Isolation and pathogenicity evaluation of Escherichia coli O157:H7 from common carp, Cyprinus carpio.

Escherichia coli O157:H7 is the primary serotype of enterohaemorrhagic E. coli (EHEC), which can cause diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. It is considered as a major health concern due to it being a zoonotic disease that is transmitted through food. In this study, a pathogenic bacterium was isolated from infected carp, which identified as E. coli O157:H7 named X21 through genetic sequencing, phylogenetic analysis, physiological and biochemical tests. In the experiment, crucian carp was used as a model to study the pathogenicity of the isolate, the pathological histological observations and cytokines expression of fish tissues were determined after bacterial challenge. The results showed that severe pathological damage observed in the liver, spleen, headkidney of fish infected with isolate X21. Besides, we found that accumulation of IgT+ B cells in the lamina propria of intestine, and up-regulation of SUCH-r, IL-1ß, IL-10, IL-11, MyD88, and TNF-α gene in various tissues. After challenged, the survivability of crucian carp infected with isolate X21 stands at a mere 14.27%. To our knowledge, this is the first report that E. coli O157:H7 infected the freshwater fish C. carpio, which indicates that this bacterium is a potential threat to public health and freshwater fish aquaculture.

Effect of in ovo administration of Newcastle disease vaccine conjugated with Astragalus polysaccharide on growth performance, intestinal development, and mucosal immunity in broiler chickens.

This study was conducted to investigate the effect of in ovo administration of a mixture of Astragalus polysaccharide (APS) and Newcastle disease vaccine (NDV) on growth performance, intestinal development, and mucosal immunity in newly hatched chicks. Six hundred specific-pathogen-free (SPF) Leghorn fertilised eggs were incubated in a commercial hatchery and divided into four groups: (a) control group injected with 1 ml of 0.9% physiological saline, (b) APS group injected with 1 ml of 1 mg/ml APS solution, and (c) NDV group injected with 1 ml of 104.0 EID50 /dose of NDV solution, and (d) APS + NDV group injected with a mixture of 0.5 ml of 2 mg/ml APS plus 0.5 ml 104.0 EID50 /dose ND vaccine (NDV) on Day 18.5 of incubation. The results showed that in ovo injection of APS or the mixture of APS and NDV increased the body weight at 1 day (IW) and final weight (FW) at 28 days and increased the feed conversion ratio (FCR) at 1-7, 8-14, 15-21, and 1-28 days of age. The villus height (VH) was increased (p < 0.05), and the crypt depth (CD) was decreased (p < 0.05) in the duodenum compared with the control group. The VH/CD ratios were increased (p < 0.05) in the APS + NDV group compared with controls, NDV group, and APS group on d3. The levels of slgA in washings were increased (p < 0.05) on Days 3, 7, 14, 21, and 28, and the number of IgA+ cells in the duodenum was increased on Days 7, 14, 21, and 28. In addition, the IgA+ cells were promoted from the villus root to the apex in the APS + NDV group. It can be concluded that in ovo administration of NDV conjugated with APS compared with NDV alone may be more effective in promoting growth performance and intestinal mucosal immunity.

The Low Dose of Saccharomyces cerevisiae Is Beneficial for Rumen Fermentation (Both In Vivo and In Vitro) and the Growth Performance of Heat-Stressed Goats.

This study aimed to investigate the effects of Saccharomyces cerevisiae on rumen fermentation and the growth performance of heat-stressed goats. The fermentation experiment was conducted using Saccharomyces cerevisiae added at 0‱ (HS1), 0.30‱ (SC1), 0.60‱ (SC2), and 1.20‱ (SC3) of the dry matter (DM) weight of the basal diet. The results showed that supplementing with 0.60‱ (SC2) could increase the pH, acetic acid to propionic acid ratio, the concentrations of ammonia nitrogen, total volatile fatty acids, acetic acid, propionic acid, butyric acid, and the degradability of DM, neutral detergent fiber, and acid detergent fiber in rumen fluids of heat-stressed goats. In the feeding experiment, twelve heat-stressed goats were assigned to a 4 × 4 Latin square experimental design, and the Saccharomyces cerevisiae supplement levels are similar to the fermentation experiment above. Similar effects on rumen fermentation and digestibility parameters were obtained with a supplement with 0.60‱ of Saccharomyces cerevisiae (SC2A) compared to the fermentation trial. Moreover, in the SC2A group, the DM intake and average daily gain also increased significantly compared with other groups. These results suggested that a low dose of Saccharomyces cerevisiae can still effectively improve the rumen fermentation and growth performance of heat-stressed goats.

