Gnotobiotic Silage

Selected strains of lactic acid bacteria isolated from grass silage were found to flourish when inoculated into irradiation-sterilized forage under gnotobiotic conditions. The acid content and pH of these silages resembled naturally fermented silage. Inoculation of gnotobiotic silage with Clostridium sporogenes and C. tyrobutyricum failed to cause any noticeable deterioration of silage quality.     

Inhibition of Listeria monocytogenes by defined lactic microflora in gnotobiotic silages of lucerne,fescue, ryegrass and maize — Influence of dry matter and temperature

The development of L. monocytogenes was studied in gnotobiotic silages of four plant species (ryegrass, fescue, lucerne, maize) with various dry matter concentrations and incubation temperatures. The microflora of these silages was perfectly defined and included Lactobacillus plantarum, Lactobacillus brevis, Lactobacillus casei, Pediococcus spp., Enterobacter liquefaciens and Clostridium tyrobutyricum. The results show that in silages of good quality not only is there no development of L. monocytogenes, but the cells initially present are eliminated. It is therefore most unlikely that a silage of good quality with a pH lower than 5 plays a contaminating role. The hypothesis according to which silages, especially those of bad quality, contain factors favouring contamination seems to be more plausible.     

Characterization,Identification and Application of Lactic Acid Bacteria Isolated from Forage Paddy Rice Silage

There has been growing interest to develop forage rice as a new feed resource for livestock. This study was to characterize the natural population of lactic acid bacteria (LAB) and select potentially excellent strains for paddy rice silage preparation in China. One hundred and twenty-six strains were isolated and screened from paddy rice silage prepared using a small-scale fermentation system, and ninety-nine of these isolates were considered to be LAB based on their Gram-positive and catalase-negative morphology and the production of most of their metabolic products as lactic acid. These isolates were divided into eight groups (A-H) on the basis of their morphological and biochemical characteristics. The Group A to H strains were identified as Lactobacillus (L.) plantarum subsp. plantarum (species ratio: 8.1%), L. casei (5.1%), Leuconostoc (Ln.) pseudomesenteroides (11.1%), Pediococcus (P.) pentosaceus (24.2%), Enterococcus (E.) mundtii (12.1%), Lactococcus (Lc.) garvieae (15.2%), E. faecium (9.1%) and Lc. lactis subsp. lactis (15.2%) based on sequence analyses of their 16S rRNA and recA genes. P. pentosaceus was the most abundant member of the LAB population in the paddy rice silage. A selected strain, namely L. casei R 465, was found to be able to grow under low pH conditions and to improve the silage quality with low pH and a relatively high content of lactic acid. This study demonstrated that forage paddy rice silage contains abundant LAB species and its silage can be well preserved by inoculation with LAB, and that strain R 465 can be a potentially excellent inoculant for paddy rice silage.     

Influence of Lactobacillus spp. from an Inoculant and of Weissella and Leuconostoc spp. from Forage Crops on Silage Fermentation

