Invited review: "Probiotic" approaches to improving dairy production: Reassessing "magic foo-foo dust".

The gastrointestinal microbial consortium in dairy cattle is critical to determining the energetic status of the dairy cow from birth through her final lactation. The ruminant's microbial community can degrade a wide variety of feedstuffs, which can affect growth, as well as production rate and efficiency on the farm, but can also affect food safety, animal health, and environmental impacts of dairy production. Gut microbial diversity and density are powerful tools that can be harnessed to benefit both producers and consumers. The incentives in the United States to develop Alternatives to Antibiotics for use in food-animal production have been largely driven by the Veterinary Feed Directive and have led to an increased use of probiotic approaches to alter the gastrointestinal microbial community composition, resulting in improved heifer growth, milk production and efficiency, and animal health. However, the efficacy of direct-fed microbials or probiotics in dairy cattle has been highly variable due to specific microbial ecological factors within the host gut and its native microflora. Interactions (both synergistic and antagonistic) between the microbial ecosystem and the host animal physiology (including epithelial cells, immune system, hormones, enzyme activities, and epigenetics) are critical to understanding why some probiotics work but others do not. Increasing availability of next-generation sequencing approaches provides novel insights into how probiotic approaches change the microbial community composition in the gut that can potentially affect animal health (e.g., diarrhea or scours, gut integrity, foodborne pathogens), as well as animal performance (e.g., growth, reproduction, productivity) and fermentation parameters (e.g., pH, short-chain fatty acids, methane production, and microbial profiles) of cattle. However, it remains clear that all direct-fed microbials are not created equal and their efficacy remains highly variable and dependent on stage of production and farm environment. Collectively, data have demonstrated that probiotic effects are not limited to the simple mechanisms that have been traditionally hypothesized, but instead are part of a complex cascade of microbial ecological and host animal physiological effects that ultimately impact dairy production and profitability.

Schizochytrium sp. and lactoferrin supplementation alleviates Escherichia coli K99-induced diarrhea in preweaning dairy calves.

Calf diarrhea, a common disease mainly induced by Escherichia coli infection, is one of the main reasons for nonpredator losses. Hence, an effective nonantibacterial approach to prevent calf diarrhea has become an emerging requirement. This study evaluated the microalgae Schizochytrium sp. (SZ) and lactoferrin (LF) as a nutrient intervention approach against E. coli O101:K99-induced preweaning calve diarrhea. Fifty 1-d-old male Holstein calves were randomly divided into 5 groups (n = 10): (1) control, (2) blank (no supplement or challenge), (3) 1 g/d LF, (4) 20 g/d SZ, or (5) 1 g/d LF plus 20 g/d SZ (LFSZ). The experimental period lasted 14 d. On the morning of d 7, calves were challenged with 1 × 1011 cfu of E. coli O101:K99, and rectum feces were collected on 3, 12, 24, and 168 h postchallenge for the control, LF, SZ, and LFSZ groups. The rectal feces of the blank group were collected on d 14. Data were analyzed using the mixed procedure of SAS (version 9.4; SAS Institute Inc.). The E. coli K99 challenge decreased the average daily gain (ADG) and increased feed-to-gain ratio (F:G) and diarrhea frequency (control vs. blank). Compared with the control group, the LFSZ group had a higher ADG and lower F:G, and the LFSZ and SZ groups had lower diarrhea frequency compared with the control group. In addition, the LFSZ and SZ groups have no differences in diarrhea frequency compared with the blank group. Compared with the control group, the blank group had lower serum nitric oxide (NO), endothelin-1, d-lactic acid (D-LA), and lipopolysaccharide (LPS) concentrations, as well as serum IgG, IL-1ß, IL-6, IL-10, and TNF-α levels on d 7 and 14. On d 7, compared with the control group, all treatment groups had lower serum NO level, the SZ group had a lower serum D-LA concentration, and the LF and LFSZ groups had lower serum LPS concentration. On d 14, compared with the control group, the fecal microbiota of the blank group had lower Shannon, Simpson, Chao1, and ACE indexes, the LFSZ group had lower Shannon and Simpson indexes, the SZ and LFSZ groups had a higher Chao1 index, and all treatment groups had a higher ACE index. In fecal microbiota, Bifidobacterium and Actinobacteria were negatively associated with IL-10 and d-lactate, while Akkermansia was negatively associated with endothelin-1 and positively correlated with LPS, fecal scores, and d-lactate levels. Our results indicated that LF and SZ supplements could alleviate E. coli O101:K99-induced calf diarrhea individually or in combination. Supplementing 1 g/d LF and 20 g/d SZ could be a potential nutrient intervention approach to prevent bacterial diarrhea in calves.

