Technical note: Infusion,sampling, and vacuum-assisted collection devices for use in ruminally cannulated calves

Calves can be ruminally cannulated at young ages, but equipment size limitations preclude use of an infusion and sampling device in these small animals. Likewise, a procedure to easily evacuate rumen contents in young calves has not been described. Overcoming these technical complications related to assessment of ruminal passage kinetics, nutrient digestion, and volatile fatty acid absorption would aid in future studies advancing our knowledge of dairy calf nutrition. The first objective was to design and fabricate 2 devices (one device for infusion and sampling, and another for vacuum-assisted collection) suitable for use in young ruminally cannulated dairy calves. The second objective was to test the utility of these tools when performing procedures commonly used in ruminant nutrition research. A single weaned 62-d-old ruminally cannulated calf was used to evaluate the ability to infuse a solution of LiCoEDTA and sample rumen contents through the cannula cap over a period of 2 h to assess the rumen liquid passage rate (procedure 1). The device was capable of infusing the LiCoEDTA and sampling the rumen fluid, as evidenced by the presence of elevated Co concentrations in the sampled rumen fluid. Using the fluid samples obtained, liquid passage rate within the calf was estimated to be 40.2% of ruminal fluid/h. The second procedure tested the vacuum-assisted collection device and consisted of evacuating and weighing the rumen contents, which is considered a key preparatory step in washed reticulorumen technique experiments that aim to measure nutrient absorption. In agreement with existing literature, evacuated rumen contents represented approximately 4% of the calf's body weight. In conclusion, custom-built devices for infusion, sampling, and vacuum-assisted collection were efficacious when tested in a 62-d-old ruminally cannulated calf fed a diet of 100% texturized starter (18% crude protein, as-fed). Fellow scientists may employ and further modify these techniques to suit their needs when assessing passage kinetics, nutrient digestion, and volatile fatty acid absorption in calves.     

Effects of lasalocid on selected ruminal and blood metabolites in young calves.

Twelve Holstein bull calves were ruminally cannulated at 5 d of age and assigned to 0 or 1 mg of lasalocid/kg of BW daily, administered postruminally via milk replacer or into the ruminal cannula. Calves were fed milk replacer for 8 wk and calf starter for 12 wk. Lasalocid administration was terminated at weaning in calves fed lasalocid in milk replacer. Ruminal pH tended to be higher in calves fed lasalocid ruminally than in calves on control treatment and averaged 5.9 and 5.6 and 5.4 and 5.1 during wk 1 to 8 and 9 to 12, respectively. Molar proportion of ruminal butyrate tended to be lower when lasalocid was added to the rumen, particularly after weaning. Blood beta-hydroxybutyrate and acetoacetate were lower when lasalocid was administered into the rumen after weaning and averaged .897 and .646 and .026 and .015 mM in calves on control and ruminal treatments, respectively. No effects of lasalocid administered via the milk replacer were observed, except for plasma NEFA, which were reduced postweaning. These data suggest that lasalocid reduces blood beta-hydroxybutyrate by changes in ruminal fermentation and subsequent metabolism of butyrate by ruminal epithelium.     

Effects of supplemental butyrate and weaning on rumen fermentation in Holstein calves

The objectives of this study were to determine the effects of the weaning transition and supplemental rumen-protected butyrate on subacute ruminal acidosis, feed intake, and growth parameters. Holstein bull calves (n = 36; age = 10.7 ± 4.1 d; ± standard deviation) were assigned to 1 of 4 treatment groups: 2 preweaning groups, animals fed milk replacer only (PRE-M) and those fed milk replacer, calf starter, and hay (PRE-S); and 2 postweaning groups, animals fed milk replacer, calf starter, and hay without supplemental rumen-protected butyrate (POST-S) or with supplemental rumen-protected butyrate at a rate of 1% wt/wt during the 2-wk weaning transition (POST-B). Milk replacer was provided at 1,200 g/d; starter, water, and hay were provided ad libitum. Weaning took place over 14 d by reducing milk replacer provision to 900 g/d in wk 7, 600 g/d in wk 8, and 0 g/d in wk 9. Rumen pH was measured continuously for 7 d during wk 6 for PRE-S and PRE-M and during wk 9 for POST-S and POST-B. After rumen pH was measured for 7 d, calves were euthanized, and rumen fluid was sampled and analyzed for volatile fatty acid (VFA) profile. Individual feed intake was recorded daily, whereas, weekly, body weights were recorded, and blood samples were collected. Compared with PRE-M, PRE-S calves tended to have a greater total VFA concentration (35.60 ± 11.4 vs. 11.90 ± 11.8 mM) but mean rumen pH was unaffected (6.25 ± 0.22 vs. 6.17 ± 0.21, respectively). Between PRE-S (wk 6) and POST-S (wk 9), calf starter intake increased (250 ± 219 vs. 2,239 ± 219 g/d), total VFA concentrations increased (35.6 ± 11.4 vs. 154.4 ± 11.8 mM), but mean rumen pH was unaffected (6.25 ± 0.22 vs. 6.40 ± 0.22, respectively). Compared with POST-S, POST-B calves had greater starter intake in wk 7, 8, and 9, but POST-B tended to have lower total VFA concentration (131.0 ± 11.8 vs. 154.4 ± 11.8 mM) and lower mean ruminal pH (5.83 ± 0.21 vs. 6.40 ± 0.22). In conclusion, the weaning transition does not appear to affect rumen pH and VFA profile, but supplementing rumen-protected butyrate during the weaning transition increased starter intake and average daily gain. Further, these data suggest that the ability of the rumen to manage rumen pH changes fundamentally postweaning. Why weaned calves with lower rumen pH can achieve higher calf starter intakes is unclear; these data suggest the effect of rumen pH on feed intake differs between calves and cows.     