Lactobacillus spp. from an inoculant and Weissella and Leuconostoc spp. from forage crops were characterized, and their influence on silage fermentation was studied. Forty-two lactic acid-producing cocci were obtained from forage crops and grasses. All isolates were gram-positive, catalase-negative cocci that produced gas from glucose, and produced more than 90% of their lactate in the d-isomer form. These isolates were divided into groups A and B by sugar fermentation patterns. Two representative strains from the two groups, FG 5 and FG 13, were assigned to the species Weissella paramesenteroides and Leuconostoc pseudomesenteroides, respectively, on the basis of DNA-DNA relatedness. Strains FG 5, FG 13, and SL 1 (Lactobacillus casei), isolated from a commercial inoculant, were used as additives to alfalfa and Italian ryegrass silage preparations. Lactic acid bacterium counts were higher in all additive-treated silages than in the control silage at an early stage of ensiling. During silage fermentation, inoculation with SL 1 more effectively inhibited the growth of aerobic bacteria and clostridia than inoculation with strain FG 5 or FG 13. SL 1-treated silages stored well. However, the control and FG 5- and FG 13-treated silages had a significantly (P < 0.05) higher pH and butyric acid and ammonia nitrogen contents and significantly (P < 0.05) lower lactate content than SL 1-treated silage. Compared with the control silage, SL 1 treatments reduced the proportion of d-(−)-lactic acid, gas production, and dry matter loss in two kinds of silage, but the FG 5 and FG 13 treatments gave similar values in alfalfa silages and higher values (P < 0.05) in Italian ryegrass silage. The results confirmed that heterofermentative strains of W. paramesenteroides FG 5 and L. pseudomesenteroides FG 13 did not improve silage quality and may cause some fermentation loss.Silage is now the most common preserved cattle feed in many countries, including Japan. It is well established that lactic acid bacteria (LAB) play an important role in silage fermentation. Epiphytic microflora, the microorganisms naturally present on forage crops, are responsible for silage fermentation and also influence silage quality (3, 11, 15). Lactobacilli and lactic acid-producing cocci, e.g., leuconostocs, lactococci, streptococci, pediococci, and Weissella species, are major components of the microbial flora in various types of forage crops (3). Stirling and Whittenbury (21) reported that leuconostocs were the most numerous and widely distributed on forages and that lactobacilli occurred mostly on grasses. Cai et al. (3) examined a large number of forage crops and grasses and also found that the predominant LAB were lactic acid-producing cocci and that lactobacilli were the least numerous and mostly homofermentative. Ruser (17) found that although all LAB groups were present in chopped-maize samples, homofermentative lactobacilli and heterofermentative leuconostocs were present in the highest numbers.In order to improve silage quality, many LAB-containing biological additives have been developed and are currently available (13, 20, 25). These inoculants may inhibit the growth of harmful bacteria and enhance lactic acid fermentation during ensiling periods. The epiphytic LAB influence the effectiveness of silage inoculants because the introduced bacteria must compete with these LAB (12). Therefore, the LAB species and their characteristics in the silage environment require further study. However, while an increasing number of studies have reported positive benefits from using some bacterial inoculants as silage additives, relatively few have reported the effect of epiphytic LAB, especially Leuconostoc and Weissella species, on silage fermentation. In the present study, the characterization of Leuconostoc and Weissella species isolated from forage crops and their influence on silage fermentation were examined.     

Assessment of Pediococcus acidilactici as a Potential Silage Inoculant

Eighteen Pediococcus strains were screened for their potential as silage inoculants. Pediococcus acidilactici G24 was found to be the most suitable, exhibiting a short lag phase on both glucose and fructose, a rapid rate of acid production, a high sugar-to-lactate conversion efficiency, no detectable breakdown of proteins or lactic acid, and the ability to grow within a broad range of pH and temperature. When tested in laboratory silos using grass with a water-soluble carbohydrate content of 24 g/kg of aqueous extract, P. acidilactici G24 stimulated the natural Lactobacillus plantarum population and accelerated the rates of lactic acid production and pH decrease. After 6 days of fermentation, the inoculated silage exhibited a 12% decrease in ammonia nitrogen and an 11% increase in crude protein levels compared with uninoculated controls. The use of an L. plantarum inoculant at a rate of 10⁴ bacteria per g of grass in conjunction with P. acidilactici G24 produced no additional beneficial effect. Inoculation of grass with a water-soluble carbohydrate level of 8 g/kg of aqueous extract with P. acidilactici G24 led to no acceleration in the rate of L. plantarum growth or pH decrease. However, after 7 days of fermentation the inoculated silage had a 14% lower ammonia nitrogen protein content than did uninoculated controls. The results suggest that P. acidilactici G24 may be useful as a silage inoculant for crops with a sufficiently high water-soluble carbohydrate level.     

Biological ensiling of sugarcane tops

The following lactic acid bacteria were isolated from sugarcane tops silage:Lactobacillus plantarum, Lactobacillus buchneri, Lactobacillus brevis, Lactobacillus delbrueckii, Pediococcus cerevisiae, Leuconostoc mesenteroides andStreptococcus lactis. The isolates were grown in a synthetic medium and the final pH, sugar uptake and the effect of adding CaCO₃ and L1, L2 and L3 factors was observed. The importance of the buffering capacity of the ensiled material for the silage process with a view to the occurrence of lactic acid bacteria is discussed.     

