negative bacteria (or their products) and contributes greatly to host defense against invading pathogens. Though TLR4 overexpressing transgenic sheep, resistant to certain diseases related with gram-negative bacteria, had been bred in our previous research, the effects of overexpression of TLR4 on innate immune response remained unclear. In this study, TLR4 overexpressing ovine macrophages were obtained from peripheral blood, and it was found that the overexpression of TLR4 initially promoted the production of proinflammatory cytokines TNFα and IL-6 by activating TLR4-mediated IRAK4-dependent NF-κB and MAPK (JNK and ERK1/2) signaling following LPS stimulation. However, this effect was later impaired due to increased internalization of TLR4 into endosomal compartment of the macrophages. Then the overexpression of TLR4 triggered TBK1-dependent interferon-regulatory factor-3 (IRF-3) expression, which in turn led to the induction of IFN-β and IFN-inducible genes (i.e.IP10, IRG1 and GARG16). Understandably, an increased IFN-β level facilitated phosphorylation of STAT1 to induce expression of innate antiviral genes Mx1 and ISG15, suggesting that TLR4 overexpressing macrophages were equipped better against viral infection. Correspondingly, the bacterial burden in these macrophages, after infection with live S. Typhimurium, was decreased significantly. In summary, the results indicated that overexpression of TLR4 could enhance innate inflammatory responses, initiate the innate antiviral immunity, and control effectively S. Typhimurium growth in ovine macrophages.     

Effects of structured lipids containing eicosapentaenoic or docosahexaenoic acid and caprylic acid on serum and liver lipid profiles in rats

The physiological activity and effect on lipid metabolism of four types of structured lipids (SLs), that contain caprylic acid (C8) and either eicosapentaenoic (EPA) or docosahesaenoic acid (DHA), were evaluated in male Wistar rats fed experimental diets containing 7% (wt %) of each SL and 3% (wt %) soybean oil for 28 days. Control rats were fed a diet containing 10% (wt %) soybean oil. The relative perirenal adipose tissue weights of rats fed D-8-8 and 8-D-8 diets were significantly lower than those of other groups. We observed significantly lower serum cholesterol concentrations in rats fed SLs than those of control group over experimental period. The serum lipids concentrations in rats fed diets containing SLs were significantly lower P < 0.05) than those of soybean oil group. The fatty acid compositions of WAT did not reflect the structural differences in the triglyceride. These results suggest that the physiological effects of the SLs used in this study were due to the fatty acids rather than the structural specificity. Therefore, further study will be needed to ascertain the most desirable structural configuration.     

A proteomic-based approach to study underlying molecular responses of the small intestine of Wistar rats to genetically modified corn (MON810)

A genetically modified (GM) commercial corn variety, MON810, resistant to European corn borer, has been shown to be non-toxic to mammals in a number of rodent feeding studies carried out in accordance with OECD Guidelines. Insect resistance results from expression of the Cry1Ab gene encoding an insecticidal Bt protein that causes lysis and cell death in susceptible insect larvae by binding to midgut epithelial cells, which is a key determinant of Cry toxin species specificity. Whilst whole animal studies are still recognised as the ‘gold standard’ for safety assessment, they only provide indirect evidence for changes at the cellular/organ/tissue level. In contrast, omics-based technologies enable mechanistic understanding of toxicological or nutritional events at the cellular/receptor level. To address this important knowledge-gap and to gain insights into the underlying molecular responses in rat to MON810, differential gene expression in the epithelial cells of the small intestine of rats fed formulated diets containing MON810, its near isogenic line, two conventional corn varieties, and a commercial (Purina?) corn-based control diet were investigated using comparative proteomic profiling. Pairwise and five-way comparisons showed that the majority of proteins that were differentially expressed in the small intestine epithelial cells in response to consumption of the different diets in both 7-day and 28-day studies were related to lipid and carbohydrate metabolism and protein biosynthesis. Irrespective of the diet, a limited number of stress-related proteins were shown to be differentially expressed. However these stress-related proteins differed between diets. No adverse clinical or behavioural effects, or biomarkers of adverse health, were observed in rats fed GM corn compared to the other corn diets. These findings suggest that MON810 has negligible effects on the small intestine of rats at the cellular level compared with the well-documented toxicity observed in susceptible insects.