Abrupt weaning reduces postweaning growth and is associated with alterations in gastrointestinal markers of development in dairy calves fed an elevated plane of nutrition during the preweaning period

The benefits of feeding elevated quantities of milk to dairy calves have been well established. However, there is a reluctance to adopt this method of feeding in commercial dairy production because of concerns around growth, health, and ruminal development during weaning. The objective of this study was to characterize the effect of an abrupt (0 d step-down) or gradual (12 d step-down) feeding scheme when calves are fed an elevated plane of nutrition (offered 1.35 kg of milk replacer/d). For this experiment, a total of 54 calves were randomly assigned to an abrupt or a gradual weaning protocol before weaning at 48 d of life. Calves were housed and sampled in individual pens for the duration of the experiment, and milk, starter, and straw intake were measured on a daily basis. Body weight was measured every 6 d, whereas blood, rumen fluid, and fecal samples were collected on d 36 (pre-step-down), 48 (preweaning), and 54 (postweaning) of the experiment. Although the growth rates of the step-down calves were lower from d 37 to weaning (0.62 ± 0.04 vs. 1.01 ± 0.04 kg/d), the postweaning average daily gain was greater compared with the group that was abruptly weaned (0.83 ± 0.06 vs. 0.22 ± 0.06 kg/d). Total ruminal volatile fatty acid was greater in the step-down group on the day of weaning (d 48; 59.80 ± 2.25 vs. 45.01 ± 2.25 mmol), whereas the fecal starch percentage was lower during postweaning compared with the abruptly weaned calves (d 54; 3.31 ± 0.76 vs. 6.34 ± 0.76%). Analysis of the digestive tract of bull calves on d 55 revealed minimal differences between gross anatomy measurements of gut compartments as well as no morphological differences in rumen papillae development, yet the total mass of rumen when full of contents was larger in the step-down calves (7.83 ± 0.78 vs. 6.02 ± 0.78 kg). Under the conditions of this study, the results showcase the benefits of a step-down feeding strategy from an overall energy balance standpoint, due to increased adaptation of the gastrointestinal tract preweaning.     

Technical note: A simple rumen collection device for calves: An adaptation of a manual rumen drenching system

A limited amount of research is available related to the rumen microbiota of calves, yet there has been a recent spike of interest in determining the diversity and development of calf rumen microbial populations. To study the microbial populations of a calf's rumen, a sample of the rumen fluid is needed. One way to take a rumen fluid sample from a calf is by fistulating the animal. This method requires surgery and can be very stressful on a young animal that is trying to adapt to a new environment and has a depressed immune system. Another method that can be used instead of fistulation surgery is a rumen pump. This method requires a tube to be inserted into the rumen through the calf's esophagus. Once inside the rumen, fluid can be pumped out and collected in a few minutes. This method is quick, inexpensive, and does not cause significant stress on the animal. This technical note presents the materials and methodology used to convert a drenching system into a rumen pump and its respective utilization in 2 experiments using dairy bull calves.     

Performance and ruminal function development of young calves fed diets with Aspergillus oryzae fermentation extract.

Neonatal Holstein heifer (n = 72) and bull (n = 40) calves were used to study the effects of Aspergillus oryzae fermentation extract (Amaferm) on their performance and on rumen development. The starter diets were formulated to achieve Amaferm consumption of 0, .5, 1, or 3 g per calf daily. Calves were fed milk daily and allowed to consume starter and a mixture of alfalfa and bromegrass hay ad libitum. Weaning was when calves consumed 550 g of starter on 2 consecutive d. Weight gain and feed consumption were recorded weekly. Forty of the heifer calves, 10 from each treatment, were selected randomly to study the effects of Amaferm on ruminal fermentative development. Ruminal fluid samples were collected for pH, ruminal fermentation products, and for bacterial enumerations. Overall, Amaferm-supplemented calves were weaned 1 wk earlier than unsupplemented calves. They had higher total VFA, propionate, and acetate concentrations in the rumen than unsupplemented calves. Total anaerobic, hemicellulolytic, and pectinolytic bacterial counts were higher; cellulolytic bacterial counts tended to be higher for the Amaferm-supplemented calves than for controls. In general, Amaferm-supplemented calves had greater ruminal microbial activity than those not fed Amaferm.     

