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Self-ordered sequencing is an important executive function involving planning and executing a series of steps to achieve goal-directed outcomes. The lateral frontal cortex is implicated in this behavior, but downstream striatal outputs remain relatively unexplored. We trained marmosets on a three-stimulus self-ordered spatial sequencing task using a touch-sensitive screen to explore the role of the caudate nucleus and putamen in random and fixed response arrays. By transiently blocking glutamatergic inputs to these regions, using intrastriatal CNQX microinfusions, we demonstrate that the caudate and putamen are both required for, but contribute differently to, flexible and fixed sequencing. CNQX into either the caudate or putamen impaired variable array accuracy, and infusions into both simultaneously elicited greater impairment. We demonstrated that continuous perseverative errors in variable array were caused by putamen infusions, likely due to interference with the putamen's established role in monitoring motor feedback. Caudate infusions, however, did not affect continuous errors, but did cause an upward trend in recurrent perseveration, possibly reflecting interference with the caudate's established role in spatial working memory and goal-directed planning. In contrast to variable array performance, while both caudate and putamen infusions impaired fixed array responding, the combined effects were not additive, suggesting possible competing roles. Infusions into either region individually, but not simultaneously, led to continuous perseveration. Recurrent perseveration in fixed arrays was caused by putamen, but not caudate, infusions. These results are consistent overall with a role of caudate in planning and flexible responding and the putamen in more rigid habitual or automatic responding., (Copyright © 2024 Gould et al.)

Much is known about the biomechanical performance of various types of suture anchors commonly used for labral fixation in the shoulder; however, similar studies in the hip are less common. We sought to compare all-suture and polyether ether ketone small-diameter anchors in the setting of labral repair during hip arthroscopy, with and without acetabuloplasty. We hypothesized that the biomechanical properties of the all-suture group when compared to polyether ether ketone anchors would be similar amongst native acetabula and significantly less following acetabuloplasty and that pullout forces would be reduced in the anterior and inferior regions of the acetabulum compared to the superior region. Bone density was measured in nine matched pairs of fresh-frozen cadaveric acetabula in the superior, anterosuperior, and anterior regions. Acetabuloplasty was performed in all three regions, while the contralateral acetabulum was left in situ as a control. Suture anchors were placed such that one each of two different types was placed within each region. Specimens were tested in cyclic fatigue and loaded to failure. The all-suture group had significantly higher cyclic displacement compared to the polyether ether ketone, but there was no significant difference in ultimate load, regardless of acetabuloplasty. Amongst all non-resected specimens, the lowest bone density was observed consistently in the inferior region. Our results indicate that, with or without acetabuloplasty, a small-diameter polyether ether ketone anchor appears to be more stable than an all-suture anchor, which needs to be set first., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: All authors declare study sponsorship from Smith & Nephew in the form of direct funding as well as in-kind donation of cadaveric specimens. Dr. Emblom declares payments unrelated to the study from Arthrex for royalties, consulting, and other services. Dr. Walters declares payments unrelated to the study from Arthrex for non-consulting services. Dr. Ryan declares payments unrelated to the study from Arthrex for non-consulting services. This does not alter our adherence to PLOS ONE policies on sharing data and materials., (Copyright: © 2023 Emblom et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Neurones are highly polarized cells with extensive axonal projections that rely on transport of proteins, RNAs, and organelles in a bidirectional manner to remain healthy. This process, known as axonal transport, can be imaged in real time through epifluorescent imaging of fluorescently labeled proteins, organelles, and other cargoes. While this is most conveniently done in primary neuronal cultures, it is more physiologically relevant when carried out in the context of a developed nerve containing both axons and glia. Here we outline how to image axonal transport ex vivo in sciatic and optic nerves, and the fimbria of the fornix. These methods could be altered to image other fluorescently labeled molecules, as well as different mechanisms of intracellular transport., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Activation of the pro-degenerative protein SARM1 after diverse physical and disease-relevant injuries causes programmed axon degeneration. Original studies indicate that substantially decreased SARM1 levels are required for neuroprotection. However, we demonstrate, in Sarm1 haploinsufficient mice, that lowering SARM1 levels by 50% delays programmed axon degeneration in vivo after sciatic nerve transection and partially prevents neurite outgrowth defects in mice lacking the pro-survival factor NMNAT2. In vitro, the rate of degeneration in response to traumatic, neurotoxic, and genetic triggers of SARM1 activation is also slowed. Finally, we demonstrate that Sarm1 antisense oligonucleotides decrease SARM1 levels by more than 50% in vitro, which delays or prevents programmed axon degeneration. Combining Sarm1 haploinsufficiency with antisense oligonucleotides further decreases SARM1 levels and prolongs protection after neurotoxic injury. These data demonstrate that axon protection occurs in a Sarm1 gene dose-responsive manner and that SARM1-lowering agents have therapeutic potential, making Sarm1-targeting antisense oligonucleotides a promising therapeutic strategy., Competing Interests: Declaration of interests K.L., P.J.-N., and F.R. are employees of Ionis Pharmaceuticals, Inc., and part of this study was jointly funded by Takeda (Cambridge) Ltd., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Chemotherapy-induced peripheral neuropathy (CIPN) is a common dose-limiting side effect of cancer treatment, often associated with degeneration of sensory axons or their terminal regions. Presence of the slow Wallerian degeneration protein (WLD S ), or genetic deletion of sterile alpha and TIR motif containing protein 1 (SARM1), which strongly protect axons from degeneration after injury or axonal transport block, alleviate pain in several CIPN models. However, oxaliplatin can cause an acute pain response, suggesting a different mechanism of pain generation. Here, we tested whether the presence of WLD S or absence of SARM1 protects against acute oxaliplatin-induced pain in mice after a single oxaliplatin injection. In BL/6 and Wld S mice, oxaliplatin induced significant mechanical and cold hypersensitivities which were absent in Sarm1 -/- mice. Despite the presence of hypersensitivity there was no significant loss of intraepidermal nerve fibers (IENFs) in the footpads of any mice after oxaliplatin treatment, suggesting that early stages of pain hypersensitivity could be independent of axon degeneration. To identify other changes that could underlie the pain response, RNA sequencing was carried out in DRGs from treated and control mice of each genotype. Sarm1 -/- mice had fewer gene expression changes than either BL/6 or Wld S mice. This is consistent with the pain measurements in demonstrating that Sarm1 -/ - DRGs remain relatively unchanged after oxaliplatin treatment, unlike those in BL/6 and Wld S mice. Changes in levels of four transcripts - Alas2, Hba-a1, Hba-a2, and Tfrc - correlated with oxaliplatin-induced pain, or absence thereof, across the three genotypes. Our findings suggest that targeting SARM1 could be a viable therapeutic approach to prevent oxaliplatin-induced acute neuropathic pain., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Previous research indicated that phytase may release less phosphorus (P) from phytate when it is evaluated using diets with P levels above requirement as compared with diets below requirement. The objectives of this experiment were to further test the hypothesis that the P release values determined for phytase are higher when pigs are fed diets that are deficient (DE) in P compared with when they are fed diets that are adequate (AD) in P, and that phytase will increase the digestibility of dry matter (DM), gross energy (GE), nitrogen (N), and calcium (Ca) independent of dietary P status. Twenty-four barrows (body weight: 23.2 ± 1.8 kg) were randomly assigned to one of eight dietary treatments and housed in individual pens for 21 d and then moved to metabolism crates for 9 d, with the collection of urine and feces occurring on the final 5 d. A basal corn-soybean meal diet (P-AD) was formulated at 0.36% standardized total tract digestible (STTD) P and total calcium:STTD P (Ca:STTD P) of 2:1. A P-DE diet was also formulated to maintain a constant Ca:STTD P of 2:1 in both basal diets. Phytase was added to AD and DE diets at 350, 600, 1,000 phytase units (FYT)/kg. Pig was the experimental unit; diet (P-AD or P-DE), phytase level, and replicate were fixed effects. Orthogonal polynomial contrasts were used to test linear and quadratic effects of phytase within P-AD and P-DE diets. Phytase improved apparent total tract digestibility (ATTD) and STTD of P in both P-AD (linear P < 0.001) and P-DE diets (quadratic P < 0.001). Estimates for STTD P release were 0.07%, 0.09%, and 0.09% for 350, 600, and 1,000 phytase units (FYT)/kg in P-DE diets, and 0.02%, 0.03%, and 0.05% in P-AD diets, respectively. In P-DE diets, phytase improved absorption and retention of P and increased urinary excretion of P (quadratic P < 0.001). In P-AD diets, phytase improved absorption of P (linear P = 0.066), tended to improve retention (linear P = 0.066), and increased urinary excretion of P (quadratic P = 0.021). Phytase improved ATTD of Ca in P-DE diets (quadratic P = 0.002) but not in P-AD diets (P > 0.1). In conclusion, the release of P by phytase is lower in diets that are AD in P than those which are DE. Phytase increased the availability of Ca only in the diets DE in P. Finally, phytase increased the ATTD of DM and tended to increase the ATTD of energy, independent of dietary P status., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Society of Animal Science.)

