Forages and pastures symposium: forage biodegradation: advances in ruminal microbial ecology.

The rumen microbial ecosystem provides ruminants a selective advantage, the ability to utilize forages, allowing them to flourish worldwide in various environments. For many years, our understanding of the ruminal microbial ecosystem was limited to understanding the microbes (usually only laboratory-amenable bacteria) grown in pure culture, meaning that much of our understanding of ruminal function remained a "black box." However, the ruminal degradation of plant cell walls is performed by a consortium of bacteria, archaea, protozoa, and fungi that produces a wide variety of carbohydrate-active enzymes (CAZymes) that are responsible for the catabolism of cellulose, hemicellulose, and pectin. The past 15 years have seen the development and implementation of numerous next-generation sequencing (NGS) approaches (e.g., pyrosequencing, Illumina, and shotgun sequencing), which have contributed significantly to a greater level of insight regarding the microbial ecology of ruminants fed a variety of forages. There has also been an increase in the utilization of liquid chromatography and mass spectrometry that revolutionized transcriptomic approaches, and further improvements in the measurement of fermentation intermediates and end products have advanced with metabolomics. These advanced NGS techniques along with other analytic approaches, such as metaproteomics, have been utilized to elucidate the specific role of microbial CAZymes in forage degradation. Other methods have provided new insights into dynamic changes in the ruminal microbial population fed different diets and how these changes impact the assortment of products presented to the host animal. As more omics-based data has accumulated on forage-fed ruminants, the sequence of events that occur during fiber colonization by the microbial consortium has become more apparent, with fungal populations and fibrolytic bacterial populations working in conjunction, as well as expanding understanding of the individual microbial contributions to degradation of plant cell walls and polysaccharide components. In the future, the ability to predict microbial population and enzymatic activity and end products will be able to support the development of dynamic predictive models of rumen forage degradation and fermentation. Consequently, it is imperative to understand the rumen's microbial population better to improve fiber degradation in ruminants and, thus, stimulate more sustainable production systems.

Forage degradation in the rumen is critical to producing ruminant animals. For many years, scientists were limited to biochemical techniques to understand how ruminal microbes degraded forage, impairing our understanding of which microbes were involved with degrading which forage components. However, we have understood that as the ruminant opened up plant cells to microbial activity, a succession of microbes was involved in colonizing and breaking fiber into increasingly smaller pieces. The recent development of sequencing techniques has allowed a more detailed understanding of changes in the microbial population of the rumen during forage degradation and the types of degradative enzymes produced by this complex microbial ecosystem. We described the enzymes involved in the degradation of specific forage components, how their end products impact the microbial population through cross-feeding interactions, and how fermentation products can impact food animal production.

Voluntary intake and digestibility by sheep of alfalfa ensiled at different moisture concentrations following fertilization with dairy slurry.

Dairy slurry is used commonly as an animal-sourced fertilizer in agronomic production. However, residual effects of slurry application on intake and digestibility of alfalfa (Medicago sativa L.) silage from subsequent harvests are not well known. The objective of this study was to determine if moisture concentration of alfalfa silage and timing of dairy slurry application relative to subsequent harvest affected intake and digestibility by sheep. Katahdin crossbred ewes (n = 18; 48 ± 5.3 kg) in mid-gestation were stratified by BW and allocated randomly in each of two periods to one of six treatments arranged in a two × three factorial arrangement. Treatments consisted of recommended (RM; 46.8%) or low (LM; 39.7%) moisture at baling after either no slurry application (NS), slurry application to stubble immediately after removal of the previous cutting (S0), or slurry application 14 d after removal of the previous cutting (S14). Silages were chopped through a commercial straw chopper, packed into plastic trash cans, and then offered to ewes within 4 d of chopping. Period 1 of the intake and digestion study consisted of a 14-d adaptation followed by a 7-d fecal collection period. Period 2 followed period 1 after a 4-d rest and consisted of an 11-d adaptation followed by 7 d of fecal collection. Ewes were housed individually in 1.4 × 4.3-m pens equipped with rubber mat flooring. Feces were swept from the floor twice daily, weighed, and dried at 50 °C. Ewes had ad libitum access to water and were offered chopped silage for a minimum of 10% refusal (DM). Blood samples were collected immediately prior to feeding, and 4 and 8 h after feeding on the day prior to the end of each period. Organic matter intake (g/kg BW) and OM digestibility tended (P < 0.10) to be, and digestible OM intake (g/kg BW) was reduced by slurry application. Lymphocytes (% of total white blood cells) were greater (P < 0.05) from LM vs. RM and from NS vs. S0 and S14. Red blood cell concentrations were greater (P < 0.05) from S14 vs. S0 and from S0 and S14 vs. NS. Serum urea N concentrations did not differ (P > 0.17) across treatments. Therefore, moisture concentration of alfalfa silage within the range used in this study may not affect voluntary intake or digestibility, but slurry application may have an effect on digestible OM intake. Also, moisture concentration of alfalfa silage and time of dairy slurry application may affect specific blood hemograms.