Short communication: Comparison of pH,volatile fatty acids,and microbiome of rumen samples from preweaned calves obtained via cannula or stomach tube

The objective of this study was to compare rumen samples from young dairy calves obtained via a stomach tube (ST) or a ruminal cannula (RC). Five male Holstein calves (46 ± 4.0 kg of body weight and 11 ± 4.9 d of age) were ruminally cannulated at 15 d of age. Calves received 4 L/d of a commercial milk replacer (25% crude protein and 19.2% fat) at 12.5% dry matter, and were provided concentrate and chopped oats hay ad libitum throughout the study (56 d). In total, 29 paired rumen samples were obtained weekly throughout the study in most of the calves by each extraction method. These samples were used to determine pH and volatile fatty acids (VFA) concentration, and to quantify Prevotella ruminicola and Streptococcus bovis by quantitative PCR. Furthermore, a denaturing gradient gel electrophoresis was performed on rumen samples harvested during wk 8 of the study to determine the degree of similarity between rumen bacteria communities. Rumen pH was 0.30 units greater in ST compared with RC samples. Furthermore, total VFA concentrations were greater in RC than in ST samples. However, when analyzing the proportion of each VFA by ANOVA, no differences were found between the sampling methods. The quantification of S. bovis and P. ruminicola was similar in both extraction methods, and values obtained using different methods were highly correlated (R² = 0.89 and 0.98 for S. bovis and P. ruminicola, respectively). Fingerprinting analysis showed similar bacteria band profiles between samples obtained from the same calves using different extraction methods. In conclusion, when comparing rumen parameters obtained using different sampling techniques, it is recommended that VFA profiles be used rather than total VFA concentrations, as total VFA concentrations are more affected by the method of collection. Furthermore, although comparisons of pH across studies should be avoided when samples are not obtained using the same sampling method, the comparison of fingerprinting of a bacteria community or a specific rumen bacterium is valid.     

Changes in the rumen and colon microbiota and effects of live yeast dietary supplementation during the transition from the dry period to lactation of dairy cows

The first objective of this study was to evaluate the dynamics and their potential association with animal performance of the microbiota in both the rumen and colon of dairy cows as they move from a nonlactation to a lactation ration. The second objective was to assess the potential effects on the microbiota of live yeast supplementation. Twenty-one Holstein cows were split in 2 treatments consisting of 1 × 10¹⁰ cfu/d of live yeast (LY; n = 10) or no supplementation (control; n = 11) starting 21 d before until 21 d after calving. At 14 d before and 7 and 21 d after calving, samples of rumen and colon digesta were obtained from each cow using an endoscope. Total DNA was extracted and submitted to high-throughput sequencing. Shannon diversity index, in both the rumen and colon, was unaffected by LY; however, in the rumen it was lowest 7 d after calving and returned to precalving values at 21 d in milk, whereas in the colon it was greatest 14 d before calving but decreased after calving. In the rumen, LY supplementation increased the relative abundance (RA) of Bacteroidales (group UCG-001), Lachnospiracea (groups UCG-002 and UCG-006), and Flexilinea 14 d before calving, and increased RA of Streptococcus 21 d after calving compared with control cows. However, changes in the ruminal microbiota were more drastic across days relative to calving than as influenced by the dietary treatment, and the effect of LY in the colon was milder than in the rumen. The ruminal RA of several genera was associated with postcalving DMI, and that of Gastranaerophilales was the only order positively associated with milk yield. Several genera were positively correlated with feed efficiency, with Clostridiales (unclassified) being the only genus negatively associated with feed efficiency. In the colon, Prevotellaceae (group Ga6A1) was the only genus positively associated with feed efficiency. The ruminal RA of Prevotella 7 and Ruminobacter 14 d precalving was negatively correlated with dry matter intake and milk yield postcalving. The RA of Parabacteroides in the colon 14 d before calving was negatively correlated with milk yield, whereas the RA of Eggerthellaceae (unclassified) and Erysipelotrichaceae (groups c and unclassified) were positively correlated with feed efficiency. Interestingly, LY supplementation doubled the RA of Eggerthellaceae (unclassified) in the colon. It is concluded that microbial diversity in the rumen experiences a transient reduction after calving, whereas in the colon, the reduction is maintained at least until 21 d in milk. Most of the effects of LY on rumen microbiota were observed before calving, whereas in the colon, LY effects were more moderate but consistent and independent of the stage of production. The microbial community of the rumen after calving is more associated with feed intake, milk yield, and feed efficiency than that of the colon. However, the colon microbiota before calving is more associated with feed efficiency after calving than that of the rumen.     

