The Optimal Supplementation of Fermented Product Produced by Bacillus subtilis Strain LYS1 with High Surfactin Yield for Improving Growth Performance, Intestinal Villi Morphology, and Tibial Bone Strength in Broilers.

This study aimed to investigate the physiochemical characterizations of the fermented product (FP) produced by the high-yield surfactin strain Bacillus subtilis LYS1 (LYS1), as well as its effects on growth performance, carcass traits, intestinal morphology, tibial bone characteristics, and clinical blood biochemistry in broilers. Accordingly, the optimal supplementation of FP for improving growth performance, intestinal villi development, and tibial bone strength in broilers was elucidated using a broken-line quadratic (BLQ) ascending model. Three hundred and sixty 0-day-old Ross 308 broiler chicks, with equal numbers of both sexes, were randomly assigned to dietary supplementation of 2.5% fish meal or 0, 1, 1.5, 2, or 2.5% FP. Each treatment had six replicates, and the experimental period was 5 wk. The LYS1 count, surfactin content, and surfactin composition of the FP were 9.1 log CFU/g, 11.23 mg/g, and C12 to C18, respectively. The FP-supplemented groups improved feed intake, weight gain (WG), and production efficiency factor at 0 to 5 weeks old (p < 0.05) compared with the 0% group. The villus height/crypt depth (V/C) in the jejunum and ileum of the FP-supplemented groups was higher than in the 0% group (p < 0.05). The tibiotarsal index, Ca, and P in the tibia showed a linear effect with increased FP supplementation (p < 0.05). Moreover, the tibiotarsus weight/length index (TWLI) showed a quadratic effect with increased FP supplementation (p < 0.05). The optimal supplementation of FP for WG, V/C in the jejunum and ileum, and TWLI was 1.8, 1.9, and 1.6%, respectively. In conclusion, dietary supplementation with 1 to 2.5% LYS1 FP in broilers can improve their growth performance and the development of intestinal villi. Moreover, 1.9% is the optimal supplementation of LYS1 FP in the diet, based on the fitting results obtained with the BLQ model.

The fermented product of high-yield surfactin strain Bacillus subtilis LYS1 improves the growth performance and intestinal villi morphology in broilers.

This study aimed to select Bacillus spp. for surfactin production by solid-state fermentation and to investigate the physiochemical characterizations of the fermented product (FP) and its effect on growth performance, carcass trait, intestinal morphology, and clinical blood biochemistry of broilers. Accordingly, the correlations between the functional components of FP and the growth performance of broilers are elucidated. Four hundred eighty 1-day-old Ross 308 broiler chicks were randomly assigned to dietary supplementation of 2.5% fish meal, 2.5% unfermented product, or 2.5% FP produced by Bacillus subtilis LYS1 (LYS1), Bacillus amyloliquefaciens Da16, B. subtilis Lo6 (Lo6), B. subtilis NSN7, B. subtilis subsp. natto N21, or B. subtilis N12. Each treatment had 6 replicates. The experimental period was 5 wk. Results showed that the Lo6 showed the highest protease activity among all fermented groups. The LYS1 showed the highest surfactin yields (10.69 mg/g) among all fermented groups (P < 0.05). The weight gain (WG), feed conversion ratio (FCR), and production efficiency factor (PEF) of LYS1 group were significantly better than unfermented group at 0 to 3 and 0 to 5-wk-old (P < 0.05). The Bacillus-like counts and surfactin content of FP were moderately correlated to WG (0.7 > r > 0.3), FCR (-0.3 > r > -0.7), and PEF (0.7 > r > 0.3) at 0 to 3 and 0 to 5-wk-old (P < 0.05). The protease activity of FP was moderately correlated to WG (0.7 > r > 0.3), FCR (-0.3 > r > -0.7), and PEF (0.7 > r > 0.3) at 0 to 3-wk-old (P < 0.05). The villus height to crypt depth ratio in duodenum and jejunum of fish meal group and LYS1 group were higher than unfermented group (P < 0.05). In conclusion, LYS1 shows the highest surfactin yields. Diets supplemented with 2.5% LYS1 FP can improve the growth performance and the development of intestinal villi in broilers. Moreover, this study proves that the surfactin content, Bacillus-like counts, and protease activity of FP show a correlation to the growth performance of broilers.

Bacillus amyloliquefaciens CU33 fermented feather meal-soybean meal product improves the intestinal morphology to promote the growth performance of broilers.

