Effect of a synbiotic supplement as an antibiotic alternative on broiler skeletal, physiological, and oxidative parameters under heat stress.

The aim of this study was to examine if synbiotics can function as alternatives to antibiotics in broiler production under heat stress (HS). Day-old broiler chicks (528 birds) were randomly placed in floor pens within 2 identical temperature-controlled rooms (11 birds/pen and 24 pens/room). The pens of each room were evenly divided among 3 treatments (n = 8): basal diet (CON), the basal diet mixed with 50 ppm of bacitracin methylene disalicylate (BMD) or a synbiotic (50 ppm of PoultryStar meUS, SYN). From d 15, room 2 was under thermoneutral (TN) conditions (TN-CON, TN-BMD, and TN-SYN), while HS was applied to room 1 at 32oC for 9 hrs/d (0800 to 1700) (HS-CON, HS-BMD, and HS-SYN). Treatment effects on footpad dermatitis and gait score were measured on 5 birds/pen, and latency to lie (LTL) test was measured on 2 birds/pen at d 27 and d 41; and 1 broiler/pen was sampled on d 28 and d 42, respectively. Body, liver, and spleen weight were determined. Plasma levels of interleukins (IL), heat shock protein 70, immunoglobulin (Ig)Y, liver superoxide dismutase (SOD) and glutathione peroxidase (GPx) enzyme activities were examined. Heat stress suppressed BW and IgY concentrations on both d 28 and d 42, while suppressed plasma IL-6 concentrations, SOD activities, and LTL duration on d 28 only (P < 0.05). Among all treatments, SYN birds had the best foot and skeletal health scores on both d 27 and d 41 (P < 0.05). On d 42, SYN increased BW, and TN-SYN birds had higher relative spleen weight than both TN-BMD and TN-CON birds (P < 0.05). Antibiotic BMD increased BW (P < 0.05) but decreased SOD activities (P < 0.05) on d 42. These results indicate that the SYN supplementation decreases HS negative effect on broilers by improving BW, foot, and skeletal health, while BMD improves BW but also increases oxidative stress in broilers. The data suggest that synbiotic supplement may function as an alternative to antibiotics in broiler production during summer seasons, especially in the tropical and subtropical regions.

Effects of dietary supplementation of a probiotic (Bacillus subtilis) on bone mass and meat quality of broiler chickens.

The aim of this study was to investigate the effect of a dietary probiotic supplement on bone mass and meat quality of broiler chickens. Two hundred ten 1-day-old male Ross 708 broiler chicks were divided among 21 floor pens (10 chicks per pen). The pens were randomly distributed to 1 of 3 dietary treatments containing a probiotic, Bacillus subtilis, at 0 (control), 0.25 (0.25X), and 0.5 (0.5X) g/kg (n = 7). Gait score, footpad dermatitis (FPD), leg straightness, and hock burn (HB) were examined at day 33, and a latency-to-lie test was performed at day 34. At the end of the experiment (day 35), plasma, right leg, and litter samples were collected for mineral contents, meat quality, bone morphometric parameters, and litter quality assessments. The results indicated that probiotic-fed birds stood much longer during the latency-to-lie test with a greater tibial length, weight, and strength as well as higher plasma levels of calcium and phosphorus compared with the controls. In addition, probiotic-fed birds' leg muscle had higher color lightness at both 30 min and 5 h postmortem and greater water-holding capacity with a trend for less cooking loss (P = 0.056) and lower pH values (P < 0.05) at 5 h postmortem. Probiotic-fed birds' leg meat was tastier (P < 0.05) at 24 h after slaughter. These probiotic effects were greater in the 0.5X group than in the 0.25X group. There were no treatment effects on other measured parameters including gait score, HB, FPD, tibial lateral and medial wall thickness, diaphysis and medullary canal diameters, robusticity and tibiotarsal indexes, plasma magnesium concentrations, and litter moisture and pH values (P > 0.05). These findings indicate that the probiotic supplement could be a useful management tool for improving broiler production and welfare by enhanced bone mass and meat quality.

Pathophysiological characteristics and gene transcriptional profiling of bone microstructure in a low calcium diet fed laying hens.