Effects of corn processing on growth characteristics, rumen development, and rumen parameters in neonatal dairy calves

Neonatal Holstein calves were fed texturized calf starters containing 33% whole (WC), dry-rolled (DRC), roasted-rolled (RC), or steam-flaked (SFC) corn to investigate how corn processing method affects intake, growth, rumen and blood metabolites, and rumen development. In the first experiment, 92 Holstein calves (52 male and 40 female) were started at 2 +/- 1 d of age and studied for 42 d. Starter dry matter (DM) intake was measured and fecal scoring conducted daily. Growth and blood parameter measurements were conducted weekly. A subset of 12 male calves (3/treatment) was euthanized at 4 wk of age and rumen tissue sampled for rumen epithelial development measurements. Experiment 2 consisted of 12 male Holstein calves ruminally cannulated at 7 +/- 1 d of age. Rumen fluid and blood samples were collected during wk 2 to 6. In the first experiment, postweaning and overall starter and total DM intake were significantly higher in calves fed starter with DRC than RC or SFC. Postweaning and overall starter and total DM intake were significantly higher in calves fed starter with WC than SFC. Postweaning average daily gain was significantly greater in calves fed starter with DRC than SFC. Blood volatile fatty acid concentrations were significantly higher in calves fed starter with SFC than in calves fed all other treatments. Papillae length and rumen wall thickness at 4 wk were significantly greater in calves fed starter with SFC than DRC and WC, respectively. In experiment 2, calves fed starter with WC had higher rumen pH and lower rumen volatile fatty acid concentrations than calves fed all other starters. Rumen propionate production was increased in calves receiving starter with SFC; however, rumen butyrate production was higher in calves fed starter with RC. Results indicate that the type of processed corn incorporated into calf starter can influence intake, growth, and rumen parameters in neonatal calves. Calves consuming starter containing RC had similar body weight, feed efficiency, and rumen development but increased structural growth and ruminal butyrate production when compared with the other corn processing treatments.     

Effects of antibiotic residues in milk on growth,ruminal fermentation,and microbial community of preweaning dairy calves

The aim of this study was to evaluate the effects of antibiotic residues in milk on growth, ruminal fermentation, and microbial community of dairy calves in their first 35 d of age. Twenty newborn Holstein bull calves were assigned to 1 of 2 treatments equally: milk replacer without antibiotics (control) and milk replacer plus 4 antibiotics: 0.024 mg/L of penicillin, 0.025 mg/L of streptomycin, 0.1 mg/L of tetracycline, and 0.33 mg/L of ceftiofur (ANT). Starter intake and fecal consistency scores of each calf were recorded on a daily basis. Body weight, withers height, body length, and heart girth were measured on d 1, 7, 14, 21, 28, and 35 before feeding in the morning. Rumen fluid was collected on d 15, 25, and 35 to determine ruminal pH, volatile fatty acids (VFA), and NH₃-N concentrations. A total of 10 (5 per treatment) samples of rumen fluid taken on d 35 were analyzed for microbial community. Rumen tissues from the cranial ventral sac and cranial dorsal sac were collected from 8 calves of each group for morphology analysis on d 35 after being harvested. The results showed that calves in 2 treatments had similar starter intake, body weight, withers height, body length, heart girth, and average daily gain. The ANT group showed a lower diarrhea frequency in wk 4, and no differences were found for other weeks. Calves in the ANT group exhibited a greater concentration of acetic acid in the rumen and no differences for other VFA, total VFA, rumen pH, or NH₃-N. As for rumen morphology, the length of papillae from cranial ventral sac of the ANT group was longer than that of the control group. The results of ruminal microbial community showed that antibiotic residues had minor effects on bacteria phyla and bacteria diversity. At the genus level, calves in the ANT group showed lower richness of Prevotella and higher richness of Acetitomaculum. In conclusion, antibiotic residues stimulated the development of ruminal papillae and increased the production of acetic acid in rumen, which might be caused by the influence of antibiotics on the ruminal microbial community.     

Replacing concentrates with a high-quality hay in the starter feed of dairy calves: II. Effects on the development of chewing and gut fermentation,and selected systemic health variables