Efficacy and safety of 6 or 8 weeks of simeprevir, daclatasvir, sofosbuvir for HCV genotype 1 infection.

The phase 2, open-label ACCORDION (ClinicalTrials.gov: NCT02349048) study investigated the efficacy, safety and pharmacokinetics of a 6- or 8-week regimen of simeprevir, daclatasvir and sofosbuvir in treatment-naïve patients with chronic hepatitis C virus (HCV) genotype (GT) 1 infection and either early-stage fibrosis or compensated cirrhosis. Patients were assigned to treatment groups according to their fibrosis stage. Early-stage fibrosis: simeprevir 150 mg, daclatasvir 60 mg, sofosbuvir 400 mg once daily for 6 weeks; compensated cirrhosis: same regimen for 8 weeks. The primary endpoint was sustained virologic response 12 weeks after the end of treatment (SVR12). Safety, tolerability and pharmacokinetics of simeprevir, daclatasvir and sofosbuvir were investigated. Sixty-eight patients were treated (6-week group: n = 59; 8-week group: n = 9). SVR12 was achieved by 86.4% (51/59) of patients with early-stage fibrosis and by 100% (9/9) of patients with cirrhosis. The main reason for not achieving SVR12 in the 6-week group was viral relapse (11.9%; 7/59). One patient had on-treatment failure due to an early withdrawal (lost to follow-up due to incarceration). One patient with SVR12 in the 6-week group had a late viral relapse at post-treatment week 24. No clinically significant drug-drug interactions were observed. Adverse events were reported in 63.2% of patients (43/68) and were mainly grade 1/2. None of these led to treatment discontinuation. The 3 direct-acting antiviral regimens of simeprevir, daclatasvir and sofosbuvir were safe and well tolerated in treatment-naïve, HCV GT1-infected patients with early-stage fibrosis or compensated cirrhosis.

Effects of rotating antibiotic and ionophore feed additives on volatile fatty acid production, potential for methane production, and microbial populations of steers consuming a moderate-forage diet.

Ionophores and antibiotics have been shown to decrease ruminal methanogenesis both in vitro and in vivo but have shown little evidence toward a sustainable means of mitigation. Feed additive rotation was proposed and investigated for methane, VFA, and microbial population response. In the present study, cannulated steers ( = 12) were fed a moderate-forage basal diet in a Calan gate facility for 13 wk. In addition to the basal diet, steers were randomly assigned to 1 of 6 treatments: 1) control, no additive; 2) bambermycin, 20 mg bambermycin/d; 3) monensin, 200 mg monensin/d; 4) the basal diet + weekly rotation of bambermycin and monensin treatments (B7M); 5) the basal diet + rotation of bambermycin and monensin treatments every 14 d (B14M); and 6) the basal diet + rotation of bambermycin and monensin treatments every 21 d (B21M). Steers were blocked by weight in a randomized complete block design where the week was the repeated measure. Rumen fluid was collected weekly for analysis ( = 13), and results were normalized according to individual OM intake (OMI; kg/d). Potential activity of methane production was not significantly different among treatments ( > 0.05). However, treatment tended to affect the CH-to-propionate ratio ( = 0.0565), which was highest in the control and lowest in the monensin, B21M, and B14M treatments (0.42 vs. 0.36, 0.36, and 0.33, respectively). The CH:propionate ratio was lowest in wk 2 and 3 ( < 0.05) but the ratio in wk 4 to 12 was not different from the ratio in wk 0. Week also affected total VFA, with total VFA peaking at wk 3 and plummeting at wk 4 (4.02 vs. 2.86 m/kg OMI; < 0.05). A significant treatment × week interaction was observed for the acetate-to-propionate (A:P) ratio, where bambermycin- and rotationally fed steers did not have a reduced A:P ratio compared with monensin-fed steers throughout the feeding period ( < 0.0001). Microbial analysis revealed significant shifts, but several predominant classes showed adaptation between 4 and 6 wk after additive initiation. There was no significant evidence to suggest that rotations of monensin and bambermycin provided additional benefits to steers consuming a moderate-forage diet at the microbial/animal and environmental level versus those continuously fed.

