转植酸酶基因玉米在蛋鸡上的饲用安全性和有效性评价
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摘要
本论文通过3个试验系统研究了转植酸酶基因玉米(Phytase transgenic corn, PTC)对蛋鸡产蛋性能、蛋品质、血清生化、免疫反应、组织发育、肠道微生物区系和肠道消化能力的影响,并对饲喂PTC产蛋鸡消化道内食糜、血液、组织和鸡蛋中外源植酸酶基因phyA2及其表达蛋白进行了检测；同时,还对比了PTC和外源商业微生物植酸酶在低磷蛋鸡日粮上的饲喂效果,以评价PTC作为蛋鸡饲料原料的安全性和有效性。
试验一本试验分析对比了PTC及其亲本玉米(Isogenic conventional corn, CC)的营养成分,并评定了两种玉米在公鸡上的真代谢能、氨基酸和磷的真消化率。在公鸡绝食48h后,每只公鸡通过插管强饲法分别强饲50g CC、PTC、无氮日粮或无磷日粮,随后收集强饲后48h排泄物。无氮日粮和无磷日粮,分别用来测定公鸡内源氨基酸和内源磷的排泄量。结果表明,PTC的概略养分、氨基酸和脂肪酸含量与CC相比均无显著性差异(P>0.05)。PTC中植酸酶活性极显著高于CC,植酸磷含量极显著低于CC(P<0.001)。两种玉米在公鸡上的真代谢能、氨基酸真消化率均无显著性差异(P>0.05)。PTC在鸡上磷的真消化率,极显著高于CC(37.92vs.55.85%；P<0.001)。结果提示,PTC的营养成分含量及其在鸡上的能量和氨基酸利用率与CC无显著差异,PTC在鸡上的有效磷含量极显著高于其CC。
试验二本试验旨在研究长期饲喂PTC日粮对蛋鸡产蛋性能、蛋品质、血清生化指标、组织发育、肠道微生物区系和组织病理结构的影响,并对外源基因片段phyA2及其表达蛋白在产蛋鸡肠道内容物、血液、组织及鸡蛋中的存在情况进行了检测。选取50周龄的健康海兰褐产蛋鸡144只,随机分为2个处理,分别饲喂含62.4%的PTC或CC日粮。每个处理8个重复,每个重复9只鸡。预饲期2周,试验期16周。结果表明,与CC组相比,PTC日粮未对动物的产蛋性能和鸡蛋品质产生不利影响(P>0.05),但会降低蛋黄颜色(P<0.05)。长期饲喂转基因玉米日粮未对产蛋鸡血清生化指标、组织器官重量、免疫机能、组织病理结构及肠道微生物区系等指标产生不利影响。玉米外源植酸酶基因phyA2及其表达蛋白均可在动物肠道内被完全降解,未在产蛋鸡直肠食糜、血液、心脏、肝脏、脾脏、肾脏、胸肌和鸡蛋中检测到玉米外源基因片段phyA2及其表达蛋白。结果提示,与CC组相比,给蛋鸡长期饲喂含62.4%PTC日粮,未见对产蛋鸡的健康造成不利影响；外源转基因片段 phyA2不会迁移到蛋鸡组织和鸡蛋中。在本试验条件下,PTC作为饲料原料与其CC相比,在蛋鸡上有同等的饲用安全性。
试验三本试验通过转基因玉米来源植酸酶与两种商业微生物来源植酸酶(PA和PB)的对比试验,研究了转基因玉米来源植酸酶在低磷日粮中对蛋鸡产蛋性能、蛋品质、肠道食糜植酸酶残留酶活和植酸磷残留量、胫骨发育及营养物质回肠表观利用率的影响。将576只50周龄海兰褐产蛋鸡随机分为8个处理。分别为：正对照组(PC,0.32%NPP)、负对照组(NC,0.10%NPP),然后在NC基础上分别添加3种不同来源植酸酶,每种植酸酶分为两个添加水平(500和5,000(?)TU/kg)。每个处理8个重复,每个重复9只鸡。预饲期2周,试验期16周。结果表明,给蛋鸡长期饲喂低磷日粮(NC,0.10%NPP)会显著降低蛋鸡的产蛋率、产蛋量、采食量、体增重、蛋壳厚度和蛋壳强度,并提高料蛋比和破畸形蛋率(p<0.05)。在NC基础上分别添加3种不同来源的植酸酶,均可显著改善蛋鸡的产蛋性能和蛋品质(P<0.05),并可达到PC组相同的水平。蛋鸡各肠段内容物中植酸酶的残留活性,随着日粮植酸酶添加水平的增加而显著增加(P<0.01)。转基因玉米来源植酸酶在蛋鸡嗉囊、腺胃和肌胃、空肠和回肠食糜中的残留活性显著高于相同剂量的PA和PB添加组(P<0.05)。与NC组相比,日粮添加500～5,000FTU/kg任一来源的植酸酶,均可显著降低各肠段内容物中的植酸磷的含量(P<0.01)。各肠段中的植酸磷含量,随着肠道消化时间的延长逐渐降低。与NC组相比,日粮分别添加3种不同来源的植酸酶均可显著提高蛋鸡回肠磷的表观消化率、胫骨灰分含量和胫骨强度(P<0.05)。日粮磷在蛋鸡上的回肠表观消化率均随着3种植酸酶添加量的增加而显著增加(P<0.05)。结果提示,在低磷日粮中添加转基因玉米来源植酸酶可显著改善蛋鸡的产蛋性能和蛋品质,并显著提高蛋鸡各肠段中的植酸酶活性和植酸磷的降解率,改善蛋鸡骨骼质量。转基因玉米来源植酸酶与两种商业微生物植酸酶(PA和PB)相比,在低磷蛋鸡日粮中的饲喂效果相当。
总之,本研究通过一系列试验研究表明,以PTC作为饲料原料未对产蛋鸡的健康造成不利影响。外源植酸酶基因片段phyA2及其表达蛋白可在产蛋鸡消化道内被完全降解,未发现外源植酸酶基因phyA2及其表达蛋白在产蛋鸡血液、组织和鸡蛋中发生迁移和残留。与CC相比,PTC在蛋鸡上有同等的饲用安全性。与相同剂量的外源商业微生物植酸酶相比,转基因玉米来源植酸酶在低磷蛋鸡日粮上同等有效。