Early development of the rumen, rumination, and fermentation is highly important in dairy calves. Yet, common rearing practices with feeding of concentrate-rich starters may jeopardize them because of lacking physically effective fiber (peNDF). The main objective of this study was to establish the influence of the composition of the calf starter feed (only forage with 2 different qualities or concentrate-rich starter diet) on chewing behavior, rumen development, rumen and hindgut fermentation, and selected systemic health and stress variables of dairy calves. The experiment was carried out with 40 newborn Holstein-Friesian calves, randomly assigned to 4 different solid feed treatments: MQH = 100% medium-quality hay (9.4 MJ metabolizable energy, 149 g of crude protein, and 522 g of neutral detergent fiber/kg of dry matter); HQH = 100% high-quality hay (11.2 MJ of metabolizable energy, 210 g of crude protein, 455 g of neutral detergent fiber/kg of dry matter); MQH+C = 30% MQH + 70% starter concentrate; HQH+C = 30% HQH + 70% starter concentrate). All calves were up to 14 wk in the trial and received acidified whole milk ad libitum in the first 4 wk of life, thereafter in reduced quantity until weaning on 12 wk of age. Water and the solid feed treatments were available ad libitum throughout the trial. Chewing activity was measured in wk 4, 6, 10, and 12 using RumiWatch halters. Until wk 3, rumen fluid, feces and blood were sampled weekly, thereafter every 2 wk. Rumen mucosal thickness (RMT) was measured on the same days with rumen fluid samples. Data showed that calves fed the HQH diet consumed more peNDF and this was associated with longer rumination time (591 min/d) and more ruminating boli (709 boli/d) than calves fed concentrate-rich diets (MQH+C: 430 min/d, 518 boli/d; HQH+C: 430 min/d, 541 boli/d), whereas the MQH group was intermediate (539 min/d, 644 boli/d). Ruminal and fecal pH were higher in calves fed only hay (especially MQH) compared with calves with concentrate supplementation. In both hay-fed groups, ruminal and fecal short-chain fatty acids were shifted toward acetate, whereas only the HQH diet increased the butyrate proportion in the ruminal short-chain fatty acids profile. Ruminal ammonia concentration was at a high level only in the first 3 wk and decreased thereafter. Feeding HQH tended to increase ruminal ammonia, likely because of its high crude protein content and ruminal degradability as well as lower assimilation from rumen microbes. The RMT similarly, though nonlinearly, increased in all groups over the course of the experiment. When using RMT as an indicator of rumen development in dairy calves in the practice, our data suggest an RMT of 1.7 mm and >2 mm at wk 5 and 10 of life, respectively. Feeding did not affect the blood levels of aspartate aminotransferase, gamma glutamyl transferase, glutamate dehydrogenase, and cortisol. In conclusion, feeding high-quality hay, instead of concentrate-rich starter feeds, resulted in improved rumination and ruminal fermentation profile, without affecting ruminal pH and systemic and stress health variables.     

Preweaning to postweaning rumen papillae structural growth,ruminal fermentation characteristics,and acute-phase proteins in calves

This study evaluated pre- to postweaning ruminal structural development, fermentation characteristics, and acute-phase protein levels in calves with a high milk replacer (MR) feeding rate prior to weaning. Six ruminally cannulated Holstein bull calves were fed MR (150 g/L) at 15% of body weight (BW) in 2 equal volumes daily. Volumes were adjusted weekly based on BW. Calves were weaned using a 1-step weaning method, with MR decreased by 50% at the end of wk 5 and full weaning at the end of wk 6. Calf starter, chopped straw, and water were offered ad libitum. Intake was recorded daily, and BW was recorded weekly. From wk 5 to 12, ruminal pH was continuously measured using a ruminal pH bolus. Ruminal fluid was collected weekly from wk 5 to 12 for measurement of short-chain fatty acid concentrations and quantification of total bacteria and protozoa. Rumen papillae were obtained at wk 5, 6, 7, 8, and 12 for histological analysis. Serum amyloid A and lipopolysaccharide-binding protein were measured weekly. Data were analyzed using GLIMMIX procedure of SAS (SAS Institute Inc., Cary, NC), with week as a fixed effect and calf as a random effect. During the weaning step-down, starter intake was 3-fold higher and continued to increase until wk 12. Body weight increased from birth to wk 12; however, BW did not change during wk 6, 7, and 8, possibly due to low metabolizable energy intake caused by the weaning strategy. Preweaning ruminal pH was below 5.8 for approximately 936.3 ± 125.99 min/d, implying ruminal acidosis. Furthermore, ruminal pH below 5.8 reached a peak at wk 8 with 1,203.9 ± 227.65 min/d below pH 5.8 and slowly decreased to 388.1 ± 189.82 min/d below pH 5.8 at wk 12. Papillae surface area, length, and width increased during wk 12 compared with wk 5. Corneum thickness increased by week, whereas spinosum/basale thickness only increased during wk 8 compared with wk 5. The acute-phase protein concentration was highest at wk 1 and then decreased and remained constant until wk 12. In conclusion, even before step-down weaning, calves experienced ruminal acidosis despite low starter intake. Further, the observed prolonged ruminal pH depression suggests that dietary rumen adaptation after weaning can take several weeks in calves with a high MR feeding rate preweaning. The prolonged depressed ruminal pH did not affect acute-phase proteins and this finding, along with the other results, suggests that rumen epithelium barrier integrity is not compromised during weaning.     

Fecal shedding and rumen growth of Escherichia coli O157:H7 in fasted calves.