Calcium depletion is a valuable non-invasive tool for studying skeletal system disorders. A low calcium diet (LCD) was used to examine the pathophysiological characteristics and molecular mechanisms of osteoporotic damage in caged laying hens. Sixty 64-wk-old laying hens were randomly housed in single-bird cages, and the cages were divided into 2 treatments: fed a regular calcium diet (RCD, 3.69%) or a LCD (1.56%) for 8 wk. The diet-induced changes of serum bone remodeling indicators, bone strength, microstructure of the distal femur, and the gene expression profiling of keel bone were measured. Compared to RCD hens, LCD hens had higher activity of serum alkaline phosphatase and tartrate resistant acid phosphatase but lower serum calcium concentrations with reduced tibial and femoral mass, width, and strength (P < 0.05). In addition, LCD hens had greater densities of osteoclasts, osteoblasts, connective tissue cells, and osteoid in the trabecular bone (P < 0.05). The transcriptome analysis revealed that 563 unigenes were differentially expressed in keel bone between LCD and RCD hens. The Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that these differentially expressed genes were mainly associated with the osteoporosis-related signaling pathways involved in the biological functions of the bone cellular and extracellular structural modulations. The real-time PCR analysis further confirmed that the LCD enhanced the mRNA expression of collagen type 1 alpha 2, integrin-binding sialoprotein and periostin, but inhibited sclerostin expression. These findings indicate that LCD hens have a higher bone turnover and micro-architectural damage compared to RCD hens. The results further evidence dietary supplement of calcium is a critical nutrient strategy for improving hen skeletal health.

Effects of probiotic supplementation on performance traits, bone mineralization, cecal microbial composition, cytokines and corticosterone in laying hens.

Recent researches have showed that probiotics promote bone health in humans and rodents. The objective of this study was to determine if probiotics have the similar effects in laying hens. Ninety-six 60-week-old White Leghorn hens were assigned to four-hen cages based on their BW. The cages were randomly assigned to 1 of 4 treatments: a layer diet mixed with a commercial probiotic product (containing Enterococcus faecium, Pediococcus acidilactici, Bifidobacterium animalis and Lactobacillus reuteri) at 0, 0.5, 1.0 or 2.0 g/kg feed (Control, 0.5×, 1.0× and 2.0×) for 7 weeks. Cecal Bifidobacterium spp. counts were higher in all probiotic groups (P0.05). In addition, the plasma concentrations of cytokines (interleukin-1ß, interleukin-6, interleukin-10, interferon-γ and tumor necrosis factor-α) and corticosterone as well as the levels of heterophil to lymphocyte ratio were similar between the 2.0× group and the control group (P>0.05). In line with these findings, no differences of cecal tonsil mRNA expressions of interleukin-1ß, interleukin-6 and lipopolysaccharide-induced tumor necrosis factor-α factor were detected between these two groups (P>0.05). These results suggest that immune cytokines and corticosterone may not involve in the probiotic-induced improvement of eggshell quality and bone mineralization in laying hens. In conclusion, the dietary probiotic supplementation altered cecal microbiota composition, resulting in reduced shell-less egg production and improved bone mineralization in laying hens; and the dietary dose of the probiotic up to 2.0× did not cause negative stress reactions in laying hens.

Supplementation of Bacillus subtilis-based probiotic reduces heat stress-related behaviors and inflammatory response in broiler chickens.