Three experiments were conducted to evaluate safety and efficiency of phytase transgenic corn (PTC) used as a feed ingredient in laying hens. The effect of PTC on the laying performance, egg quality, organ development, serum parameters, immune response and intestinal microflora of laying hens were studied and transgenic corn DNA fragments and proteins were detected in digesta, blood, tissues, and eggs of laying hens. Furthermore, we compared the effect of transgenic corn derived-phytase and two microbial phytases on the laying performance, egg quality, ileum nutrient digestibility, and bone mineralization in laying hens fed a low-P corn-soybean diet.
Exp.l was conducted to evaluate the compositional and nutritional equivalence of PTC and isogenic conventional corn (CC) in roosters. Following a48h fasting period, sixteen roosters were given50g of each sample via crop intubation and excreta were collected for48h. N-free and P-free diets were used to evaluate endogenous amino acid and endogenous P losses, respectively. Chemical composition was not different between PTC and CC, whereas the phytase activity for PTC was greater than CC(P<0.001). No difference was observed in the true metabolizable energy (TME) and true amino acid availability (TAAA) values between the PTC and CC in roosters. The true P utilization for PTC was greater than CC (37.92vs.55.85%;P<0.001), and CC and PTC contained0.13and0.18%available P (AP, DM basis,P<0.001), respectively. The results of this study indicated that the chemical composition, TME and TAAA in PTC are essentially equivalent to that in CC, and the true P utilization for roosters is higher in PTC than in CC.
Exp.2was conducted to assess the effects of long-term feeding with PTC to hens on laying performance, egg quality, organ development, serum parameters, immune response and intestinal microflora, and investigate the fate of transgenic DNA and protein in digesta, blood, tissues, and eggs. Fifty-week old laying hens (n=144) were fed with a diet containing62.4%PTC or CC for16weeks. We observed that feeding PTC to laying hens had no adverse effect on laying performance or egg quality (P>0.05) except on yolk color (P<0.05). There were no observed adverse effects of the PTC on the serum parmeters, organ development, immune response, intestinal microbial community in laying hens (P>0.05). Transgenic phyA2gene and protein were rapidly degraded in the digestive tract and were not detected in blood, heart, liver, spleen, kidney, breast muscle, and eggs of laying hens fed with diet containing PTC. It was concluded that performance of hens fed diets containing PTC, as measured by egg production and egg quality, was similar to that of hens fed diets formulated with CC. There was no evidence of phyA2gene or protein translocation to the blood, tissues, and eggs of laying hens.
