Transformation of 19-norgestagens to estrogens by rumen microorganisms

The 19-norgestagens norethisterone acetate (17 alpha-ethinyl-4-oestren-17 beta-ol-3-on-17-acetate), ethinodiol diacetate (17 alpha-ethinyl-4-oestren-3 beta, 17 beta-diol-3, 17-diacetate), and norgestrol (17 alpha-ethinyl-18-methyl-4-oestren-17 beta-ol-3-on) are transformed to ethinyloestradiol or 18-methyl homologue by microorganisms of cattle rumen. Such transformation of steroid gestagens to oestrogens is likely to offer an explanation for the occurrence of oestrogen effects which had been observed during synchronised oestrus of cattle following oral application of 19-norgestagens.     

Effects of halogenated hydrocarbons on rumen microorganisms

Halogenated hydrocarbons such as polychlorinated biphenyls (PCB's), heptachlor (HEP), 1,1,1-trichloro-2,2-bis(p-chlorophenyl)-ethane (DDT), and pentachlorophenol (PCP) are environmental contaminants and, at times, can bioaccumulate in the food chain. Cattle have been contaminated in a variety of ways, but generally it is believed that they are only affected by high concentrations of the chemicals. Rumen microorganisms, however, may be affected at lower doses, thus possibly affecting the cow's growth and milk production. Polychlorinated biphenyls, HEP, DDT, and PCP were tested by a 1-stage in vitro fermentation procedure. Substrate utilization was determined by measuring percent dry matter disappearance. Four concentrations (0, 10, 50, and 100 ppm) were studied, and in vitro incubations were conducted for 24 and 48 hr. Samples were removed from 48-hr incubations to determine if the chlorinated hydrocarbons were metabolized during fermentation. Dry matter disappearance proved to be a reliable method to determine microbial activity in the presence of chemicals. Substrate dry matter disappearance for controls and all concentrations of PCB's, HEP, and DDT was approximately 50 and 80% at 24 and 48 hr, respectively. The PCP significantly (P less than 0.05) depressed the percent dry matter disappearance in 50- and 100-ppm cultures to 45 and 30% at 24 hr and 70 and 50% at 48 hr, respectively. Metabolic changes in the test chemicals were not detected by gas chromatographic analysis.     

Production of acids from inulin by a mixed culture of rumen microorganisms

The production of volatile fatty acids (VFA) and lactic acid from inulin (plant fructan with 2-1 bonds) in in vitro rumen fermentations was followed. The experiments were performed with inocula from wethers receiving two rations, with two pH regimes and using as inocula either whole rumen contents or the corresponding rumen fluid. The following results were obtained: The VFA production was higher when using inocula from hay and concentrate fed wethers. In contrast to this, the lactic acid production was higher when using inocula from hay fed wethers. The VFA production tends to be higher in a weakly acid medium than in a neutral medium. Acetate-to-propionate molar ratio was lower at lower pH. The amounts and composition of the VFA were not different when using both types of inocula. Thus, we believe that micro-organisms colonising plant fibres obviously do not play an important role in inulin splitting.     

苜蓿根瘤菌对不同形态磷利用效率的研究

为研究酸性土地区耐酸苜蓿根瘤菌对难溶性磷的利用,采用固体培养和液体培养方法在含不同形态磷(酵母膏有机磷、无机可溶性磷、磷酸钙、磷酸铝和磷酸铁),初始pH分别为4.1,7.0和9.0的培养基中进行了苜蓿根瘤菌的培养,观察并测定了不同固体培养基上的菌落形态和大小,液体培养基中根瘤菌的吸光度、培养液中有效磷含量和培养液pH。结果显示,在各pH条件下,无磷对照CK中的根瘤菌生长最差,其固体培养基上的菌落直径显著低于酵母膏有机磷对照(CK_YE-P)和无机磷对照(CK_P),同时,无磷对照液体培养中根瘤菌吸光度也远低于CK_YE-P和CK_P,说明在本试验条件下有效磷是影响根瘤菌生长的关键因素。固体培养基上4个不同难溶性磷处理中,植酸磷处理(T_Phy-P)菌落直径在各pH条件下皆为最大,磷酸钙处理(T_Ca-P)次之,而磷酸铝处理(T_Al-P)和磷酸铁处理(T_Fe-P)在各条件下的菌落直径都较小。根瘤菌经液体培养后,T_Phy-P和T_Ca-P的培养液在酸性和中性条件下有效磷含量和根瘤菌吸光度较高,但碱性条件下则较低;T_Al-P和T_Fe-P在各pH条件下的有效磷含量和根瘤菌吸光度皆较低。根瘤菌的接种培养使所有处理的pH下降,其中中性和酸性条件下的T_Ca-P以及中性条件下的T_Phy-P培养液pH变化相对较小。未接种对照中,T_Phy-P和T_Ca-P的有效磷含量在酸性条件下最大,随pH值升高而迅速降低,碱性条件下植酸钙镁几乎不溶;T_Al-P和T_Fe-P在3种pH条件下的有效磷没有明显变化。与未接种对照比,除酸性条件下的T_Phy-P和T_Ca-P外,其他接种处理的有效磷含量皆有不同程度的升高,碱性处理的升高幅度最大。苜蓿根瘤菌能较有效的利用植酸钙镁和磷酸钙中的磷,但在碱性条件下,对二者的利用大幅下降;根瘤菌很难利用磷酸铝和磷酸铁中的磷。培养液pH对植酸钙镁和磷酸钙的有效性影响较大,而对磷酸铝和磷酸铁影响较小。根瘤菌的生长会导致培养液pH的降低,促进难溶性磷中磷的释放,提高有效磷含量,但提高程度随磷形态而异。     