Survival of Campylobacter jejuni during in vitro culture with mixed bovine ruminal microorganisms in the presence of methanogen inhibitors.

Foodborne pathogen Campylobacter jejuni has been associated with ruminants. The objectives of this experiment were to determine C. jejuni survivability in mixed in vitro rumen microbial populations and the impact on methane production with or without methane inhibitors 2-bromosulfonate (BES) and/or sodium nitrate. When inoculated into rumen microbial populations without or with 0.5 mM BES, 5.0 mM nitrate or their combination, C. jejuni viability decreased from 4.7 ± 0.1 log10 colony forming units (CFU)/mL after 24 h. Loss of C. jejuni viability was greater (P < 0.05) when incubated under 100% CO2 compared to 50% H2:50% CO2, decreasing 1.46 versus 1.15 log units, respectively. C. jejuni viability was also decreased (P < 0.05) by more than 0.43 log units by the anti-methanogen treatments. Rumen microbial populations produced less methane (P = 0.05) when incubated with than without C. jejuni regardless of whether under 100% CO2 or 50% H2:50% CO2. For either gas phase, nitrate was decreased (13.2 versus 37.9%) by the anti-methanogen treatments versus controls although not always significant. C. jejuni-inoculated populations metabolized 16.4% more (P < 0.05) nitrate under H2:CO2 versus 100% CO2. Apparently, C. jejuni can compete for H2 with methanogens but has limited survivability under rumen conditions.

Effects of cashew nut shell extract supplementation on production, rumen fermentation, metabolism, and inflammatory biomarkers in transition dairy cows.

Cashew nut shell extract (CNSE) is a byproduct of the cashew nut industry, containing bioactive compounds that alter rumen fermentation patterns. Therefore, study objectives were to evaluate the effects of CNSE (59% anacardic acid and 18% cardol) on production, rumen fermentation variables, metabolism, and inflammation in transition dairy cows. A total of 51 multiparous Holstein cows were used in a randomized design and assigned to treatment based on their previous 305-d mature equivalent milk and parity. Cows were assigned to 1 of 2 treatments 21 d before expected calving: (1) CON (control diet; n = 17) or (2) CNSE-5.0 (control diet and 5.0 g/d CNSE granule [containing 50% CNSE]; n = 34). Following parturition, 17 cows (preselected at initial treatment assignment) from the CNSE-5.0 treatment were reallocated into a third treatment group: CNSE-2.5 (control diet and 2.5 g/d CNSE granule; n = 17), resulting in 3 total treatments postpartum: (1) CON, (2) CNSE-2.5, and (3) CNSE-5.0. Prepartum rumen pH was unaltered by treatment; however, postpartum rumen pH was increased (0.31 units) in CNSE cows relative to CON. Prepartum rumen ammonia N concentration tended to be decreased (34%) in CNSE-5.0 cows compared with CON, and there tended to be a quadratic effect on postpartum ammonia N, as it was decreased in CNSE-2.5 compared with CON and CNSE-5.0. Prepartum dry matter intake (DMI) was unaffected by treatment; however, postpartum DMI was increased (8%) in CNSE cows relative to CON. No treatment differences were observed in pre- or postpartum digestibility measurements. Milk and protein yields from cows fed CNSE tended to be increased (6% and 7%, respectively) relative to CON. No treatment differences were detected for energy-corrected milk, feed efficiency, body weight, body condition score, energy balance, milk composition, milk urea nitrogen, or somatic cell count. Prepartum fecal pH decreased (0.12 units) in CNSE-5.0 cows relative to CON cows but was similar between treatments postpartum. Supplementing CNSE did not affect prepartum glucose, nonesterified fatty acids (NEFA), ß-hydroxybutyrate (BHB), or insulin. However, prepartum circulating blood urea nitrogen tended to be decreased and glucagon was decreased in CNSE-5.0 cows compared with CON (9 and 20%, respectively). Additionally, CNSE supplementation decreased glucose and insulin concentrations postpartum relative to CON cows (6% and 20%, respectively). Quadratic effects were detected for postpartum circulating NEFA and BHB such that their levels were increased in CNSE-2.5 cows relative to CON and CNSE-5.0. Pre- and postpartum circulating serum amyloid A, lipopolysaccharide-binding protein, and haptoglobin were unaffected by treatment. Overall, CNSE influenced some key rumen fermentation variables, altered postabsorptive metabolism, and increased production parameters in transition dairy cows.

