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1.
为探讨牛支原体氟喹诺酮类药物耐药靶位的突变情况,对分离自我国多个省份的牛支原体进行氟喹诺酮类药物耐药性检测和耐药株筛选,通过对临床分离敏感株、耐药株及体外诱导高度耐药株的氟喹诺酮类药物耐药决定区(QRDR)进行测序分析,发现分离菌株中有19%(6/32株)对氟喹诺酮类药物耐药,其QRDR中均存在gyr A(Ser83Phe)或par C(Ser80Ile)的氨基酸突变;但在体外诱导的高度耐药株中QRDR突变类型则以gyr A(Ser83Phe/Tyr或Glu87Lys)和par C(Ser80Ile或Asp84Asn/Tyr)的氨基酸发生突变为主,以上靶位突变在介导牛支原体对氟喹诺酮类药物耐药水平方面是否起着决定性作用,还有待进一步证明。  相似文献   

2.
本试验采用药敏纸片法对2014年从13个不同规模化鸡场分离的135株鸡源大肠杆菌进行常用12种抗菌药物敏感性试验,结果表明,头孢西丁和阿莫西林/棒酸耐药率较低,分别为17.8%、18.5%,而氟喹诺酮类的环丙沙星、诺氟沙星和左氧氟沙星的耐药率较高,分别为68.9%、65.2%和63.7%;进一步对高耐药率的氟喹诺酮类耐药菌株测定环丙沙星MIC值为4~256μg/ml;PCR扩增相关耐药基因(gyr A和par C),并测序分析耐药决定区突变及与MIC对应关系表明,耐药菌株突变主要发生在gyr A基因的Ser83密码子和Asp87密码子,及par C基因的Ser80密码子和Glu84密码子上,不同突变类型分别对应一定MIC值范围,其中73.1%(57/78)突变类型为gyr A基因的C(Ser83)→T(Leu)和G(Asp87)→A(Asn)突变,及par C基因的G(Ser80)→T(Ile)突变,其MIC值在8~256μg/ml之间;本研究提示规模化鸡场在防治大肠杆菌等细菌病时,应根据药敏结果选用药物,少用或不用氟喹诺酮类药物。  相似文献   

3.
为了解大肠杆菌对氟喹诺酮类药物的耐药机制和喹诺酮耐药决定区GyrA、GyrB、ParC、ParE四种基因的流行情况,采用聚合酶链式反应(PCR)技术对30株虎源大肠杆菌的耐药菌株进行了氟喹诺酮类药物耐药基因的检测,并对目的片段进行测序分析。结果表明:GyrA、GyrB、ParC、ParE阳性率分别为40.00%、63.33%、63.33%、40.00%;GyrA亚基发生Ser83→Leu、Asp87→Asn、Glu214→Gly的突变,GyrB亚基发生Ser195→Asn的突变,ParC亚基上氨基酸未发生取代,ParE亚基发生Ser85→Ala的突变。说明GyrA、GyrB亚基上发生的氨基酸替代是耐药菌对氟喹诺酮类药物产生耐药性的主要机制之一。  相似文献   

4.
15株动物源性耐氟喹诺酮类药物大肠杆菌进行PCR检测、测序、WDNASIS软件分析gyrA基因中的氟喹诺酮耐药决定区(QRDR)、AcrA以及编码与质粒介导的氟喹诺酮类药物耐药机制相关的qnrA、qnrB、qnrS、qepA和aac(6′)-Ib-cr基因。结果表明,15株耐药菌中,QRDR基因在其编码第72、75、83位或第87位氨基酸均发生突变;AcrA基因未检测到氨基酸的突变;qnrS、qepA和aac(6′)-Ib-cr耐药基因阳性菌各检测到1株,序列分析表明不存在氨基酸突变。QRDR基因编码的氨基酸4个位点发生突变,其中Ser83→Leu和Asp87→Asn 2个基因的突变均与文献报道的突变相同,双突变的7个菌株均表现为高度耐氟喹诺酮类抗生素,表明gyrA基因为大肠杆菌耐氟喹诺酮类抗生素的一个重要机制。高度耐氟喹诺酮类抗生素的菌株中有2株没有检测到氨基酸突变的存在,但是aac-(6′)-Ib-cr基因和qnrS检测为阳性,表明质粒介导的喹诺酮类耐药也可单独导致菌株的耐药。存有一个菌株gyrA基因编码的氨基酸发生突变Ser83→Leu,AcrA基因和qnrA、qnrB、qnrS、qepA和aac(6′...  相似文献   