The objective of this experiment was to evaluate the growth performance and bone mineral content (BMC) of nursery pigs in response to increasing total calcium (Ca) to available phosphorus (aP) ratios in diets containing phytase (250 FTU/kg; Natuphos E, BASF, Florham Park, NJ). A total of 480 nursery pigs (body weight (BW) = 5.7 ± 0.6 kg) with 10 pigs per pen and 7 pens per treatment (6 pens fed 2.75:1 diet) were allotted to seven treatments consisting of increasing ratios of calcium to available phosphorus (Ca:aP): 1.25, 1.50, 1.75, 2.00, 2.25, 2.50, and 2.75. From day -7 to 0, pigs were fed a common diet. They were then fed the treatment diets during two experimental phases from day 1 to 14 and 15 to 28, respectively. Available P was formulated to 0.33% and 0.27% (approximately 90% of requirement) in dietary phases 1 and 2, respectively. BW, average daily gain (ADG), average daily feed intake (ADFI), and gain-to-feed ratio (G:F) were determined. BMC of the femur was measured on day 28 on one pig per pen using dual x-ray absorptiometry. Data were analyzed as a linear mixed model using PROC MIXED (SAS, 9.3). Orthogonal polynomial contrasts were used to determine the linear and quadratic effects of increasing the Ca:aP. Over the 28-d experimental period, increasing Ca:aP resulted in a linear decrease in ADG (353, 338, 328, 304, 317, 291, and 280 g/d; P < 0.01), ADFI (539, 528, 528, 500, 533, 512, and 489 g/d; P < 0.05), and G:F (0.68, 0.66, 0.64, 0.62, 0.61, 0.59, and 0.58; P < 0.01). Increasing Ca:aP also resulted in decreased BW on days 14 and 28 (P < 0.01). The BMC of the femur decreased with increasing Ca:aP (6.2, 6.3, 5.7, 5.9, 5.5, 5.6, and 5.3 g; P < 0.05). Regression analysis explained the impact of Ca:aP as follows on ADG (ADG [g/d] = 339 - 36x; r2 = 0.81), G:F (G:F = 0.61 - 0.03x; r2 = 0.72), and BMC (BMC [g] = 6.4 - 0.27x; r2 = 0.43), where x is the Ca:aP. In conclusion, all outcomes indicated that any level of calcium above the minimum used in this experiment impaired growth performance and skeletal development. Further research using even lower levels of dietary Ca is warranted., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science.)

The objective of this study was to determine the impact of reducing the mean particle size (PS) of corn distillers dried grains with solubles (DDGS) with a hammermill (HM) or with a roller mill (RM) on the apparent total tract digestibility (ATTD) of dry matter (DM), gross energy (GE), N, acid hydrolyzed ether extract (AEE), and fiber components in growing and finishing pigs. Twenty-four growing barrows were housed in individual pens and were randomly assigned to a 3 × 2 factorial design ( n = 8): three grinding methods [either corn DDGS ground with an HM to a PS of 450 μm; corn DDGS ground with an RM to a PS of 450 μm; and corn DDGS with a PS of 670 μm (not further ground)] and two body weight (BW) periods (growing pigs with an average initial BW of 54.7 ± 0.9 kg, and finishing pigs with an average initial BW of 107.8 ± 1.5 kg BW). Fecal samples were collected for each BW period in the last 3 d of an 11-d feeding period. Titanium dioxide was used as an indigestible marker. Digestibility data were analyzed using the MIXED procedure of SAS. Results showed that finishing pigs tended to have better ATTD of DM than growing pigs ( P = 0.09) and had increased ATTD of GE and N than growing pigs ( P = 0.03 and P < 0.01, respectively). On the other hand, growing pigs had better ATTD of AEE than finishing pigs ( P = 0.01). Pig BW period did not affect the ATTD of neutral detergent fiber (NDF), acid detergent fiber (ADF), and hemicellulose. Reducing the mean PS of corn DDGS with either HM or RM (from 670 to 450 µm) improved the ATTD of DM and GE ( P < 0.01 and P < 0.01), tended to improve the ATTD of N ( P = 0.08), and improved the ATTD of AEE ( P < 0.01). No effect of reducing PS was observed for the ATTD of NDF, ADF, or hemicellulose. There were no differences between HM and RM in any of the ATTD variables tested. In conclusion, reducing PS of corn DDGS from 670 to 450 μm either with an HM or with an RM improved the digestibility of DM, GE, and AEE and modestly improved the digestibility of N in growing and finishing pigs. However, reducing the PS of corn DDGS did not affect the digestibility of fiber components., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science.)