Feeding distillers' grains, soybean hulls, or a mixture of both to cows as a forage replacement: Effects on intake, digestibility, and ruminal fermentation characteristics.

Coproduct feedstuffs offer a unique and potentially profitable avenue for cattle feeding strategies. However, research is lacking in the evaluation of varying coproducts on ruminal fermentation and digestive characteristics when included as the major component of the diet of cows. Our objective was to determine the effect of coproduct feedstuffs as a forage replacement on digestive and fermentative characteristics of cows. Eight ruminally fistulated cows (672 ± 32.0 kg initial BW and approximately 9 yr of age) were stratified by BW and randomly allocated to 1 of 4 diets (2 cows∙diet∙period) in a 2-period study: soybean hulls (SH), distillers' dried grains with solubles (DG), an isoenergetic mixture of soybean hulls and distillers' dried grains with solubles (MX), or ad libitum hay plus 0.9 kg/d of an isoenergetic mixture of soybean hulls and distillers' dried grains with solubles (HY). Diets were formulated to meet the ME requirements of a similar, companion study. Coproduct amounts were increased over a 14-d period. This was followed by a 14-d adaptation to diet and facilities and 5 d of total fecal collections. On the final day of fecal collections, rumen fluid was sampled immediately prior to feeding and 2, 4, 6, 8, 10 and 12 h after feeding for measurement of rumen VFA and ammonia concentrations. Intake of DM and OM was not different ( ≥ 0.28) among treatments, but digestibilities of DM, OM, NDF, and ADF were improved ( < 0.05) by coproduct feeding and by MX vs. the mean of SH and DG. Ruminal DM and OM fill were greater ( < 0.05) for cows offered HY than for cows offered the coproduct diets, greater for cows offered SH than for cows offered DG, and for the mean of SH and DG vs. MX. Ruminal retention time was greater ( < 0.05) for HY vs. the coproduct diets and for SH vs. DG. Apparent N absorption tended ( < 0.10) to be greater for cows offered the coproduct diets than for cows offered HY and greater for cows offered DG than for cows offered SH. Total VFA averaged across sampling times were greatest ( < 0.05) for cows offered SH, and ruminal ammonia N was greatest ( < 0.05) for cows offered either DG or MX at all sampling times. Based on these data, coproduct feedstuffs may be fed to meet the energy requirement of cows without negative effects on digestion or ruminal fermentation.

The clinical spectrum of omega-5-gliadin allergy.

BACKGROUND: IgE-mediated allergy to the wheat protein omega-5-gliadin (O5G) is associated with wheat-dependent exercise-induced anaphylaxis (WDEIA), where exercise acts as a cofactor, triggering anaphylaxis after wheat ingestion. The wider application of O5G-specific IgE (sIgE) testing has revealed that the manifestations of O5G allergy extend beyond WDEIA. AIMS: This study documents clinical manifestations in a large series of patients with sIgE to O5G. METHODS: A retrospective clinical audit was performed on adult patients with a positive O5G sIgE (>0.35kU/L) between 2007 and 2013 compared with a group who had negative O5G sIgE. Clinical characteristics and skin prick test (SPT) results were examined. RESULTS: Sixty-seven patients were characterised, 26 of whom presented with food-dependent exercise-induced allergy, whilst others presented with exercise-induced symptoms without apparent food association (16/67), idiopathic anaphylaxis (10/67), food-induced allergic symptoms without exercise (10/67) or recurrent acute urticaria (5/67). Specific IgE to O5G had 91% sensitivity and 92% specificity for wheat-related allergic symptoms. SPT had sensitivity of 92% and specificity of 84%. CONCLUSION: WDEIA is the most common manifestation of O5G allergy, but patients may present with a variety of allergic manifestations, and wheat allergy is not always obvious on history. Non-exercise cofactors or a lack of cofactors were identified in many patients. A distinctive feature of this allergy is that despite regular wheat ingestion, allergic reactions to wheat occur infrequently. Testing for sIgE to O5G should be considered in patients presenting with exercise-induced urticaria/anaphylaxis, idiopathic anaphylaxis and recurrent acute (but not chronic) urticaria.

