Responsive changes of rumen microbiome and metabolome in dairy cows with different susceptibility to subacute ruminal acidosis

Subacute ruminal acidosis(SARA)represents one of the most important digestive disorders in intensive dairy farms,and dairy cows are individually different in the severity of SARA risk.The objectives of the current study were to investigate differences in the ruminal bacterial community and metabolome in dairy cattle with different susceptibility to SARA.In the present study,12 cows were initially enrolled in the experiment.Based on average ruminal pH,4 cows with the lowest ruminal pH were assigned to the susceptible group(SUS,pH=5.76,n=4)and 4 cows with the highest ruminal pH assigned to the tolerant group(TOL,pH=6.10,n=4).Rumen contents from susceptible(SUS,n=4)and tolerant(TOL,n=4)dairy cows were collected through rumen fistula to systematically reveal the rumen microbial and metabolic alterations of dairy cows with different susceptibility to SARA using multi-omics approaches(16S and 18S rRNA gene sequencing and metabolome).The results showed that despite being fed the same diet,SUS cows had lower ruminal pH and higher concentrations of total volatile fatty acids(VFA)and propionate than TOL cows(P<0.05).No significant differences were observed in dry matter intake,milk yield,and other milk compositions between the SUS and TOL groups(P>0.05).The principal coordinates analysis based on the analysis of molecular variance indicated a significant difference in bacterial composition between the two groups(P=0.01).More specifically,the relative abundance of starch-degrading bacteria(Prevotella spp.)was greater(P<0.05),while the proportion of fiber-degrading bacteria(unclassified Ruminococcaceae spp.,Ruminococcus spp.,Papillibacter,and unclassified Family_-XIII)was lower in the rumen of SUS cows compared with TOL cows(P<0.05).Community analysis of protozoa showed that there were no significant differences in the diversity,richness,and community structure(P>0.05).Metabolomics analysis revealed that the concentrations of organic acids(such as lactic acid),biogenic amines(such as histamine),and bacterial degradation products(such as hypoxanthine)were significantly higher in the SUS group compared to the TOL group(P<0.05).These findings revealed that the higher proportion of starch-degrading bacteria/lower fiber-degrading bacteria in the rumen of SUS cows resulted in higher VFA-producing capacity,in particular propionate.This caused a disruption in metabolic homeostasis in the rumen which might be the reason for the higher susceptibility to SARA.Overall,these findings enhanced our understanding of the ruminal microbiome and metabolic changes in cows susceptible to SARA.     

Infectious pancreatic necrosis: first isolation of virus from fish in Norway.

Infectious pancreatic necrosis (IPN) is a disease of salmonid fishes. It has been reported in many countries throughout the world (M’Gonigle 1940, Wood et al. 1955, Besse & Kinkelin 1965, Vestergård Jørgensen & Bregnballe 1969, Sano 1971, Ball et al 1971, Ljungberg & Vestergård Jørgensen 1972, Schlotfeldt et al. 1975). Outbreaks of the disease can cause serious losses in populations of hatchery reared salmonids, the mortality rate varying between 10 and 90 % (Vestergård Jørgensen & Kehlet 1971). There are at least four different serotypes of the virus distinguished by neutralization tests (Wolf et al. 1968). Twenty-five isolates of IPN virus in Denmark proved to represent only two serotypes (Sp and Ab) (Vestergård Jørgensen & Kehlet). The present paper reports the first isolation of IPN virus from the stock at a fish farm in Norway.     

Studies of Aspergillus fumigatus; casein precipitating and proteolytic effects of mycelial filtrate