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.

Effects of abomasally infused rumen fluid from corn-challenged donor cows on production, metabolism, and inflammatory biomarkers in healthy recipient cows.

Subacute rumen acidosis may cause postruminal intestinal barrier dysfunction, but this does not appear to be due to increased hindgut fermentation. Alternatively, intestinal hyperpermeability may be explained by the plethora of potentially harmful substances (e.g., ethanol, endotoxin, and amines) produced in the rumen during subacute rumen acidosis, which are difficult to isolate in traditional in vivo experiments. Therefore, objectives were to evaluate whether abomasal infusion of acidotic rumen fluid collected from donor (Donor) cows elicits systemic inflammation or alters metabolism or production in healthy recipients. Ten rumen-cannulated lactating dairy cows [249 ± 63 d in milk; 753 ± 32 kg of body weight (BW)] were randomly assigned to 1 of 2 abomasal infusion treatments: (1) healthy rumen fluid (HF; 5 L/h; n = 5) or (2) acidotic rumen fluid (AF; 5 L/h; n = 5) infused. Eight rumen-cannulated cows [4 dry, 4 lactating (lactating = 391 ± 220 d in milk); 760 ± 70 kg of BW] were used as Donor cows. All 18 cows were acclimated to a high-fiber diet (46% neutral detergent fiber; 14% starch) during an 11-d prefeeding period during which rumen fluid was collected for the eventual infusion into HF cows. During period (P) 1 (5 d), baseline data were obtained and on d 5 Donor were corn-challenged (2.75% BW ground corn after 16 h of 75% feed restriction). Cows were fasted until 36 h relative to rumen acidosis induction (RAI), and data were collected through 96 h RAI. At 12 h RAI, an additional 0.50% BW of ground corn was added, and acidotic fluid collections began (7 L/Donor every 2 h; 6 M HCl was added to collected fluid until pH was between 5.0 and 5.2). On d 1 of P2 (4 d), HF/AF cows were abomasally infused with their respective treatments for 16 h, and data were collected for 96 h relative to the first infusion. Data were analyzed in SAS (SAS Institute Inc.) using PROC MIXED. Following the corn challenge in the Donor cows, rumen pH only mildly decreased at nadir (pH = 5.64 at 8 h RAI) and remained above the desired threshold for both acute (5.2) and subacute (5.6) acidosis. In contrast, fecal and blood pH markedly decreased to acidotic levels (nadir = 4.65 and 7.28 at 36 and 30 h RAI, respectively), and fecal pH remained below 5 from 22 to 36 h RAI. In Donor cows, dry matter intake remained decreased through d 4 (36% relative to baseline) and serum amyloid A and lipopolysaccharide-binding protein markedly increased by 48 h RAI in Donor cows (30- and 3-fold, respectively). In cows that received the abomasal infusions, fecal pH decreased in AF from 6 to 12 h relative to the first infusion (7.07 vs. 6.33) compared with HF; however, milk yield, dry matter intake, energy-corrected milk, rectal temperature, serum amyloid A, and lipopolysaccharide-binding protein were unaffected. Overall, the corn challenge did not cause subacute rumen acidosis but markedly decreased fecal and blood pH and stimulated a delayed inflammatory response in the Donor cows. Abomasal infusion of rumen fluid from corn-challenged Donor cows decreased fecal pH but did not cause inflammation, nor did it create an immune-activated phenotype in recipient cows.

Association of pre-treatment somatic cell counts with bacteriological cure following diagnosis of intramammary infection.