Metabolite Profiles of Lactic Acid Bacteria in Grass Silage

The metabolite production of lactic acid bacteria (LAB) on silage was investigated. The aim was to compare the production of antifungal metabolites in silage with the production in liquid cultures previously studied in our laboratory. The following metabolites were found to be present at elevated concentrations in silos inoculated with LAB strains: 3-hydroxydecanoic acid, 2-hydroxy-4-methylpentanoic acid, benzoic acid, catechol, hydrocinnamic acid, salicylic acid, 3-phenyllactic acid, 4-hydroxybenzoic acid, (trans, trans)-3,4-dihydroxycyclohexane-1-carboxylic acid, p-hydrocoumaric acid, vanillic acid, azelaic acid, hydroferulic acid, p-coumaric acid, hydrocaffeic acid, ferulic acid, and caffeic acid. Among these metabolites, the antifungal compounds 3-phenyllactic acid and 3-hydroxydecanoic acid were previously isolated in our laboratory from liquid cultures of the same LAB strains by bioassay-guided fractionation. It was concluded that other metabolites, e.g., p-hydrocoumaric acid, hydroferulic acid, and p-coumaric acid, were released from the grass by the added LAB strains. The antifungal activities of the identified metabolites in 100 mM lactic acid were investigated. The MICs against Pichia anomala, Penicillium roqueforti, and Aspergillus fumigatus were determined, and 3-hydroxydecanoic acid showed the lowest MIC (0.1 mg ml⁻¹ for two of the three test organisms).     

Prevention of Aerobic Spoilage of Maize Silage by a Genetically Modified Killer Yeast, Kluyveromyces lactis, Defective in the Ability To Grow on Lactic Acid

In this study, we propose a new process of adding a genetically modified killer yeast to improve the aerobic stability of silage. Previously constructed Kluyveromyces lactis killer strain PCK27, defective in growth on lactic acid due to disruption of the gene coding for phosphoenolpyruvate carboxykinase, a key enzyme for gluconeogenesis, inhibited the growth of Pichia anomala inoculated as an aerobic spoilage yeast and prevented a rise in pH in a model of silage fermentation. This suppressive effect of PCK27 was not only due to growth competition but also due to the killer protein produced. From these results, we concluded that strain PCK27 can be used as an additive to prolong the aerobic stability of maize silage. In the laboratory-scale experiment of maize silage, the addition of a killer yeast changed the yeast flora and significantly reduced aerobic spoilage.     

Dynamic variation of the microbial community structure during the long-time mono-fermentation of maize and sugar beet silage

This study investigated the development of the microbial community during a long-term (337 days) anaerobic digestion of maize and sugar beet silage, two feedstocks that significantly differ in their chemical composition. For the characterization of the microbial dynamics, the community profiling method terminal restriction fragment length polymorphism (TRFLP) in combination with a cloning-sequencing approach was applied.Our results revealed a specific adaptation of the microbial community to the supplied feedstocks. Based on the high amount of complex compounds, the anaerobic conversion rate of maize silage was slightly lower compared with the sugar beet silage. It was demonstrated that members from the phylum Bacteroidetes are mainly involved in the degradation of low molecular weight substances such as sugar, ethanol and acetate, the main compounds of the sugar beet silage. It was further shown that species of the genus Methanosaeta are highly sensitive against sudden stress situations such as a strong decrease in the ammonium nitrogen (NH₄⁺-N) concentration or a drop of the pH value. In both cases, a functional compensation by members of the genera Methanoculleus and/or Methanosarcina was detected. However, the overall biomass conversion of both feedstocks proceeded efficiently as a steady state between acid production and consumption was recorded, which further resulted in an equal biogas yield.     

Potential To Reduce Escherichia coli Shedding in Cattle Feces by Using Sainfoin (Onobrychis viciifolia) Forage, Tested In Vitro and In Vivo

