The effectiveness of an Escherichia coli phytase in improving phosphorus and calcium bioavailabilities in poultry and young pigs.

The effectiveness of an Escherichia coli phytase in comparison with a commercially available Aspergillus phytase in improving the bioavailability of phosphorus in broilers, layers and young pigs was studied in three separate experiments. Three basal diets, marginally deficient in dietary P mainly provided as phytate, were formulated. Both phytases were added to the diets at the rate of 500 U/kg diet. The phytases significantly (P < or = 0.05) improved the availability of phytate P to broilers, layers and young pigs. Aspergillus and E. coli phytases enhanced the pre-caecal digestibility of P by 11 and 29% for broilers and 18 and 25% for layers, respectively. Total tract digestibility of P (P balance) was also enhanced but with smaller magnitude. In pigs, total tract digestibility of P was improved by 33 and 34% by Aspergillus and E. coli phytases, respectively. Under the conditions of this study, it was observed that E. coli consistently, though with small magnitude in layers and pigs, enhanced the availability of phytate P at the same range or slightly better than Aspergillus phytase. It was only in pigs that the availability of Ca was significantly (P < or = 0.05) improved by addition of both phytases. It can be concluded that E. coli phytase is highly effective in improving the bioavailability of phytate P to broilers, layers and young pigs. This seems to be based on the high proteolytic stability of the enzyme in the digestive tract, as shown recently.     

Escherichia coli sepsis and endotoxemia in conscious young pigs

In conscious pigs the influence of intravenous infusion of live E. coli (7×10⁸/kg), of the equivalent amount of endotoxin (20 g/kg) or of a high dose of endotoxin (2.5 mg/kg) on the hemodynamic, clinical and pathological parameters and on survival rate was studied. E. coli and endotoxin infusion resulted in pulmonary hypertension, systemic arterial hypotension, a decrease in cardiac output and an increase in heart rate. Clinical signs were characterized by respiratory and nervous disturbances, whereas necropsy revealed hemorrhages and edema in several organs. Although these findings were similar in the three groups, a marked difference in lethality was observed. Infusion of E. coli or of the high dose of endotoxin resulted in a significant mortality, whereas all pigs survived the infusion of the low dose of endotoxin. This suggests that the lethal pathophysiological mechanisms may only become activated when a sufficient amount of endotoxin is released into the circulation.     

An Escherichia coli phytase expressed in yeast effectively replaces inorganic phosphorus for finishing pigs and laying hens

Two experiments determined the efficacy of an Escherichia coli phytase (ECP) added to P-deficient, corn-soybean meal diets fed to finishing pigs and second-cycle laying hens. Sixty finishing pigs (49 +/- 0.9 kg) were formed into blocks within sex based on weight and ancestry and allotted to a P-deficient diet unsupplemented or supplemented with 0.10% inorganic P (iP) from KH2PO4 or ECP at 250, 500, 1,000, or 10,000 phytase units (FTU)/kg. Individually fed pigs were allowed ad libitum access to the experimental diets until a BW of 120 +/- 3 kg was achieved, at which time the pigs were euthanized and the left fibula and fourth metatarsal were excised for determination of bone ash. Pigs were fed a 2-phase diet program for early- and late-finishing pigs; available P in the basal diets was set 0.10% below the requirement. Dietary supplementation of iP or ECP increased weight gain (P < 0.10) and G:F (P < 0.01); performance was not different (P > 0.13) among the phytase-supplemented groups. Fibula ash was greatest (P < 0.01) for pigs fed diets containing 10,000 FTU of ECP/kg. Two hundred forty second-cycle hens were allotted to a P-deficient diet or a P-deficient diet supplemented with 0.10% iP or ECP at 150, 300, or 10,000 FTU/kg for a 12-wk experiment. The basal diet was a corn-soybean meal diet with no added iP (17% CP, 3.8% Ca, 0.10% available P). Hens fed the P-deficient diet were removed from the experiment after 4 wk due to poor egg production. Supplementation of iP or ECP resulted in increased (P < 0.01) feed intake, egg weight, and egg production during the first 4 wk. During the entire 12-wk period, there were no differences (P > 0.28) between the iP- and ECP-supplemented groups in feed intake, egg weight, or egg production. These experiments reveal that ECP was as efficacious as supplemental iP and that supplementation of an excess dose of ECP was efficacious and without negative effects in finishing pigs and laying hens.     