The objective of this experiment was to determine the impact of diets containing increasing amounts of enzymatically treated soybean meal (ESBM) but decreasing amounts of soybean meal (SBM) on growth performance, intestinal structure, and barrier integrity, inflammation, and oxidative status in weaned pigs. A total of 480 pigs [6.3 ± 1.2 kg body weight (BW)] were blocked by initial BW and pens ( n = 12 per treatment) were randomly allotted to one of four dietary treatments. Diets were fed in three phases (days 0-14, 14-28, and 28-35) over a 35-d period. The four dietary treatments consisted of a negative control diet (NC): the NC with 7.0% ESBM (ESBM1), the NC with 14.0% ESBM (ESBM2), and the NC with 21.0% ESBM (ESBM3). Soybean meal was reduced proportionately in each treatment. In phase 2, ESBM inclusion was decreased by 50% (3.5%, 7.0%, and 10.5% ESBM, respectively); phase 3 was a common diet and contained no ESBM. Fecal score was visually ranked weekly using a four-point scale. Intestinal tissue, digesta, and blood samples were collected from 48 pigs (1 per pen) on day 10. Data were analyzed using PROC MIXED of SAS (9.4) with pen as the experimental unit; diet and block were considered fixed effects. Linear and quadratic contrasts were used to determine the effect of increasing ESBM. Overall, ESBM2 and ESBM3 decreased final BW, average daily gain, and average daily feed intake compared to NC and ESBM1 (diet, P < 0.05; linear, P < 0.05). Overall fecal score (diet, P < 0.05) and fecal dry matter ( P < 0.05) were improved by feeding ESBM diets compared to NC. Volatile fatty acid (VFA) concentration of acetate, propionate, butyrate, and total VFA in ileal contents increased as ESBM inclusion increased ( P < 0.05). Colonic VFA concentration was not impacted ( P > 0.10). Total antioxidant capacity was increased by ESBM ( P < 0.05). The concentration of mucosal interleukin-4 increased as the inclusion of ESBM increased (linear, P < 0.05). Messenger ribonucleic acid abundance of occludin and zonula-occludens-1 in ileal tissue was increased by ESBM1 or ESBM2 ( P < 0.05). In conclusion, increasing the dietary levels of ESBM over 7% had a negative impact on nursery pig performance, but ESBM positively impacted fecal score. Feeding ESBM improved oxidative status and intestinal barrier integrity while increasing ileal VFA production but had minimal impact on intestinal inflammation or morphology. Further research is needed to determine the optimal inclusion level of ESBM., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science.)

The objective of this experiment was to compare the effects of spray-dried plasma protein (SDPP) and dried egg protein (DEP), without (AB-) or with (AB+) in-feed antibiotics, on growth performance and markers of intestinal health in nursery pigs raised in commercial conditions. This 42-d experiment utilized 1,230 pigs (4.93 ± 0.04 kg body weight; approximately 15-18 d of age). Pigs were randomly assigned to one of six dietary treatments that were arranged as a 2 × 3 factorial of in-feed antibiotics (AB- vs. AB+) and a specialty protein additive (none [CON], porcine SDPP, or DEP). Diets were fed in four phases with phases 3 and 4 as a common diet across all treatments. Specialty protein additives were fed in phases 1 (0-13 d; 3% SDPP, and 0.20% DEP) and 2 (13-26 d; 2% SDPP, and 0.10% DEP). Antibiotics were fed in phases 1-3 (662 mg chlortetracycline [CTC]/kg, 28 mg carbadox/kg, and 441 mg CTC/kg, respectively). Ileal tissue and blood samples were collected from 48 pigs (8 per treatment) on d 20. Data were analyzed using PROC MIXED of SAS (9.4) with pen as the experimental unit; protein additives, antibiotics, and their interaction were fixed effects and block was a random effect. The pigs experienced naturally occurring health challenges in weeks 2 and 4. In the AB- diets, SDPP and DEP increased average daily gain (ADG; P = 0.036) and average daily feed intake (ADFI; P = 0.040) compared to CON; in the AB+ diets, neither SDPP nor DEP increased ADG or ADFI compared to CON but SDPP did increase these parameters over DEP. The SDPP and DEP diets decreased the number of individual medical treatments compared to CON ( P = 0.001). The AB+ increased ileal mucosal interleukin (IL)-1 receptor antagonist ( P = 0.017). Feeding DEP reduced the concentration of mucosal IL-1β compared to CON, but not SDPP ( P = 0.022). There was a trend for SDPP and DEP to increase villus height:crypt depth compared to CON ( P = 0.066). Neither antibiotics or protein additive affected serum malondialdehyde concentration or ileal mRNA abundance of claudin-3 or 4 , occludin , or zonula occludens-1 ( P > 0.10). In conclusion, SDPP and DEP improved growth performance of weaned pigs in the absence of antibiotics but neither improved growth compared to CON when feeding standard antibiotic levels. The specialty proteins had a positive effect on health; specialty proteins and antibiotics were able to modulate some markers of intestinal inflammation and morphology., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science.)