Probiotics reduce stress-related inflammation and abnormal behaviors in humans and rodents via regulation of the microbiota-gut-brain axis. The objective of this study was to determine if probiotic, Bacillus subtilis, has similar functions in broiler chickens under heat stress (HS). Two hundred forty 1-d-old broiler chicks were assigned to 48 pens with 4 treatments: Thermoneutral (TN)-RD (regular diet), TN-PD (the regular diet mixed with 1 × 106 CFU/g feed probiotic), HS-RD and HS-PD. Probiotic (Sporulin) was fed from day 1; and HS at 32°C for 10 h daily was initiated at day 15. The data showed that final BW, average daily gain , and feed conversion efficiency were improved in PD groups as compared to RD groups regardless of the ambient temperature (P < 0.01). Heterophil to lymphocyte ratio was affected by treatment and its value was in the order of HS-RD > HS-PD > TN-RD > TN-PD birds (P < 0.01). Compared to TN birds, HS birds spent more time in wing spreading, panting, squatting close to the ground, drinking, sleeping, dozing, and sitting but spent less time in eating, standing, and walking (P < 0.05 or 0.01). In addition, HS birds had greater levels of hepatic IL-6, IL-10, heat shock protein (HSP)70, and HSP70 mRNA expression (P < 0.01) and greater levels of cecal IgA and IgY (P < 0.01) compared to TN birds. Within TN groups, TN-PD birds had greater concentrations of hepatic IL-10 (P < 0.05) and cecal IgA (P < 0.01) than TN-RD birds. Within HS groups, HS-PD birds spent less time in wing spreading, panting, squatting close to the ground, drinking, sleeping, dozing, and sitting but spent more time in eating, foraging, standing, and walking than HS-RD birds (P < 0.05 or 0.01). The HS-PD birds also had lower concentrations of hepatic IL-6 and HSP70 (P < 0.01), whereas greater levels of IL-10 (P < 0.05) and lower concentrations of cecal IgA and IgY (P < 0.01). These results indicate that broilers fed the probiotic, B. subtilis, are able to cope with HS more effectively by ameliorating heat-induced behavioral and inflammatory reactions through regulation of microbiota-modulated immunity.

Acute brief heat stress in late gestation alters neonatal calf innate immune functions.

Heat stress, as one of the environmental stressors affecting the dairy industry, compromises the cow milk production, immune function, and reproductive system. However, few studies have looked at how prenatal heat stress (HS) affects the offspring. The objective of this study was to evaluate the effect of HS during late gestation on calf immunity. Calves were born to cows exposed to evaporative cooling (CT) or HS (cyclic 23-35°C) for 1 wk at 3 wk before calving. Both bull and heifer calves (CT, n=10; HS, n=10) were housed in similar environmental temperatures after birth. Both CT and HS calves received 3.78 L of pooled colostrum within 12 h after birth and were fed the same diet throughout the study. In addition to tumor necrosis factor α, IL-1ß, IL-1 receptor antagonist (IL-1RA), and toll-like receptor (TLR)2, and TLR4 mRNA expression, the expression of CD14(+) and CD18(+) cells, and DEC205(+) dendritic cells were determined in whole blood samples at d 0, 3, 7, 14, 21, and 28. The neutrophil to lymphocyte ratio, differential cell counts, and the hematocrit were also determined. During late gestation, the HS cows had greater respiration rates, rectal temperatures, and tended to spend more time standing compared with the CT cows. The HS calves had less expression of tumor necrosis factor-α and TLR2 and greater levels of IL-1ß, IL-1RA, and TLR4 compared with CT calves. The HS calves also had a greater percentage of CD18(+) cells compared with the CT calves. Additionally, a greater percentage of neutrophils and lesser percentage of lymphocytes were in the HS calves compared with the CT calves. The results indicate that biomarkers of calves' immunity are affected in the first several weeks after birth by HS in the dam during late gestation.

Effects of early serotonin programming on behavior and central monoamine concentrations in an avian model.

Serotonin (5-HT) acts as a neurogenic compound in the developing brain; however serotonin altering drugs such as SSRIs are often prescribed to pregnant and lactating mothers. Early agonism of 5-HT receptors could alter the development of serotonergic circuitry, altering neurotransmission and behaviors mediated by 5-HT signaling, including memory, fear and aggression. This study was designed to investigate the effects of early serotonin agonism on later behaviors. An extremely aggressive White leghorn strain (15I5) was used in the study. The chicks were injected with 5-MT (a serotonin agonist) at 2.5mg/kg (low dose), 10mg/kg (high dose) or saline (control) on the day of hatch and a second dose 24h later (n=9/sex/trt). Chicks' fear response and memory were tested at 2 weeks of age. In the fear test, chicks were subjected to a social isolation test for 20min, time to first vocalization and numbers of vocalizations were recorded. In the memory test, chicks were placed in a running wheel and presented with an imprinted object (white box with a red light) and a similar shaped novel object (blue box with a white light), respectively. The distance traveled in the wheel toward each object was measured. At 10 weeks of age birds were tested for aggression and concentrations of catecholamines were determined from the raphe nucleus and hypothalamus by HPLC (n=12). Expression of 5-HT1A and 5-HT1B receptor genes were measured by RT-PCR. Both high and low dose chicks tended to have shorter latency to first vocalization and a greater number of vocalizations compared with control chicks. Memory test showed that chicks from all groups traveled a similar distance toward a familiar object. However, control chicks walked the least toward a novel object, low dose chicks tended to walk further, and high dose chicks walked significantly further for a novel object. In aggression tests, both high and low dose males exhibited greater frequency of aggressive behaviors compared to controls, while no difference in aggression was evident in the females. Norepinephrine concentrations were also reduced in the low dose birds in the hypothalamus and in the raphe nucleus. Serotonin concentrations tended to be lower only in the both hypothalamus and raphe nucleus of the low dose birds. 5-HT1A expression was greatest in the hypothalamus and raphe nucleus of low dose birds. The agonism of the serotonin system during neural development of birds genetically predisposed to aggression alters both the dopaminergic and serotonergic systems further increasing their aggressiveness.