This study is aimed to select optimum keratin degradation ability from Bacillus strains for feather meal-soybean meal fermentation, and favorably water content for the strain during fermentation of feather meal-soybean meal, and finally investigate the effects of the fermented feather meal-soybean meal product (FFSMP) on growth performance, carcass trait, clinical blood biochemistry, and intestinal morphology of broilers. Thirty-six bacteria strains from soil, sewage pool, and feather waste were screened and selected Bacillus subtilis var. natto N21 (N21), B. subtilis CU14 (CU14), and B. amyloliquefaciens CU33 (CU33) with better keratinase activity and feather-degrading rate. The result of trial 1 showed that the FFSMP produced by CU33 had the optimum physiochemical characterizations, amino acid composition and feeding performance for broilers. Hence the effects of water content (45, 50, 55, and 60%) on FFMSP fermentation of CU33 were investigated in trial 2. Result showed that pH value, counts of Bacillus-like bacteria, γ-PGA, viscosity, surfactin yield and odor all significantly increased according to the water content (P < 0.05). The protease activity reached significantly highest in the 55% and 60% water content groups (P < 0.01). The broilers performance of 55% and 60% water content group were significantly higher than control group (P < 0.05) in weight gain (WG), feed intake (b) at 0 to 21-d-old and the WG, feed conversion ratio (FCR), and production efficiency factor at 0 to 35-d-old, and could reach the similar growth performance as fish meal group (P > 0.05). The fermentation groups significantly decreased urea nitrogen (P < 0.05) and increased creatinine (P < 0.05) in the blood. The fermentation groups also significantly decreased the crypt depth in the duodenum (P < 0.05) and increased villus height to crypt depth ratio of the duodenum (P < 0.05). In conclusion, CU33 shows the best degradation rate for feather and keratinase activity, and the FFSMP with a water content of 50% to 60% during fermentation is suggested. Diets supplemented with 5% FFSMP can promote the growth of broilers by improving the morphology of the duodenum and achieve the feeding effect of high-quality fish meal.

Dietary Supplementation of Two-Stage Fermented Feather-Soybean Meal Product on Growth Performance and Immunity in Finishing Pigs.

This study investigates the effects of two-stage fermented feather meal-soybean meal product (TSFP) on growth performance, blood characteristics, and immunity of finishing pigs. Firstly, feather meal-soybean meal is subjected to aerobic fermentation with Bacillus subtilis var. natto N21, B. subtilis Da2 and Da15, B. amyloliquefaciens Da6, Da16 for two days, and anaerobic fermentation with B. coagulans L12 for three days. Then, the fermented product is air-dried into an end product-TSFP. Eighty hybrid pigs (Duroc x KHAPS) with equal numbers of both sexes are randomly assigned into 3% fish meal, 0%, 2.5%, or 5.0% TSFP groups with five replicates per group. Our results show that the average daily feed intake and feed conversion rate of TSFP groups are significantly better than the other groups at 0-3 weeks (p < 0.05). The 5% TSFP group significantly increased HDL-C in the blood (p < 0.05), and decreased LDL-C and blood urea nitrogen content (p < 0.05). The lipopolysaccharide (LPS) and concanavalin A (ConA) in 5% TSFP group and interferon-γ (IFN-γ) content in 2.5% and 5% TSFP groups are significantly higher than the other groups (p < 0.05). The phagocytic oxygen burst capacity and serum IgA content of the 5% TSFP group are significantly higher than those of the fishmeal group (p < 0.05). The CD3, CD4, and CD4 + CD8 + T cells subsets in 2.5% and 5% TSFP groups are significantly higher than the control group (p < 0.05). In conclusion, TSFP has a positive effect on the growth performance and immunity of finishing pigs with the best performance on 5% TSFP.

Screening lactic acid bacteria to manufacture two-stage fermented feed and pelleting to investigate the feeding effect on broilers.

Bacillus subtilis var. natto N21 (BS) and different lactic acid bacteria were applied to produce two-stage fermented feeds. Broilers were fed these feeds to select the best fermented feed. The selected fermented feed was pelleted and investigated for its effects on growth performance, carcass traits, intestinal microflora, serum biochemical constituents, and apparent ileal nutrient digestibility. Trial 1 involved three hundred thirty-six 1-d-old broilers with equal numbers of each sex, randomly assigned into control, BS + Bacillus coagulans L12 (BBC), BS + Lactobacillus casei (BLC), BS + Lactobacillus acidophilus (BLA), BS + Lactobacillus acidophilus L15 (BLA15), BS + Lactobacillus delbruekckii (BLD), and BS + Lactobacillus reuteri P24 (BLR24) groups with 3 replicates per group. Trial 2 involved two hundred forty 1-d-old broilers with equal numbers of each sex, randomly assigned into control, BBC, and pelleted BS + Bacillus coagulans L12 fermented feed (PBBC) groups with 4 replicates per group. Trial 3 involved sixteen 21-d-old male broilers randomly assigned into control and PBBC groups with 4 replicates per group for a nutrient digestibility trial. The feed conversion ratio (FCR) in the BBC group was better than the control (P < 0.05), and the production efficiency factor (PEF) was the best. However, weight gain (WG), feed intake (FI), and PEF were the lowest in the BLD group (P < 0.05). The WG during 0 to 21 d and 0 to 35 d in the PBBC groups were higher than the control (P < 0.05). The relative weight of the proventriculus + gizzard in the BBC and PBBC groups were higher than the control (P < 0.05). The digestible amino acid content in the PBBC group increased significantly (P < 0.05). Bacillus coagulans L12 is the best lactic acid bacteria for second stage fermentation. PBBC improved broiler growth performance, which may be due to the higher digestible amino acid content, it has the potential to become a commercial feed.