Post-weaning diarrhea caused by enterotoxigenic E. coli (ETEC) causes significant economic losses for pig producers. This study was to test the hypotheses that an ETEC challenge disrupts intestinal microbial homeostasis and the inclusion of dietary soluble (10% sugar beet pulp) or insoluble fiber (15% corn distillers dried grains with solubles) with or without exogenous carbohydrases will protect or restore the gut microbial homeostasis in weaned pigs. Sixty crossbred piglets (6.9 ± 0.1 kg) were blocked by body weight and randomly assigned to one of six treatments ( n = 10), including a non-challenged control (NC), ETEC F18-challenged positive control (PC), ETEC-challenged soluble fiber without (SF-) or with carbohydrases (SF+), and ETEC-challenged insoluble fiber without (IF-) or with carbohydrases (IF+). Pigs were housed individually and orally received either ETEC inoculum or PBS-sham inoculum on day 7 post-weaning. Intestinal contents were collected on day 14 or 15. The V4 region of the bacterial 16S rRNA was amplified and sequenced. High-quality reads (total 6,671,739) were selected and clustered into 3,330 OTUs. No differences were observed in α-diversity among treatments. The ileal microbiota in NC and PC had modest separation in the weighted PCoA plot; the microbial structures were slightly altered by SF+ and IF- compared with PC. The PC increased ileal Escherichia-Shigella ( P < 0.01) and numerically decreased Lactobacillus compared to NC. Predicted functional pathways enriched in the ileal microbiota of PC pigs indicated enhanced activity of Gram-negative bacteria, in agreement with increased Escherichia-Shigella . The SF+ tended to decrease ( P < 0.10) ileal Escherichia-Shigella compared to PC. Greater abundance of ileal Streptococcus , Turicibacter , and Roseburia and colonic Prevotella were observed in SF- and SF+ than PC ( P < 0.05). Pigs fed IF + had greater Lactobacillus and Roseburia than PC pigs ( P < 0.05). The ETEC challenge reduced total volatile fatty acid (VFA) compared with NC ( P < 0.05). The SF+ tended to increase ( P < 0.10) and SF- significantly increased ( P < 0.05) colonic total VFA compared with PC. Collectively, ETEC challenge disrupted gut microbial homeostasis and impaired microbial fermentation capacity. Soluble fiber improved VFA production. Dietary fiber and carbohydrases altered microbiota composition to maintain or restore microbial homeostasis., (Copyright © 2020 Li, Peng, Burrough, Sahin, Gould, Gabler, Loving, Dorman and Patience.)

Feed grains are processed to improve their value in pig diets by exposing kernel contents to enzymatic and microbial action. The objective of this study was to quantify the effect of reducing mean particle size (PS) of wheat grain ground with two different grinding methods (GMs) on the apparent total tract digestibility (ATTD) of nutrients and energy in growing and finishing pigs. Forty-eight barrows were housed in individual pens for 11 d for two periods. Pigs were randomly assigned to a 3 × 2 × 2 factorial experimental design: three target mean PS of wheat grain (300, 500, and 700 µm), two GMs (roller mill and hammermill), and two body weight (BW) periods (growing period; initial BW of 54.9 ± 0.6 kg and finishing period; initial BW of 110.7 ± 1.4 kg). Diets contained one of six hard red wheat grain samples, vitamins, minerals, and titanium dioxide as an indigestible marker. Feed allowance provided 2.5 (for the two lightest pigs in each treatment) or 2.7 (for the remaining six pigs in each treatment) times the estimated daily maintenance energy requirement for each growth stage. Fecal samples were collected for the last 3 d of each period. Data were analyzed as a linear mixed model with pig as a random effect and PS, GM, and BW period and their interactions as fixed effects utilizing the MIXED procedure of SAS. Growing pigs had greater ( P < 0.05) ATTD of dry matter (DM), gross energy (GE), N, acid hydrolyzed ether extract (AEE), and neutral detergent fiber (NDF) by lowering mean PS from 700 to 500 μm using either a roller mill or a hammermill. However, digestibility did not increase when PS was reduced from 500 to 300 μm, except for AEE ( P < 0.05). Finishing pigs had greater ATTD of DM, GE, N, AEE, and NDF by lowering mean PS with a hammermill from 700 to 500 μm ( P < 0.05), but it was greater for 500 μm than for 300 μm ( P < 0.05). Using a roller mill reduced the ATTD of DM and NDF by lowering PS from 700 to 300 μm ( P < 0.05). The ATTD of GE decreased by lowering PS from 700 to 500 μm with a roller mill ( P < 0.05) for finishing pigs. The ATTD of N and AEE for finishing pigs were similar from 700 to 300 μm when ground by a roller mill. These data suggest that the PS that maximized digestibility for a hammermill is 500 μm for both growing and finishing pigs. However, for the roller mill, the PS resulting in the best digestibility were 500 and 700 μm for growing and finishing pigs, respectively., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science.)

The objective of this experiment was to investigate the impact of an F18 enterotoxigenic Escherichia coli (ETEC) challenge on growth performance, aspects of intestinal function, and selected immune responses of piglets, as well as to evaluate potential protective effects of direct-fed microbial (DFM) blends. Seventy-two weaned piglets (6.4 ± 0.2 kg body weight [BW]; ~21 d of age) were assigned to one of four treatments: 1) NC: Nonchallenged (n = 10), 2) positive challenged control (PC): F18 ETEC-challenged (n = 10), 3) PC + DFM1 (n = 8; three strains of Bacillus amyloliquefaciens; 7.5 × 105 colony-forming units [cfu]/g), or 4) PC + DFM2 (n=8; 2 strains of B. amyloliquefaciens and one strain of Bacillus subtilis; 1.5 × 105 cfu/g). Feed intake and BW were recorded on day 0, 7, and 17. Pigs were sham-infected either with 6 mL phosphate-buffered saline or inoculated with 6 mL F18 ETEC (~1.9 × 109 cfu/mL) on day 7 (0 d postinoculation [dpi]). All ETEC-challenged pigs were confirmed to be genetically susceptible to F18. Pigs had ad libitum access to feed and water throughout the 17-d trial. Fecal scores were visually ranked and rectal temperatures were recorded daily. To evaluate ETEC shedding, fecal swabs were collected on dpi 0, 1, 2, 3, 5, 7, and 10. Blood samples were collected on dpi 0, 1, 2, 4, 7, and 10. Ileal tissues were collected at necropsy on dpi 10. All challenged treatments had lower final BW, decreased average daily gain (ADG), and average daily feed intake (ADFI) during the 10-d postchallenge period (P < 0.01). The DFM2 treatment increased E. coli shedding on dpi 2 and decreased iton dpi 7 (P < 0.05) compared with the PC. Rectal temperature decreased across all challenged treatments (P < 0.01). Ileal mRNA abundance of occludin (OCLN) and zonula occludens-1 (ZO-1) decreased in PC and DFM1 compared with NC (P < 0.05). Pigs fed DFM2 had intermediate ileal mRNA abundance of OCLN and increased ZO-1 mRNA compared with pigs in PC (P < 0.05). Interleukin 8 (IL-8) increased in the plasma of PC and DFM2 on dpi 2 compared with NC (P < 0.05). Mucosal IL-8 increased in PC compared with NC (P < 0.05). All challenged treatments tended to have elevated tumor necrosis factor-α (TNF-α) mRNA abundance compared with NC (P < 0.10). Challenged pigs had reduced secretory immunoglobulin A and villus height compared with NC pigs (P < 0.05). The impact of an ETEC challenge on intestinal function and the immune system has been revealed, information critical to developing improved treatment regimes., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science.)