Schäffer (1900) and Butkewitch (1903) seem to have been the first to focus attention on the proteolytic activity of micro-fungi. The occurrence and properties of proteolytic enzymes from various fungus species were subsequently studied by several authors. Literature in this field was reviewed by e.g. Ito (1950), Gorbach & Koch (1955), Koch & Dedic (1957), Hagihara (1960), Davies (1963) and Roper & Fennell (1965).In connection with investigation of the proteolytic activity of several fungus species, a gelatin hydrolyzing effect of Aspergillus fumigatus (AF) was reported to be exerted by living organisms in pure culture (Jensen 1931), extracted mycelial material (Maxwell 1950, Dingle & Solomons 1952), and by fluid culture medium in which AF was cultured (Dion 1950a, b, Dingle & Solomons). Ay res & Tobie (1943) demonstrated moderate casein hydrolyzing activity in extracted mycelial material from 4 AF strains, and Amatayakul (1955) observed low fibrinolytic activity in 1 strain.Proteolytic activity measured by breakdown of gelatin and casein was demonstrated by Jonsson & Martin (1964) in culture medium in which AF had been cultured. Three activity optima were observed at pH values around 3, 6.5 and 10. A subsequent study indicated that the activity in neutral and alkaline environment was caused by the same enzyme (Martin & Jönsson 1965).By means of electrophoretic separation combined with reactions for enzyme characterization, Tran Van Ky et al. (1966) demonstrated proteolytic activity in mycelial extracts of 21-day AF cultures. A casein precipitating enzyme (CP enzyme) was demonstrated by Sandvik (1967) in the fluid phase of frozen and thawed skimmilk agar cultures from e.g. AF.In addition to haemolysin and toxin (Rutqvist 1965, 1968, Rutqvist & Persson 1966), mycelial filtrates of AF have proved to contain a proteolytic enzyme. An account is given in the following of a study of this enzyme with respect to (1) casein precipitating ability, (2) casein and gelatin hydrolyzing effect, and (3) relation to toxin and haemolysin.     

Comparative study on the aflatoxin B1 degradation ability of rumen fluid from Holstein steers and Korean native goats

The aflatoxin B1 degrading abilities of two different ruminants were compared in this study. One set of experiments evaluated the aflatoxin B1 degradation ability of different rumen fluid donors (steers vs. goats) as well as the rumen fluid filtration method (cheese cloth filtered vs. 0.45 µm Millipore) in a 2 × 2 factorial arrangement. Additional studies examined aflatoxin B1 degradation by collecting rumen fluid at different times (0, 3, 6, 9 and 12 h) after feeding. Cannulated Holstein steers (740 ± 10 kg bw) and Korean native goats (26 ± 3 kg bw) were fed a 60% timothy and 40% commercial diet with free access to water. Rumen fluid from Korean native goats demonstrated higher (p < 0.01) aflatoxin B1 degradability than Holstein steers. However, filtration method had no significant influence on degradability. In addition, aflatoxin degradation did not depend upon rumen fluid collection time after feeding, as no significant differences were observed. Finally, a comparison of two types of diet high in roughage found aflatoxin degradability in goats was higher with timothy hay opposed to rice straw, although individual variation existed. Thus, our findings showed the aflatoxin degradability is comparatively higher in goats compared to steers.     

Anti-lipopolysaccharide antibody administration mitigates ruminal lipopolysaccharide release and depression of ruminal pH during subacute ruminal acidosis challenge in Holstein bull cattle

The effects of anti-lipopolysaccharide (LPS) antibody on rumen fermentation and LPS activity were investigated during subacute ruminal acidosis (SARA) challenge. Eleven Holstein cattle (164 ± 14 kg) were used in a 3 × 3 Latin square design. Cattle were fed a roughage diet on days −11 to −1 (pre-challenge) and day 2 (post-challenge), and a high-grain diet on days 0 and 1 (SARA challenge). For 14 days, 0-, 2-, or 4-g of anti-LPS antibody was administered once daily through a rumen fistula. Ruminal pH was measured continuously, and rumen fluid and blood samples were collected on days −1, 0, 1, and 2. Significantly lower ruminal LPS activity on day 1 was observed in the 2- and 4-g groups than those in the 0-g group. In addition, significantly higher 1-hr mean ruminal pH on SARA challenge period (days 0 and 1) was identified in the 4-g group than in the 0-g group. However, rumen fermentation measurements (total volatile fatty acid [VFA], VFA components, NH₃-N and lactic acid) and peripheral blood metabolites (glucose, free fatty acid, beta-hydroxybutyrate, total cholesterol, blood urea nitrogen, aspartate aminotransferase and gamma-glutamyl transferase) were not different among the groups during the experimental periods. Therefore, anti-LPS antibody administration mitigates LPS release and pH depression without the depression of rumen fermentation and peripheral blood metabolites during SARA challenge in Holstein cattle.     