     

Effects of cloned-cattle meat on reproductive physiology in rats

The influence of the cloned-cattle meat diets upon reproduction in mammals was rarely studied. This study was performed to analyze the effects of the diets containing cloned-cattle (Korean native beef, Hanwoo) meat on the reproductive physiology in rats. The male and female rats were fed with the diets containing 5% or 10% of normal- (N-5 or N-10) or cloned- (C-5 or C-10) cattle meat during test periods. The rats fed with commercial pellets were used as control. Lower food consumption in normal- and cloned-cattle meat diet groups is detected in both male and female rats compared with that of control (P < 0.05, 0.01 and 0.001). No signs of cloned-cattle meat diets on male reproductive parameters are found in all groups, except for lower sperm deformity in C-5 group (P < 0.05) and higher testosterone concentration in C-10 group (P < 0.05), respectively. There are no significant test substance-related differences of Caesarean section and delivery in dams and external examination and physiological development test in neonate compared with control and normal meat groups. Based on these results, it can be postulated that there are no obvious negative effects on the reproductive physiology in rats fed with cloned-cattle meat diets compared to their comparators.     

Effect of feeding genetically modified Bt MON810 maize to ∼40-day-old pigs for 110 days on growth and health indicators

A total of 72 male weaned pigs were used in a 110-day study to investigate the effect of feeding genetically modified (GM) Bt MON810 maize on selected growth and health indicators. It was hypothesised that in pigs fed Bt maize, growth and health are not impacted compared with pigs fed isogenic maize-based diets. Following a 12-day basal period, pigs (10.7 ± 1.9 kg body weight (BW); ∼40 days old) were blocked by weight and ancestry and randomly assigned to treatments: (1) non-GM maize diet for 110 days (non-GM), (2) GM maize diet for 110 days (GM), (3) non-GM maize diet for 30 days followed by GM maize diet up to day 110 (non-GM/GM) and (4) GM maize diet for 30 days followed by non-GM maize diet up to day 110 (GM/non-GM). BW and daily feed intake were recorded on days 0, 30, 60 and 110 (n = 15). Body composition was determined by dual energy X-ray absorptiometry (n = 10) on day 80. Following slaughter on day 110, organs and intestines were weighed and sampled for histological analysis and urine was collected for biochemical analysis (n = 10). Serum biochemistry analysis was performed on days 0, 30, 60, 100 and 110. Growth performance and serum biochemistry were analysed as repeated measures with time and treatment as main factors. The slice option of SAS was used to determine treatment differences at individual time points. There was no effect of feeding GM maize on overall growth, body composition, organ and intestinal weight and histology or serum biochemistry on days 60 and 100 and on urine biochemistry on day 110. A treatment × time interaction was observed for serum urea (SU; P < 0.05), creatinine (SC; P < 0.05) and aspartate aminotransferase (AST; P < 0.05). On day 30, SU was lower for the non-GM/GM treatment compared with the non-GM, GM and GM/non-GM treatments (P < 0.05). On day 110, SC was higher for the non-GM/GM and GM/non-GM treatments compared with non-GM and GM treatments (P < 0.05). Overall, serum total protein was lower for the GM/non-GM treatment compared with the non-GM/GM treatment (P < 0.05). The magnitude of change observed in some serum biochemical parameters did not indicate organ dysfunction and the changes were not accompanied by histological lesions. Long-term feeding of GM maize to pigs did not adversely affect growth or the selected health indicators investigated.     

Macrophage ABCA1 reduces MyD88-dependent Toll-like receptor trafficking to lipid rafts by reduction of lipid raft cholesterol

We previously showed that macrophages from macrophage-specific ATP-binding cassette transporter A1 (ABCA1) knockout (Abca1^-M/-M) mice had an enhanced proinflammatory response to the Toll-like receptor (TLR) 4 agonist, lipopolysaccharide (LPS), compared with wild-type (WT) mice. In the present study, we demonstrate a direct association between free cholesterol (FC), lipid raft content, and hyper-responsiveness of macrophages to LPS in WT mice. Abca1^-M/-M macrophages were also hyper-responsive to specific agonists to TLR2, TLR7, and TLR9, but not TLR3, compared with WT macrophages. We hypothesized that ABCA1 regulates macrophage responsiveness to TLR agonists by modulation of lipid raft cholesterol and TLR mobilization to lipid rafts. We demonstrated that Abca1^-M/-M vs. WT macrophages contained 23% more FC in isolated lipid rafts. Further, mass spectrometric analysis suggested raft phospholipid composition was unchanged. Although cell surface expression of TLR4 was similar between Abca1^-M/-M and WT macrophages, significantly more TLR4 was distributed in membrane lipid rafts in Abca1^-M/-M macrophages. Abca1^-M/-M macrophages also exhibited increased trafficking of the predominantly intracellular TLR9 into lipid rafts in response to TLR9-specific agonist (CpG). Collectively, our data suggest that macrophage ABCA1 dampens inflammation by reducing MyD88-dependent TLRs trafficking to lipid rafts by selective reduction of FC content in lipid rafts.     