The experimental objective was to determine the role of mean particle size (PS), grinding method, and body weight (BW) category on nutrient, fiber, and energy digestibility of corn. A total of 48 barrows were housed in individual pens and randomly assigned to one of six dietary treatments for 11 d at two BW categories (55 kg and 110 kg). The six treatments consisted of corn ground at three different targeted mean PSs (300, 500, and 700 µm) using either a roller mill or a hammermill. Fecal samples were collected for the last 3 d of each feeding period. Titanium dioxide was used as an indigestible marker. Digestibility data were analyzed as a linear mixed model using the MIXED procedure of SAS. Finishing pigs had greater apparent total tract digestibility (ATTD) of dry matter (DM), gross energy (GE), and N than growing pigs ( P = 0.02, P = 0.01, and P <0.01, respectively). The ATTD of DM, GE, and N was similar in pigs fed hammermilled corn across all PS treatments. However, in roller-milled corn, they increased as PS was reduced ( P < 0.05). The ATTD of acid-hydrolyzed ether extract (AEE) in growing pigs was similar between corn ground at 700 and 500 µm, but it was increased by further reducing PS to 300 µm ( P < 0.05). In finishing pigs, the ATTD of AEE increased as mean PS decreased from 700 to 300 µm ( P < 0.05). The ATTD of AEE was similar in hammermilled corn at all three PS treatments. On the other hand, the ATTD of AEE was similar in corn ground in a roller mill to 700 and 500 µm, but it increased when PS was reduced to 300 µm ( P < 0.05). In conclusion, reducing PS of corn with a roller mill increased digestibility of energy and nutrients, but there was less effect using a hammermill. It is possible that differences in SD, distribution, chemical composition, and the shape of the particles resulting from the two grinding processes help to explain the different response., (© The Author(s) 2019. Published by Oxford University Press on behalf of the American Society of Animal Science.)

The objective of this study was to evaluate the relative bioavailability (RBV) of l-Lys sulfate in comparison to l-Lys HCl based on the growth performance response from approximately 26 to 48 kg and from approximately 68 to 114 kg. The effect of Lys source on blood urea nitrogen (BUN), digestibility of dry matter (DM) and sulfur (S), as well as carcass characteristics was determined. A total of 280 growing pigs (25.9 ± 0.25 kg BW) were randomly assigned to one of seven dietary treatments in 56 pens, with five pigs per pen. The diets included a Lys-deficient basal diet (65% of requirement) and the basal diet supplemented with three graded levels of Lys (75%, 85%, and 95% of requirement), as either l-Lys HCl (78.8% Lys purity) or l-Lys sulfate (54.6% Lys purity). The experiment lasted for 112 d, with four dietary phases: Phase 1 lasted for 4 wk (BW: 25.9 to 47.5 kg), Phase 2 lasted for 3 wk (common commercial diet as washout period), Phase 3 lasted for 5 wk (BW: 67.5 to 98.2 kg), and Phase 4 lasted for 3 or 4 wk to reach an average market weight of 114.2 kg. Fresh fecal samples of pigs fed the highest levels of Lys (both Lys sources) were collected on 7 to 10 days after the beginning of Phase 3 for digestibility assay. Blood samples were collected on day 21 and day 81 to determine BUN. Carcass data were collected at a commercial packing plant. Data were analyzed using PROC GLM of SAS (9.4) according to a completely randomized design with pen as the experimental unit. The RBV of l-Lys sulfate was determined using the multiple regression slope-ratio method. Increasing levels of Lys, independent of source, increased ( P < 0.05) BW, ADG, and feed efficiency during Phases 1, 3, and 4; market BW increased linearly ( P < 0.01) and backfat and BUN decreased linearly ( P < 0.01). Lysine source had no impact on growth performance, carcass characteristics, BUN, or digestibility of S and DM. The RBV of l-Lys sulfate compared with l-Lys HCl was also not different based upon ADG or G:F during Phase 1, 3, or 4. These data suggest that the bioavailability of Lys in l-Lys sulfate and l-Lys HCl is at least equivalent for growing-finishing pigs., (© The Author(s) 2019. Published by Oxford University Press on behalf of the American Society of Animal Science.)