Antibiotic administration is crucial to ensure the health and productivity of dairy cattle. Mastitis is a disease that is typically a result of an intramammary infection (IMI), and antibiotic regimens are implemented to aid in curing IMI. Diagnosis is usually by detection of elevated milk somatic cell counts (SCC) and/or presence of culturable pathogens in the milk. Antibiotic treatment success is associated with the SCC at the time of treatment, though this correlation is still poorly understood. The objective of this project was to evaluate pre-treatment SCC and its association with IMI cure incidence following a standard antibiotic treatment. We hypothesized that pre-treatment SCC would be significantly lower in cases where the IMI ultimately cured compared to cases where the IMI failed to cure. Milk samples were collected aseptically from lactating cow quarters experiencing clinical or subclinical mastitis (n = 52). Clinical mastitis was diagnosed by a trained milking technician and subclinical mastitis was diagnosed at the quarter level as a SCC > 200,000 cells/mL and presence of bacterical growth in milk at time of treatment. After collection of the day 0 (D0) milk samples, the SCC was enumerated, and the milk sample cultured. Intramammary antibiotic therapy Cetftiofur hydrochloride (Spectramast® LC) was administered once/day for 5 days. Post-treatment samples were collected 14 d (D14) and 28 d (D28) later. A bacteriological cure was confirmed when both the D14 and D28 samples were free of culturable pathogens. The overall cure rate was 46.2%. Interestingly, the cure rates of antibiotic therapy decreased as pre-treatment SCC increased. Quarters that experienced bacteriological cure demonstrated a lower pre-treatment SCC (507,041 cells/mL ± 127.86 SEM, P = 0.01) compared to cows that did not cure, which had high pre-treatment SCC (1,640,392 cells/mL ± 333.28 SEM). Quarters that failed to cure had higher SCC values 28 days post-treatment in comparison to quarters that cured (P < 0.001). Future studies should investigate whether we can develop unique SCC-dependent mastitis treatment protocols which increase mastitis cure rates and enhance overall mammary health.

Effects of close-up dietary energy level and supplementing rumen-protected lysine on energy metabolites and milk production in transition cows.

The objective of this study was to investigate the effects of dietary energy levels and rumen-protected lysine supplementation on serum free fatty acid levels, ß-hydroxybutyrate levels, dry matter (DM) intake, and milk production and composition. Treatments were arranged in a 2 × 2 factorial design with 2 dietary energy levels [high net energy for lactation (NEL) = 1.53 Mcal/kg of DM vs. low NEL = 1.37 Mcal/kg of DM; HE vs. LE) fed either with rumen-protected lysine (bypass lysine; 40 g/cow per day) or without rumen-protected lysine (control). Sixty-eight third-lactation Holstein dairy cows entering their fourth lactation were randomly allocated to 4 treatments groups: HE with bypass lysine, HE without bypass lysine, LE with bypass lysine, and LE without bypass lysine. Groups were balanced based upon their expected calving date, previous milk yields, and body condition score. All cows were fed the same diet (NEL = 1.34 Mcal/kg of DM) during the dry period prior to the trial. Rumen-protected lysine was top-dressed on a total mixed ration to deliver 9.68 g/d of metabolizable lysine to pre- and postpartum cows. After calving, all cows received the same TMR (1.69 Mcal/kg of DM). Blood samples were collected at -21, -14, -7, 0, 3, 7, 14, and 21 d relative to calving, and free fatty acids and ß-hydroxybutyrate concentrations were measured. Amount of feed offered and orts were collected and measured for individual cows 4 d/wk. Milk samples were collected once per week following calving, and milk composition was analyzed. Feeding high NEL to close-up cows decreased the concentrations of free fatty acid and ß-hydroxybutyrate in prepartum cows but not in postpartum cows. Addition of rumen-protected lysine increased postpartum DM intake, and decreased serum free fatty acid and ß-hydroxybutyrate concentrations. Neither energy nor rumen-protected lysine supplementation nor their interaction affected milk yield or fat or lactose yields. However, cows in the group receiving HE with bypass lysine tended to produce more milk compared with other groups and had a lower blood ß-hydroxybutyrate concentration postpartum. These results indicate that feeding a high-energy diet together with rumen-protected lysine improved DM intake and lowered serum free fatty acid and ß-hydroxybutyrate concentrations in transition cows.

Isolation and screening of aflatoxin-detoxifying yeast and bacteria from ruminal fluids to reduce aflatoxin B1 contamination in dairy cattle feed.

AIMS: To obtain yeast and bacteria from ruminal fluids that possess aflatoxin B1 (AFB1 ) detoxifying ability for use in animal feed. METHODS AND RESULTS: Sources of isolation were ruminal fluids of three nonfistulated dairy cows, fed diets containing cassava pulp, rice straw or distillery yeast sludge. The isolation was carried out to screen for the isolates that were active in both anaerobic and aerobic conditions. Three yeast and three bacteria isolates were selected. Up to 85% of AFB1 was detoxified by yeast isolates and up to 60% AFB1 reduction was evident by bacteria isolates. Two yeast isolates were identified as Kluyveromyces marxianus and one isolate as Pichia kudriavzevii. The three bacteria isolates were identified as Enterococcus faecium, Corynebacterium phoceense and C. vitaeruminis. All strains showed high biomass production when cultivated in medium with 80 g l-1 glucose. CONCLUSIONS: The isolated yeast and bacteria with AFB1 detoxifying ability showed a good potential to be applied as an aflatoxin-detoxifying agent to ingredients used to feed dairy cattle. SIGNIFICANCE AND IMPACT OF THE STUDY: The abilities of isolates to survive and be active in anaerobic and aerobic conditions rendered them to be active in cattle's rumen. Their biomass could be produced in bulk and used as feed supplement for aflatoxin detoxification in dairy cattle.