There is a growing concern about the presence of pathogens in cattle manure and its implications on human and environmental health. The phytochemical-rich forage sainfoin (Onobrychis viciifolia) and purified phenolics (trans-cinnamic acid, p-coumaric acid, and ferulic acid) were evaluated for their ability to reduce the viability of pathogenic Escherichia coli strains, including E. coli O157:H7. MICs were determined using purified phenolics and acetone extracts of sainfoin and alfalfa (Medicago sativa), a non-tannin-containing legume. Ground sainfoin or pure phenolics were mixed with fresh cattle feces and inoculated with a ciprofloxacin-resistant strain of E. coli, O157:H7, to assess its viability at −20°C, 5°C, or 37°C over 14 days. Forty steers were fed either a sainfoin (hay or silage) or alfalfa (hay or silage) diet over a 9-week period. In the in vitro study, the MICs for coumaric (1.2 mg/ml) and cinnamic (1.4 mg/ml) acids were 10- to 20-fold lower than the MICs for sainfoin and alfalfa extracts. In the inoculated feces, the −20°C treatment had death rates which were at least twice as high as those of the 5°C treatment, irrespective of the additive used. Sainfoin was less effective than coumaric acid in reducing E. coli O157:H7 Cip^r in the inoculated feces. During the animal trial, fecal E. coli numbers declined marginally in the presence of sainfoin (silage and hay) and alfalfa silage but not in the presence of hay, indicating the presence of other phenolics in alfalfa. In conclusion, phenolic-containing forages can be used as a means of minimally reducing E. coli shedding in cattle without affecting animal production.     

Natural populations of lactic acid bacteria associated with silage fermentation as determined by phenotype, 16S ribosomal RNA and recA gene analysis

One hundred and fifty-six strains isolated from corn (Zea mays L.), forage paddy rice (Oryza sativa L.), sorghum (Sorghum bicolor L.) and alfalfa (Medicago sativa L.) silages prepared on dairy farms were screened, of which 110 isolates were considered to be lactic acid bacteria (LAB) according to their Gram-positive and catalase-negative characteristics and, mainly, the lactic acid metabolic products. These isolates were divided into eight groups (A-H) based on the following properties: morphological and biochemical characteristics, γ-aminobutyric acid production capacity, and 16S rRNA gene sequences. They were identified as Weissella cibaria (36.4%), Weissella confusa (9.1%), Leuconostoc citreum (5.3%), Leuconostoc lactis (4.9%), Leuconostoc pseudomesenteroides (8.0%), Lactococcus lactis subsp. lactis (4.5%), Lactobacillus paraplantarum (4.5%) and Lactobacillus plantarum (27.3%). W. cibaria and W. confusa were mainly present in corn silages, and L. plantarum was dominant on sorghum and forage paddy rice silages, while L. pseudomesenteroides, L. plantarum and L. paraplantarum were the dominant species in alfalfa silage. The corn, sorghum and forage paddy rice silages were well preserved with lower pH values and ammonia-N concentrations, but had higher lactic acid content, while the alfalfa silage had relatively poor quality with higher pH values and ammonia-N concentrations, and lower lactic acid content. The present study confirmed the diversity of LAB species inhabiting silages. It showed that the differing natural populations of LAB on these silages might influence fermentation quality. These results will enable future research on the relationship between LAB species and silage fermentation quality, and will enhance the screening of appropriate inoculants aimed at improving such quality.     

Effects of formic acid,molasses and inoculant additives on corn silage composition,organic matter digestibility and microbial protein synthesis in sheep

This study examined the production of com silages with low or high lactic acid concentrations, provided by the addition of formic acid (0.5%), molasses (5%) or microbial inoculant (homofermentative lactic acid bacteria, 10 g/tonne). After the fermentation period, sheep were fed the silages to determine true and apparent digestibility of the organic matter and microbial protein synthesis. The experiment were carried out with four KıvırcıkxMorkaraman sheep, 1.5 years old, fixed with cannula in their rumen and duodenum.Lactic acid concentrations were significantly higher in silages treated with enzyme or molasses compared to other specific treatments. Acetic acid concentration was highest in silage treated with formic acid, and lowest in silage treated with molasses (P < 0.05). The by-pass of crude protein was highest in silage treated with formic acid.     