This study aimed to evaluate the effects of a source of dietary soluble (SF) and insoluble fiber (IF) without or with exogenous carbohydrases (xylanase, β-glucanase, and pectinase) on diarrhea incidence, selected immune responses, and growth performance in enterotoxigenic Escherichia coli (ETEC)-challenged pigs. Sixty weaned pigs (6.9 ± 0.1 kg BW, ~23 d of age) were blocked by initial BW and placed in individual pens. Pens were randomly assigned to one of six treatments (n = 10 per treatment), including a nonchallenged control (NC), a positive challenge control (PC), the PC + a soluble fiber diet (10% sugar beet pulp) without (SF-) or with carbohydrases (SF+), and PC + an IF diet (15% corn distillers dried grains with solubles) without (IF-) or with carbohydrases (IF+). The control diet was primarily based on corn and soybean meal with 13.5% whey powder. The two sources of fiber were added at the expense of cornstarch in the control diet. Pigs were orally inoculated with 6 mL hemolytic F18 ETEC (~3.5 × 109 cfu/mL) or sham infected with 6 mL phosphate-buffered saline on day 7 (0 d postinoculation, dpi) postweaning. All ETEC challenged pigs were confirmed to be genetically susceptible to F18 ETEC. Pigs had free access to feed and water throughout the 14-d trial. Pig BW and feed intake were recorded on dpi -7, 0, and 7 or 8. Fecal swabs were collected on dpi -7, 0, 1, 2, 3, 5, and 7 or 8 to evaluate hemolytic E. coli shedding. Fecal score was visually ranked daily postchallenge to evaluate diarrhea incidence. Blood samples were collected on dpi -1, 3, and 7 or 8 at necropsy and intestinal tissues were collected at necropsy. Pigs on PC had lower dpi 1 to 7 ADG and ADFI than those on NC (P < 0.05). Compared with PC pigs, SF+ pigs had greater ADG during both pre- and postchallenge period (P < 0.05). The IF- increased postchallenge diarrhea incidence compared with PC (P < 0.05). Pigs on SF- had lower ileal E. coli attachment than PC (P < 0.05). The SF+ reduced haptoglobin and IF+ reduced C-reactive protein on dpi 3 compared with PC (P < 0.05). Compared with PC pigs, SF+ pigs tended to have lower ileal tumor necrosis factor alpha and greater ileal occludin (OCLN) mRNA (P < 0.10) and had greater (P < 0.05) colonic OCLN mRNA levels. Collectively, IF- increased incidence of diarrhea and fecal E. coli shedding compared with PC. The SF+ pigs had improved growth compared with PC pigs, likely due in part to a reduction in inflammatory intermediates., (© The Author(s) 2019. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Although the impact of carbohydrases on performance and nutrient utilization has been well studied, their effects on immune status and intestinal microbiota are less known in pigs. This study aimed to evaluate the impact of xylanase (X) and a carbohydrase enzyme blend (EB; cellulase, ß-glucanase, and xylanase) on the immune profile of the intestine and peripheral system as well as intestinal microbes and microbial metabolites of weaned pigs fed higher fiber diets. Pigs (n = 460; 6.43 ± 0.06 kg BW; F25 × 6.0 Genetiporc) were blocked by initial BW. Pens (n = 48; 12 per treatment; 9 or 10 pigs per pen) were randomly assigned to 1 of 4 dietary treatments, including a higher fiber control diet (CON) and the CON supplemented with 0.01% X, 0.01% EB, or both enzymes (X + EB), arranged in a 2 × 2 factorial. The diets were based on corn, soybean meal, corn distillers dried grains with solubles, and wheat middlings. After 7-d adaptation to the environment, pigs were fed experimental diets ad libitum for 28 d. Blood samples were collected from the same pig within each pen on days 0, 7, 14, and 28. Intestinal tissues and digesta were collected on day 28. Bacteria 16S rRNA gene copy numbers were quantified using qPCR. The mRNA levels of colonic IL-17, occludin (OCLN), and claudin 3 (CLDN3) were greater in pigs fed diets with X + EB, but not X or EB, compared with those fed CON (P < 0.05). The EB in the diet reduced plasma IL-8 over the 28-d trial compared with diets without EB (P < 0.05). There was an X × EB interaction on plasma tumor necrosis factor α and IL-1ß (P < 0.05); their levels were decreased when X and EB were added together, but not individually, compared with CON. The EB decreased cecal propionate, butyrate, and total volatile fatty acids (P < 0.05). Pigs fed X had lower ileal Lactobacillus and greater ileal and cecal Enterobacteriaceae compared with those fed unsupplemented diets (P < 0.05). The EB decreased Lactobacillus (P < 0.05) and tended to decrease (P = 0.065) Enterobacteriaceae in the colon compared with diets without EB. In conclusion, the addition of X and EB together decreased systemic markers of immune activation, potentially diverting energy and nutrients towards growth. The EB reduced colonic Lactobacillus and cecal total volatile fatty acids, probably due to improved prececal fiber and starch degradation and thus reduced substrate availability in the large intestine. These data corroborated previously observed enhanced growth in pigs fed EB-supplemented diets., (© The Author(s) 2018. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Microbial phytase is widely used to enhance digestibility of phytate-P. By tradition, diets with P content well below requirement are used to quantify phytate-P release by phytase, but P-adequate diets may be more physiologically relevant. The objective of this study was to investigate the effects of phytase on P digestion and metabolism and develop a P release curve for phytase in P-adequate diets (above requirement according to NRC, 2012), and to compare these effects in a P-deficient diet. Three replicates of 24 barrows each (BW = 23.0 ± 1.8 kg) were randomly assigned to 1 of 8 dietary treatments, housed in individual pens for 21 d, then moved to metabolism crates for 5 d urine and fecal collections. A basal corn-soybean meal diet (P-adequate, A) was formulated at 0.36% standardized total tract digestible (STTD) P and total Ca:STTD P of 1.83. Phytase was added to A at 200 (A200), 400 (A400), 600 (A600), and 800 (A800) phytase units (FTU)/kg. A positive control diet (PC) was formulated using monocalcium phosphate (MCP) to increase STTD P by 0.16% to 0.52%, the expected STTD P release of 800 FTU/kg. A P-deficient diet (D) was formulated by reducing MCP to achieve 0.21% STTD P, and 200 FTU phytase/kg was added to D for D200. Pig was the experimental unit, and replicate and dietary treatment were fixed effects. Orthogonal polynomial contrasts were used to test linear and quadratic effects of phytase within A, A200, A400, A600, and A800. Phytase increased percent apparent total tract digestibility (ATTD) and STTD of P (quadratic P < 0.001), and quantity of absorbed P (linear P < 0.001; quadratic P = 0.069). Urinary P increased linearly with phytase (P < 0.001) and retained P also increased (linear P = 0.001, quadratic P = 0.094). Phytate-P release was estimated to be 0.049, 0.080, 0.093, and 0.09% STTD P for 200, 400, 600, and 800 FTU/kg, respectively. It appears that the effect of phytase may be lower in P-adequate diets as compared to P-deficient diets, given that there was a 12% improvement for A200 versus A, and a 28% improvement in STTD P for D200 versus D. In conclusion, phytase improved P digestibility and retention in P-adequate diets, and P digestibility was used to estimate the quantity of P released by phytase. Further research investigating P release by phytase in P-adequate diets, rather than P-deficient diets, may be preferable.