The migration test on circulating bovine leukocytes and its possible application in the diagnosis of Johne's disease

The leukocyte migration test for in vitro studies of delayed type hypersensitivity has recently been reviewed (Søborg & Ben-dixen 1967; Bendixen & Søborg 1969). Søborg & Bendixen applied the test to circulating leukocytes in man and thereby widely increased the potentialities of this test. They obtained high leukocyte yields with only moderate erythrocyte admixture by harvesting the supernatant plasma after sedimentation of the erythrocytes for 60 min. at 37°C in the normal gravitational field (1 × g)- Their procedure was unsuitable for the present investigation because bovine erythrocytes sediment so slowly. Sedimentation after clumping at the interphase of aqueous solutions of polymers, dextran and methylcellulose, in combination with metrizoic acid (Böyum 1968) was tried without success because the vast majority of the leukocytes sedimented together with the erythrocytes. Separation of leukocytes from erythrocytes could not be achieved by differential centrifugation.     

Mycoplasmosis: cervical and uterine inoculation of heifers with a Danish strain of Mycoplasma bovigenitalium

The “Κ” strain of Mycoplasma bovigenitalium has proved to be pathogenic for the mammary gland of cows, for the genital tract of bulls, and for calves (Εrnø 1967, Blom & Εrnø 1967, Εrnø 1969). Antibodies can be demonstrated in the blood following infection of the mammary gland, and after intravenous and intraperitoneal inoculation of calves with or without clinical signs of infection.     

Ruminal metabolism and methemoglobin-forming effect of dinobuton in sheep

Many aromatic nitro compounds are reduced to the corresponding amines in the rumen by the activity of the rumen flora, and this may influence the effect of such compounds when given orally to ruminants. The dinitrophenol herbicide DNBP (2-(1-methyl-n-propyl)-4,6-dinitrophenol, dinoseb) is a good example of a compound with this property, as DNBP has a methemoglobin-forming effect in ruminants (Frøslie & Karlog 1970). These authors connected this phenomenon with the ruminai metabolism of DNBP, because it is reduced to 6-ANBP (2-(1-methyl-n-propyl)-4-nitro-6-aminophenol), and 6-ANBP successively is reduced to DABP (2-(1-methyl-n-propyl)-4,6-diaminophenol) in the rumen.     

Comparison of two ruminoscopy techniques in calves

Two different techniques were tested in nine calves to describe endoscopically the visible structures of the rumen and the reticulum. Ruminoscopy using an orally introduced flexible endoscope proved to be unsatisfactory, but when the endoscope was introduced through a ruminal fistula in the left flank into the dorsal and ventral ruminal sacs, the blind sacs, ruminal pillars, the atrium of the rumen, the reticulum and the reticular groove could all be displayed effectively. Indentations of the ruminal wall caused by the left kidney, the spleen and the abomasum could be seen with the endoscope as could reticulo-ruminal contractions. The technique allowed visual evaluation of the mucosal surfaces and movement of the rumen, reticulum and reticular groove. The advantages and disadvantages of ruminoscopy in bovine medicine are discussed.     

Occurrence of gaffkaemia in Lobsters in Norway.

Gaffkaemia is a bacterial disease which causes periodic, lethal epizootics in holding facilities for live American (Homarus americanus) and European (Homarus vulgaris) lobsters (Snieszko & Taylor 1947, Roskam 1957). Gaffkaemia, with the causative agent Aerococcus viridans, has been diagnosed in many countries in Europe and elsewhere. Roskam (1957, 1958) reported the disease in lobsters imported to the Netherlands from Norway. In Norway, however, the disease has never before been reported, though an experiment has shown that gaffkaemia could be induced in lobsters from a Norwegian area with injections of the A.T.C.C. type strain no. 10400 of the causative agent (Møllerud 1971).     