Effects of Agave salmiana Otto Ex Salm-Dyck silage as forage on ruminal fermentation and growth in goats

Ensiling of Agave salmiana Otto Ex Salm-Dyck, a widespread plant in Mexico, as a viable preservation method to create a potential animal feed resource for ruminants was investigated. Fresh A. salmiana with 205 g dry matter (DM)/kg and wilted alfalfa with 602 g DM/kg were ensiled in combinations (DM:DM) of 1000:0, 500:500 and 350:650, to evaluate feeding value of agave:alfalfa silages on ruminal fermentation and growth of goats. Chemical composition and in situ ruminal disappearance of three total mixed rations (TMRs), which included 240 g/kg DM of each silage (1000:0, 500:500 and 350:650) were determined. The TMR were used to assess ruminal fermentation and growth of 15 goats (20 ± 2.2 kg body weight (BW)). Silage pH (≤4), lactate (>25 g/kg DM) and ammonia (<50 g/kg total N) concentrations indicate that silage quality was good. Lactic acid was the main acid in all silages, acetic acid concentrations were relatively low, and butyrate was only detected in only the 1000:0 agave:alfalfa silage. Potential DM disappearance of the TMR increased quadratically as the amount of alfalfa included in the silage mixture increased. The BW gain and feed efficiency were not changed by treatment, even though DM intake decreased and aNDF intake increased linearly as the amount of alfalfa included in the silage mixture increased. Ruminal pH and butyrate increased, and ammonia N, lactate and propionate decreased linearly as alfalfa proportion of alfalfa in the silage mixture was increased. The TMR ingredient selectivity by the goats may have limited goat performance when alfalfa was included in agave silage mixtures. Because the agave:alfalfa blend improved nutritional quality, ruminal digestibility and intake of agave silage, alfalfa inclusion may improve nutritional characteristics of agave plants silages for ruminants.     

Replacement of grass and maize silages with lucerne silage: effects on performance,milk fatty acid profile and digestibility in Holstein-Friesian dairy cows

In total, 20 multiparous Holstein-Friesian dairy cows received one of four diets in each of four periods of 28-day duration in a Latin square design to test the hypothesis that the inclusion of lucerne in the ration of high-yielding dairy cows would improve animal performance and milk fatty acid (FA) composition. All dietary treatments contained 0.55 : 0.45 forage to concentrates (dry matter (DM) basis), and within the forage component the proportion of lucerne (Medicago sativa), grass (Lolium perenne) and maize silage (Zea mays) was varied (DM basis): control (C)=0.4 : 0.6 grass : maize silage; L20=0.2 : 0.2 : 0.6 lucerne : grass : maize silage; L40=0.4 : 0.6 lucerne : maize silage; and L60=0.6 : 0.4 lucerne : maize silage. Diets were formulated to contain a similar CP and metabolisable protein content, with the reduction of soya bean meal and feed grade urea with increasing content of lucerne. Intake averaged 24.3 kg DM/day and was lowest in cows when fed L60 (P<0.01), but there was no effect of treatment on milk yield, milk fat or protein content, or live weight change, which averaged 40.9 kg/day, 41.0, 30.9 g/kg and 0.16 kg/day, respectively. Milk fat content of 18:2 c9 c12 and 18:3 c9 c12 c15 was increased (P<0.05) with increasing proportion of lucerne in the ration. Milk fat content of total polyunsaturated fatty acids was increased by 0.26 g/100 g in L60 compared with C. Plasma urea and β-hydroxybutyrate concentrations averaged 3.54 and 0.52 mmol/l, respectively, and were highest (P<0.001) in cows when fed L60 and lowest in C, but plasma glucose and total protein was not affected (P>0.05) by dietary treatment. Digestibility of DM, organic matter, CP and fibre decreased (P<0.01) with increasing content of lucerne in the diet, although fibre digestibility was similar in L40 and L60. It is concluded that first cut grass silage can be replaced with first cut lucerne silage without any detrimental effect on performance and an improvement in the milk FA profile, although intake and digestibility was lowest and plasma urea concentrations highest in cows when fed the highest level of inclusion of lucerne.     

Effect of inoculation of herbage prior to ensiling with Streptomyces achromogenes ISP 5028 on chemical composition of silage