Natural Killer Cell-Dependent Anti-Fibrotic Pathway in Liver Injury via Toll-Like Receptor-9

The toll-like receptor-9 (TLR9) agonist cytosine phosphate guanine (CpG), activates hepatic stellate cells (HSCs) and mediates fibrosis. We investigated the TLR9 effects on lymphocyte/HSCs interactions. Liver fibrosis was induced in wild-type (WT) mice by intra-peritoneal carbon-tetrachloride (CCl₄) induction for 6 weeks. Fibrotic groups were intravenously treated by a vehicle versus CpG along last 2 weeks. Compared to vehicle-treated fibrotic WT, the in-vivo CpG-treatment significantly attenuated hepatic fibrosis and inflammation, associated with decreased CD8 and increased NK liver cells. In-vitro, co-cultures with vehicle-treated fibrotic NK cells increased HSCs proliferation (P<0.001) while their CpG-treated counterparts achieved a significant decrease. To investigate the role of lymphocytes, TLR9^-/- mice induced-hepatic fibrosis were used. Although TLR9^-/- mice manifested lower fibrotic profile as compared to their wild-type (WT) counterparts, senescence (SA-β-Gal activity) in the liver and ALT serum levels were significantly greater. In an adoptive transfer model; irradiated WT and TLR9^-/- recipients were reconstituted with naïve WT or TLR9^-/- lymphocytes. The adoptive transfer of TLR9^-/- versus WT lymphocytes led to increased fibrosis of WT recipients. TLR9^-/- fibrotic recipients reconstituted with TLR9^-/- or WT lymphocytes showed no changes in hepatic fibrosis severity or ALT serum levels. TLR9 activation had inconsistent effects on lymphocytes and HSCs. The net balance of TLR9 activation in WT, displayed significant anti-fibrotic activity, accompanied by CD8 suppression and increased NK-cells, activity and adherence to HSCs. The pro-fibrotic and pro-inflammatory properties of TLR9^-/- lymphocytes fail to activate HSCs with an early senescence in TLR9^-/- mice.      

A 90-Day Dietary Toxicity Study of Genetically Modified Rice T1C-1 Expressing Cry1C Protein in Sprague Dawley Rats

In a 90-day study, Sprague Dawley rats were fed transgenic T1C-1 rice expressing Cry1C protein and were compared with rats fed non-transgenic parental rice Minghui 63 and rats fed a basal diet. No adverse effects on animal behavior or weight gain were observed during the study. Blood samples were collected and analyzed, and standard hematological and biochemical parameters were compared. A few of these parameters were found to be significantly different, but were within the normal reference intervals for rats of this breed and age, and were thus not considered to be treatment-related. Following sacrifice, a large number of organs were weighed, and macroscopic and histopathological examinations were performed with no changes reported. The aim of this study was to use a known animal model to determine the safety of the genetically modified (GM) rice T1C-1. The results showed no adverse or toxic effects due to T1C-1 rice when tested in this 90-day study.     

Using mixed silages of sweet sorghum and alfalfa in total mixed rations to improve growth performance,nutrient digestibility,carcass traits and meat quality of sheep