Effect of acupuncture on young pigs with induced enteropathogenic Escherichia coli diarrhea

Thirty-four preweaning pigs with induced enteropathogenic Escherichia coli diarrhea were treated with electroacupuncture, traditional acupuncture, or neomycin. In the group treated with electroacupuncture, points GV-1, bilateral ST-36, and Bai-hui were stimulated electrically. In the group treated with traditional acupuncture, points GV-1, bilateral ST-36, BL-20, bulb points, bilateral ear tip, and Shan-gen were used. Acupuncture points CV-12 and bilateral ST-25 also were treated with moxibustion (applying heat generated by a burning herb, Artemisia argyi). Hemoacupuncture also was applied to Shan-gen, bilateral ear tip, and bulb points. Pigs in the third group were given neomycin orally. Five pigs were inoculated with E coli, but were not treated and served as nontreated controls. At postinoculation day 5, 60% of control pigs and greater than 80% of pigs in treated groups recovered from diarrhea. However, at postinoculation day 3, recovery rates for pigs in the control and group treated with electroacupuncture were only 20 and 27.3%, respectively; whereas pigs treated with acupuncture or neomycin attained 81.8 and 71.4% of recovery rates, respectively. Seemingly, traditional acupuncture, but not electroacupuncture, was effective in controlling induced E coli diarrhea in pigs at its early stage.     

Escherichia coli phytase improves growth performance of starter, grower, and finisher pigs fed phosphorus-deficient diets

Corn-soybean meal-based diets, consisting of a high-P control (HPC) containing supplemental dicalcium phosphate (DCP), a basal diet containing no DCP, and the basal diet plus Escherichia coli phytase at 500 or 1,000 phytase units per kilogram (FTU/kg; as-fed basis) were fed to evaluate growth performance in starter, grower, and finisher pigs. Pigs were blocked by weight and gender, such that average weight across treatments was similar, with equal numbers of barrows and gilts receiving each treatment in each block. In Exp. 1, 48 pigs with an average initial BW of 11 kg, housed individually, with 12 pens per diet, were used to evaluate growth performance over 3 wk. Overall ADG and G:F were increased linearly (P < 0.05) by dietary phytase addition. Final BW and plasma P concentrations at 3 wk also increased linearly (P < 0.05). In Exp. 2, 128 pigs with an average initial BW of 23 kg, housed four pigs per pen, with eight pens per diet, were used to evaluate growth performance over 6 wk. A linear increase in response to phytase was noted for ADG and G:F in all three 2-wk periods, as well as overall (P < 0.05). Percentage of bone ash also showed a linear increase (P < 0.01). In Exp. 3, 160 pigs (53 kg), housed five pigs per pen, with eight pens per diet, were used to evaluate growth performance over 6 wk. A linear increase was detected for final BW, as well as ADG and G:F in the first and second 2-wk periods, and overall (P < 0.01). Twenty-four 15-kg individually housed pigs were used to evaluate total-tract nutrient digestibility in Exp. 4. Daily absorption of P linearly increased (P < 0.05) with phytase supplementation. Results of this research indicate that E. coli phytase is effective in liberating phytate P for uptake and utilization by starter, grower, and finisher pigs.     

Efficacy of an E. coli phytase expressed in yeast for releasing phytate-bound phosphorus in young chicks and pigs

Four chick trials and one pig trial were conducted to investigate the phosphorus-releasing efficacy oftwo commercial phytase enzymes (Natuphos and Ronozyme) and an experimental E. coli phytase enzyme (ECP) when added to corn-soybean meal diets containing no supplemental inorganic P (iP). In the 13- or 14-d chick trials, three or four graded levels of iP (0, 0.05,0.10,0.15%) from KH2PO4 were added to the basal diet to construct standard curves from which bioavailable P release could be calculated for the phytase treatments. In all cases, phytase supplementation levels were based on an assessment of phytase premix activity (i.e., P release from Na phytate at pH 5.5). Linear (P < 0.01) responses in tibia ash and weight gain resulted from iP supplementation in all assays. In the first chick trial, supplementation of 500 phytase units (FTU)/kg of ECP resulted in superior (P < 0.01) weight gain and tibia ash values compared with 500 FTU/kg of Natuphos. Results of the second chick trial revealed P-release values of 0.032 and 0.028% for 500 FTU/kg Natuphos and Ronozyme, respectively, and these were lower (P < 0.01) than the 0.125% P-release value for 500 FTU/kg of ECP. Tibia ash responded quadratically (P < 0.05) in response to graded levels of ECP up to 1,500 FTU/kg in the third chick trial. Combining Natuphos with either Ronozyme or ECP in Chick Trial 4 revealed no synergism between phytases with different initiation sites of P removal. The pig trial involved 10 individually fed weanling pigs per diet, and and phytase enzymes were supplemented to provide 400 FTU/kg in diets containing 0.60% Ca. Based on the linear regression of fibula ash on supplemental iP intake (r2 = 0.87), P-release values were 0.081% for Natuphos, 0.043% for Ronozyme, and 0.108% for ECP. These trials revealed an advantage of the E. coli phytase over the commercial phytases in young chicks.     