Resistance of feed enzymes to proteolytic inactivation by rumen microorganisms and gastrointestinal proteases.

Potential feed enzyme additives for ruminants were tested in vitro for their stability to ruminal microbial and gastrointestinal proteolysis. Four commercial preparations from Trichoderma longibrachiatum (A, B, C, and D) and one from an undisclosed source (E) were incubated up to 6 h with ruminal fluid taken from four lactating dairy cows before or 2 h after feeding. The stability of preparation B was also tested in the presence of pepsin at pH 3 and pancreatin at pH 7. Cellulase (EC 3.2.1.4), cellulose 1,4-beta-cellobiosidase (EC 3.2.1.91), beta-glucanase (EC 3.2.1.6), xylanase (EC 3.2.1.8), beta-glucosidase (EC 3.2.1.21), and beta-xylosidase (EC 3.2.1.37) activities were monitored throughout the incubations. Polysaccharidase activities of all enzyme preparations were remarkably stable in ruminal fluid taken after feeding. Ruminal fluid obtained before feeding inactivated the polysaccharidases in preparations B and D to a greater extent than ruminal fluid obtained after feeding. Cellulase and cellulose 1,4-beta-cellobiosidase activities were the least stable, declining (P < 0.05) by 35 and 60% for preparations B and D, respectively. Xylanase activity of preparation D decreased (P < 0.05) by up to 30% after 6 h of incubation, whereas beta-glucanase activity was not affected. The ability to degrade exogenous enzymes also differed among cows (P < 0.05). Pepsin and acid (pH 3.0) did not affect polysaccharidases in preparation B but decreased glycosidase activities by 10 to 15% (P < 0.05) after 1 h of incubation. Pancreatin, at the maximum concentration used, inactivated cellulase, cellulose 1,4-beta-cellobiosidase, and xylanase activities at a rate of 0.55, 1, and 0.45%/min, respectively. beta-Glucosidase and beta-xylosidase activities decreased by 1 and 0.75%/min, respectively. Partial proteolysis of cellulase, cellulose 1,4-beta-cellobiosidase, and xylanase by pancreatin produced a transient increase in activity. This twofold increase for cellulase and fourfold increase for cellulose 1,4-beta-cellobiosidase was directly proportional to pancreatin concentration. These results suggest that the enzyme feed additives tested were stable in the rumen of animals after feeding. Exogenous enzymes are likely to be more susceptible to the host gastrointestinal proteases in the abomasum and intestines than to ruminal proteases. However, exogenous polysaccharidases may survive for a considerable period of time in the small intestine and they probably maintain activity against target substrates in this environment.     

瘤胃微生物来源的纤维降解酶及其应用

瘤胃微生物含有种类丰富的纤维降解酶系统。通过研究纤维降解酶活性在瘤胃内的分布和各种瘤胃微生物降解酶系统等有助于这些酶的开发利用。系统生物学研究的快速发展为大规模克隆更多的纤维降解酶基因提供了可能。文中还综述了纤维降解酶在畜牧生产中的一些应用途径。     

Production of aromatic amino acids and related compounds from p-hydroxyphenylacetic acid by rumen microorganisms in vitro