Combining sweet sorghum (SS) with alfalfa for ensiling has the potential to make a high-quality silage with relatively a better nutrient balance. However, the existing data are insufficient about how changes in ratios of SS to alfalfa in different silage mixtures affect animal performance and meat quality. Therefore, the objective of this study was to determine the effect of feeding total mixed rations containing mixed silages of SS and alfalfa on growth performance, nutrient digestibility, carcass traits and meat quality of Karakul sheep. Five total mixed rations were formulated with different SS proportions at 100%, 80%, 60%, 40%, and 20% on a fresh weight basis. Thirty 4-month old male Karakul sheep with 25.5 ± 1.4 kg BW were randomly allocated into five treatment groups, each with six lambs for this experiment. The results indicated that the Karakul sheep consuming total mixed rations containing SS-Alfalfa (SS-AF) silage mixtures with a lower proportion of SS tended to increase nutrient digestibility, growth performance, carcass traits, and meat quality. Feeding diets with SS at 40% and 20% inclusion rate showed a significant linear increase in the apparent digestibility of DM, CP and NDF, as well as subcutaneous fat thickness and water holding capacity (P < 0.5). Moreover, reduction in SS inclusion rate in these diets caused significant (P < 0.05) linear and quadratic increases in DM intake, final BW, average daily gain, carcass weight, and the amino acid contents of meat (P < 0.05). Conversely, feeding diet with lower SS inclusion rate led to decrease in feed conversion ratio and shear force significantly (P < 0.05) for these sheep. It appears that 40% inclusion of SS was the optimal rate in making the SS-AF silage mixture for lambs to achieve a superior production performance and high-quality meat products in Karakul sheep. However, further research is needed to investigate the effect of feeding SS-AF silage mixtures alongside contrasting ingredients on rumen function, ruminal microorganisms and digestive enzyme activity of sheep and other ruminant animals.     

Fourteen-week feeding test of meat and milk derived from cloned cattle in the rat

Agricultural application of cloned livestock produced by nuclear transfer requires public and governmental understanding of food-safety issues. To determine whether physiological effects occurred in animals fed products derived from cloned cattle, we conducted long-term (14 week) trials feeding Crj:CD(SD)IGS rats meat and milk from cloned cattle. Diets containing meat and milk were equal in nutritional value to the basal diet (AIN93G). Urinalysis was performed at Weeks 4, 8 and 12; at the end of the feeding period, blood sampling and autopsies were conducted. During the feeding periods, there were no significant differences in general condition, death loss, growth, battery of functional observational tests and estrous cycles among groups given diets containing meat and milk powder from non-clone, embryonic clone and somatic clone cattle. Furthermore, no significant changes attributed to consumption of clone meat or milk were detected in urinalysis, hematological and blood chemical, gross pathological or histological examinations. Therefore, we concluded that the physiologic conditions of the rats were not affected by consumption of meat and milk from bovine clones.     

Lipopolysaccharides-Induced Suppression of Innate-Like B Cell Apoptosis Is Enhanced by CpG Oligodeoxynucleotide and Requires Toll-Like Receptors 2 and 4

Innate-like B lymphocytes play an important role in innate immunity in periodontal disease through Toll-like receptor (TLR) signaling. However, it is unknown how innate-like B cell apoptosis is affected by the periodontal infection-associated innate signals. This study is to determine the effects of two major TLR ligands, lipopolysaccharide (LPS) and CpG-oligodeoxynucleotides (CpG-ODN), on innate-like B cell apoptosis. Spleen B cells were isolated from wild type (WT), TLR2 knockout (KO) and TLR4 KO mice and cultured with E. coli LPS alone, P. gingivalis LPS alone, or combined with CpG-ODN for 2 days. B cell apoptosis and expressions of specific apoptosis-related genes were analyzed by flow cytometry and real-time PCR respectively. P. gingivalis LPS, but not E. coli LPS, reduced the percentage of AnnexinV⁺/7-AAD^- cells within IgM^highCD23^lowCD43^-CD93^- marginal zone (MZ) B cell sub-population and IgM^highCD23^lowCD43⁺CD93⁺ innate response activator (IRA) B cell sub-population in WT but not TLR2KO or TLR4KO mice. CpG-ODN combined with P. gingivalis LPS further reduced the percentage of AnnexinV⁺/7-AAD^- cells within MZ B cells and IRA B cells in WT but not TLR2 KO or TLR4 KO mice. Pro-apoptotic CASP4, CASP9 and Dapk1 were significantly down-regulated in P. gingivalis LPS- and CpG-ODN-treated B cells from WT but not TLR2 KO or TLR4 KO mice. Anti-apoptotic IL-10 was significantly up-regulated in P. gingivalis LPS- and CpG-ODN-treated B cells from WT and TLR2 KO but not TLR4 KO mice. These results suggested that both TLR2 and TLR4 signaling are required for P. gingivalis LPS-induced, CpG-ODN-enhanced suppression of innate-like B cell apoptosis.     