Prophylactic Feeding of Clostridium butyricum and Saccharomyces cerevisiae Were Advantageous in Resisting the Adverse Effects of Heat Stress on Rumen Fermentation and Growth Performance in Goats.

This study aimed to investigate the effect of the prophylactic feeding of Clostridium butyricum (CB), Saccharomyces cerevisiae (SC), and their mixture before the onset of heat stress on the rumen fermentation and growth performance of goats, and subsequently, on heat stress status. Forty-eight male Macheng Black × Boer crossed goats (22.25 ± 4.26 kg) were divided into four groups­the control group (fed the basal diet), and the CB (0.05% CB added to the basal diet), SC (0.60% SC added to the basal diet), and Mix (0.05% CB and 0.60% SC added to the basal diet) groups­and fed for fourteen days. Then, these goats were kept in a heat stress environment (with a temperature−humidity index of 87.04) for fourteen days. Then, the parameters of rumen fermentation and growth performance were measured. The results showed that the pH values, the activities of cellulolytic enzymes (avicelase, CMCaes, cellobiase, and xylanase), and the concentrations of ammonia-N, total volatile fatty acid, acetic acid, propionic acid, and butyric acid were significantly increased (p < 0.05) in the rumens of the CB, SC, and Mix groups compared to those of the control group. Moreover, the average daily gain and the digestibility of dry matter, neutral detergent fiber, and acid detergent fiber were significantly increased (p < 0.05) in the CB, SC, and Mix groups compared to those of the control group. These results suggest that these two probiotics and their mixture effectively alleviate the adverse effects of heat stress on rumen fermentation and growth performance via prophylactic feeding.

The construction of recombinant Lactobacillus casei expressing hemagglutinin-neuraminidase protein and its immune response in chickens.

Newcastle disease virus (NDV) is one of the most important diseases in poultry. The present study generated recombinant surface-displayed Lactobacillus casei (L. casei) expressing the hemagglutinin-neuraminidase (HN) of NDV (Lc-pPG-HN) and a live pPG vector (Lc-pPG) and evaluated their immunogenicity. A 1670 bp HN gene fragment was successfully amplified and cloned into a prokaryotic protein expression system. Protein expression in the resulting recombinant Lc-pPG-HN (surface displayed) strain was verified using Western blotting and indirect immunofluorescence. A single band was observed on the Western blots, and the molecular weight of the corresponding protein was 63 kDa. A fluorescent signal for Lc-pPG-HN was observed using fluorescence microscopy. A total of 270 healthy chicks were divided into three treatment groups. Five replicates were used for each treatment, while six chicks were used per replicate. The following three treatment groups were used: physiological saline group (Control), Lc-pPG group and recombinant vaccine group (Lc-pPG-HN). The primary immunization and booster immunization of the chicks were performed via oral administration on 1 and 10 days old. Tissue and blood samples were collected from chickens that received oral recombinant L. casei strains on 1, 7, 14 and 21 days post-immunization for immune-related index analyses. Chickens orally immunized with Lc-pPG-HN showed significantly increased body weights and immune organ indices. Oral immunization with Lc-pPG-HN also enhanced the concentrations of serum interleukin-2 (IL-2), interferon-γ (IFN-γ), intestinal lavage fluid secretory immunoglobulin A (SIgA) and histomorphological development of the small intestine. Our results also indicated that recombinant L. casei significantly increased Lactobacillus and Bifidobacterium colonization and decreased the relative abundance of Escherichia coli (E. coli) in the chicken caecum. Similar enhancement effects from hemagglutination inhibition were also observed in the antibody titers. Oral administration of Lc-pPG-HN effectively protected against NDV and alleviated the symptoms of the NDV challenge. In summary, recombinant L. casei had positive impacts on the performance, immunological function, gut development, and microbiota of growing chicks and may be a potential therapeutic candidate against NDV.