5.
为了探讨耐喹诺酮类决定区(QRDR)、外排泵负调控基因(acrR、marR和soxR)突变对临床分离株氟喹诺酮(FQs)高水平耐药的影响,本研究对临床分离的18株FQs耐药大肠杆菌(E.coli),采用PCR方法检测QRDR、acrR、marR和soxR的突变情况;通过RT-PCR的方法检测外排泵及膜孔蛋白相关基因的表达水平。结果显示,QRDR的突变主要集中在常规突变GyrA(Ser83Leu 和 Asp87Asn)和ParC(Ser80Ile),同时也检测出稀有突变ParC Glu84Gly、Glu84Lys、Glu84Val和Glu84Ala,ParE Ser458Ala等。ED28在acrR基因存在777 bp插入序列;12株菌(包括ATCC25922)在MarR存在Gly103Ser和Tyr137His双突变,其中EP26和EG42存在插入片段;ED40在SoxR存在Thr38Ser、Gly74Arg氨基酸替换。在多突变药菌株中,AcrAB的表达水平明显升高,OmpC和OmpF表达量降低、甚至缺失。  相似文献   

6.
为了解近年来鸭源鸡杆菌(Gallibacterium anatis)的药物敏感性及其对氟喹诺酮类药物的耐药机制,以204株G.anatis为对象,选取11种抗菌药,用纸片法检测其药物敏感性,随后从中随机选取30株进行gyrA、parC基因的PCR扩增和测序,并分析喹诺酮耐药决定区编码氨基酸的突变情况。结果显示,试验菌株对复方新诺明、四环素、氟喹诺酮类药物的耐药率达86.76%以上,对头孢曲松与头孢噻肟的耐药率较低,分别为11.27%和7.35%;99.02%的菌株耐3种以上药物,92.16%的菌株耐6种以上药物,其中有6株对11种药物全部耐药;30株G.anatis全部扩增出parC基因,其中22株扩增出gyrA基因,GyrA亚基存在Ser83→Phe、Asp87→Ala/Tyr/Asn和Asp139→Tyr 3个位点氨基酸置换,ParC亚基存在Thr84→Ile、Glu88→Gly、Ala94→Val和Gly179→Val 4个位点氨基酸置换,敏感菌株不存在氨基酸位点突变,中度敏感菌株发生了3处氨基酸的替换,94.7%的耐药菌株发生了4处及以上氨基酸位点的置换。结果表明,G.anat...  相似文献   

7.
按常规方法提取鸡毒支原体参考株(S6-10)、疫苗株(F14-6)及在氟喹诺酮类药物压力下体外筛选出来的耐药株的基因组DNA,扩增各菌株DNA旋转酶gyrA基因片段,并克隆、测序,利用DNASIS分析软件对测序结果进行分析。结果表明,S6-10和F14-6在恩诺沙星或环丙沙星压力下均筛选出不同程度的耐药菌,而且恩诺沙星致鸡毒支原体发生耐药性突变的机率高于环丙沙星,S6-10株耐药突变频率高于F14-6。S6-10筛选出来的耐药菌株发生突变的位置在DNA旋转酶gyrA亚基的第83位(相对于大肠杆菌gyrA亚基中的位置,以下同)和87位上,取代模式为Ser83→Ile、Glu87→Gln,高水平耐药株(Se32M,MIC≥128)除了在第83位发生突变外,在第82位还增加一突变位点(Asp→Gly);F14-6筛选出来的高水平耐药菌株(Fe32M)在DNA旋转酶gyrA亚基的第83、87位发生了双位点突变(Ser83→Ile,Glu87→Gly),而低水平耐药株(Fe8M、Fc8M)在gyrA亚基未发生任何位点突变。  相似文献   