Silibinin inhibits VEGF secretion and age-related macular degeneration in a hypoxia-dependent manner through the PI-3 kinase/Akt/mTOR pathway.

BACKGROUND AND PURPOSE: Hypoxia-mediated neovascularization plays an important role in age-related macular degeneration (AMD). There are few animal models or effective treatments for AMD. Here, we investigated the effects of the flavonoid silibinin on hypoxia-induced angiogenesis in a rat AMD model. EXPERIMENTAL APPROACH: Retinal pigmented epithelial (RPE) cells were subjected to hypoxia in vitro and the effects of silibinin on activation of key hypoxia-induced pathways were examined by elucidating the hypoxia-inducible factor-1 alpha (HIF-1α) protein level by Western blot. A rat model of AMD was developed by intravitreal injection of VEGF in Brown Norway rats, with or without concomitant exposure of animals to hypoxia. Animals were treated with oral silibinin starting at day 7 post-VEGF injection and AMD changes were followed by fluorescein angiography on days 14 and 28 post-injection. KEY RESULTS: Silibinin pretreatment of RPE cells increased proline hydroxylase-2 expression, inhibited HIF-1α subunit accumulation, and inhibited VEGF secretion. Silibinin-induced HIF-1α and VEGF down-regulation required suppression of hypoxia-induced phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (mTOR) pathway. In the rat model of AMD, silibinin administration prevented VEGF- and VEGF plus hypoxia-induced retinal oedema and neovascularization. CONCLUSION AND IMPLICATIONS: The effects of silibinin, both in vitro and in vivo, support its potential as a therapeutic for the prevention of neovascular AMD.

Effects of selective serotonin antagonism on central neurotransmission.

Aggression and cannibalism in laying hens can differ in intensity and degree due to many factors, including genetics. Previous behavioral analysis of 2 strains of White Leghorns, DeKalb XL (DXL) and HGPS (a group-selected line for high group productivity and survivability), revealed high and low aggressive phenotypes, respectively. However, the exact genetic mechanisms mediating aggressiveness are currently unknown. Analysis of serotonin (5-HT) mediation of aggression in subordinate hens of these strains revealed increases in aggression in DXL hens following antagonism of the 5-HT1A receptor and in HGPS hens following antagonism of the 5-HT1B receptor. Here, we investigate the different neurotransmitter response in the hypothalamus and raphe nucleus mediating these aggressive responses to receptor antagonism. Elevated aggressive response to 5-HT1B antagonism by HGPS hens was also accompanied by a decrease in raphe nucleus dopamine (DA) and an increase in DA turnover. Increased aggressiveness in DXL hens did not coincide with a reduction in raphe nucleus 5-HT or turnover (as indicated by 5-hydroxyindoleacetic acid levels) following 5-HT1A antagonism. A reduction in 5-hydroxyindoleacetic acid (but not 5-HT) was seen in HGPS hens treated with 5-HT1A antagonist; however, these hens exhibited no change in aggressive behaviors. Our data show evidence of different heritable mechanisms of neurotransmitter regulation of aggressive response, specifically heritable differences in the interaction between 5-HT and catecholamines in regulating aggression.

Determining potential indicators of Cryptosporidium oocysts throughout the wastewater treatment process.