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.

Transferability of antimicrobial resistance from multidrug-resistant Escherichia coli isolated from cattle in the USA to E. coli and Salmonella Newport recipients.

OBJECTIVES: This study aimed to evaluate conjugative transfer of cephalosporin resistance among 100 strains of multidrug-resistant Escherichia coli (MDRE) to Salmonella enterica serotype Newport and E. coli DH5α recipients. METHODS: Phenotypic and genotypic profiles were determined for MDRE as well as for Salmonella Newport (trSN) and E. coli DH5α (trDH) transconjugants. RESULTS: Of 95 MDRE donor isolates, 26 (27%) and 27 (28%) transferred resistance to trSN and trDH recipients, respectively. A total of 27 MDRE (27%) were confirmed as extended-spectrum ß-lactamase (ESBL)-producers based on the double-disk synergy assay and whole-genome sequencing (WGS). WGS was performed on 25 of the ESBL-producing isolates, showing that 2 isolates carried blaCTX-M-6, 22 possessed blaCTX-M-32 and 1 was negative for blaCTX-M genes. Fourteen of the ESBLs sequenced were qnrB19. Differential transfer of IncA/C and IncN from MDRE32 was observed between trSN32 and trDH32. IncN-positive trDH32 displayed an ESBL phenotype, whereas IncA/C-positive trSN32 displayed an AmpC phenotype. The rate of ESBL transfer to trSN and trDH recipients was 11% and 96%, respectively. CONCLUSIONS: Twenty-seven MDRE were phenotypically identified as ESBL-producers. WGS of 25 MDRE revealed that 2 and 22 isolates carried blaCTX-M-6 and blaCTX-M-32, respectively. One multidrug-resistant isolate exhibited conversion from an AmpC phenotype to an ESBL phenotype with the transfer of only the IncN plasmid. The rate of resistance transfer to Salmonella or E. coli recipients was nearly identical. However, the ESBL phenotype was transferred with significantly greater prevalence to E. coli compared with Salmonella Newport (96% and 11%, respectively).

Effect of monensin withdrawal on intake, digestion, and ruminal fermentation parameters by Bos taurus indicus and Bos taurus taurus steers consuming bermudagrass hay.

Effects of monensin withdrawal and cattle subspecies on the utilization of bermudagrass hay (14.3% CP, 72.3% NDF, and 36.9% ADF) were evaluated using ruminally cannulated steers (5 [BI] and 5 [BT]). Subspecies were concurrently subjected to a 2-period, 2-treatment crossover design. Treatments consisted of either 0 mg·steer·d-1 monensin with no previous monensin feeding (CON) or withdrawal from 200 mg·steer·d-1 monensin (MON) fed individually in 0.91 kg dried distillers' grains with solubles for 42 d. Withdrawal was evaluated for a 28-d period. Ruminal fluid was collected 2 h after feeding on d 0, 1, 4, 7, 14, and 21 after withdrawal for determination of pH, VFA, ruminal NH-N (RAN), rate of NH production, and CH production rate. Hay, ort, and fecal grab samples were collected d 23 through 28 after withdrawal for determination of intake and digestion. No subspecies × monensin, subspecies × day, or subspecies × monensin × day interactions were observed ( ≥ 0.11). An effect of day after monensin withdrawal was observed ( < 0.01) for total VFA concentration, with an increase following withdrawal followed by a decrease and then stabilization. Monensin × day after monensin withdrawal interactions ( ≤ 0.01) were observed for the acetate:propionate (A:P) ratio and molar percent of acetate and propionate. There was a decrease in molar percent of propionate between d 1 and 4 from 19.1 to 18.0; however, it remained greater ( ≤ 0.10) for MON than CON through d 7. Withdrawal increased molar percent of acetate from 68.3 to 69.8 between d 0 and 4 for MON steers. The A:P ratio was less ( ≤ 0.01) on d 0 for MON than for CON (3.4 vs. 4.0), but by d 4, it increased to 3.8 and was not different ( = 0.14) from CON. By d 14, no differences ( ≥ 0.88) remained for acetate, propionate, or the A:P ratio. After monensin withdrawal, monensin reduced ( < 0.01) RAN by 12.3% (2.09 vs. 1.83 m for CON and MON, respectively). Monensin withdrawal and cattle subspecies had no effect ( ≥ 0.23) on rate of NH production or CH production rate. Monensin withdrawal had no effect ( ≥ 0.45) on intake or digestibility parameters. Greater forage OM intake ( = 0.09; 21.2 vs. 19.2 g/kg BW) and OM digestibility ( < 0.01; 72.4 vs. 63.0%) resulted in greater ( < 0.01) total digestible OM intake (16.8 vs. 13.2 g/kg BW) in BT steers than in BI steers. These results suggest that BT steers are better able to utilize bermudagrass hay than BI steers. Upon monensin withdrawal, steers previously fed monensin continue to have a reduced A:P ratio for at least 7 d.