The potential application of plant wax markers from alfalfa for estimating the total feed intake of sheep

Estimating the feed intake of grazing herbivores is critical for determining their nutrition, overall productivity and utilization of grassland resources. A 17-day indoor feeding experiment was conducted to evaluate the potential use of Medicago sativa as a natural supplement for estimating the total feed intake of sheep. A total of 16 sheep were randomly assigned to four diets (four sheep per diet) containing a known amount of M. sativa together with up to seven forages common to typical steppes. The diets were: diet 1, M. sativa + Leymus chinensis + Puccinellia distans; diet 2, species in diet 1 + Phragmites australis; diet 3, species in diet 2 + Chenopodium album + Elymus sibiricus; and diet 4, species in diet 3 + Artemisia scoparia + Artemisia tanacetifolia. After faecal marker concentrations were corrected by individual sheep recovery, treatment mean recovery or overall recovery, the proportions of M. sativa and other dietary forages were estimated from a combination of alkanes and long-chain alcohols using a least-square procedure. Total intake was the ratio of the known intake of M. sativa to its estimated dietary proportion. Each dietary component intake was obtained using total intake and the corresponding dietary proportions. The estimated values were compared with actual values to assess the estimation accuracy. The results showed that M. sativa exhibited a distinguishable marker pattern in comparison to the other dietary forage species. The accuracy of the dietary composition estimates was significantly (P < 0.001) affected by both diet diversity and the faecal recovery method. The proportion of M. sativa and total intake across all diets could be accurately estimated using the individual sheep or the treatment mean recovery methods. The largest differences between the estimated and observed total intake were 2.6 g and 19.2 g, respectively, representing only 0.4% and 2.6% of the total intake. However, they were significantly (P < 0.05) biased for most diets when using the overall recovery method. Due to the difficulty in obtaining individual sheep recovery under field conditions, treatment mean recovery is recommended. This study suggests that M. sativa, a natural roughage instead of a labelled concentrate, can be utilized as a dietary supplement to accurately estimate the total feed intake of sheep indoors and further indicates that it has potential to be used in steppe grassland of northern China, where the marker patterns of M. sativa differ markedly from commonly occurring plant species.     

High-throughput sequence-based analysis of the intestinal microbiota of weanling pigs fed genetically modified MON810 maize expressing Bacillus thuringiensis Cry1Ab (Bt maize) for 31 days

The objective of this study was to investigate if feeding genetically modified (GM) MON810 maize expressing the Bacillus thuringiensis insecticidal protein (Bt maize) had any effects on the porcine intestinal microbiota. Eighteen pigs were weaned at ~28 days and, following a 6-day acclimatization period, were assigned to diets containing either GM (Bt MON810) maize or non-GM isogenic parent line maize for 31 days (n = 9/treatment). Effects on the porcine intestinal microbiota were assessed through culture-dependent and -independent approaches. Fecal, cecal, and ileal counts of total anaerobes, Enterobacteriaceae, and Lactobacillus were not significantly different between pigs fed the isogenic or Bt maize-based diets. Furthermore, high-throughput 16S rRNA gene sequencing revealed few differences in the compositions of the cecal microbiotas. The only differences were that pigs fed the Bt maize diet had higher cecal abundance of Enterococcaceae (0.06 versus 0%; P < 0.05), Erysipelotrichaceae (1.28 versus 1.17%; P < 0.05), and Bifidobacterium (0.04 versus 0%; P < 0.05) and lower abundance of Blautia (0.23 versus 0.40%; P < 0.05) than pigs fed the isogenic maize diet. A lower enzyme-resistant starch content in the Bt maize, which is most likely a result of normal variation and not due to the genetic modification, may account for some of the differences observed within the cecal microbiotas. These results indicate that Bt maize is well tolerated by the porcine intestinal microbiota and provide additional data for safety assessment of Bt maize. Furthermore, these data can potentially be extrapolated to humans, considering the suitability of pigs as a human model.     