Most research on wastewater treatment efficiency compliance focuses on physicochemical and microbial indicators; however, very little emphasis has been placed so far on determining suitable indicator organisms to predict the discharge level of pathogens from treatment plants. In this study, raw wastewater, activated sludge, and the resulting final effluents and biosolids in four municipal wastewater treatment plants (WWTPs A, B, C and D) were seasonally investigated for human-virulent water-borne pathogens Cryptosporidium parvum/hominis and Giardia duodenalis, and microsporidia (e.g. Encephalitozoon hellem, E. intestinalis, and Enterocytozoon bieneusi) between 2008 and 2009. A suite of potential microbial indicators for human-virulent protozoa and microsporidia was also determined. A combination of multiple fluorescent in situ hybridization and immunofluorescent antibody assays were applied to detect Cryptosporidium oocysts, Giardia cysts, and microsporidian spores. Escherichia coli, enterococci and Clostridium perfringens spores were cultivated in selective media. Positive correlations were found between the abundance of enterococci and E. coli and abundance of Cryptosporidium oocysts (r(s) > 0.47, p < 0.01) and Giardia cysts (r(s) > 0.44, p < 0.01) at WWTPs A-D. Cryptosporidium perfringens spores were positively correlated to Cryptosporidium oocysts (r(s) = 0.40, p < 0.01) and Giardia cysts (r(s) = 0.46, p < 0.01). There was a strong positive correlation between abundance of Giardia cysts and that of Cryptosporidium oocysts (r(s) > 0.89, p < 0.01). To sum up, a suite of faecal indicator bacteria can be used as indicators for the presence of Cryptosporidium oocysts and Giardia cysts in these activated-sludge systems (WWTPs A, B and C). Overall, Giardia duodenalis was noted to be the best Cryptosporidium indicator for human health in the community-based influent wastewater and throughout the treatment process.

The dopaminergic system and aggression in laying hens.

The dopaminergic system is involved in the regulation of aggression in many species, especially via dopamine (DA) D1 and D2 receptor pathways. To investigate heritable differences in this regulation, 2 high aggressive strains [Dekalb XL (DXL) and low group egg productivity and survivability (LGPS)] and one low aggressive strain (low group egg productivity and survivability; HGPS) of laying hens were used in the study. The HGPS and LGPS lines were diversely selected using group selection for high and low group production and survivability. The DXL line is a commercial line selected through individual selection based on egg production. Heritable differences in aggressive propensity between the strains have been previously assessed. The birds were pair housed within the same strain and labeled as dominant or subordinate based on behavioral observation. For both experiments 1 and 2, behavioral analysis was performed on all 3 strains whereas neurotransmitter analysis was performed only on the most aggressive (DXL) and least aggressive (HGPS) strains. In experiment 1, the subordinate birds were treated with D1 agonist, D2 agonist, or saline controls (n = 12). In experiment 2, the dominant birds from a separate flock were treated with D1 antagonist, D2 antagonist, or saline controls (n = 12). Treatment-associated changes in aggressive behaviors and central neurotransmitters were measured. Aggression was increased in all strains in response to D1 agonism but increased only in the less aggressive HGPS birds with D2 agonism. Aggression was decreased and hypothalamic serotonin and epinephrine were increased in birds from all strains treated with D2 receptor antagonist. The D1 receptor antagonism elicited different behavioral and neurotransmitter responses based on the aggressive phenotype of the genetic strains. Aggressive strains DXL and LGPS but not the HGPS strain decreased aggressiveness following antagonism of the D1 receptor. The data show evidence for distinct neurotransmitter regulation of aggression in high and low aggressive strains of hens through different receptor systems. These chicken lines could provide new animal models for the biomedical investigation of the genetic basis of aggression.

[Locating the impairment of human cognitive function during hypoxia].

Objective. To find the location where the human brain cognitive function impairs under hypoxia. Method. 14 healthy males, aged 18-20 years, performed auditory Oddball test of two different intensities (55 dB, 80 dB) during exposure to 5000 m by breathing low oxygen mixture. EEG and Reaction Time (RT) were recorded. P3, the component of ERP, was extracted from EEG. P3 latency and RT were used as indices of the experiment. Additive Factors Method was used to analyse the result of the experiment. Result. Interaction between hypoxia and stimulus intensity was found for P3 latency and RT. Conclusion. Acute hypoxia influences the preprocessing stage of information processing.

