No abstract

beta-1,4-Mannanases (mannanases), which hydrolyse mannans and glucomannans, are located in glycoside hydrolase families (GHs) 5 and 26. To investigate whether there are fundamental differences in the molecular architecture and biochemical properties of GH5 and GH26 mannanases, four genes encoding these enzymes were isolated from Cellvibrio japonicus and the encoded glycoside hydrolases were characterized. The four genes, man5A, man5B, man5C and man26B, encode the mannanases Man5A, Man5B, Man5C and Man26B, respectively. Man26B consists of an N-terminal signal peptide linked via an extended serine-rich region to a GH26 catalytic domain. Man5A, Man5B and Man5C contain GH5 catalytic domains and non-catalytic carbohydrate-binding modules (CBMs) belonging to families 2a, 5 and 10; Man5C in addition contains a module defined as X4 of unknown function. The family 10 and 2a CBMs bound to crystalline cellulose and ivory nut crystalline mannan, displaying very similar properties to the corresponding family 10 and 2a CBMs from Cellvibrio cellulases and xylanases. CBM5 bound weakly to these crystalline polysaccharides. The catalytic domains of Man5A, Man5B and Man26B hydrolysed galactomannan and glucomannan, but displayed no activity against crystalline mannan or cellulosic substrates. Although Man5C was less active against glucomannan and galactomannan than the other mannanases, it did attack crystalline ivory nut mannan. All the enzymes exhibited classic endo-activity producing a mixture of oligosaccharides during the initial phase of the reaction, although their mode of action against manno-oligosaccharides and glucomannan indicated differences in the topology of the respective substrate-binding sites. This report points to a different role for GH5 and GH26 mannanases from C. japonicus. We propose that as the GH5 enzymes contain CBMs that bind crystalline polysaccharides, these enzymes are likely to target mannans that are integral to the plant cell wall, while GH26 mannanases, which lack CBMs and rapidly release mannose from polysaccharides and oligosaccharides, target the storage polysaccharide galactomannan and manno-oligosaccharides.

We evaluated the effects of 3 additives, sodium butyrate (AC), avilamycin (AB), and a combination of plant extracts (XT), on the productive performance and the intestinal environment of the early-weaned pig. The XT was a standardized mixture with 5% (wt/wt) carvacrol (from Origanum spp.), 3% cinnamaldehyde (from Cinnamonum spp.), and 2% capsicum oleoresin (from Capsicum annum). Pigs (n = 32) weaned at 18 to 22 d of age with an initial BW of 6.0 +/- 0.10 kg were allocated to 8 pens that, in turn, were allocated to 4 treatments. The treatments included a basal diet (CT) or the basal diet supplemented with 0.3% of AC, 0.04% of AB, or 0.03% of XT. Productive performance was determined during the initial 14 d postweaning. On d 19 and 21 of the experiment, the pigs were killed to allow collection of digesta and intestinal tissue to evaluate variables indicative of aspects of the gastrointestinal environment. Treatments AB and AC improved G:F (P = 0.012 and 0.003, respectively) compared with the CT. Butyrate included in the diet was only detected in the stomach but not in cranial jejunum. When compared with CT, AC produced a lower ileal starch digestibility (P = 0.002) and a lower whole-tract OM and starch digestibility (P = 0.001 and 0.003, respectively), related to a lower VFA concentration in the cranial colon (P = 0.082) and a numerically reduced branched VFA percentage in the rectum. The AB treatment diminished propionate production in caudal colon (P = 0.002) and rectum (P = 0.012) compared with CT. The AC group exhibited deeper crypt depth in the jejunum without variations in villus height compared with CT (P = 0.042). The AC and AB groups also increased goblet cell presence in the colon (P = 0.001 and 0.032, respectively). On the other hand, AB and XT diminished intraepithelial lymphocytes in the jejunum (P = 0.003 and 0.034, respectively). The XT increased lymphocyte presence in the colon (P = 0.003). These results show the important influence of AB and AC on productive performance and on pig gut dynamics. The intestinal modifications observed for AB and AC compared with CT suggest distinct modes of action for each additive.