Efficacy and equivalency of an Escherichia coli-derived phytase for replacing inorganic phosphorus in the diets of broiler chickens and young pigs

Two studies were conducted to determine the efficacy of an Escherichia coli-derived phytase (ECP) and its equivalency relative to inorganic phosphorus (iP) from monosodium phosphate (MSP). In Exp. 1, one thousand two hundred 1-d-old male broilers were used in a 42-d trial to assess the effect of ECP and iP supplementation on growth performance and nutrient digestibility. Dietary treatments were based on corn-soybean meal basal diets (BD) containing 239 and 221 g of CP, 8.2 and 6.6 g of Ca, and 2.4 and 1.5 g of nonphytate P (nPP) per kg for the starter and grower phases, respectively. Treatments consisted of the BD; the BD + 0.6, 1.2, or 1.8 g of iP from MSP per kg; and the BD + 250, 500, 750, or 1,000 phytase units (FTU) of ECP per kg. Increasing levels of MSP improved gain, gain:feed, and tibia ash (linear, P < 0.01). Increasing levels of ECP improved gain, gain:feed, tibia ash (linear, P < 0.01), apparent ileal digestibility of P, N, Arg, His, Phe, and Trp at d 21 (linear, P < 0.05), and apparent retention of P at d 21 (linear, P < 0.05). Increasing levels of ECP decreased apparent retention of energy (linear, P < 0.01). Five hundred FTU of ECP per kg was determined to be equivalent to the addition of 0.72, 0.78, and 1.19 g of iP from MSP per kg in broiler diets based on gain, feed intake, and bone ash, respectively. In Exp. 2, forty-eight 10-kg pigs were used in a 28-d trial to assess the effect of ECP and iP supplementation on growth performance and nutrient digestibility. Dietary treatments consisted of a positive control containing 6.1 and 3.5 g of Ca and nPP, respectively, per kg; a negative control (NC) containing 4.8 and 1.7 g of Ca and nPP, respectively, per kg; the NC diet plus 0.4, 0.8, or 1.2 g of iP from MSP per kg; and the NC diet plus 500, 750, or 1,000 FTU of ECP per kg. Daily gain improved (linear, P < 0.05) with ECP addition, as did apparent digestibility of Ca and P (linear, P < 0.01). Five hundred FTU of ECP per kg was determined to be equivalent to the addition of 0.49 and 1.00 g of iP from MSP per kg in starter pigs diets, based on ADG and bone ash, respectively.     

Flow paths of plant tissue residues and digesta through gastrointestinal segments in Spanish goats and methodological considerations

A sequence of eight twice-daily meals, each marked with different rare earth elements, was fed to 24 Spanish goats (BW = 20.6 +/- 1.94 kg) to produce meal-based profiles of rare earth markers within segments of the gastrointestinal digesta on subsequent slaughter. Accumulative mean residence time and time delay of rare earths and segmental and accumulative mean residence times of indigestible NDF (IDF) were estimated for each sampled segment. Diets consisted of ad libitum access to bermudagrass hay with a limit feeding of one of four supplements: 1) minerals (basal, B); 2) B + energy (E); 3) B + CP (CP); or 4) B + E + CP for 84 d. Mean daily intake (g/kg of BW) during the 5 d before slaughter differed (P < 0.05) via diet for DM but not for IDF (8.0 +/- 0.35 g/kg of BW). Larger estimates of cumulative mean residence time for IDF vs. rare earths were suggested to be the consequence of a meal-induced bias in the single measurement of IDF pool size by anatomical site. The rare earth compartment method was considered more reliable than the IDF pool dilution method because it yielded flow estimates based on the flux of eight meal-dosed rare earth markers over 4 d and was independent of anatomical definitions of pool size. Statistically indistinguishable estimates for gastrointestinal mean residence times for IDF and rare earths conform to assumed indelibility for the specifically applied rare earths and indigestibility of IDF. The potentially digestible NDF (PDF):IDF ratio of dietary fragments (0.8) progressively decreased in the following order: caudodorsal reticulorumen (0.390) > crainodorsal reticulorumen (0.357) approximately reticulum (0.354) > mid-dorsal reticulorumen (0.291) approximately ventral reticulorumen (0.286), to that within the omasal folds and in the abomasum (0.259). Such a gradient of progressively aging mixture of plant tissue fragments is consistent with age-dependent flow paths established in the reticulorumen and flowing to the omasum and abomasum. Such heterogeneity of fragment ages within the reticulorumen is also indicated by the superior fit of marker dose site double dagger marker sampling site model assumptions. Additionally, cyclic meal- and rumination-induced variations in escape rate occur. Estimates of mean escape rates over days, needed for the practice of ruminant nutrition, must consider the complex interactions among plant tissues and the dynamics of their ruminal digestion of PDF.     