Enzymes derived from bacteria and fungi have been used as additives in silage production. Increasing attention has been given to the use of cell-wall degrading enzymes such as cellulases, hemicellulases and peroxidases. However, little information has been published on the potential role of actinomycetes in silage production. The experiment reported in this paper examined the changes in chemical composition and predicted nutritive value of herbage inoculated with Streptomyces achromogenes ISP 5028 during 60 days of ensiling. Changes in chemical composition and nutritive value of herbage were examined after 1, 2, 3, 5, 10, 30 and 60 days ensiling. Colonisation of inoculated silage by S. achromogenes ISP 5028 occurred rapidly and after 30 days of ensiling the difference in viable spores was 1000-fold between the control and treated silages (P<0.001). Inoculation reduced the concentration of NDF (P<0.01) and crude hemicellulose (P<0.001) and accelerated the rate of loss of NDF (P<0.05) but not crude hemicellulose (P>0.05) from the cell wall. The changes in the composition of the cell wall facilitated an increase in the concentration of WSC (P<0.001), thus allowing greater concentrations of lactic (P<0.001) and acetic (P<0.01) acids to be formed. Elevated concentrations of various isomeric forms of butyric and valeric acid were also observed in the treated silage compared to the control. Inoculation of herbage with S. achromogenes ISP 5028 prior to ensiling altered the chemical and nutritive composition of silage, however it is not known whether the risks associated with other actinomycetes (respiratory disorders and toxin release) apply to the test organism in these experiments.     

Antagonistic activity of lactobacilli isolated from natural ecotopes

Lactobacilli are widely used in silage production, for fermentation of foodstuffs, and as probiotics. Their therapeutic effect in preparations is based on their antagonistic activity against pathogens. In this work, antagonistic activity of Lactobacillus strains isolated from silage and fermented plant-derived foodstuffs was studied in order to select the strains promising for industry, agriculture, and medicine. Twenty Lactobacillus strains were ranked according to the intensity and rate of acid production and antibiotic resistance. Lactobacillus sp. Cа9L was selected as a promising starter culture strain for biotechnology based on the optimal combination of acid production, rate of acidification, and antibiotic resistance.     

Evaluation of the factors affecting silage intake of dairy cows: a revision of the relative silage dry-matter intake index

An evaluation of the factors affecting silage dry-matter intake (SDMI) of dairy cows was conducted based on dietary treatment means. The data were divided into six subsets based on the silage treatments used in the experiments: concentration of digestible organic matter in dry matter (D-value) influenced by the maturity of grass ensiled (n = 81), fermentation quality influenced by silage additives (n = 240), dry matter (DM) concentration influenced by wilting of grass prior to ensiling (W; n = 85), comparison of silages made from primary growth or regrowth of grass (n = 46), and replacement of grass silage with legume (L; n = 53) or fermented whole-crop cereal (WC; n = 37) silages. The data were subjected to the mixed model regression analysis. Both silage D-value and fermentation quality significantly affected SDMI. The average effects of D-value and total acid (TA) concentration were 17.0 g and − 12.8 per 1 g/kg DM, respectively. At a given D-value, silage neutral-detergent fibre (NDF) concentration tended to decrease SDMI. Silage TA concentration was the best fermentation parameter predicting SDMI. Adding other parameters into the multivariate models did not improve the fit and the slopes of the other parameters remained insignificant. Total NDF intake was curvilinearly related to silage D-value the maximum intake being reached at a D-value of 640 g/kg DM. Results imply that physical fill is not limiting SDMI of highly digestible grass silages and that both physical and metabolic factors constrain total DM intake in an interactive manner. Silage DM concentration had an independent curvilinear effect on SDMI. Replacing primary growth silage with regrowth, L or WC silages affected SDMI significantly, the response to regrowth silage being linearly decreasing and to L and WC quadratically increasing. The outcome of factors affecting SDMI was used to update the relative SDMI index as follows: SDMI index = 100+10 × [(D-value − 680) × 0.0170 − (TA − 80) × 0.0128+(0.0198 ×  (DM − 250) − 0.00002364 × (DM² − 250 ²)) − 0.44 × a+4.13 × b − 2.58 × b²+5.90 × c − 6.14 × c² − 0.0023 × (550 − NDF)], where a, b and c represent the proportions (0–1) of regrowth, L or WC silages from total silage DM. For the whole data set, one index unit corresponded to the default value of 0.10 kg in SDMI. The SDMI index explained proportionally 0.852 of the variation in SDMI with 0.34 kg DM per day residual. The updated SDMI index provides improved basis for the practical dairy cow ration formulation and economic evaluation.     