Icatibant in angiotensin-converting enzyme (ACE) inhibitor-associated angioedema.

BACKGROUND: Angioedema occurs in up to 2% of those taking angiotensin-converting enzyme (ACE) inhibitors. Upper airway angioedema may potentially require endotracheal intubation or cricothyrotomy, and is usually unresponsive to adrenaline. The bradykinin receptor antagonist icatibant is proven to be effective in the treatment of acute attacks of hereditary angioedema, and has also been reported effective in the treatment of angioedema associated with ACE inhibitors. AIM: To describe the use of icatibant for ACE inhibitor-associated airway angioedema. METHODS: We treated 13 consecutive emergency department (ED) patients, who had not improved with adrenaline and/or corticosteroids, with icatibant 30 mg subcutaneously for ACE inhibitor-associated upper respiratory tract angioedema according to an agreed protocol. RESULTS: Four patients were intubated in the ED either before or after receiving icatibant; three of these were extubated within 24 h of treatment. Eight patients received early icatibant and did not require intubation. The time from onset of airway angioedema to ED presentation ranged from 1 h to 3 days (median 4 h); from ED presentation to receiving icatibant, from 30 minutes to 3 days (median 3 h); and to onset of symptom improvement after icatibant, 15 minutes to 7 h (median 2 h). One patient received a second dose of icatibant. CONCLUSION: All patients improved after receiving icatibant, consistent with its bradykinin receptor blocking mechanism. Icatibant rapidly reversed symptoms, and appeared to avert the need for intubation or expedite extubation. Timely use of icatibant in ACE inhibitor-associated angioedema may avert the need for invasive airway procedures and intensive care unit admission.

Repeat treatment of acute hereditary angioedema attacks with open-label icatibant in the FAST-1 trial.

Hereditary angioedema (HAE) is characterized by potentially life-threatening recurrent episodes of oedema. The open-label extension (OLE) phase of the For Angioedema Subcutaneous Treatment (FAST)-1 trial (NCT00097695) evaluated the efficacy and safety of repeated icatibant exposure in adults with multiple HAE attacks. Following completion of the randomized, controlled phase, patients could receive open-label icatibant (30 mg subcutaneously) for subsequent attacks. The primary end-point was time to onset of primary symptom relief, as assessed by visual analogue scale (VAS). Descriptive statistics were reported for cutaneous/abdominal attacks 1-10 treated in the OLE phase and individual laryngeal attacks. Post-hoc analyses were conducted in patients with ≥ 5 attacks across the controlled and OLE phases. Safety was evaluated throughout. During the OLE phase, 72 patients received icatibant for 340 attacks. For cutaneous/abdominal attacks 1-10, the median time to onset of primary symptom relief was 1·0-2·0 h. For laryngeal attacks 1-12, patient-assessed median time to initial symptom improvement was 0·3-1·2 h. Post-hoc analyses showed the time to onset of symptom relief based on composite VAS was consistent across repeated treatments with icatibant. One injection of icatibant was sufficient to treat 88·2% of attacks; rescue medication was required in 5·3% of attacks. No icatibant-related serious adverse events were reported. Icatibant provided consistent efficacy and was well tolerated for repeated treatment of HAE attacks.

PDE5 inhibitors: considerations for preference and long-term adherence.