Establishment and application of an analytical in-house database (IHDB) for rapid discrimination of Bacillus subtilis group (BSG) using whole-cell MALDI-TOF MS technology.

Members of the Bacillus subtilis group (BSG) possess industrial applicability; unfortunately, B. subtilis and its phylogenetically closest species are indistinguishable from one another using 16S rDNA sequencing, physiological and biochemical tests. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is a relatively novel technique for the fast and reliable identification of microorganisms. The aim of this study was to construct a unique analytical in-house database (IHDB) for BSG discrimination based on whole-cell protein fingerprinting using MALDI-TOF MS, as well as to discover biomarkers from the MS peaks to generate a classification model for further differentiation using the ClinProTools software. Type strains of 12 species (included five subspecies) of the BSG were used to build a main spectrum profile (MSP) to create an IHDB under the optimized parameters. The BSG isolates obtained from partial recA gene sequencing were used for IHDB validation. A total of 84 (100%) isolates were correctly identified to the species level and had high score values (mean score: 2.52). However, the IHDB had ambiguous identification at the subspecies level of Bacillus amyloliquefaciens. After implementation of the classification models, the strains could be clearly differentiated. We have successfully developed a rapid, accurate and cost-effective platform for the species- and subspecies-level discrimination of BSG based on the implementation of the IHDB and coupled with ClinProTools, which can be employed as an alternative technology to DNA sequencing and applied for efficient quality control of the microbial agent.

Salmonella Serovars and Vaccination Effect on the Immune Responses of Male and Female Layers.

Salmonella Enteritidis, S. Gallinarum and S. Pullorum are common serovars to infect poultry and cause diseases differently. The antibody production and cellular immune responses of male and female layers were evaluated before and after inoculation. Before inoculation, S. Gallinarum and S. Pullorum could survive and grow in 10% sera from 6-week-old layers, and S. Enteritidis and E. coli were completely eliminated. The weights of the male and female layers were increased the lowest by inoculation with S. Gallinarum, followed by S. Pullorum, and S. Enteritidis. Inoculation with S. Enteritidis, S. Gallinarum and S. Pullorum increased the antibody titer in the males depending on the serovars and maintained same higher antibody level in females. Furthermore, an increased anti-Salmonella IgG titer was associated with bactericidal ability and the level was reduced by serovars and complemente. Despite the vaccination and serovars, the male layers expressed more IgG2a than IgG1, indicating preferential activation of the Th1 pathway. The inoculation number affected the expression level of IFN-γ and IL-12 in the blood not in the secretion of the peripheral blood mononucleated cells (PBMCs) and more inoculations increased the expression of both cytokines. Inoculation increased more reactive oxygen species (ROS) production in polymorphonuclear (PMN) cells, not the PBMCs. ROS production was greater in cells from the males than from the females and greater in the cells treated with S. Enteritidis than S. Gallinarum and S. Pullorum. These three serovars and their vaccinations differed in sera killing and immune responses.

Gender, caponization and exogenous estrogen effects on lipids, bone and blood characteristics in Taiwan country chickens.

This study investigated gender, caponization and exogenous estrogen effects on lipids, bone and blood characteristics in Taiwan country chickens. Thirty male chickens were caponized at 8 weeks (capons); 15 capons were injected with estrogen (5 mg/bird estradiol 3-benzoate) every 2 weeks from 8 to 28 weeks, and 15 sham-operated male (shams) chickens and 15 females were selected for this trial. The results showed that the shams had lower relative abdominal and chest subcutaneous fat than females (P < 0.05). The estrogen-treated capons had greater relative abdominal and chest subcutaneous fat than shams and capons (P < 0.05), which might result from higher blood very low-density lipoproteins and triacylglycerol concentrations (P < 0.05). Caponization could dramatically increase relative abdominal fat (506%; P < 0.05). The shams had higher tibia weight and biomechanical properties, such as maximum bone strength and bending moment values than the capons (P < 0.05). Tibia biomechanical properties were reduced by estrogen treatment (P < 0.05). The females obtained the lowest biomechanical value in all treatments (P < 0.05). Histological examination revealed cavity formation in the cortical bone of estrogen-treated capons and female chickens, which suggested that estrogen reduced bone biomechanical properties by destroying its structural integrity.

Prevalence and antimicrobial susceptibility of serogroup D nontyphoidal Salmonella in a university hospital in Taiwan.

The incidence of serogroup D Salmonella has been increasing in Taiwan. Most of these isolates belonged to Salmonella enterica serovar Enteritidis and showed a relatively higher rate of resistance to sulfamethoxazole-trimethoprim than to other antimicrobial agents. The results of molecular experiments indicated that genes responsible for the resistance were located on plasmids. The resistance may occur via horizontal gene transfer. Furthermore, the first identification of ciprofloxacin and ceftriaxone resistance in serogroup D Salmonella in our hospital is also than they did to other antimicrobial agents cause for concern.