Suspensions of mixed rumen bacteria (B), protozoa (P), and mixed rumen microorganisms (BP) prepared from rumen contents of fistulated goats were anaerobically incubated with 1 mM p‐hydroxyphenylacetic acid (HPA) at 39°C for 24 h. Tyrosine (Tyr), phenylalanine (Phe), tryptophan (Trp) and other related compounds in both supernatants and hydrolyzates of microbial cells in all incubations were analyzed by HPLC. Large amounts of Tyr (32.1, 42.7 and 36.1% of disappeared HPA in B, P and BP, respectively) were produced from HPA during a 12 h incubation period. The formation of Tyr in P (178.6 µmol/g MN) was 1.5 and 2 times higher than in B and BP, respectively. Phe (7–11% of the disappeared HPA) and Trp (3–6% of the disappeared HPA) were also synthesized from HPA in B, P, and BP. Phe synthesis in P (46.3 µmol/g MN) was 1.7 times higher than in B but, in contrast, Trp synthesis in B, was 1.6 times higher than in P. The metabolites p‐hydroxyphenylpyruvic acid (in the range of 5–14% of disappeared HPA), phenylacetic acid (1–11%), p‐hydroxybenzoic acid (3–7%) and benzoic acid (1–6%) were produced from HPA in B, P and BP. Phenylpropionic acid (6% of the disappeared HPA) was produced only in B and BP.     

荷斯坦奶牛瘤胃微生物分离鉴定及多样性分析

研究为分离获得荷斯坦奶牛瘤胃内容物中的细菌,建立系统进化树,获取有益菌种。采用培养组学技术和16S rDNA分子鉴定方法相结合,对3头健康荷斯坦奶牛瘤胃内容物中细菌进行分离培养。共分离得到105株细菌,包括肠球菌属(Enterococcus)共15株14.29%,芽孢杆菌属(bacillus)共11株10.48%,不动杆菌属(Acinetobacter)共14株13.33%,葡萄球菌属(Staphylococcus)共22株20.95%,梭菌属(Clostridium)共1株0.95%,狭义梭菌属(Clostridium sensu stricto)共2株1.90%,短杆菌科(Brevibacteriaceae)共2株1.90%,链球菌属(Streptococcus)共11株10.48%,气球菌属(Aerococcus)共4株3.81%,柠檬酸杆菌属(Citrobacter)共14株13.33%,杆菌属(Brachybacterium)共1株0.95%,克雷伯氏菌属(Klebsiella)共1株0.95%,普罗维登斯菌属(Providencia)共3株2.86%,沙雷氏菌属(Serratia)共4株3.81%。结果中所占比例最高的菌属是葡萄球菌属;系统进化树分析和GenBank中的同源性比对结果发现,从细菌门、纲、目、科、属、种分析,分支明确;26个菌种同源性都在95.01%~100%之间。分离纯化出11株芽孢杆菌属(Bacillus)细菌具有潜在益生菌活性。可作为饲料添加剂饲喂荷斯坦奶牛。     

Sulfur influences on rumen microorganisms in vitro and in sheep and calves

When continuously cultured ruminal microbes were given orchardgrass hay and sufficient sulfuric acid or hydrochloric acid to maintain a pH of 5.5, fermentation and numbers of protozoa were reduced compared with cultures whose pH was controlled with phosphoric acid. Likewise, when sulfur-deficient, purified diets were supplied to cultures, less methane (mmol X liter-1 X d-1), 3.2 vs 32.6, was produced and fewer cellulolytic bacteria (log10/ml), 5.8 vs 7.2 were present than when cultures were given the same diet supplemented with .3% elemental sulfur. The rumen of sheep fed the .04% sulfur diet had reduced digesta weights (1.69 vs 3.2 kg) compared with sheep fed the diet with .34% sulfur at the same intake. There also was reduced methanogenesis 12.3 vs 25.8 mmol X liter-1 X d-1) and reduced numbers of cellulolytic bacteria (7.4 vs 8.4 log10/ml) in sulfur-deficient sheep in comparison to sulfur-supplemented sheep. In growing calves, the same types of bacteria predominated in the rumen, but more facultative anaerobic bacteria were isolated from calves fed .04% sulfur than from calves fed diets with .34 to 1.72% sulfur. None of the dietary levels of sulfur appeared toxic. Regardless of treatment, volatile fatty acids were more predominant than lactic acid as end-products of fermentation of ruminal microbes in fermenters, sheep and calves. The greater methanogenesis and the greater cellulolytic bacterial numbers of sulfur-supplemented sheep compared with sulfur-deficient in vitro cultures, is interpreted to be the result of recycling of sulfur to the rumen in sheep where it is efficiently scavengered by ruminal bacteria.     

Influence of redox potential on metabolism of glucose in mixed cultures of rumen microorganisms

The rumen inocula taken from cows and sheep were incubated in pH controlled anaerobic batch cultures and the effect of the elevated redox potential (Eh) on metabolism of glucose was examined. In Eh regulated cultures (Eh = +100 mV) the VFA production and production of methane were decreased with a concomitant increase of production of lactate. In these cultures counts of lactilytic bacteria tended to be lower and counts of lactic acid bacteria higher than in cultures with a normal Eh. The elevated Eh also exerted an inhibitory effect on the rumen protozoa. The relevance of these changes to the rumen metabolism is discussed.