8.
对 5株临床分离的耐氟喹诺酮类药物的金黄色葡萄球菌 (MIC值范围 :4~≥ 12 8mg/ L) ,提取各细菌染色体DNA,对 gyr A和 grl A基因进行 PCR扩增 ,产物与 p MD18- T载体连接 ,转化至大肠杆菌 JM10 9感受态细胞 ,筛选阳性克隆 ,测序。序列分析结果表明 ,5株菌均发生了基因突变。突变位点 grl A:Ser80 (TCC)→ Phe(TGC) ,Ser80 (TCC)→ Tyr(TAC) ;gyr A:Ser84 (TCA)→ L eu(TTA )、Ser84 (TCA)→ Ala(GCA)、Glu88(GAA)→ L ys(AAA) ,其中 2株对氟喹诺酮类药物的 MIC值低 (4~ 6 4 m g/ L) ,均发生 grl A单一点突变 ,其余 3株在 gyr A和 grl A上产生多突变位点 ,且对氟喹诺酮类药物的 MIC值较高 (8~≥ 12 8mg/ L )。说明单一的点突变只能引起低水平或中等水平的耐药 ,高水平的耐药需要多位点的突变。 5株耐药菌株均发生 grl A Ser80→ Tyr(Phe)点突变 ,提示环丙沙星、诺氟沙星等氟喹诺酮类药物作用于金黄色葡萄球菌的首要靶酶是拓扑异构酶 Iv  相似文献   

9.
用微量稀释法测定盐酸环丙沙星等5种氟喹诺酮类抗菌药物对20株耐氟喹诺酮类药物的动物源致病性大肠杆菌的最低抑菌浓度(MIC),PCR扩增gyrA基因的喹诺酮耐药决定区(quinolone resistance determining regions,QRDR),扩增的片断长度为496 bp,PCR产物直接测序,用DNAStar软件分析氨基酸序列。结果显示,盐酸环丙沙星、恩诺沙星、氧氟沙星、甲磺酸培氟沙星和烟酸诺氟沙星的MIC范围分别为64~>512μg/mL、16~256μg/mL、16~128μg/mL、64~>512μg/mL和64~>512μg/mL。gyrA基因的突变位点均位于83和87位氨基酸位点,主要的变异方式为Ser83→Leu(15/20),其次是Asp87→Asn(13/20),其他的为Asp87→Tyr(6/20),Ser83→Trp(4/20),Ser83→Ala(1/20)和Asp87→Gly(1/20)。说明试验用菌株对氟喹诺酮类药物均表现为多重耐药,其gyrA基因QRDR的突变表现为多种形式。  相似文献   

10.
利用PCR结合变性高效液相色谱法快速检测大肠埃希菌喹诺酮耐药决定区gyrA基因和gyrB基因突变。方法:对22株喹诺酮耐药大肠埃希菌进行2种基因的PCR扩增,产物经DNA杂交形成同源和异源双链后进行变性高效液相色谱法检测和序列测定。结果:20株试验菌gyrA基因出现异常峰型,14株试验菌gyrB基因出现异常峰型。测序证实所有异常峰型的样本均存在突变位点,正常单峰则无突变位点。其中gyrA基因存在Ser83Leu和Asp87Asn突变,突变率分别为90.9%和86.4%;gyrB基因存在Glu185Asp突变,突变率为4.5%,两种基因均存在多个位点的同义突变。结论:DHPLC方法可用于快速检测大肠埃希菌gyrA和gyrB基因突变。  相似文献   

11.
试验选用96头平均体重14.82 kg左右的杜×长×大断奶仔猪,随机分成4组,每组3栏,每栏8头(公母各半)。对照组饲喂基础日粮,试验1、2、3组分别添加1%80目白术、0.2%白术多糖和1%微米白术。试验期30 d。结果表明:在生长性能方面,与对照组相比,1%微米白术添加组可显著提高日增重(P0.05)、降低饲料增重比和腹泻率,而且效果优于1%80目白术组和0.2%白术多糖组,在肠道形态和肠道微生态区系方面,与对照组相比,日粮添加1%80目白术、0.2%白术多糖、1%微米白术均可不同程度的提高十二指肠和空肠的绒毛高度,加深十二指肠和空肠的隐窝深度,并且增加肠道微生态区系的多样性,其中以1%微米白术添加组的效果最佳。  相似文献   

