The efficacy of 2 phytases on inositol phosphate degradation in different segments of the gastrointestinal tract, calcium and phosphorus digestibility, and bone quality of broilers.

The anti-nutritional effects of dietary inositol phosphates (IP6 through IP3) have been recognized in broiler chickens; however, inositol hexaphosphate (IP6) is more potent than the lower IP esters. The efficacies of 2 commercial phytases, a Buttiauxella sp. phytase (BSP) and a Citrobacter braakii phytase (CBP) at 500 and 1,000 FTU/kg, were studied on IP6-3 concentrations in the crop, proventriculus + gizzard, and distal ileum digesta, and ileal IP6 disappearance in broilers at day 22. Apparent ileal P and Ca digestibility, and bone quality at days 22 and 33 were also measured. Female Ross 308 broilers (n = 1,890; 30 birds × 7 diets × 9 replicates) were fed corn-soy-based crumbled diets. The 7 diets included a primary breeder recommendation-based positive control diet (PC); the PC marginally reduced in available P by 0.146% and Ca by 0.134% of the diet, (NC1) or moderately reduced by 0.174 and 0.159% of the diet, respectively (NC2). Other diets were the NC1 + BSP or CBP at 500 FTU/kg (NC1+500BSP and NC1+500CBP) and the NC2 + BSP or CBP at 1,000 FTU/kg (NC2+1,000BSP and NC2+1,000CBP). Each of the NC1 and NC2 had distal ileum IP6 disappearance similar to that of PC, but each had lower P digestibility and the majority of measured bone quality parameters than the PC. The ileal IP6 levels were decreased by 52.0 and 32.7% for NC1+500BSP and NC1+500CBP, respectively, relative to NC1 and by 73.6 and 50.9% for NC2+1,000BSP and NC2+1,000CBP, respectively, relative to NC2 (P < 0.001), with a similar effect for distal ileum IP6 disappearance. Overall, phytase in the NC diets increased P digestibility, and femur breaking strength and cortical bone mineral density at days 22 and 33. Overall, each of the phytases at each dose degraded IP6-3 across the gastrointestinal tract segments to increase P digestibility and the P and Ca utilization in bone. However, dietary BSP at 1,000 FTU/kg was most effective. Supplemental phytase degrades phytate to decrease the anti-nutritional effects in a dose- and phytase-dependent manner.

Long-term effects of Buttiauxella sp. phytase on performance, eggshell quality, apparent ileal Ca and P digestibility, and bone properties of white egg layers.

Adequate dietary Ca and available phosphorus (avP) are essential to long-term egg production and bone health in laying hens. The effects of dietary Ca and avP levels and Buttiauxella sp. phytase (BSP) were studied in Lohmann LSL Lite hens from 30 to 70 wk of age (woa). Hens (n = 456; 4 per cage) were fed either a primary breeder recommendation-based diet (positive control; PC); the PC with avP and Ca levels reduced by 0.146 and 0.134% of the diet, respectively, without (NC) or with 300 FTU/kg BSP (NC+BSP). Egg production, BW, feed intake, FCR, and eggshell quality from 30 to 70 woa, and apparent ileal digestibility of P (AIDP) and Ca (AIDCa), and bone quality at 32, 48, and 70 woa were measured. The avP and Ca levels in the NC diet were not clinically deficient, as most parameters were unaffected by diet. Hen BW from 34 to 70 woa tended to be 2.9% greater (P = 0.076) for PC and NC+BSP compared to NC. Mid-diaphysis cortical bone mineral content (CBMC) tended to be 10% and 9% higher (P = 0.065) in the NC+BSP hens than in NC hens at 48 and 70 woa, respectively. AIDP of NC+BSP was 24% greater (P = 0.034) than of NC at 32 woa and tended to be 18% greater (P = 0.082) than AIDP of PC at 48 woa, and 25% lower than of NC and PC at 70 woa (P = 0.028). AIDCa was 25% lower for NC+BSP than PC at 48 woa only (P = 0.037). The avP and Ca sufficiency in the NC diet limited the opportunity to determine a phytase effect. Although the supplemental BSP tended to increase BW and 48 and 70 woa CBMC, and increased 32 woa AIDP, the efficacy of BSP could not be determined due to the lack of an NC effect on most parameters. Commercial laying hens can maintain health and productivity at lower than recommended levels of dietary Ca and avP; phytase supplementation may allow for even further reductions.