Nine weaned calves aged from 8 to 12 weeks were fitted with rumen cannulas and were inoculated by cannula with 10(10) CFU of a five-strain mixture of nalidixic acid-resistant Escherichia coli O157:H7. Six calves were fasted for 48 h on days 15 and 16 and days 22 and 23 after inoculation. Samples of rumen contents and feces were obtained daily to enumerate E. coli O157:H7 populations and to determine rumen volatile fatty acid (VFA) concentrations and rumen pH. Fasting resulted in a marked decrease in rumen VFA concentrations from a mean of 135 mmol/liter before the fast to a mean of 35 mmol/liter during the second day of the fast. However, there was no correlation between daily VFA concentration and daily rumen or fecal numbers of E. coli O157:H7 in any of the calves. Fasting generally had no significant effect on the rumen or fecal numbers of E. coli O157:H7. The exception was a single fasted calf that experienced a 3-log(10) CFU/g increase in fecal shedding during and after the first fast. Despite the consistent changes in VFA concentrations in fasted calves, the fluctuations in rumen numbers of E. coli O157:H7 in the rumen of fasted calves were minimal. At the end of the experiment, E. coli O157:H7 was detected in either the rumen or omasum in two of three control calves at necropsy and in either the rumen or reticulum in five of six fasted calves. E. coli O157:H7 was detected in the colon in two of three control calves and in six of six fasted calves at necropsy. These results suggest that in cattle already shedding E. coli O157:H7, feed withdrawal and the associated changes in rumen pH and VFA concentrations have little effect on fecal shedding and rumen proliferation of E. coli O157:H7.     

Changes in gene expression in the rumen and colon epithelia during the dry period through lactation of dairy cows and effects of live yeast supplementation

The objectives of this study were (1) to use endoscopy to collect biopsies from the rumen and colon epithelia to describe changes in gene expression in these 2 tissues as cows move from a dry to a lactation ration and (2) to evaluate the potential influence that supplementation of live yeast could exert on these 2 epithelia. Twenty-one Holstein cows were split into 2 treatments and received either 300 g/d of corn containing 1 × 10¹⁰ cfu/d of live yeast (LY; n = 10) or 300 g/d of corn with no supplementation (control; n = 11) starting 21 ± 2.6 d (average ± SD) before until 21 d after calving. At 14 ± 2.6 d before the expected calving date, and exactly at 7 and 21 d after calving, rumen and colon biopsies were obtained from each cow using an endoscope. Total RNA was extracted from rumen and colon tissues, and the expression of IL10, TNFA, TLR4, IL1B, PCNA, MKI67, SGLT1, BAX, CASP3, OCLN, CLDN4, HSPA1A, HSPB1, DEFB1, and MCT1 (the latter only in rumen samples) was quantified by quantitative PCR. Overall, fluctuations in expression of the selected genes in the colon between the 2 stages of production and the 2 treatments were smaller than those found in the rumen. In the rumen epithelium, expression of TLR4 and DEFB1 was greatest before calving, with LY cows having a greater expression of TLR4 than control cows. Similarly, expression of IL10 was greatest in LY cows before calving. Expression of TNFA in the rumen epithelium of control cows was lowest at 21 DIM but in LY cows was kept steady among production stages. The expression of PCNA and MKI67 in the rumen epithelium was greatest at 7 DIM, indicating a high proliferation rate of this epithelium after calving. In the colon mucosa, expression of TLR4 and DEFB1 was greater than in the rumen, and DEFB1 expression was greater in LY cows than in control cows. The use of an endoscope allowed us to study the dynamics of rumen epithelium adaptation to increased supply of concentrate after calving, consisting of increased epithelia remodeling, reduction of the TLR4, and increased IL10 expression. Furthermore, the rumen epithelium of dry cows responded rapidly to live yeast, with changes in the expression of genes involved in the immune response becoming evident after 7 d of exposure to yeast. The expression of genes related to the immune response (mainly TLR4 and DEFB1) in the colon mucosa was greater than in the rumen, and the expression of DEFB1 was further stimulated by live yeast. It is concluded that the use of an endoscope allows the study of gene expression patterns in the rumen and hindgut epithelia. We report marked changes in the rumen wall and more modest changes in the colon when transitioning from a dry to a lactation ration. Furthermore, supplementation of live yeast fostered and increased expression of genes regulating inflammation and epithelial barrier in the rumen, and in the colon it increased the expression of DFEB1 coding for an antimicrobial peptide.     

Longitudinal assessment revealed the shifts in rumen and colon mucosal-attached microbiota of dairy calves during weaning transition

The objectives of this study were to investigate the shifts in rumen and colon mucosa-associated microbiota in dairy calves fed a high milk replacer feeding rate before and after weaning and to determine whether such shifts are associated with tissue physiological measures. Longitudinal biopsy was performed to collect rumen and colon mucosal tissues of 4 ruminally cannulated Holstein dairy bull calves (weaned at 6 wk of age) at the end of wk 5 (before weaning), 7 (weaning adaptation) and 12 (after weaning), and were used to assess mucosa-associated microbiota and their changes using amplicon sequencing. Both rumen and colon mucosa-associated bacterial communities shifted during the weaning process, as evidenced by their clear separation among 3 different weaning periods and increased α diversity (Shannon and Chao1 indices) during weaning transition. Among the 3 dominant bacterial phyla identified (relative abundance >1.0%), the relative abundance of Proteobacteria and Bacteroidetes decreased in the rumen mucosa, whereas the relative abundance of Firmicutes increased in both rumen and colon mucosa during weaning transition. In the rumen mucosa, Campylobacter (0.6–22.1%) gradually became prevalent during weaning transition, whereas Succinivibrio (6.2–10.3%) and Prevotella 1 (4.7–10.5%) were dominant regardless of weaning transition. In the colon mucosa, Bacteroides (12.8–25.4%) was dominant during weaning transition, although its relative abundance decreased after weaning. In the meantime, relative abundance of uncultured Lachnospiraceae increased from 2.2% to 25.7% during this period. In addition, genera Pyramidobacter (in the rumen mucosa) and Lachnoclostridium (in the colon mucosa) were positively correlated with rumen papilla surface area and colon mucosal thickness, respectively. Moreover, genera Ruminococcaceae UCG-005 and Sharpea in the rumen mucosa were positively correlated with the molar proportion of propionate and butyrate, respectively. Overall, our findings revealed that rumen and colon mucosa-associated bacterial communities altered in response to the weaning transition, and some bacterial taxa in these communities may have positive effects on rumen and colon mucosa development during this period.     