12.
Sissay, M.M., Uggla, A. and Waller, P.J., XXXX. Prevalence and seasonal incidence of nematode parasites and fluke infections of sheep and goats in eastern Ethiopia. Tropical Animal Health and Production, XXXX. A 2-year abattoir survey was carried out to determine the prevalence, abundance and seasonal incidence of gastro-intestinal (GI) nematodes and trematodes (flukes) of sheep and goats in the semi-arid zone of eastern Ethiopia. During May 2003 to April 2005, viscera including liver, lungs and GI tracts were collected from 655 sheep and 632 goats slaughtered at 4 abattoirs located in the towns of Haramaya, Harar, Dire Dawa and Jijiga in eastern Ethiopia. All animals were raised in the farming areas located within the community boundaries for each town. Collected materials were transported within 24 h to the parasitology laboratory of Haramaya University for immediate processing. Thirteen species belonging to 9 genera of GI nematodes (Haemonchus contortus, Trichostrongylus axei, T. colubriformis, T. vitrinus, Nematodirus filicollis, N. spathiger, Oesophagostomum columbianum, O. venulosum, Strongyloides papillosus, Bunostomum trigonocephalum, Trichuris ovis, Cooperia curticei and Chabertia ovina), and 4 species belonging to 3 genera of trematodes (Fasciola hepatica, F. gigantica, Paramphistomum {Calicohoron} microbothrium and Dicrocoelium dendriticum) were recorded in both sheep and goats. All animals in this investigation were infected with multiple species to varying degrees. The mean burdens of adult nematodes were generally moderate in both sheep and goats and showed patterns of seasonal abundance that corresponded with the bi-modal annual rainfall pattern, with highest burdens around the middle of the rainy season. In both sheep and goats there were significant differences in the mean worm burdens and abundance of the different nematode species between the four geographic locations, with worm burdens in the Haramaya and Harar areas greater than those observed in the Dire Dawa and Jijiga locations. Similar seasonal variations were also observed in the prevalence of flukes. But there were no significant differences in the prevalence of each fluke species between the four locations. Overall, the results showed that Haemonchus, Trichostrongylus, Nematodirus, Oesophagostomum, Fasciola and Paramphistomum species were the most abundant helminth parasites of sheep and goats in eastern Ethiopia.  相似文献   

13.
The purpose of this study was to determine the frequency of different tumor types within a large cohort of cats with intracranial neoplasia and to attempt to correlate signalment, tumor size and location, and survival time for each tumor. Medical records of 160 cats with confirmed intracranial neoplasia evaluated between 1985 and 2001 were reviewed. Parameters evaluated included age, sex, breed, FeLV/FIV status, clinical signs, duration of signs, number of tumors, tumor location(s), imaging results, treatment, survival times, and histopathologic diagnosis. Most of the cats were older (11.3 +/- 3.8 years). Primary tumors accounted for 70.6% of cases. Metastasis and direct extension of secondary tumors accounted for only 5.6 and 3.8% of cases, respectively. Twelve cats (7.5%) had 2 or more discrete tumors of the same type, whereas 16 cats (10.0%) had 2 different types of intracranial tumors. The most common tumor types were meningioma (n = 93, 58.1%), lymphoma (n = 23, 14.4%), pituitary tumors (n = 14, 8.8%), and gliomas (n = 12, 7.5%). The most common neurological signs were altered consciousness (n = 42, 26.2%), circling (n = 36, 22.5%), and seizures (n = 36, 22.5%). Cats without specific neurological signs were common (n = 34, 21.2%). The tumor was considered an incidental finding in 30 (18.8%) cats. In addition to expected relationships (eg, meninges and meningioma, pituitary and pituitary tumors), we found that lesion location was predictive of tumor type with diffuse cerebral or brainstem involvement predictive of lymphoma and third ventricle involvement predictive of meningioma.  相似文献   

14.
Genetic variations in chromosome Y are enabling researchers to identify paternal lineages, which are informative for introgressions and migrations. In this study, the male‐specific region markers, sex‐determining region‐Y (SRY), amelogenin (AMELY) and zinc finger (ZFY) were analysed in seven Turkish native goat breeds, Angora, Kilis, Hair, Honaml?, Norduz, Gürcü and Abaza. A SNP in the ZFY gene defined a new haplotype Y2C. All domestic haplogroups originate from Capra aegagrus, while the finding of Y1A, Y1B, Y2A and Y2C in 32, 4, 126 and 2 Turkish domestic goats, respectively, appears to indicate a predomestic origin of the major haplotypes. The occurrence of four haplotypes in the Hair goat and, in contrast, a frequency of 96% of Y1A in the Kilis breed illustrate that Y‐chromosomal variants have a more breed‐dependent distribution than mitochondrial or autosomal DNA. This probably reflects male founder effects, but a role in adaptation cannot be excluded.  相似文献   