Previous research indicates that "super-dosing" phytase may improve pig growth performance by improved nutrient use, although the benefits appear to be more consistent in nursery than in grow-out pigs. Therefore, two experiments were conducted to determine if performance could be improved by feeding phytase at super-dosed levels, and whether this response would be different if energy and amino acid (AA) were limiting. Experiment 1 involved 440 weaned pigs (6.27 ± 0.01 kg) in a factorial arrangement of treatments comparing the main effects of diet (positive control [PC] balanced for all nutrients vs. a negative control [NC]: 10% lower standardized ileal digestible (SID) lysine with relative reduction of all other essential AA and 1% reduced fat) and phytase levels (0 vs. 2,500 FTU Quantum Blue 5G phytase/kg). Pigs were assigned to pen according to a randomized complete block design based on body weight (BW). Feed and water were provided ad libitum across four dietary phases: 3 × 1 wk plus 1 × 2 wk. The average daily gain (ADG) and gain to feed ratio (G:F) were improved in the PC relative to the NC ( P < 0.05) indicating success in formulating a diet limiting in energy and/or AA. Phytase improved ADG and G:F, regardless of diet composition ( P < 0.05). Thus, super-dosing phytase improved nursery pig growth performance, irrespective of diet nutrient adequacy or deficit. Experiment 2 involved 2,200 growing pigs (36.6 ± 0.30 kg) allotted to five treatments: a balanced PC (250 FTU Quantum Blue 5G phytase/kg), an NC (PC with 15% less SID lysine and 1.5% lower net energy [NE]), and three super-dosing phytase treatments applied to the NC totaling 1,000, 1,750, and 2,500 FTU phytase/kg. Feed and water were available ad libitum. At trial completion (approximately 122 kg), the PC pigs were heavier and more efficient than the NC pigs ( P < 0.05) indicating success in formulating an NC treatment. Super-dosing phytase had no effect on whole body ADG or average daily feed intake ( P > 0.10) but tended to improve G:F and feed energy efficiency ( P < 0.10). Super-dosing phytase improved carcass-based feed and feed energy efficiency ( P < 0.05) and tended to improve ADG ( P < 0.10). Supplying phytase at "super-dosed" levels-above that required to meet the phosphorus requirement-improved growth performance in nursery pigs (6 to 22 kg BW) and provided smaller benefits in grow-finish pigs (37 to 122 kg BW). The improvement during the nursery period was independent of energy and AA levels in the diet., (© The Author(s) 2018. Published by Oxford University Press on behalf of the American Society of Animal Science.)

The objective of this study was to evaluate the effects of dietary xylanase (X) and a carbohydrase enzyme blend (EB: cellulase, β-glucanase, and xylanase) on nutrient digestibility, intestinal barrier integrity, inflammatory status, and growth performance in weaned piglets fed higher fiber diets. A total of 460 pigs (6.43 ± 0.06 kg BW; F25 × 6.0 Genetiporc) were blocked by initial BW and pens (n = 12 per treatment) were randomly assigned to 1 of 4 dietary treatments. The diets included a higher fiber unsupplemented control diet (CON) and the CON supplemented with 0.01% X, 0.01% EB, or both enzymes, arranged in a 2 × 2 factorial. The diets were based on corn, soybean meal, corn distillers dried grains with solubles (DDGS), and wheat middlings. Pigs had 7 d to adapt to the environment and consumed the same commercial diet. Pigs were fed the experimental diets for 28 d with free access to feed and water. Body weight and feed disappearance were recorded weekly. One pig with BW closest to the pen average from each pen was selected and moved to metabolism crates on day 16 and intragastric gavaged a solution of lactulose and mannitol on day 22 followed by 12-h urine collection. Feces were collected from day 23 to 25. Intestinal tissues and mucosal scrapings were collected on day 28. Data were analyzed using PROC MIXED of SAS (9.4). Xylanase, EB, and their interaction were fixed effects and block was a random effect. The EB, but not X, increased pig BW and improved ADG over 28 d (P < 0.05). Neither carbohydrase impacted ADFI, G:F, or apparent total tract digestibility (ATTD) of DM, GE, or CP. The EB improved ATTD of ADF (32.45 vs. 26.57%; P < 0.01), but had no effect on NDF. Unexpectedly, X reduced ATTD of NDF and ADF (P < 0.01). The EB reduced urinary lactulose:mannitol and increased ileal claudin-3 mRNA abundance (P < 0.05), indicating improved small intestinal barrier integrity. There was a X × EB interaction on ileal secretory immunoglobulin A (sIgA) concentration (P < 0.05); in the absence of X, EB decreased sIgA compared to CON, but this effect disappeared in the presence of X. The EB also reduced ileal IL-22 mRNA abundance (P < 0.05), probably indicating decreased immune activation. In conclusion, EB but not X enhanced growth rate of weaned pigs fed higher fiber diets, which may be partly explained by the improved small intestinal barrier integrity and reduced immune activation, rather than improvement in nutrient digestibility.