Form of calf diet and the rumen. I: Impact on growth and development

Preweaning diet is known to affect rumen tissue appearance at the gross level. The objectives of this experiment were to investigate effects of different preweaning diets on the growth and development of the rumen epithelium and on putative rumen epithelial stem and progenitor cell measurements at the gene and cell levels. Neonatal Holstein bull calves (n = 11) were individually housed and randomly assigned to 1 of 2 diets. The diets were milk replacer only (MRO; n = 5) or milk replacer with starter (MRS; n = 6). Diets were isoenergetic (3.87 ± 0.06 Mcal of metabolizable energy per day) and isonitrogenous (0.17 ± 0.003 kg/d of apparent digestible protein). Milk replacer was 22% crude protein, 21.5% fat (dry matter basis). The textured calf starter was 21.5% crude protein (dry matter basis). Water was available ad libitum and feed and water intake were recorded daily. Putative stem and progenitor cells were labeled by administering a thymidine analog (5-bromo-2′-deoxyuridine, BrdU; 5 mg/kg of body weight in sterile saline) for 5 consecutive days and allowed a 25-d washout period. Calves were killed at 43 ± 1 d after a 6 h exposure to a defined concentration of volatile fatty acids. We obtained rumen tissue from the ventral sac and used it for immunohistochemical analyses of BrdU (putative stem and progenitor cells) and Ki67 (cell proliferation), gene expression analysis, and morphological measurements via hematoxylin and eosin staining. Epithelial stem and progenitor cell gene markers of interest, analyzed by real-time quantitative PCR, were β1-integrin, keratin-14, notch-1, tumor protein p63, and leucine-rich repeat-containing G protein-coupled receptor 5. Body growth did not differ by diet, but empty reticulorumens were heavier in MRS calves (MRS: 0.67 ± 0.04 kg; MRO: 0.39 ± 0.04 kg). The percentage of label-retaining BrdU basale cells was higher in MRO calves than in MRS calves (2.0 ± 0.3% vs. 0.3 ± 0.2%, respectively). We observed a higher percentage of basale cells undergoing proliferation in MRS calves than in MRO calves (18.4 ± 2.6% vs. 10.8 ± 2.8%, respectively). Rumen epithelial gene expression was not affected by diet, but the submucosa was thicker in MRO calves and the epithelium and corneum/keratin layers were thicker in MRS calves. Presumptive stem and progenitor cells in the rumen epithelium were identifiable by their ability to retain labeled DNA in the long term, changed proliferative status in response to diet, and likely contributed to observed treatment differences in rumen tissue thickness.     

Effects of forage source and forage particle size as a free-choice provision on growth performance,rumen fermentation,and behavior of dairy calves fed texturized starters

We investigated the interactive effects of forage source and forage particle size (PS) as a free-choice provision on growth performance, rumen fermentation, and behavior of dairy calves fed texturized starters. Forty-eight Holstein calves (42 ± 3 kg of body weight) were randomly assigned (n = 12 calves per treatment) in a 2 × 2 factorial arrangement of treatments with the factors of forage source [alfalfa hay (AH) and wheat straw (WS)] and forage PS [(AH: medium = 1.96 mm or long = 3.93 mm) and (WS: medium = 2.03 mm or long = 4.10 mm), as geometric mean diameters]. The treatments were (1) AH with medium PS (AH-MPS), (2) AH with long PS (AH-LPS), (3) WS with medium PS (WS-MPS), and (4) WS with long PS (WS-LPS). Regardless of forage PS, the preweaning starter intake, dry matter intake, metabolizable energy intake, weaning body weight, and forage intake were greater for AH calves than WS calves. Average daily gain, average daily gain/metabolizable energy intake, feed efficiency, and final body weight of the calves did not differ among groups. An interaction of forage source and forage PS influenced acetate, propionate, and acetate-to-propionate ratio in the rumen on d 35, with the greatest acetate proportion and acetate-to-propionate ratio, but the least propionate proportion for AH-MPS calves than the other calves. The total volatile fatty acid concentration and the rumen proportions of propionate (d 70), butyrate (d 35), and valerate (d 35) were greater in AH-MPS calves than in AH-LPS calves. Calves fed AH had greater total volatile fatty acid concentration (d 35 and 70) and propionate proportion (d 70), but lesser ruminal proportions of butyrate (d 35 and 70), valerate (d 35 and 70), and acetate-to-propionate ratio (d 70) compared with calves fed WS. The ruminal valerate proportion (d 70) was greatest in WS-MPS calves than the other calves. An interaction of forage source and forage PS influenced preweaning standing time and starter eating time, with the least standing time for WS-MPS calves and the greatest eating starter time for AH-LPS calves. Calves fed AH spent less time for rumination, but devoted more time to non-nutritive oral behaviors than WS calves. Calves fed forage with long PS spent more time for rumination, eating forage, and spent less time lying and non-nutritive oral behaviors than medium PS. In conclusion, forage source and PS interacted, affecting behavior and rumen fermentation when calves were fed texturized starters. In addition, a desirable ruminal pH in dairy calves can be obtained with texturized starters.     