Coliform Bacteria for Bioremediation of Waste Hydrocarbons.

Raw, domestic sewage of Kuwait City contained about 106 ml-1 colony forming units of Enterobacter hormaechei subsp. oharae (56.6%), Klebsiella spp. (36%), and Escherichia coli (7.4%), as characterized by their 16S rRNA-gene sequences. The isolated coliforms grew successfully on a mineral medium with crude oil vapor as a sole source of carbon and energy. Those strains also grew, albeit to different degrees, on individual n-alkanes with carbon chains between C9 and C36 and on the individual aromatic hydrocarbons, toluene, naphthalene, phenanthrene, and biphenyl as sole sources of carbon and energy. These results imply that coliforms, like other hydrocarbonoclastic microorganisms, oxidize hydrocarbons to the corresponding alcohols and then to aldehydes and fatty acids which are biodegraded by ß-oxidation to acetyl CoA. The latter is a well-known key intermediate in cell material and energy production. E. coli cells grown in the presence of n-hexadecane (but not in its absence) exhibited typical intracellular hydrocarbon inclusions, as revealed by transmission electron microscopy. Raw sewage samples amended with crude oil, n-hexadecane, or phenanthrene lost these hydrocarbons gradually with time. Meanwhile, the numbers of total and individual coliforms, particularly Enterobacter, increased. It was concluded that coliform bacteria in domestic sewage, probably in other environmental materials too, are effective hydrocarbon-biodegrading microorganisms.

Effect of calcium level and phytase addition on ileal phytate degradation and amino acid digestibility of broilers fed corn-based diets.

This study investigated the effect of dietary Ca to available P (AvP) ratio and phytase supplementation on bone ash, ileal phytate degradation, and nutrient digestibility in broilers fed corn-based diets. The experimental design was a 4 × 2 factorial arrangement of treatments evaluating 4 Ca:AvP ratios (1.43, 2.14, 2.86, and 3.57) and 2 levels of phytase (0 and 1,000 phytase units/kg of feed). The 4 Ca:AvP ratios were achieved by formulating all diets to a constant AvP level of 0.28% and varying Ca levels (0.4, 0.6, 0.8, and 1.0%). Each treatment was fed to 6 cages of 8 male Ross 308 broilers from 5 to 21 d. At 21 d, digesta from the terminal ileum was collected and analyzed for energy, phytate, P, Ca, and amino acids (AA) to determine digestibility. Digesta pH was measured in each segment (crop, gizzard, duodenum, and ileum) of the digestive tract. Data were analyzed by 2-way analysis of covariance. There was a significant interaction between dietary Ca:AvP ratio and phytase supplementation for weight gain (WG), feed intake (FI), and feed conversion ratio (FCR). In diets with no phytase, Ca:AvP ratio had a greater effect on WG, FI, and FCR compared with those fed diets without phytase. The orthogonal polynomial contrasts showed that the increase in dietary Ca:AvP ratio significantly decreased WG and FI in a quadratic manner, whereas FCR increased (P < 0.05) linearly with higher dietary Ca:AvP ratio. Increasing dietary Ca:AvP ratio led to a significant quadratic decrease in phytate degradation and significant linear decreases in P digestibility and bone ash. Phytase addition increased (P < 0.05) phytate degradation and improved (P < 0.05) energy, AA, and P digestibility at all levels of Ca:AvP with no interaction (P > 0.05) between the main factors. Digestibility of AA was positively correlated (P < 0.05) with the degree of phytate degradation. Increasing dietary Ca:AvP ratio significantly increased gizzard pH in a linear manner. In conclusion, phytase (1,000 phytase units/kg of feed) improved phytate, and P and AA digestibility at all Ca:AvP ratios evaluated in this study.