Exp.3was compared the efficacy of transgenic corn-derived phytase (TCDP) and two other commercial microbial phytases (PA and PB) in a long-term feeding study of laying hens. The treatments consisted of a positive control (PC) diet adequate in P (0.32%NPP); a negative control (NC) diet low in P (0.10%NPP); and an NC supplemented diet containing three phytase sources (TCDP, PA and PB) at two supplemental levels (500and5,000FTU/kg of diet). Eight diets were fed to Hy-line hens (n=576) from50to66weeks of age. We found that with a reduction in dietary P in the NC diet, egg production, egg mass, feed intake, final BW, BW gain, shell thickness, and eggshell strength of laying hens decreased (P<0.05). In addition, the number of soft-shelled, cracked and broken eggs increased (P<0.05) in the NC group. The addition of TCDP, PA or PB significantly increased laying production and egg quality (P<0.05), and performed similarly in hens fed the PC diet. Hens fed each source of phytase had greater ileal P digestibility, tibia ash, and bone breaking strength than hens fed the NC diet (P<0.05). The residual phytase activities along the GIT had increased (P<0.01) with the addition of TCDP, PA and PB to the NC diets. The TCDP had higher residual activity (P<0.05) in the crop, proventriculus and gizzard, jejunum, and ileum as compared to the PA and PB activity. There was a decrease (P<0.01) in the phytate P content of the digesta from all sources of phytase supplementation in the NC diets. Residual phytate P content decreased caudally along the GIT of hens. The ileum P digestibility increased as phytase level increased from500to5,000FTU/kg of diet (P<0.05). Results from this study indicate that the addition of a novel TCDP to a P-deficient diet improves laying performance, egg quality, ileum P utilization, and bone mineralization, and TCDP is as efficacious as two commercial microbial phytases when P-deficient diets for laying hens were supplemented with it.
In conclusion, no adverse effect was found when the phytase transgenic corn was used in laying hens diet in the series of experiments. There was no evidence of phyA2gene or protein translocation to the blood, tissues, and eggs of laying hens. TCDP is as efficacious as two commercial microbial phytases in P-deficient diets for laying hens.