The dietary overlap between red kangaroos (Macropus rufus) and sheep (Ovis aries) in the arid rangelands of Australia

Abstract The interaction between red kangaroos and sheep in the more arid areas of Australia was examined by means of a large-scale manipulative experiment. The diets of red kangaroos and sheep grazing together and separately in large paddocks were studied to assess whether competition for food might influence diet selection in these species. Diets and the amount of intra- and interspecific dietary overlap varied in accordance with pasture conditions. At pasture biomasses ranging from 50 to 200 g m^?2 (dry weight [wt]), the diets of red kangaroos and sheep overlapped by 58–73%, with forbs and grasses being the major items in the diets of both species. The diets of both sheep and red kangaroos under different experimental regimes were similar. In dry times (pasture biomass 40–50 g m^?2 (dry wt) more shrubs were eaten by both species and the amount of dietary overlap tended to be lower (52–66%). The diets of kangaroos were similar between paddocks. However, in paddocks containing both species sheep consumed proportionately more chenopodaceous shrubs and fewer grasses than those in ‘kangaroo-free’ paddocks.     

Leukocyte- and platelet-derived microvesicle interactions following in vitro and in vivo activation of toll-like receptor 4 by lipopolysaccharide

Background

Pro-coagulant membrane microvesicles (MV) derived from platelets and leukocytes are shed into the circulation following receptor-mediated activation, cell-cell interaction, and apoptosis. Platelets are sentinel markers of toll-like receptor 4 (TLR4) activation. Experiments were designed to evaluate the time course and mechanism of direct interactions between platelets and leukocytes following acute activation of TLR4 by bacterial lipopolysaccharide (LPS).

Methodology/Principal Findings

Blood from age-matched male and female wild type (WT) and TLR4 gene deleted (dTLR4) mice was incubated with ultra-pure E. coli LPS (500 ng/ml) for up to one hour. At designated periods, leukocyte antigen positive platelets, platelet antigen positive leukocytes and cell-derived MV were quantified by flow cytometry. Numbers of platelet- or leukocyte-derived MV did not increase within one hour following in vitro exposure of blood to LPS. However, with LPS stimulation numbers of platelets staining positive for both platelet- and leukocyte-specific antigens increased in blood derived from WT but not dTLR4 mice. This effect was blocked by inhibition of TLR4 signaling mediated by My88 and TRIF. Seven days after a single intravenous injection of LPS (500 ng/mouse or 20 ng/gm body wt) to WT mice, none of the platelets stained for leukocyte antigen. However, granulocytes, monocytes and apoptotic bodies stained positive for platelet antigens.

Conclusions/Significance

Within one hour of exposure to LPS, leukocytes exchange surface antigens with platelets through TLR4 activation. In vivo, leukocyte expression of platelet antigen is retained after a single exposure to LPS following turn over of the platelet pool. Acute expression of leukocyte antigen on platelets within one hour of exposure to LPS and the sustained expression of platelet antigen on leukocytes following a single acute exposure to LPS in vivo explains, in part, associations of platelets and leukocytes in response to bacterial infection and changes in thrombotic propensity of the blood.     

A 90-Day Feeding Study in Rats to Assess the Safety of Genetically Engineered Pork

Our laboratory recently produced genetically engineered (GE) Meishan pigs containing a ZFN-edited myostatin loss-of-function mutant. These GE pigs develop and grow as normal as wild type pigs but produce pork with greater lean yield and lower fat mass. To assess any potential subchronic toxicity risks of this GE pork, a 90-day feeding study was conducted in Sprague-Dawley rats. Rats were randomly divided into five groups, and fed for 90 days with basic diet and basic diets formulated with low dose and high dose pork prepared from wild type pigs and GE pigs, respectively. Animal behaviors and clinical signs were monitored twice daily, and body weight and food consumption were measured and recorded weekly. At days 45 and 90, blood tests (lipid panel, electrolytes, parameters related to liver and kidney functions, and complete blood counts) were performed. Additionally, gross pathology and histopathological analyses were performed for major organs in each group. Data analysis shows that there were no significant differences in growth rate, food consumption, and blood test parameters between rat groups fed with GE pork and wild type pork. Although differences in some liver function parameters (such as aspartate aminotransferase, total proteins, albumin, and alkaline phosphatase) and white blood cell counts (such as lymphocyte percentage and monocyte percentage) were observed between rats fed with high dose GE pork and basic diet, all test results in rats fed with GE pork are in the normal range. Additionally, there are no apparent lesions noted in all organs isolated from rats in all five feeding groups on days 45 and 90. Overall, our results clearly indicate that food consumption of GE pork produced by ZFN-edited myostatin loss-of-function mutant pigs did not have any long-term adverse effects on the health status in rats.