INTRODUCTION: Erectile dysfunction (ED) is a highly prevalent condition affecting nearly one in five men worldwide. The advent of phosphodiesterase type 5 inhibitors (PDE5i) has revolutionised the ED treatment landscape and provided effective, minimally invasive therapies to restore male sexual function. MATERIALS AND METHODS: A pubmed search was performed of all English language articles from 1996 to present reviewing PDE5i, including pharmacokinetics, efficacy profiles and comparisons, where available. RESULTS: Currently available PDE5i in the United States include sildenafil, vardenafil, tadalafil and avanafil, each of which has unique side effect, pharmacokinetic and outcome profiles. Sildenafil is associated with increased rate of visual changes, vardenafil with QT prolongation and tadalafil with lower back pain. Avanafil and vardenafil orodispersible tablet rapidly achieve peak plasma concentration, which results in faster onset of action, whereas tadalafil exhibits the longest half-life. First time response to PDE5i is approximately 60-70%, with no significant differences in efficacy noted among therapies. The literature does not clearly demonstrate a preference for one drug. High-treatment success rates (89%) were reported when patients were prescribed all available PDE5i. Daily dosing with tadalafil is associated with improved erectile function (EF) over time. Finally, novel modes of patient-provider interaction, including internet-based education, communication and prescribing, may also improve long-term adherence. CONCLUSIONS: PDE5i represent first line therapy for ED with excellent overall efficacy and satisfactory side effect profiles. Enhanced communciation, coupled with increased knowledge of drug characteristics, comparative treatment regimens and optimal prescribing patterns, offer compelling tools to improve long-term treatment success.

Clinical and laboratory characteristics of 19 patients with Churg-Strauss syndrome from a single South Australian centre.

BACKGROUND: Churg-Strauss syndrome (CSS) is a rare, idiopathic systemic vasculitis. There is emerging evidence of an association between the presence or absence of antineutrophil cytoplasmic antibodies (ANCA) and clinical phenotype. Thromboembolism is an increasingly recognised complication of the disease. AIMS: Given the paucity of Australian data, the aim of this study was to examine the clinical and laboratory features of CSS in a single Australian centre. METHODS: We performed a retrospective review of all patients who fulfilled the American College of Rheumatology classification criteria for CSS managed at the Department of Immunology, Royal Adelaide Hospital between 2002 and 2008. RESULTS: Nineteen patients were included. All patients had asthma and most had upper airway involvement. Peripheral nerve, musculoskeletal, gastrointestinal and cutaneous involvement was common. Renal and cardiac involvement was uncommon in this series. Histological confirmation was obtained in 15 patients (78.9%). Ten patients (52.6%) were ANCA+, and these were more likely to have musculoskeletal involvement, such as arthralgia or myalgia (odds ratio 57, P = 0.005). Thrombosis was a feature at diagnosis in six patients (31.6%); two of these recurred with relapse. Sixteen patients (84.2%) were followed up; five died, and mean survival was 8.9 years. CONCLUSIONS: This is the first Australian study to focus on CSS. Our results demonstrate similar presentation and prognosis of CSS to previous descriptions; however, we noted that musculoskeletal involvement was more common in ANCA+ patients. In our series, thrombosis was a significant complication and we suggest that thromboprophylaxis may be warranted.

Oral drug challenges in non-steroidal anti-inflammatory drug-induced urticaria, angioedema and anaphylaxis.

BACKGROUND: Urticaria, angioedema and anaphylaxis are common adverse reactions to non-steroidal anti-inflammatory drugs (NSAIDs). AIM: To investigate the clinical characteristics of NSAID-induced acute hypersensitivity reactions with structured oral drug challenges. METHODS: Patients with NSAID-induced urticaria, angioedema or anaphylaxis were challenged with either the homologous NSAID to confirm diagnosis or a heterologous NSAID to investigate cross-reactivity. Data were analysed retrospectively and supplemented by a telephone questionnaire. RESULTS: Sixty-eight patients (mean age 48.3, 53 females) reported a total of 75 instances of NSAID-induced reactions of which 64% were purely cutaneous and 36% were systemic anaphylaxis. Ibuprofen was the most frequent cause of reactions (35%), however, diclofenac was the most frequent cause of anaphylaxis (48%). Seventeen out of 40 (43%) homologous NSAID challenges were positive; presentation with anaphylaxis or reaction to diclofenac predicted a positive challenge. Only 7 of 28 (25%) of heterologous NSAID challenges were positive. Structured challenges enabled us to identify 23 (34%) patients with selective reactivity to a single NSAID, 19 (28%) patients with cross-reactivity to multiple NSAIDs and 23 (34%) patients in whom NSAID hypersensitivity was not reproduced. Selective reactors presented most often with anaphylaxis and some had a background of beta-lactam antibiotic allergy. Cross-reactive patients often had a background of chronic urticaria and presented with milder reactions. CONCLUSION: In the absence of a reliable in vitro test, structured drug challenges allow identification of selective and cross-reactive NSAID hypersensitivity syndromes. NSAID-induced anaphylaxis is often associated with selective hypersensitivity and patients may not need to avoid other NSAIDs.