试验一本试验分析对比了PTC及其亲本玉米(Isogenic conventional corn, CC)的营养成分,并评定了两种玉米在公鸡上的真代谢能、氨基酸和磷的真消化率。在公鸡绝食48h后,每只公鸡通过插管强饲法分别强饲50g CC、PTC、无氮日粮或无磷日粮,随后收集强饲后48h排泄物。无氮日粮和无磷日粮,分别用来测定公鸡内源氨基酸和内源磷的排泄量。结果表明,PTC的概略养分、氨基酸和脂肪酸含量与CC相比均无显著性差异(P>0.05)。PTC中植酸酶活性极显著高于CC,植酸磷含量极显著低于CC(P<0.001)。两种玉米在公鸡上的真代谢能、氨基酸真消化率均无显著性差异(P>0.05)。PTC在鸡上磷的真消化率,极显著高于CC(37.92vs.55.85%；P<0.001)。结果提示,PTC的营养成分含量及其在鸡上的能量和氨基酸利用率与CC无显著差异,PTC在鸡上的有效磷含量极显著高于其CC。
试验二本试验旨在研究长期饲喂PTC日粮对蛋鸡产蛋性能、蛋品质、血清生化指标、组织发育、肠道微生物区系和组织病理结构的影响,并对外源基因片段phyA2及其表达蛋白在产蛋鸡肠道内容物、血液、组织及鸡蛋中的存在情况进行了检测。选取50周龄的健康海兰褐产蛋鸡144只,随机分为2个处理,分别饲喂含62.4%的PTC或CC日粮。每个处理8个重复,每个重复9只鸡。预饲期2周,试验期16周。结果表明,与CC组相比,PTC日粮未对动物的产蛋性能和鸡蛋品质产生不利影响(P>0.05),但会降低蛋黄颜色(P<0.05)。长期饲喂转基因玉米日粮未对产蛋鸡血清生化指标、组织器官重量、免疫机能、组织病理结构及肠道微生物区系等指标产生不利影响。玉米外源植酸酶基因phyA2及其表达蛋白均可在动物肠道内被完全降解,未在产蛋鸡直肠食糜、血液、心脏、肝脏、脾脏、肾脏、胸肌和鸡蛋中检测到玉米外源基因片段phyA2及其表达蛋白。结果提示,与CC组相比,给蛋鸡长期饲喂含62.4%PTC日粮,未见对产蛋鸡的健康造成不利影响；外源转基因片段 phyA2不会迁移到蛋鸡组织和鸡蛋中。在本试验条件下,PTC作为饲料原料与其CC相比,在蛋鸡上有同等的饲用安全性。
试验三本试验通过转基因玉米来源植酸酶与两种商业微生物来源植酸酶(PA和PB)的对比试验,研究了转基因玉米来源植酸酶在低磷日粮中对蛋鸡产蛋性能、蛋品质、肠道食糜植酸酶残留酶活和植酸磷残留量、胫骨发育及营养物质回肠表观利用率的影响。将576只50周龄海兰褐产蛋鸡随机分为8个处理。分别为：正对照组(PC,0.32%NPP)、负对照组(NC,0.10%NPP),然后在NC基础上分别添加3种不同来源植酸酶,每种植酸酶分为两个添加水平(500和5,000(?)TU/kg)。每个处理8个重复,每个重复9只鸡。预饲期2周,试验期16周。结果表明,给蛋鸡长期饲喂低磷日粮(NC,0.10%NPP)会显著降低蛋鸡的产蛋率、产蛋量、采食量、体增重、蛋壳厚度和蛋壳强度,并提高料蛋比和破畸形蛋率(p<0.05)。在NC基础上分别添加3种不同来源的植酸酶,均可显著改善蛋鸡的产蛋性能和蛋品质(P<0.05),并可达到PC组相同的水平。蛋鸡各肠段内容物中植酸酶的残留活性,随着日粮植酸酶添加水平的增加而显著增加(P<0.01)。转基因玉米来源植酸酶在蛋鸡嗉囊、腺胃和肌胃、空肠和回肠食糜中的残留活性显著高于相同剂量的PA和PB添加组(P<0.05)。与NC组相比,日粮添加500～5,000FTU/kg任一来源的植酸酶,均可显著降低各肠段内容物中的植酸磷的含量(P<0.01)。各肠段中的植酸磷含量,随着肠道消化时间的延长逐渐降低。与NC组相比,日粮分别添加3种不同来源的植酸酶均可显著提高蛋鸡回肠磷的表观消化率、胫骨灰分含量和胫骨强度(P<0.05)。日粮磷在蛋鸡上的回肠表观消化率均随着3种植酸酶添加量的增加而显著增加(P<0.05)。结果提示,在低磷日粮中添加转基因玉米来源植酸酶可显著改善蛋鸡的产蛋性能和蛋品质,并显著提高蛋鸡各肠段中的植酸酶活性和植酸磷的降解率,改善蛋鸡骨骼质量。转基因玉米来源植酸酶与两种商业微生物植酸酶(PA和PB)相比,在低磷蛋鸡日粮中的饲喂效果相当。
总之,本研究通过一系列试验研究表明,以PTC作为饲料原料未对产蛋鸡的健康造成不利影响。外源植酸酶基因片段phyA2及其表达蛋白可在产蛋鸡消化道内被完全降解,未发现外源植酸酶基因phyA2及其表达蛋白在产蛋鸡血液、组织和鸡蛋中发生迁移和残留。与CC相比,PTC在蛋鸡上有同等的饲用安全性。与相同剂量的外源商业微生物植酸酶相比,转基因玉米来源植酸酶在低磷蛋鸡日粮上同等有效。
Three experiments were conducted to evaluate safety and efficiency of phytase transgenic corn (PTC) used as a feed ingredient in laying hens. The effect of PTC on the laying performance, egg quality, organ development, serum parameters, immune response and intestinal microflora of laying hens were studied and transgenic corn DNA fragments and proteins were detected in digesta, blood, tissues, and eggs of laying hens. Furthermore, we compared the effect of transgenic corn derived-phytase and two microbial phytases on the laying performance, egg quality, ileum nutrient digestibility, and bone mineralization in laying hens fed a low-P corn-soybean diet.