15.
16.
Infectious diseases have always been a terrible scourge for humans. The appearance of these plagues, as they were called without distinction, was generally connected to various conditions: asters, climatic changes or religious reasons. The concept of contagious, and then infectious, diseases came slowly. Variolation, i.e. transmission of ‘virulent’ matter to induce a natural disease and the immunity against it, was brought from Constantinople to England by Lady Montague, in 1721. This ‘variolation’ technique was also often performed in veterinary medicine against diseases like sheep-pox or pleuropneumonia. As ‘vaccination’ is the term generally accepted for ‘immunisation’, variolation can be the word designating such a technique. The second period of the history of immunisation began, in 1880, with the studies of Pasteur and his collaborators. A great number of bacterial vaccines were developed: dead, live but attenuated or only parts of pathogens. The viruses were produced in animals, then in eggs and at last, in tissue cultures. Second generation vaccines appeared with genetic engineering: recombinant vaccines, vector vaccines, nucleic acids vaccines, and markers vaccines, among others. These novel technologies can permit the development of new ones and improve the quality of the vaccines already existing.  相似文献   

17.
The ash, silica and certain important micronutrients were estimated in conventional and unconventional feed and fodder resources available in Southern India. Commonly used dry roughages, such as paddy straw, ragi straw, maize kadbi/stalk, jowar kadbi/stalk, bajra stalk and wheat straw, were high in ash (9.9% +/- 0.77%) and silica (6.4% +/- 0.65%) and low in most of the other micronutrients, except iron, with paddy straw containing most silica (>9%). Cultivated non-leguminous (maize, jowar) and improved green crops (hybrid napier, guinea, green panic, NB-21, CO-1) were also high in ash (10.5% +/- 0.60% and 12.5% +/- 0.51%) but were moderate sources of P, Mg and Cu and good sources of Zn (98 +/- 13.8 ppm and 55 +/- 6.7 ppm). Leguminous green fodders (stylosanthus, lucerne, cow pea, soyabean) were excellent sources of Ca (1.9% +/- 0.16%), Mg (0.40% +/- 0.05%), Cu (30 +/- 5.2 ppm), Zn (121 +/- 14.7 ppm) and Fe (1234 +/- 166 ppm) and moderate sources of P. Mixed local grasses and weeds were high in silica (6.9% +/- 1.00%) but were good sources of Cu, Zn and Fe. Cereal grains (maize, wheat, rice, ragi) were low in ash (2.9% +/- 0.33%) and were relatively poor sources of Ca (0.22% +/- 0.03%), Mg (0.19% +/- 0.03%) and Cu (13 +/- 3.1 ppm). Pulses were low to medium sources of most minerals and good sources of Fe (1230 +/- 293 ppm). Oil seed cake/extractions (groundnut cake, cotton seed cake, soyabean meal, sunflower cake, safflower cake) and cereal by-products (rice polish, rice bran, wheat bran) were excellent sources of P (1.1% +/- 0.47% and 2.3% +/- 0.19%) and good sources of Zn (65 +/- 3.9 ppm and 66 +/- 10.7 ppm) and Fe (938 +/- 130 ppm and 662 +/- 126 ppm). Among the unconventional feeds screened, orange peel, sunflower heads, meat meal, rubber seed cake, spirulina algae and sea weeds contained plentiful Ca, Zn and Fe: tree leaves/top feeds (mulberry, erythrina, glyricidia, banana, subabul, groundnut haulms) were excellent sources of Ca (1.5% +/- 0.13%), Zn (120 +/- 22.9 ppm) and Fe (1033 +/- 133 ppm) but relatively poor sources of P. Soyabean husk, cocoa seed husk, rubber seed cake and meat meal were moderate to good sources of P (1.0% and 0.33%). The high Zn and Fe values of most feeds/fodders were probably due to soil contamination. This account of the micronutrient content of feed/fodder resources should help in strategic supplementation intended to alleviate local deficiencies.  相似文献   