Effect of monensin inclusion on intake, digestion, and ruminal fermentation parameters by Bos taurus indicus and Bos taurus taurus steers consuming bermudagrass hay.

Effects of monensin inclusion and cattle subspecies on utilization of bermudagrass hay (13.7% CP, 77.3% NDF, and 38.8% ADF) were evaluated using ruminally cannulated steers (5 [BI] and 5 [BT]; 398 kg BW). Subspecies were concurrently subjected to a 2-period, 2-treatment crossover design. Treatments were 0 (CON) or 200 mg·steer·d monensin (MON) in 0.91 kg dried distillers' grains with solubles. Periods were 70 d in length: 20 d of adaptation, 22 d of sample collection, and 28 d for withdrawal of treatment. Steers were group housed during adaptation and moved to individual covered pens for sampling. Hay, ort, and fecal grab samples were collected d 21 through 25 for determination of intake and digestion. Ruminal fluid was collected with a suction strainer 0, 2, 4, 8, and 12 h after feeding on d 42 for pH, VFA, and ruminal NH-N (RAN) analysis. Additionally, at h 2, ruminal fluid and contents were collected for determination of rate of NH production and CH production rate. No subspecies × monensin interactions were observed ( ≥ 0.12). Monensin had no effect ( ≥ 0.16) on intake or digestibility parameters. No subspecies effect ( ≥ 0.11) was observed for forage OM intake, total OM intake, or OM digestion. Total digestible OM intake tended to be greater ( = 0.06) for BT steers than for BI steers (14.0 vs. 12.2 g/kg BW). There was an effect of hour after feeding ( ≤ 0.01) on pH, total VFA, acetate:propionate ratio, and molar percent acetate and propionate. Total VFA concentration was greater ( = 0.01) in CON steers than in MON steers (66.5 vs. 62.0 m). Monensin decreased molar percent acetate ( = 0.02) from 72.5 to 71.2% and increased molar percent propionate ( < 0.01) from 16.9 to 18.7%, resulting in a reduced ( < 0.01) acetate:propionate ratio (from 4.34 to 3.85). Although not significantly ( = 0.19), monensin numerically reduced the CH production rate by 15.8%. Greater ( = 0.07) CH production rate tended to be observed in BI steers than in BT steers (21.4 vs. 16.6 µmol CH·mL·h, respectively). Monensin had no effect ( ≥ 0.32) on pH, RAN, or rate of NH production. A subspecies × hour after feeding interaction was observed for RAN, with BT having greater RAN at h 0 and 4, whereas BI had greater RAN at h 2, 8, and 12. Overall, monensin decreased the acetate:propionate ratio and total VFA concentration but had no effect on forage utilization. steers consumed less digestible OM and had a greater CH production rate compared with BT steers, suggesting BT were better able to utilize the available forage resource than BI.

Use of a novel oleaginous microorganism as a potential source of lipids for weanling pigs.

Weanling pigs are at risk of succumbing to illness due to an immature immune system and insufficient supply of available energy at the time of weaning. This study was aimed at determining whether oleaginous bacteria could serve as a source of lipids to weanling pigs. Weanling pigs were provided a daily dose of 1×109 colony fomring unit (CFU) = kg-1 of the novel oleaginous Enterobacter cloacae strain JD6301 or JD8715 (which is a variant form of JD6301 capable of producing extracellular triglycerides) via oral gavage for 5 d. Serum was collected every 6 h and intestinal samples were collected at 6 d. Providing pigs with JD6301 or JD8715 significantly increased serum concentrations of triglycerides and non-esterified fatty acids (NEFA) within 72 h. Additionally, the JD6301 and JD8715 strains were able to survive within the gastrointestinal tract throughout the duration of the study. These results suggest that providing Enterobacter cloacae can increase the serum lipids in the pigs, thus potentially providing an additional source of energy to animals during times of stress. This could potentially help improve the metabolic response of animals during times of stress.