Exp.l was conducted to evaluate the compositional and nutritional equivalence of PTC and isogenic conventional corn (CC) in roosters. Following a48h fasting period, sixteen roosters were given50g of each sample via crop intubation and excreta were collected for48h. N-free and P-free diets were used to evaluate endogenous amino acid and endogenous P losses, respectively. Chemical composition was not different between PTC and CC, whereas the phytase activity for PTC was greater than CC(P<0.001). No difference was observed in the true metabolizable energy (TME) and true amino acid availability (TAAA) values between the PTC and CC in roosters. The true P utilization for PTC was greater than CC (37.92vs.55.85%;P<0.001), and CC and PTC contained0.13and0.18%available P (AP, DM basis,P<0.001), respectively. The results of this study indicated that the chemical composition, TME and TAAA in PTC are essentially equivalent to that in CC, and the true P utilization for roosters is higher in PTC than in CC.
Exp.2was conducted to assess the effects of long-term feeding with PTC to hens on laying performance, egg quality, organ development, serum parameters, immune response and intestinal microflora, and investigate the fate of transgenic DNA and protein in digesta, blood, tissues, and eggs. Fifty-week old laying hens (n=144) were fed with a diet containing62.4%PTC or CC for16weeks. We observed that feeding PTC to laying hens had no adverse effect on laying performance or egg quality (P>0.05) except on yolk color (P<0.05). There were no observed adverse effects of the PTC on the serum parmeters, organ development, immune response, intestinal microbial community in laying hens (P>0.05). Transgenic phyA2gene and protein were rapidly degraded in the digestive tract and were not detected in blood, heart, liver, spleen, kidney, breast muscle, and eggs of laying hens fed with diet containing PTC. It was concluded that performance of hens fed diets containing PTC, as measured by egg production and egg quality, was similar to that of hens fed diets formulated with CC. There was no evidence of phyA2gene or protein translocation to the blood, tissues, and eggs of laying hens.
Exp.3was compared the efficacy of transgenic corn-derived phytase (TCDP) and two other commercial microbial phytases (PA and PB) in a long-term feeding study of laying hens. The treatments consisted of a positive control (PC) diet adequate in P (0.32%NPP); a negative control (NC) diet low in P (0.10%NPP); and an NC supplemented diet containing three phytase sources (TCDP, PA and PB) at two supplemental levels (500and5,000FTU/kg of diet). Eight diets were fed to Hy-line hens (n=576) from50to66weeks of age. We found that with a reduction in dietary P in the NC diet, egg production, egg mass, feed intake, final BW, BW gain, shell thickness, and eggshell strength of laying hens decreased (P<0.05). In addition, the number of soft-shelled, cracked and broken eggs increased (P<0.05) in the NC group. The addition of TCDP, PA or PB significantly increased laying production and egg quality (P<0.05), and performed similarly in hens fed the PC diet. Hens fed each source of phytase had greater ileal P digestibility, tibia ash, and bone breaking strength than hens fed the NC diet (P<0.05). The residual phytase activities along the GIT had increased (P<0.01) with the addition of TCDP, PA and PB to the NC diets. The TCDP had higher residual activity (P<0.05) in the crop, proventriculus and gizzard, jejunum, and ileum as compared to the PA and PB activity. There was a decrease (P<0.01) in the phytate P content of the digesta from all sources of phytase supplementation in the NC diets. Residual phytate P content decreased caudally along the GIT of hens. The ileum P digestibility increased as phytase level increased from500to5,000FTU/kg of diet (P<0.05). Results from this study indicate that the addition of a novel TCDP to a P-deficient diet improves laying performance, egg quality, ileum P utilization, and bone mineralization, and TCDP is as efficacious as two commercial microbial phytases when P-deficient diets for laying hens were supplemented with it.
In conclusion, no adverse effect was found when the phytase transgenic corn was used in laying hens diet in the series of experiments. There was no evidence of phyA2gene or protein translocation to the blood, tissues, and eggs of laying hens. TCDP is as efficacious as two commercial microbial phytases in P-deficient diets for laying hens.
引文
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