18.
A survey of gastrointestinal parasite infections of young (<6 months old), immature (6–12 months old) and adult (>12 months old) dairy cattle on 16 farms in Kiambu District, Kenya was conducted during a dry season (September 1991 to January 1992) and during a wet season (March to July 1992). The survey was based on monthly coproparasitological examination of cohorts and worm counts in tracer calves. The effects of age, sex, farm and season on the prevalence and intensity of helminth and coccidial infections were determined. Faecal egg and oocyst counts revealed that the overall prevalences were: strongyles (including trichostrongyles) (85.5%), liver flukes (Fasciola gigantica) (34.0%), coccidia (30.9%) and tapeworms (9.6%). Eight species of the protozoan Eimeria were identified, the most prevalent species being E. bovis and E. zuernii. The most prevalent nematode genera were Haemonchus, Cooperia, Oesophagostomum and Trichostrongylus. Season, farm and age of the animals had a significant (p<0.05) influence on the intensity of infection with strongyles, liver flukes and coccidia, whereas the sex of the animals had no significant (p>0.05) effect on the prevalence or intensity of infections. A higher intensity of infection with strongyles and coccidia was found in the wet season than in the dry season (p<0.05). The age-specific intensity was in the following order: for strongyles, immature animals of 6–12 months of age had the highest egg counts, followed by young calves and adults. Calves had significantly (p<0.05) higher oocyst counts than immatures or adults. Liver fluke egg counts did not differ significant (p>0.05) between immatures and adult cattle.  相似文献   

19.
Bovine Mastitis in Selected Areas of Southern Ethiopia   总被引:3,自引:0,他引:3  
A study on bovine mastitis, designed to determine the causal agents, prevalence of infection and impact of risk factors in three cattle breeds, was conducted in selected areas of southern Ethiopia. A total of 307 lactating and non-lactating cows, of which 162 were indigenous Zebu, 85 Jersey and 60 Holstein-Friesian, were examined by clinical examination and the California mastitis (CMT) test. Of these, 40.4% were positive by CMT and bacteriology for clinical or subclinical mastitis, with prevalence rates of 37.1% and 62.9%, respectively. Out of 1133 quarters examined, 212 (18.7%) were found to be infected, 83 (39.2%) clinically and 129 (60.8%) subclinically. The prevalence of mastitis was significantly higher in Holstein-Friesian than in indigenous Zebu, in non-lactating cows than in lactating cows, in the early lactation stage than in the mid-lactation stage, in cows with lesions and/or tick infestation on skin of udder and/or teats than in cows without this factor, and in the wet season than in the dry season. Mastitis increased with parity number (R = 0.9). Of 248 CMT and clinically positive udder quarter samples analysed microbiologically, 212 were culturally positive for known mastitis pathogens and 36 were negative. Of the 199 positive samples, Staphylococcus accounted for 39.2%, Streptococcus for 23.6%, coliforms for 14.1%, Micrococcus and Bacillus species for 8.0% each and Actinomyces or Arcanobacterium (Corynebacterium) for 7.0%. It was concluded that there was a high prevalence of clinical and subclinical mastitis, mainly caused by Staphylococcus aureus, Streptococcus agalactiae and Escherichia coli, in this study area.  相似文献   

20.
不同日粮氮水平对山羊氮代谢和微生物蛋白质合成的影响   总被引:2,自引:0,他引:2  
本试验旨在探讨不同日粮氮水平对内蒙古白绒山羊氮代谢和微生物蛋白质合成(MCP)的影响,以达到提高氮利用率,减少环境污染和饲料资源的浪费。试验选用9只体况良好、体重为(43.83±2.95)kg,装有瘤胃瘘管的内蒙古白绒山羊,按体重随机分为3组,每组3只。日粮分为低氮7.5%、中氮10.5%、高氮13.5%3个氮水平。试验使用常规试验方法分析测定粪尿血液等生化指标,通过全收尿法和尿嘌呤衍生物法估测MCP。结果表明:随着氮水平增加,氨氮(NH3-N)浓度、氮摄入量、尿氮排出、尿中尿素氮(UUN)和MCP显著增加(P0.05),干物质采食量(DMI)显著降低(P0.01);沉积氮占总摄入氮的比例在中氮组最高,MCP在高氮组最高。综上所述,适当降低日粮氮水平可减少粪尿氮排放,提高反刍动物氮利用率。  相似文献   

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