Combination of capillary isoelectric focusing in a tapered capillary with MALDI-TOF MS for rapid and reliable identification of Dickeya species from plant samples.

This study was undertaken to investigate feasibility of a combination of capillary isoelectric focusing (CIEF) in a tapered fused silica (FS) capillary with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for a rapid and reliable identification of bacteria taken from plant-tissue-containing samples. Eight strains representing different species of the genus Dickeya were selected on the basis of close proximity of their isoelectric points: D. chrysanthemi, D. chrysanthemi bv. parthenii, D. chrysanthemi bv. chrysanthemi, D. dadantii, D. paradisiaca, D. solani, D. diffenbachiae, and D. dianthicola. Because the Dickeya species (spp.) cannot be easily discriminated from each other when CIEF is performed in a cylindrical FS capillary (commonly used in CIEF) even if a narrow pH gradient is used, a tapered FS capillary was employed instead, which enabled satisfactory discrimination of the examined bacteria due to enhanced separation efficiency of CIEF in the tapered FS capillary. CIEF in the tapered FS capillary was also successfully used for the detection and characterization of Dickeya spp. in a plant-tissue-containing sample. Then an off-line combination of CIEF with MALDI-TOF MS was employed for rapid and reliable identification of Dickeya spp. in the plant-tissue-containing sample. It was found that the presence of plant tissue did not affect the results, making the proposed procedure very promising with respect to the fast and reliable detection and identification of bacteria in plant-tissue-containing samples.

Proteomic profiling of Cronobacter turicensis 3032, a food-borne opportunistic pathogen.

Members of the genus Cronobacter are opportunistic pathogens for neonates and are often associated with contaminated milk powder formulas. At present little is known about the virulence mechanisms or the natural reservoir of these organisms. The proteome of Cronobacter turicensis 3032, which has recently caused two deaths, was mapped aiming at a better understanding of physiology and putative pathogenic traits of this clinical isolate. Our analyses of extracellular, surface-associated and whole-cell proteins by two complementary proteomics approaches, 1D-SDS-PAGE combined with LC-ESI-MS/MS and 2D-LC-MALDI-TOF/TOF MS, lead to the identification of 832 proteins corresponding to a remarkable 19% of the theoretically expressed protein complement of C. turicensis. The majority of the identified proteins are involved in central metabolic pathways, translation, protein folding and stability. Several putative virulence factors, whose expressions were confirmed by phenotypic assays, could be identified: a macrophage infectivity potentiator involved in C. turicensis persistence in host cells, a superoxide dismutase protecting the pathogen against reactive oxygen species and an enterobactin-receptor protein for the uptake of siderophore-bound iron. Most interestingly, a chitinase and a metalloprotease that might act against insects and fungi but no casein hydrolysing enzymes were found, suggesting that there is an environmental natural habitat of C. turicensis 3032.

Structural biology of pectin degradation by Enterobacteriaceae.

SUMMARY: Pectin is a structural polysaccharide that is integral for the stability of plant cell walls. During soft rot infection, secreted virulence factors from pectinolytic bacteria such as Erwinia spp. degrade pectin, resulting in characteristic plant cell necrosis and tissue maceration. Catabolism of pectin and its breakdown products by pectinolytic bacteria occurs within distinct cellular environments. This process initiates outside the cell, continues within the periplasmic space, and culminates in the cytoplasm. Although pectin utilization is well understood at the genetic and biochemical levels, an inclusive structural description of pectinases and pectin binding proteins by both extracellular and periplasmic enzymes has been lacking, especially following the recent characterization of several periplasmic components and protein-oligogalacturonide complexes. Here we provide a comprehensive analysis of the protein folds and mechanisms of pectate lyases, polygalacturonases, and carbohydrate esterases and the binding specificities of two periplasmic pectic binding proteins from Enterobacteriaceae. This review provides a structural understanding of the molecular determinants of pectin utilization and the mechanisms driving catabolite selectivity and flow through the pathway.