The Role of Direct-Fed Microbials in Conventional Livestock Production.

Supplementation of direct-fed microbials (DFM) as a means to improve the health and performance of livestock has generated significant interest over the past 15+ years. A driving force for this increased interest in DFM is to reduce or eliminate the use of low-dose antibiotics in livestock production. This increased attention toward DFM supplementation has generated an extensive body of research. This effort has resulted in conflicting reports. Although there has been considerable variation in the design of these studies, one of the main causes for this lack of consistency may be attributed to the variation in the experimental immune challenge incorporated to evaluate DFM supplementation. Taking into account the experimental immune challenge, there is strong evidence to suggest that DFM supplementation may have an impact on the immune response, overall health, and performance of livestock.

Dehydrated citrus pulp alters feedlot performance of crossbred heifers during the receiving period and modulates serum metabolite concentrations before and after an endotoxin challenge.

English × Continental heifers ( = 180) were sourced in 2 loads (219.3 ± 16.0 and 221.4 ± 16.4 kg, respectively) from commercial auction barns to study the effects of feeding dehydrated citrus pulp (DCP) on feedlot performance of newly received heifers. A completely randomized block design was used with BW nested within arrival load and blocked by BW into 3 dietary treatments (36 pens, 5 heifers/pen, 12 blocks, 3 pens/block, and 12 pens/treatment). Treatment diets contained 1) 0% DCP (control diet [CON]), 2) 10% DCP, or 3) 20% DCP on a DM basis. Diets containing DCP were exchanged with steam-flaked corn on a 1:1 basis. Cattle were fed a 63, 73, and 83% concentrate diet from d 0 to 28, d 28 to 42, and d 42 to 56, respectively. Over the 56-d trial period, as the amount of dietary DCP increased, DMI decreased ( = 0.01), ADG decreased ( < 0.01), and G:F decreased ( = 0.02). From d 0 to 28, there was no difference in the observed minus the predicted NEg of the diet ( = 0.73); from d 28 to 42, there was a linear increase in NEg favoring DCP treatments ( < 0.01); and from d 42 to 56, there was a linear decrease in NEg against the DCP treatments ( < 0.01). At the conclusion of the trial, a subset of heifers ( = 22; 307.89 ± 3.32 kg on d 63) were used to evaluate blood metabolite concentrations before and after a lipopolysaccharide (LPS) challenge. On d 63, heifers were fitted with jugular catheters and moved into individual stalls. On d 64, heifers were intravenously challenged with LPS (0.5 µg/kg BW), and blood samples were collected every 0.5 h from -2 to 8 h and at 24 h relative to the LPS challenge (0 h). Serum glucose, serum urea nitrogen (SUN), and NEFA concentrations were determined. Cattle lost less weight at both 24 and 72 h after the LPS challenge with increasing DCP percentage ( < 0.01). Glucose ( = 0.12) and NEFA ( = 0.13) concentrations did not differ before the LPS challenge; however, there was a treatment effect for SUN, with elevated concentrations of SUN in CON cattle ( < 0.01). After the LPS challenge, DCP-fed cattle had reduced glucose, elevated NEFA, and reduced SUN concentrations ( ≤ 0.01). Results indicate that dietary DCP modulated metabolite concentrations in heifers following an endotoxin challenge and affected feedlot performance when incorporated in receiving diets in replacement of corn. Future studies will need to address strategies to increase DMI or explore levels of DCP less than 10% in the diet of newly received heifer calves.

Meat Science and Muscle Biology Symposium: Ecological and dietary impactors of foodborne pathogens and methods to reduce fecal shedding in cattle.

Pathogenic bacteria can live asymptomatically within and on cattle and can enter the food chain but also can be transmitted to humans by fecal or direct animal contact. Reducing pathogenic bacterial incidence and populations within live cattle represents an important step in improving food safety. A broad range of preslaughter intervention strategies are being developed, which can be loosely classified as 1) directly antipathogen strategies, 2) competitive enhancement strategies (that use the microbiome's competitive nature against pathogens), and 3) animal management strategies. Included within these broad categories are such diverse methods as vaccination against foodborne pathogens, probiotics and prebiotics, bacterial viruses (i.e., bacteriophages), sodium chlorate feeding, and dietary and management changes that specifically alter the microbiome. The simultaneous application of 1 or more preharvest strategies has the potential to reduce human foodborne illnesses by erecting multiple hurdles preventing entry into humans. However, economic factors that govern producer profitability must be kept in mind while improving food safety.