The efficacy of a commercial source of mannanoligosacharides (BM), organic zinc (BP), or their combination to enhance performance, gastrointestinal health, and immune response in weaned pigs was evaluated. A total of 128 piglets, weaned at 20 +/- 2 d, were housed in 32 pens. Animals received 1 of 4 dietary treatments: a control diet (CT) to which 0.2% of BM, 80 mg/kg of Zn as BP, or both additives (BMP) were added. The experiment lasted for 5 wk including a prestarter period of 2 wk and a starter period of 3 wk. Body weight was recorded and daily feed intake was calculated. Fecal consistency was monitored for the first 21 d. After 2 wk, 32 animals were killed, digesta samples from the stomach, ileum, and cecum were collected, and pH and the short-chain fatty acid profile were determined. Microbiological counts for enterobacteria and lactobacilli were evaluated using quantitative PCR. Histological parameters in the jejunum and immunoglobulin concentrations in serum and ileal digesta were also measured. Both additives improved G:F during the starter period (0.63, 0.69, 0.67, and 0.68 for CT, BM, BP, and BMP, respectively; P < 0.04). Mean fecal score values for the first 21 d were improved by BM and BP, showing decreased values compared with the CT diet (1.22, 0.89, 0.87, and 1.06 for CT, BM, BP, and BMP, respectively; P = 0.002). The addition of BM decreased enterobacteria counts in the jejunum (9.13, 8.05, 8.87, and 7.89 log 16S rRNA gene copies/g of matter for CT, BM, BP, and BMP, respectively; P = 0.05). Empty ileal weight, defined as the segment including the continuous Peyer's patch, tended (P = 0.08) to increase with BP treatment (8.9, 9.6, 11.9, and 10.3 g/kg of BW for CT, BM, BP, and BMP, respectively). Crypt depths in the jejunum were lower in animals fed the combination of the additives (BPM) compared with those fed the control diet (281 vs. 235; P < 0.03). No significant differences were registered in pH, short-chain fatty acids, or serum and ileal immunoglobulin concentrations. The results suggest that the use of BM or BP can improve the efficiency of gain during the starter period.

An experiment was designed to evaluate the effects of 3 different additives on the gastrointestinal microbiota of early-weaned pigs. Early-weaned (18 to 22 d; n = 32) pigs (6.0 +/- 0.10 kg of BW) from 8 litters were randomly distributed into 8 pens. Each pen was assigned 1 of 4 dietary treatments: a prestarter or control diet, the control diet with 0.04% avilamycin (AB), with 0.3% sodium butyrate, or with 0.03% plant extract mixture (XT; standardized mixture with 5% (wt/wt) carvacrol extracted from Origanum spp., 3% cinnamaldehyde extracted from Cinnamonum spp., and 2% capsicum oleoresin from Capsicum annum). At the end of the experimental period, 8 pigs per treatment were killed, and samples of their intestinal content were taken. The total bacterial load along the gastrointestinal tract (GIT; stomach, jejunum, cecum, and distal colon) and the lactobacilli and enterobacteria in the jejunum and cecum were measured by quantitative PCR. The total microbial counts along the GIT did not differ among the diets, but there was an increase in the lactobacilli:enterobacteria ratio in the cecum of the piglets on the XT diet (P = 0.003). Restriction fragment length polymorphism of the PCR-amplified V3, V4, and V5 regions of the 16S rDNA gene showed changes in the structure of the microbial community in the jejunum. Dendrograms grouped animals by diets; control with 0.3% sodium butyrate was the treatment that promoted the biggest changes in the microbial ecosystem, followed by AB and then XT. Biodiversity increased when using additives compared with the control diet (P = 0.002). Microbial metabolic activity along the hindgut was studied using the concentration of purine bases and carbohydrase activities. Different patterns for purine bases were observed between diets (diet x intestinal section, P = 0.01). The control diet reached a maximum purine base concentration at the end of the colon, whereas that of the AB diet was reached at the cecum. We could not detect any cellulase or xylanase activities in animals of this age. Appreciable amylase and amylopectinase activities were found, but they did not differ between diets. The results suggest that the effects of avilamycin, butyrate, or the plant extract would not be related to a reduction in the number of total bacteria inhabiting different sections of the GIT but rather to changes in the ecological structure and metabolic activity of the microbial community.