Differential regulation of the growth-associated proteins, GAP-43 and SCG-10, in response to unilateral cortical ablation in adult rats.

Synapse replacement after brain injury has been widely documented by anatomical studies in various parts of both the developing and adult nervous system. However, the molecular events that define the specificity of the empirically derived rules of reactive synaptogenesis in different regions of the adult brain remain unclear. In this study we examined the differential regulation of the lesion-induced response of the two growth-associated proteins, superior cervical ganglia-10 and growth-associated protein-43, after unilateral cortex ablation, and determined a hierarchical order for the lesion response from remaining afferent projection neurons originating from the contralateral cortex, ipsilateral thalamus and substantia nigra. We report that in response to unilateral cortex ablation both messenger RNA, by northern blot, and protein, by western blot, for superior cervical ganglia-10 but not growth-associated protein-43 was increased in the homologous area of the contralateral cortex but not the ipsilateral thalamus or substantia nigra. In addition, the specificity of the superior cervical ganglia-10 response, assessed by combined in situ hybridization and retrograde FluoroGold labeling of striatal afferent neurons, found that superior cervical ganglia-10 messenger RNA was increased prominently in layer V pyramidal neurons of the contralateral corticostriatal pathway but was unchanged in afferent projection neurons from the thalamus and substantia nigra. Furthermore, the increase in both superior cervical ganglia-10 messenger RNA and protein seen at three days postlesion in contralateral corticostriatal neurons coincides in time with the initiation of neurite outgrowth in the deafferented striatum by contralateral corticostriatal axons described in our previous ultrastructural study. However, if cortical input to the striatum was removed bilaterally the lesion-induced response for superior cervical ganglia-10 messenger RNA shifted secondarily to thalamostriatal neurons in the ipsilateral thalamus. These data provide evidence that superior cervical ganglia-10 and growth-associated protein-43 are differentially regulated in neurons of the contralateral corticostriatal pathway in response to unilateral cortex ablation and suggests that superior cervical ganglia-10 plays a role in the regulation of neurite outgrowth in the adult striatum after brain injury. However, the specific role that superior cervical ganglia-10 may play in reactive synaptogenesis remains unclear. In addition, our data suggest that a hierarchical order exists for the reinnervation of deafferented striatal neurons after unilateral cortex ablation with preference given to homologous axons from the contralateral cortex.

Differential spine loss and regrowth of striatal neurons following multiple forms of deafferentation: a Golgi study.

Golgi-Cox method and morphometric analyses were used to study the plasticity of striatal medium spiny I neurons in 6-month-old C57BL/6N mice after unilateral or bilateral lesion of the cerebral cortex or combined lesions of the ipsilateral cerebral cortex and intralaminar thalamus. In adult mouse, unilateral lesions of the cerebral cortex did not result in a net gain or loss of linear dendritic length in a randomly selected population of striatal medium spiny I neurons. In addition, there was a well-defined time course of striatal spine loss and replacement occurring after a unilateral cortical lesion. By day 3 postlesion the average 20-microm dendritic segment had lost 30% of the unlesioned control spine value, reached its nadir, lost 45.5%, at 10 days postlesion, and recovered to 80% of unlesioned control levels by 20 days postlesion. The recovery of spines was blocked by a secondary lesion on the contralateral cortex but not on the ipsilateral intralaminar thalamus. These data suggest that striatal medium spiny I neurons of adult mice have a remarkable capacity for plasticity and reactive synaptogenesis following a decortication. The recovery of spine density is primarily induced by axonal sprouting of survival homologous afferent fibers from the contralateral cortex.

[Efficacy of Phyllanthus spp. in treating patients with chronic hepatitis B].

The efficacy of Phyllanthus amarus produced in india, P. niruri gathered from hainan province and P. urinaria from henan province was assessed in a total of 88 cases of chronic hepatitis B with 11.42 and 35 each. It was shown that P. urinaria had the effect of seroconversion on HBeAg from positive to negative as well as on HBeAb from negative to positive, while the other two herbs had not. In addition none of these three herbs had similar effect on HBsAg.