Development of a transdermal Salmonella challenge model in calves.

Recent investigations have found that Salmonella can be routinely recovered from peripheral lymph nodes (PLNs) of cattle presented for harvest. When contained within the PLNs, this foodborne pathogen is protected from currently used postharvest, inplant intervention strategies and, therefore, PLNs harboring Salmonella may be a potential contaminant of ground beef. The objective of this work was to develop a challenge model that effectively and repeatedly results in Salmonella -positive PLNs. A 10-lancet skin-allergy instrument was inoculated with Salmonella, and calves were inoculated intra- and/or transdermally by applying the device over various ventral regions of the skin. Salmonella was successfully and predictably recovered from regionspecific PLNs up to 8 days postchallenge. Furthermore, serotypes inoculated within specific regions were only recovered from the PLNs draining those regions. This model provides a method to predictably infect PLNs with Salmonella. Further, this model makes it possible to determine the duration of infection and to evaluate candidate interventions that may shorten the duration of infection.

Development of challenge models to evaluate the efficacy of a vaccine to reduce carriage of Salmonella in peripheral lymph nodes of cattle.

Because challenge models to infect peripheral lymph nodes (PLNs) with Salmonella have not been reported, we performed a series of experiments to develop and refine challenge models to evaluate an intervention applied at the animal level and to provide initial estimates of efficacy of an intervention (i.e., a vaccine) to aid in the design of future studies. In each of four experiments, steers (control or vaccinated) were inoculated with Salmonella strains Montevideo or Newport, and in experiment IV, Salmonella Senftenberg was also used. Calves were euthanized 14 to 42 days postinoculation, and PLNs were collected. In the first experiment, calves were challenged with ∼10¹° Salmonella cells, and few treatment differences were observed 14 days postchallenge. However, by day 21, Salmonella Newport was recovered from fewer vaccinated calves than control calves (P < 0.05). In experiment II, calves were challenged with ∼107 Salmonella cells and, after two necropsies (14 and 28 days postchallenge), only one lymph node was Salmonella positive; therefore, the study was terminated. In experiment III, calves were again challenged with ∼10¹° Salmonella cells, and no significant effect of vaccine was observed in calves challenged with Montevideo or Newport strains. A transdermal route of challenge was explored in experiment IV, using a 10-lancet, allergy testing instrument. Sixteen steers were challenged with either Salmonella Newport or Salmonella Montevideo (Salmonella Newport right legs; Salmonella Montevideo left legs), and all steers were challenged on the lower abdomen with Salmonella Senftenberg. Transdermal inoculation resulted in predictably Salmonella-positive PLNs, and a modest vaccine effect was detected. Because it is well tolerated by the calves and results in predictable and regionally specific Salmonella recovery from PLNs, the transdermal route of challenge may be preferred by researchers wishing to evaluate the impact of interventions designed to reduce the carriage of Salmonella in PLNs.

Board-invited review: Rumen microbiology: leading the way in microbial ecology.

Robert Hungate, considered the father of rumen microbiology, was the first to initiate a systematic exploration of the microbial ecosystem of the rumen, but he was not alone. The techniques he developed to isolate and identify cellulose-digesting bacteria from the rumen have had a major impact not only in delineating the complex ecosystem of the rumen but also in clinical microbiology and in the exploration of a number of other anaerobic ecosystems, including the human hindgut. Rumen microbiology has pioneered our understanding of much of microbial ecology and has broadened our knowledge of ecology in general, as well as improved the ability to feed ruminants more efficiently. The discovery of anaerobic fungi as a component of the ruminal flora disproved the central dogma in microbiology that all fungi are aerobic organisms. Further novel interactions between bacterial species such as nutrient cross feeding and interspecies H2 transfer were first described in ruminal microorganisms. The complexity and diversity present in the rumen make it an ideal testing ground for microbial theories (e.g., the effects of nutrient limitation and excess) and techniques (such as 16S rRNA), which have rewarded the investigators that have used this easily accessed ecosystem to understand larger truths. Our understanding of characteristics of the ruminal microbial population has opened new avenues of microbial ecology, such as the existence of hyperammonia-producing bacteria and how they can be used to improve N efficiency in ruminants. In this review, we examine some of the contributions to science that were first made in the rumen, which have not been recognized in a broader sense.