葡萄渣单宁对绵羊养分消化代谢及瘤胃发酵的影响

选用4只体重28 kg左右且安装永久瘤胃瘘管羯羊[萨福克(Suffolk)♂×小尾寒羊(small tail Han-yang)♀],采用4×4拉丁方试验设计,通过消化代谢试验,研究饲料中不同含量葡萄渣单宁对绵羊养分消化代谢及瘤胃发酵的影响。试验共设4个处理,葡萄渣缩合单宁含量分别为0.00,0.85,1.70和2.25 g/kg DM。结果表明,葡萄渣单宁对绵羊干物质、有机物、中性洗涤纤维、酸性洗涤纤维、钙和磷的表观消化率没有产生显著影响(P>0.05),但显著提高绵羊氮表观消化率和存留率(P<0.05);显著降低绵羊瘤胃液总氮和蛋白氮浓度而显著提高尿素氮浓度(P<0.05),对绵羊瘤胃液氨氮浓度没有产生显著的影响(P>0.05);显著提高绵羊瘤胃液丙酸摩尔比而显著降低丁酸摩尔比和乙丙比(P<0.05),对绵羊瘤胃液pH、总挥发性脂肪酸浓度和乙酸及其他酸摩尔比没有产生显著的影响(P>0.05)。此外,试验结果表明饲料中添加2.25 g/kg DM含量的葡萄渣单宁,能够提高绵羊对饲料蛋白质的利用率。     

Mycoplasmosis: experimental inoculation of calves with a Danish strain of Mycoplasma bovigenitalium

During the fall of 1966 several strains of Mycoplasma were isolated from semen samples of bulls. A strain, “K”, isolated from a bull with vesiculitis was selected for a series of experiments and proved to be pathogenic for the mammary gland of cows and for the genital tract of bulls (Ernø 1967, Blom & Εrnø 1967). The “K” strain has now been examined serologically by the author, and it must be considered as belonging to the species M. bovigenitalium as determined by complement fixation and indirect haemagglutination tests. This communication reports the results of experimental inoculation of calves.     

Clinical chemical comparative examination of ruminal samples collected by means of a naso-ruminal sampler

Samples of ruminal fluid collected with the COMET naso-ruminal sampler and by two other methods through fistula were subjected to comparative clinical chemical examination. It was found that the mean values of pH and buffer capacity were nearly identical for all three methods. The greatest variation between the methods were found for reduction time of methylene blue owing to different dry matter contents in the samples. The VFA contents were nearly identical in all three samples.In conclusion, it was found that it is possible to obtain ruminal samples that are representative of the biochemical status in the rumen by means of the COMET naso-ruminal sampler in non-fistulated cows.     

Toxicity of halogenated oxyquinolines to dogs. The effect of feeding fouled herring

Acute clinical toxicity of halogenated oxyquinolines in dogs has been reported (Schantz & Wikström 1965, Hangartner 1965, Müller 1967). The clinical picture was comprehensively studied (Lannek 1972 b). The dose range in the clinical material was found to vary from about 7 to 250 mg per kg body weight (Lannek 1972 a).     

Methaphylactic effect of tulathromycin treatment on rumen fluid parameters in feedlot beef cattle

The aim of this study was to evaluate the effect of tulathromycin as a bovine respiratory disease (BRD) metaphylactic treatment on rumen fluid parameters in feedlot cattle in an intensive livestock production farm. One hundred beef cattle, immediately after housing, were divided in 2 equal groups: 50 animals with metaphylactic treatment against BRD (treated group; tulathromycin at 2.5 mg/kg BW) and 50 animals with placebo treatment (control group). Rumen fluid samples were collected from each animal by rumenocentesis in 3 periods: 1 d (T1), 8 d (T8), and 15 d (T15) after treatment. Rumen pH was determined by ruminal fluid using portable pH meter. Total volatile fatty acids (total VFA) were evaluated by high performance liquid chromatography (HPLC). All animals were singularly weighed at T1 and T15. Two-way analysis of variance (ANOVA) was applied to determine significant effects of treatment (treated group versus control group) and period (T1, T8, and T15) on rumen fluid parameters and body weight. No clinical signs of BRD or other related diseases were recorded during the periods of study from any animal. Statistically significant differences (P < 0.05) were found between treated group and control group for mean values of ruminal pH (6.02 versus 5.89) and total VFA (5.84 versus 5.13) at 8 d after treatment. The weight gain (Δ) showed an average increase of 8.6 kg in treated group (P < 0.05). The trends of ruminal pH and VFA values suggest an effect of tulathromycin as BRD metaphylactic treatment on the modulation of rumen fermentation, particularly 8 d after administration.     