Growth performance,digestibility, blood metabolites,ruminal fermentation,and bacterial communities in response to the inclusion of gallic acid in the starter feed of preweaning dairy calves

The objective of this study was to evaluate the effects of feeding gallic acid on the growth, nutrient digestibility, plasma metabolites, rumen fermentation, and bacterial community in the rumen fluid and feces of preweaning calves. Thirty-six female Holstein calves with similar ages (means ± SD; 3.1 ± 1.39 d) and body weights (40.8 ± 2.87 kg) were randomly assigned to receive 3 treatments. Calves were fed 1 of 3 treatments as follows: basal diet with no gallic acid (control), 0.5 g/kg gallic acid in starter diet (low), and 1 g/kg gallic acid in starter diet (high). The results showed that feeding gallic acid increased growth by improving the starter intake and average daily gain of the calves. The fecal score tended to decrease in a linear manner with the addition of gallic acid. Total-tract apparent protein digestibility tended to increase linearly with feeding gallic acid. Feeding gallic acid led to a linear increase in the plasma total protein and β-hydroxybutyrate levels. In addition, feeding gallic acid linearly increased catalase and total antioxidant capacity levels and decreased malondialdehyde and tumor necrosis factor-α concentrations. The concentrations of total volatile fatty acids, propionate, butyrate, and valerate in the rumen fluid increased linearly with the addition of gallic acid, resulting in a linear pH reduction. Feeding gallic acid linearly increased the relative abundances of Prevotella_1, Saccharofermentans, and Prevotellaceae_UCG-001 and linearly decreased the relative abundance of Prevotella_7 in the rumen fluid. The Shannon index of ruminal bacterial communities linearly increased by feeding gallic acid. Feeding gallic acid linearly increased the relative abundances of Ruminococcaceae_UCG-005, Bacteroides, and Christensenellaceae_R-7_group in the feces. In summary, feeding gallic acid improved growth, antioxidant function, and rumen fermentation and altered the bacterial community in the rumen fluid and feces of preweaning dairy calves.     

Net flux of nutrients across the rumen wall of lactating dairy cows as influenced by dietary supplements of folic acid

The objective of the present study was to determine whether a dietary supplementation of folic acid, at levels used in our previous studies, would affect ruminal fermentation and the net flux of nutrients across the rumen wall of lactating dairy cows. Approximately 4 wk after calving, 5 lactating multiparous cows were surgically equipped with a ruminal cannula, an ultrasonic flow probe around the right ruminal artery, and indwelling catheters in the right ruminal vein and the ileocolic artery. Cows were fed a total mixed ration served in 7 equal meals per d (i.e., every 3.4 h). The experimental design was an unbalanced crossover arrangement with 3 periods of 4 wk each. The vitamin supplement, incorporated in equal amounts into each meal, was supplied at 0, 3, or 6 mg of folic acid per kg of BW per d. During the last week of each experimental period, blood samples were taken simultaneously from the 2 catheters every 30 min and rumen fluid was collected every 60 min during 2 consecutive meal intervals. Dietary supplementation with folic acid had no effect on milk production (27.2 ± 1.3 kg/d) or DMI (19.9 ± 0.7 kg/d), but milk concentrations and yields of total solids, fat, and protein increased linearly with increasing doses of folic acid ingested. Concentrations of folates in rumen fluid and arterial plasma, averaged over time, increased linearly with the dose of folic acid ingested but the net flux of folates across the rumen wall was not different from zero. Concentrations of butyrate in ruminal fluid decreased quadratically with the daily supply in folic acid. Dietary supplements of folic acid had no effect on pH and osmolality of ruminal fluid, nor on ruminal concentrations of lactate, ammonia, acetate, or propionate, total VFA, or microbial counts. The uptake of urea-N by the rumen wall tended to increase quadratically with the dose ingested but net fluxes of other nutrients were not affected by treatments. These results suggest that the effects of folic acid supplements on lactational performance cannot be explained by effects on rumen metabolism.