Beet pulp substituted for corn silage and barley grain in diets fed to dairy cows in the summer months: feed intake,total-tract digestibility,and milk production

The responses of dairy cows to the substitution of beet pulp (BP) for grain or forage are not consistent, and heat stress may affect the response of dairy cows to this substitution. The effects of substituted BP for corn silage and barley grain on feed intake, performance, and ruminal parameters were evaluated using eight multiparous Holstein cows in a duplicated 4 × 4 Latin square design with 21-day periods. Cows were in mid-lactation (45.4 ± 3.6 kg/day milk production and 116 ± 10 days in milk) with an average BW of 664 ± 41.2 kg. Dietary treatments were as follows: 1) 0% BP (0BP, control, 38.5% barley grain, and 20.3% corn silage); 2) 12% BP (12BP, 32.5% barley grain, and 14.3% corn silage); 3) 18% BP (18BP, 29.5% barley grain, and 11.3% corn silage); and 4) 24% BP (24BP, 26.5% barley grain, and 8.3% corn silage). Cows were under mild heat stress and the average temperature–humidity index was 70.5; increasing BP caused a linear decrease in respiration rate (P < 0.01). Higher BP in the diet caused a linear increase in DM intake (P = 0.01) and NDF digestibility (P = 0.03). Dry and organic matter (OM) digestibilities tended to increase linearly with higher BP (P < 0.10). Milk yield, energy-corrected milk, protein, lactose, and fat production and content were not affected by the treatments. Increasing BP in the diet caused a linear decrease in feed efficiency and rumen ammonia (P < 0.05) and a tendency to a linear decrease in milk urea nitrogen (P < 0.10). Rumen pH and acetate to propionate ratio were not affected by the replacement. Total volatile fatty acid concentration in the rumen increased linearly with increasing the BP inclusion (P = 0.04). Acetate and butyrate (P = 0.07) proportion tended to increase, whereas propionate (P = 0.06) and isovalerate (P = 0.08) proportion tended to decrease linearly as BP was substituted for corn silage and barley grain. The results indicated that under mild heat stress condition, BP can be successfully substituted for barley grain and corn silage up to 24% of the diet without any negative effect on production and ruminal pH.     

Effects of Common Forage Phenolic Acids on Escherichia coli O157:H7 Viability in Bovine Feces

Ruminant animals are carriers of Escherichia coli O157:H7, and the transmission of E. coli O157:H7 from cattle to the environment and to humans is a concern. It is unclear if diet can influence the survivability of E. coli O157:H7 in the gastrointestinal system or in feces in the environment. Feces from cattle fed bromegrass hay or corn silage diets were inoculated with E. coli O157:H7, and the survival of this pathogen was analyzed. When animals consumed bromegrass hay for <1 month, viable E. coli O157:H7 was not recovered after 28 days postinoculation, but when animals consumed the diet for >1 month, E. coli O157:H7 cells were recovered for >120 days. Viable E. coli O157:H7 cells in feces from animals fed corn silage were detected until day 45 and differed little with the time on the diet. To determine if forage phenolic acids affected the viability of E. coli O157:H7, feces from animals fed corn silage or cracked corn were amended with common forage phenolic acids. When 0.5% trans-cinnamic acid or 0.5% para-coumaric acid was added to feces from silage-fed animals, the E. coli O157:H7 death rate was increased significantly (17-fold and 23-fold, respectively) compared to that with no addition. In feces from animals fed cracked corn, E. coli O157:H7 death rates were increased significantly with the addition of 0.1% and 0.5% trans-cinnamic acid (7- and 13-fold), 0.1% and 0.5% p-coumaric acid (3- and 8-fold), and 0.5% ferulic acid (3-fold). These data suggest that phenolic acids common to forage plants can decrease viable counts of E. coli O157:H7 shed in feces.     

An integrated system for the production of gnotobiotic Anopheles quadrimaculatus

This paper reports the first successful gnotobiotic rearing of the mosquito Anopheles quadrimaculatus, a major vector of Dirofilaria immitis, the etiologic agent of canine heartworm disease in the United States. Mosquito eggs were sterilized in Zephiran, and emerging larvae were transferred to flat-sided rearing flasks which contained a sterile easily prepared formulation of commercial dietary products. Maturing pupae were withdrawn in a syringe and were injected with a small amount of water into the gnotobiotic arthropod module (GAM), a sterile Lucite capsule which allows sugar feeding at one end and blood feeding through a heated artificial membrane at the other end. The GAM provided a convenient sterilizable unit to support 12–15 gnotobiotic mosquitoes for a variety of experimental purposes.