Ruminal metabolism of DNOC and DNBP

The metabolism of DNOG and DNBP in rumen fluid was examined in vitro after elaboration of analytical methods for quantitative determination of dinitrophenols, their 6-amino derivative and diamino-phenols. It was shown that, under anaerobic conditions, the dinitrophenols were rapidly converted into 6-amino derivatives, which successively were reduced to diamino compounds. Reduction did not take place in heat-sterilized rumen specimens, and the metabolism was inhibited by high initial concentrations of the dinitro compounds in the rumen fluid (1—4 × 10⁻³M).In vivo experiments were performed for determination of the concentrations of the dinitrophenols and their metabolites in rumen fluid and blood plasma in a cow. Ruminai metabolism was rapid like in vitro, and the parent compounds were reduced in 10—30 min. The 6-amino derivatives persisted in the rumen fluid for 1 nr., and thereafter only diaminophenols could be detected for another 8—10 hrs. At least 95 % of the absorbed DNOG or DNBP were bound to plasma proteins. 6-amino-4-nitro-o-cresol w^τas found in plasma exclusively in conjugated form, while 6-amino-4-nitro-sec.-butylphenol was free. The diamino compounds were not demonstrated in blood plasma.Severe methaemoglobinaemia and haemolysis were seen after administration of DNBP. The difference in toxicity for ruminants between the two dinitrophenols is discussed on the basis of the observations reported pointing at the unconjugated 6-amino-4-nitro-sec-butylphenol as a potent inducer of methaemoglobinaemia.     

Focal Symmetrical Poliomyelomalacia in a Calf

Focal symmetrical poliomyelomalacia (FSP) is a neurological disorder mainly described in pigs in connection with experimental or spontaneous cases of selenium toxicosis (Harrison etal. 1983, Wendtetal. 1992, Wilson etal. 1983 & 1989). However, a few cases of FSP have been reported in other domestic animals including sheep, goat, and cattle (Innes & Plow-right 1955, Cordy et al. 1984, Bonniwell & Barlow 1985, Palmer et al. 1986). Common for reports on FSP in other species than pigs is an unsolved aetiology. In cattle only 1 report on FSP has been published previously describing 2 cases (Palmer et al. 1986). The present report describes a further case of FSP in cattle. A 4-month- old Red Danish × American Brown Swiss calf (♀) suddenly became lame on both fore limbs and unwilling to rise. The calf had a swelling around the coronary band and a temperature of 40°C. The calf was treated with antibiotics by a veterinary surgeon. The condition got severe during the next 24 h, and the calf became unable to stand on the fore limbs while the function of the rear limbs was normal. As the condition progressively got worse during the next week, the calf was euthanized by intravenous injection of pentobarbitone sodium and submitted for necropsy.     

滩羊羔羊瘤胃和小肠菌群多样性的比较研究

目的 研究滩羊羔羊瘤胃和小肠菌群多样性的差异。方法 选取健康的断奶滩羊公羔,屠宰后采集瘤胃、十二指肠、空肠和回肠内容物,利用16S rDNA高通量测序技术分析瘤胃和小肠菌群结构及多样性。结果 十二指肠样品Chao1指数高于瘤胃、空肠和回肠,瘤胃样品Shannon指数和Simpson指数均高于其他部位,但差异不显著（P>0.05）。瘤胃液中拟杆菌门（Bacteroidetes）、螺旋菌门（Spirochaetae）、丝状杆菌门（Fibrobacteres）的相对丰度显著（P<0.05）高于十二指肠、空肠和回肠,十二指肠中广古菌门（Euryarchaeota）的相对丰度与瘤胃相比有升高的趋势（0.05<P<0.10）,空肠和回肠中厚壁菌门（Firmicutes）的相对丰度显著（P<0.05）高于十二指肠。瘤胃中的未鉴定菌属（Unidentified）、理研菌属_RC9（Rikenellaceae_RC9_gut_group）、密螺旋体属_2（Treponema_2）、瘤胃球菌属_NK4A214（Ruminococcaceae_NK4A214_group）、普雷沃菌属_UCG-001（Prevotellaceae_UCG-001）相对丰度显著（P<0.05）高于十二指肠、空肠和回肠,空肠中Family_ⅩⅢ_AD3011_group相对丰度显著（P<0.05）高于其他部位。结论 滩羊羔羊瘤胃中细菌的多样性更丰富,瘤胃微生物与十二指肠、空肠和回肠的细菌区系显著不同。     