Pharmacological zinc levels reduce the phosphorus-releasing efficacy of phytase in young pigs and chickens

A pig trial and a chick trial were done to determine the effect of high levels of Zn and Cu on the P-releasing efficacy of phytase. Ninety-nine individually fed pigs (7.2 kg) were given ad libitum access to one of 11 experimental diets for a period of 21 d. Fibula ash (mg) was regressed against supplemental inorganic P (iP) intake (g) to establish the standard curve, from which phytase treatments were compared to determine P-releasing efficacy. The basal diet was a corn-soybean meal diet with no supplemental P (21% CP, 0.075% estimated available P, 130 mg of Zn/kg, as-fed basis). Diets included three graded levels of supplemental iP (0, 0.075, 0.150%) from reagent-grade KH2PO4, two levels of phytase (500 and 1,000 FTU/kg) from EcoPhos, 1,500 mg of Zn/kg from either Waelz ZnO or basic Zn chloride (Zn5Cl2(OH)8), and all combinations of phytase and Zn. One phytase unit (FTU) was defined as the amount of enzyme required to release 1 micromol of iP per minute from sodium phytate at 37 degrees C and pH 5.5. Phytase supplementation improved (P < 0.01) weight gain, G:F, and fibula ash (% and mg). Bone ash (mg) was highest (P < 0.01) for pigs fed diets containing 1,000 FTU/kg of phytase. Supplemental Zn had no effect (P > 0.50) on growth performance, but decreased (P < 0.05) fibula ash (mg). Comparison of the phytase treatments to the standard curve (r2 = 0.87) revealed P-release values of 0.130 and 0.195% for 500 and 1,000 FTU of phytase/kg, respectively, in the absence of Zn, whereas in the presence of Zn (pooled), P-release values were decreased (P < 0.01) to 0.092 and 0.132%, respectively. The effects of high levels of supplemental Zn (basic Zn chloride) and Cu (CuSO4 x 5H2O) on phytase efficacy also were investigated in a 12-d chick trial. Dietary treatments were arranged according to a 2(3) factorial, with two levels each of supplemental phytase (0 and 500 FTU/kg from EcoPhos), Zn (0 and 800 mg/kg), and Cu (0 and 200 mg/kg). There was a phytase x Zn interaction (P < 0.01) for tibia ash. Thus, Zn supplementation decreased tibia ash in the presence, but not in the absence, of phytase. Supplemental Cu did not affect (P > 0.30) the response to phytase. These results suggest that pharmacological levels of Zn chelate the phytate complex, thereby decreasing its availability for hydrolysis by phytase.     

Toxigenic Escherichia coli isolated from pigs in Argentina

The presence of porcine toxigenic E. coli (ETEC, VTEC) in 28 piggeries (5% of total) of the central and northeast region of Argentina was studied for a better understanding of the epidemiology of porcine strains. Samples were taken by rectal swabs from healthy piglets and from those with diarrhoea, in addition to their dams. Between 5-10 colonies were isolated from each one of 223 animals sampled from 1992 to 1997. By using specific primers each strain was screened by PCR for VT1, VT2all, VT2e, STIa, and LTI toxin genes. Only strains positive for any of the toxins mentioned above were screened for STb. Their O serogroups were determined by agglutination. All of the above enterotoxins and verocytotoxins were found in E. coli isolated from the animals. The STIa gene was detected in E. coli isolated from 27/127 piglets with diarrhoea, in comparison with LTI (4/127 pigs). No toxin gene was amplified from E. coli isolated from either healthy piglets or their dams. When strains isolated from 48 piglets without diarrhoea but showing delayed growth were analysed by PCR, their toxin profile was determined to be VT1 (1/48 piglets), VT2all (5/48), STIa (1/48), LTI (3/48) and VT2e (3/48). Serogroup O64 prevailed among ETEC; O138 prevailed for ETEC/VTEC strains. This is the first extensive study regarding porcine toxigenic E. coli in Argentina and constitutes an important database for the implementation of prevention measures.     