A pilot study of ranolazine in patients with intermittent claudication.

AIM: This pilot study provides preliminary information regarding safety and changes in exercise performance during treatment with ranolazine extended-release in patients with reproducible claudication during exercise treadmill testing (ETT). METHODS: We enrolled 45 patients with documented peripheral arterial disease, reproducible claudication on ETT, and ankle-brachial indices <0.85 at rest that decreased by at least 0.15% or 20% immediately postexercise. Randomized patients received double-blind treatment with either ranolazine 1 000 mg b.i.d. (n=22) or placebo (n=23) for 4 weeks. RESULTS: Compared with baseline, peak walking time (PWT) increased (mean+/-SEM) by 53+/-34 s with ranolazine (P=0.13) and by 41+/-33 s with placebo (P=0.22). Pain-free walking time during ETT increased by 62+/-18 s with ranolazine (P=0.002) and 36+/-18 s with placebo (P=0.045). Supplemental analyses, excluding patients with baseline exercise duration (16 min and (12 min, showed additional improvement with ranolazine on PWT. CONCLUSIONS: Ranolazine was well tolerated and these data provide a rationale for proceeding with a definitive trial.

Pseudoanaphylaxis.

Profound hypotension and cardiac arrest after commencement of combined spinal and general anaesthesia in a patient for knee replacement surgery raised the suspicion of anaphylaxis. This seemed to be confirmed when a mast cell tryptase test taken about 90 minutes after the onset of the hypotension was elevated. However, subsequent intradermal skin testing twelve weeks later did not identify a causal drug. Repeat mast cell tryptase at the time showed the same elevation, which led to the correct diagnosis of mastocytosis and a secondary diagnosis that the patient's hypotension and cardiac arrest were the result of her spinal anaesthesia. If the serum tryptase is elevated during the event but no allergic agent can be identified, a further serum tryptase should be taken several weeks later to exclude a persistent elevation due to mastocytosis.

Omapatrilat versus lisinopril: efficacy and neurohormonal profile in salt-sensitive hypertensive patients.

Omapatrilat, a vasopeptidase inhibitor, simultaneously inhibits neutral endopeptidase and ACE. The efficacy and hormonal profile of omapatrilat and lisinopril were compared in salt-sensitive hypertensive patients. On enrollment, antihypertensive medications were withdrawn, and patients received a single-blind placebo. On day 15, salt-sensitivity determinations were made. Salt-sensitive hypertensive patients returned within 5 to 10 days for baseline evaluations of ambulatory diastolic blood pressure, ambulatory systolic blood pressure, and atrial natriuretic peptide. Salt-sensitive hypertensive patients were randomized to receive double-blind omapatrilat (n=28) or lisinopril (n=33) at initial doses of 10 mg for 1 week, increasing to 40 and 20 mg, respectively, for an additional 3 weeks. Ambulatory blood pressure and urinary atrial natriuretic peptide were assessed at study termination. Both omapatrilat and lisinopril significantly reduced mean 24-hour ambulatory diastolic and systolic blood pressures; however, omapatrilat produced significantly greater reductions in mean 24-hour ambulatory diastolic blood pressure (P=0.008), ambulatory systolic blood pressure (P=0.004), and ambulatory mean arterial pressure (P=0.005) compared with values from lisinopril. Both drugs potently inhibited ACE over 24 hours. Omapatrilat significantly (P<0.001) increased urinary excretion of atrial natriuretic peptide over 0- to 24-hour (3.8-fold) and 12- to 24-hour (2-fold) intervals; lisinopril produced no change. Omapatrilat significantly (P<0.001) increased urinary excretion of cGMP over the 0- to 24- and 4- to 8-hour intervals compared with that from lisinopril. Neither drug had a diuretic, natriuretic, or kaliuretic effect. In conclusion, in salt-sensitive hypertensive patients, omapatrilat demonstrated the hormonal profile of a vasopeptidase inhibitor and lowered ambulatory diastolic and systolic blood pressures more than lisinopril.