Ultrasonography of the rumen in 30 Saanen goats

This study describes the results of ultrasonographic examination of the rumen in 30 healthy Saanen goats. A linear or convex transducer with a variable frequency of 5 to 13 MHz was used to scan standing, non-sedated goats. The location and size of the rumen, the distance between the wall of the rumen and abdominal wall and the appearance and size of the gas, fibre mat and fluid layers of the ruminal contents were assessed. The rumen was seen as a large organ medial to the left abdominal wall. The wall of the rumen appeared as a thick echogenic line. The longitudinal groove was seen as an echogenic notch, which divided the rumen into the dorsal and ventral sacs. The rumen could be visualized from the 9th to 12th intercostal space (ICS) and flank on the left side in all the goats. The rumen was largest in the 12th ICS at 41.6 ± 5.13 cm and smallest in the 8th ICS at 11.3 ± 4.29 cm. The dorsal sac of the rumen was largest in the left cranial flank (17.4 ± 4.43 cm) and the ventral sac was largest in the 12th ICS on the left (29.1 ± 6.03 cm). In the cranial left flank, the rumen was situated immediately adjacent to the abdominal wall in all the goats. The spleen was located between the rumen and abdominal wall in the 8th to 12 th ICS in many of the goats. The gas, fibre mat and fluid layers of the ruminal contents could be visualized in all the goats. The gas layer was 9.9 ± 3.05 cm, the fibre mat layer 16.0 ± 4.55 cm and the fluid layer 12.2 ± 5.57 cm.     

Effect of Saccharomyces cerevisiae on alfalfa nutrient degradation characteristics and rumen microbial populations of steers fed diets with different concentrate-to-forage ratios

Live yeast (Saccharomyces cerevisiae) constitutes an effective additive for animal production; its probiotic effect may be related to the concentrate-to-forage ratio (CTFR). The objective of this study was to assess the effects of S. cerevisiae (SC) on fiber degradation and rumen microbial populations in steers fed diets with different levels of dietary concentrate. Ten Simmental × Local crossbred steers (450 ± 50 kg BW) were assigned to a control group or an SC group. Both groups were fed the same basal diet but the SC group received SC supplementation (8 × 10⁹ cfu/h/d through the ruminal fistula) following a two-period crossover design. Each period consisted of four phases, each of which lasted 17 d: 10 d for dietary adaptation, 6 d for degradation study, and 1 d for rumen sample collection. From the 1^st to the 4^th phase, steers were fed in a stepwise fashion with increasing CTFRs, i.e., 30:70, 50:50, 70:30, and 90:10. The kinetics of dry matter and fiber degradation of alfalfa pellets were evaluated; the rumen microbial populations were detected using real-time PCR. The results revealed no significant (P > 0.05) interactions between dietary CTFR and SC for most parameters. Dietary CTFR had a significant effect (P < 0.01) on degradation characteristics of alfalfa pellets and the copies of rumen microorganism; the increasing concentrate level resulted in linear, quadratic or cubic variation trend for these parameters. SC supplementation significantly (P < 0.05) affected dry matter (DM) and neutral detergent fiber (NDF) degradation rates (c_DM, c_NDF) and NDF effective degradability (ED_NDF). Compared with the control group, there was an increasing trend of rumen fungi and protozoa in SC group (P < 0.1); copies of total bacteria in SC group were significantly higher (P < 0.05). Additionally, percentage of Ruminobacter amylophilus was significantly lower (P < 0.05) but percentage of Selenomonas ruminantium was significantly higher (P < 0.05) in the SC group. In a word, dietary CTFR had a significant effect on degradation characteristics of forage and rumen microbial population. S. cerevisiae had positive effects on DM and NDF degradation rate or effective degradability of forage; S. cerevisiae increased rumen total bacteria, fungi, protozoa, and lactate-utilizing bacteria but reduced starch-degrading and lactate-producing bacteria.