Prevalence of virulence genes in Escherichia coli strains recently isolated from young pigs with diarrhea in the US

Enterotoxigenic Escherichia coli (ETEC)-associated post-weaning diarrhea (PWD) is economically one of the most important diseases for the swine industry. Porcine ETEC strains typically express K88 or F18 fimbria and heat-labile (LT) and/or heat-stable (STa, STb) enterotoxins. However, recent studies indicate that EAST1 toxin, adhesin involved in diffuse adherence (AIDA-I) and porcine attaching and effacing-associated factor (paa) may also be expressed by ETEC strains associated with diarrhea. To better understand the virulence factors of E. coli strains that cause PWD, we applied PCR to screen for K88, F18, F41, 987P and K99 fimbrial genes; LT, STa, STb, Stx2e and EAST1 toxic genes; and AIDA-I, paa and EAE adhesin genes in E. coli strains recently isolated from young pigs with PWD in the US. Of 304 E. coli isolates from diarrheic pigs submitted for testing, 175 (57.6%) strains possessed fimbrial genes: K88 (64.6%), F18 (34.3%), F41 (0.57%), K99 (0.57%), 987P (0); toxin genes: LT (57.7%), STb (72.6%), STa (27.4%), STx2e (17.4%), EAST1 (35%); and adhesin genes: AIDA-I (26.9%), paa (60%), EAE (1.1%). All toxin genes except the EAST1 toxin gene, were almost exclusively associated with K88+ or F18+ isolates, and most of these isolates carried multiple toxin genes. The non-fimbrial adhesin paa was found present in over half of the K88+ isolates. A total of 129 (42%) isolates carried no fimbrial genes, including 66 (21.7%) isolates that did not have any of the above virulence genes. These results suggest a broad array of virulence genes associated with PWD in pigs.     

Corn expressing an Escherichia coli-derived phytase gene: a proof-of-concept nutritional study in pigs

Two experiments were conducted to investigate the concept that the addition of corn expressing an Escherichia coli-derived gene (corn-based phytase; CBP) to a P-deficient diet would improve growth performance and P utilization in pigs. An E. coli-derived microbial phytase (expressed in Pichia pastoris) sprayed onto a wheat carrier (Quantum) was included for comparison. In Exp. 1, forty-eight 10-kg pigs were blocked by BW into 6 blocks and allotted to 8 dietary treatments such that the BW among dietary treatments was similar and given free access to feed for 28 d. The dietary treatments were a negative control (NC) with no inorganic P supplementation; NC + 2, 4, or 6 g of monosodium phosphate/kg; NC + 16,500, 33,000, or 49,500 phytase units (FTU) of CBP/kg; and NC + 16,500 FTU of Quantum/kg. In Exp. 2, twenty-four 13-kg barrows were assigned to the NC, NC + 16,500 or 33,000 FTU of CBP/kg, or NC + 16,500 FTU of Quantum/kg, in a nutrient- and energy-balance study consisting of 5 d of adjustment and 5-d collection periods. The total collection method was used to determine nutrient and energy balance. Addition of CBP to the low-P NC diet linearly increased (P < 0.01) ADG, G:F, and plasma P concentration of pigs during the 28-d study. There was no difference in ADG, G:F, or plasma P concentration between pigs fed the CBP or Quantum phytase at 16,500 FTU/kg. Weight gain, G:F, and plasma P concentration of pigs increased (P < 0.01) with monosodium phosphate supplementation, confirming P deficiency of the NC diet. Linear improvements (P < 0.05) in DM digestibility and energy retention were observed with CBP supplementation of the NC diet. Although there were linear (P < 0.01) and quadratic (P < 0.05) increases in N digestibility, N retention was unaffected by CBP supplementation of the NC diet in growing pigs. Phosphorus and Ca digestibilities and retentions improved linearly and quadratically (P < 0.01) with the addition of CBP to the NC diet. There was no difference in digestive utilization of P or Ca between pigs fed CBP and Quantum phytase at 16,500 FTU/kg. The data showed that the addition of a corn expressing an E. coli-derived gene to a P-deficient diet improved growth performance and indices of P utilization in pigs, and corn expressing phytase was as efficacious as Quantum phytase when supplemented in P-deficient diets for weanling pigs.