The Effects of In Ovo Injection of Synbiotics on the Early Growth Performance and Intestinal Health of Chicks.

In this study, the effects of synbiotic inclusion at the intra-amniotic stage in layer chicks were evaluated with different parameters, such as performance, immunological function, intestinal development, and cecal microflora content. A total of 1,200 eggs with fertile embryos were allocated into four treatment groups. For every treatment, five replicates were used, and 60 eggs were included in each replicate. The following four treatment groups were established: the non-injected group, 0.9% physiological saline injection (saline) group, 1 × 106 CFU/egg Lactobacillus plantarum injection (probiotic) group, and 1 × 106 CFU/egg L. plantarum + 2 mg/egg Astragalus polysaccharide injection (synbiotic) group. In ovo injection was carried out at 18.5 days of incubation. The results showed that in ovo injection of probiotics or synbiotics did not affect the hatching or growth performance of the chicks but significantly increased their feed intake (FI), body weight (BW), and the feed conversion ratio (FCR). Additionally, in ovo injection of synbiotics enhanced the levels of serum interleukin-2 (IL-2), interferon-γ (IFN-γ), and secretory immunoglobulin A (SIgA) in intestinal lavage fluid and the histomorphological development of the small intestine. Our results also indicated that intra-amniotic synbiotic injection significantly increased Lactobacillus and Bifidobacterium colonization while decreasing the relative abundance of Escherichia coli in the chicken cecum (P < 0.05). In summary, in ovo injection of synbiotics had positive impacts on the performance, immunological function, gut development, and microbiota of growing chicks.

The immune adjuvant effect of Astragalus polysaccharide on in ovo injection of Newcastle disease vaccine.

In this study, immunogenic efficacies of in ovo administration of Astragalus polysaccharide (APS) along with live Newcastle disease vaccine (live ND vaccine) (live VG/GA strain) were evaluated. Four hundred fertilized eggs were randomly divided into four groups (n = 100/group), and vaccinated in ovo, respectively, with solutions of APS, live ND vaccine, live ND vaccine combined with APS, and 0.9% physiological saline into their amniotic fluid on d 18.5 of incubation. Significant improvement of chicks' development was displayed in those vaccinated with live ND vaccine adjuvanted with APS in ovo, manifested as enhanced hatchability and gaining weight. Moreover, in ovo administration of live NDV vaccine plus APS could significantly enhance the serum anti-NDV antibody titres and interferon gamma (IFN-γ), interleukin (IL)-2, IL-4 and IL-6 concentrations, promote lymphocyte proliferative capability as well as improve the frequencies of CD4+ and CD8+ T cells in peripheral blood. Overall results indicated in ovo administration of live ND vaccine adjuvanted with APS could stimulate stronger humoral and cellular responses in newly hatched chicks.

Dose accuracy of a durable insulin pen with memory function, before and after simulated lifetime use and under stress conditions.

OBJECTIVES: The objective of this study was to assess the dose accuracy of NovoPen® 5 in delivering low, medium and high doses of insulin before and after simulated lifetime use. A secondary objective was to evaluate the durability of the pen and its memory function under various stress conditions designed to simulate conditions that may be encountered in everyday use of an insulin pen. RESEARCH DESIGN AND METHODS: All testing was conducted according to International Organization for Standardization guideline 11608-1, 2000 for pen injectors. Dose accuracy was measured for the delivery of 1 unit (U) (10 mg), 30 U (300 mg) and 60 U (600 mg) test medium in standard, cool and hot conditions and before and after simulated lifetime use. Dose accuracy was also tested after preconditioning in dry heat storage; cold storage; damp cyclical heat; shock, bump and vibration; free fall and after electrostatic charge and radiated field test. Memory function was tested under all temperature and physical conditions. RESULTS: NovoPen 5 maintained dosing accuracy and memory function at minimum, medium and maximum doses in standard, cool and hot conditions, stress tests and simulated lifetime use. The pens remained intact and retained dosing accuracy and a working memory function at all doses after exposure to variations in temperature and after physical challenge. CONCLUSIONS: NovoPen 5 was accurate at all doses tested and under various functionality tests. Its durable design ensured that the dose accuracy and memory function were retained under conditions of stress likely to be encountered in everyday use.