Background: Burrowing is a rodent behavior validated as a robust and reproducible outcome measure to infer the global effect of pain in several inflammatory pain models. However, less is known about the effect of analgesics on burrowing in neuropathic pain models and no studies have determined burrowing performance in models of diabetes-associated neuropathic pain., Objective: To compare the sensitivity of the burrowing assay in different neuropathic pain models: mononeuropathic pain and diabetic polyneuropathy., Methods: Burrowing performance was determined by the amount of substrate left in a hollow tube by rats with chronic constriction injury (CCI). In addition, burrowing performance, locomotion and pain development was assessed in the Zucker diabetic fatty (ZDF) rat model, resembling type-2 diabetes. Efficacy of clinically-active reference drugs (opioids, gabapentin and/or pregabalin) were investigated in these models. Burrowing behavior was additionally assessed in a second model, induced by streptozotocin (STZ) treatment, resembling type-1 diabetes., Results: In the CCI model, moderate but consistent burrowing deficits were observed that persisted over a period of ≥20 days. Systemic administration of morphine, pregabalin and gabapentin reversed this deficit. In contrast, none of the reference drugs improved marked burrowing deficits detected in ZDF rats, and pregabalin did not reverse severe burrowing deficits observed in STZ rats., Conclusions: Burrowing performance cannot necessarily be used as pain-related readout across pain models and largely depends on the model used, at least in models of neuropathy. Specifically, analgesic drug effects might be masked by general diabetes-associated alteration of the animals' well-being, resulting in false negative outcomes., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: It has recently been suggested that non-reflex behavioral readouts, such as burrowing, may be used to evaluate the efficacy of analgesics in rodent models of pain., Objective: To confirm whether intraplantar Complete Freund's Adjuvant (CFA)-induced pain reliably results in burrowing deficits which can be ameliorated by clinically efficacious analgesics as previously suggested., Methods: Uni- or bilateral intraplantar CFA injections were performed in male Wistar Han rats. The time- and concentration-response of burrowing deficits and the ability of various analgesics to reinstate burrowing performance were studied. An anxiolytic was also tested to evaluate the motivational cue that drives this behavior., Results: Burrowing deficits were dependent on the concentration of CFA injected, most pronounced 24h after CFA injections and even more pronounced after bilateral compared with unilateral injections. Celecoxib and ibuprofen reversed CFA-induced burrowing deficits whereas indomethacin failed to significantly reinstate burrowing performance. Morphine and tramadol failed to reinstate burrowing performance, but sedation was observed in control rats at doses thought to be efficacious. An antibody directed against the nerve growth factor significantly improved CFA-induced burrowing deficits. Neither gabapentin nor the anxiolytic diazepam reinstated burrowing performance and the opportunity to find shelter did not modify burrowing performance., Conclusion: Burrowing is an innate behavior reliably exhibited by rats. It is suppressed in a model of inflammatory pain and differently reinstated by clinically efficacious analgesics that lack motor impairing side effects, but not an anxiolytic, suggesting that this assay is suitable for the assessment of analgesic efficacy of novel drugs., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Objective: To describe Schirmer tear test I and rebound tonometry findings in healthy bovine calves., Animals Studied: Thirty-three clinically normal dairy breed calves of mean (SD) 11 (1.1) weeks (range, 9.3-13.3 weeks) of age were evaluated., Procedures: A Schirmer tear test I was performed on each eye followed by tonometry, using a TonoVet(®) without topical anesthesia. We report means (SD) and statistical analysis of data for each assay., Results: For both Schirmer tear test (STT) I and tonometry, significant differences were not found between fellow eyes (P = 0.1536 and P = 0.83, respectively). The mean (SD) STT I value of all eyes was 20.4 (5.0) mm/min (range, 9-34 mm/min) while the mean (SD) intraocular pressure (IOP) value of all eyes was 15.2 (5.2) mmHg (range, 7-25 mmHg)., Conclusions: This study reports normal data for the STT I and rebound tonometry in calves. This data may be useful in complete ophthalmic examinations of cattle, guiding diagnosis of glaucoma, uveitis, and keratoconjunctivitis sicca. However, results of these diagnostic modalities must be interpreted in light of clinical signs, given the wide range of normal values obtained in this study., (© 2014 American College of Veterinary Ophthalmologists.)

Objective: To evaluate corneal sensitivity as measured by the corneal touch threshold in healthy bovine calves., Animals Studied: Twelve clinically normal male calves with predominantly Holstein genetics and a median age of 76.5 days (range, 67-92 days)., Procedures: Corneal touch threshold (CTT) of the central cornea was measured in both eyes of each calf using a Cochet-Bonnet aesthesiometer., Results: The mean ± standard deviation corneal touch threshold of all eyes was 1.33 ± 1.1 g/mm(2) (range, 0.62-66.15 g/mm(2) ), corresponding to a filament length of 34.56 ± 8.02 mm (range, 14-47.5 mm). There was no significant difference between fellow eyes., Conclusions: Cochet-Bonnet aesthesiometry was well tolerated in all 12 calves using a modified head restraint. Calves in this study may have a relatively sensitive central cornea compared to adult cattle and some other species; however, wide variation among individuals and eyes may be possible. Studies utilizing larger calf populations are necessary to establish reference ranges., (© 2014 American College of Veterinary Ophthalmologists.)

Context: Sidelines coverage presents unique challenges in the evaluation of injured athletes. Health care providers may be confronted with the question of when to obtain radiographs following an injury. Given that most sidelines coverage occurs outside the elite level, radiographs are not readily available at the time of injury, and the decision of when to send a player for radiographs must be made based on physical examination. Clinical tools have been developed to aid in identifying injuries that are likely to result in radiographically important fractures or dislocations., Evidence Acquisition: A search for the keywords x-ray and decision rule along with the anatomic locations shoulder, elbow, wrist, knee, and ankle was performed using the PubMed database. No limits were set regarding year of publication. We selected meta-analyses, randomized controlled trials, and survey results. Our selection focused on the largest, most well-studied published reports. We also attempted to include studies that reported the application of the rules to the field of sports medicine., Study Design: Retrospective literature review., Level of Evidence: Level 4., Results: The Ottawa Foot and Ankle Rules have been validated and implemented and are appropriate for use in both pediatric and adult populations. The Ottawa Knee Rules have been widely studied, validated, and accepted for evaluation of knee injuries. There are promising studies of decision rules for clinically important fractures of the wrist, but these studies have not been validated. The elbow has been evaluated with good outcomes via the elbow extension test, which has been validated in both single and multicenter studies. Currently, there are no reliable clinical decision tools for traumatic sports injuries to the shoulder to aid in the decision of when to obtain radiographs., Conclusion: Clinical decision tools have been developed to aid in the diagnosis and management of injuries commonly sustained during sporting events. Tools that have been appropriately validated in populations outside the initial study population can assist sports medicine physicians in the decision of when to get radiographs from the sidelines.

Predicting the bioavailability of inorganic mercury (Hg) to bacteria that produce the potent bioaccumulative neurotoxin monomethylmercury remains one of the greatest challenges in predicting the environmental fate and transport of Hg. Dissolved organic matter (DOM) affects mercury methylation due to its influence on cell physiology (as a potential nutrient) and its influence on Hg(II) speciation in solution (as a complexing agent), therefore controlling Hg bioavailability. We assessed the role of DOM on Hg(II) bioavailability to a gram-negative bacterium bioreporter under oxic pseudo- and nonequilibrium conditions, using defined media and field samples spanning a wide range of DOM levels. Our results showed that Hg(II) was considerably more bioavailable under nonequilibrium conditions than when DOM was absent or when Hg(II) and DOM had reached pseudoequilibrium (24 h) prior to cell exposure. Under these enhanced uptake conditions, Hg(II) bioavailability followed a bell shaped curve as DOM concentrations increased, both for defined media and natural water samples, consistent with bioaccumulation results in a companion paper (this issue) observed for amphipods. Experiments also suggest that DOM may not only provide shuttle molecules facilitating Hg uptake, but also alter cell wall properties to facilitate the first steps toward Hg(II) internalization. We propose the existence of a short-lived yet critical time window (<24 h) during which DOM facilitates the entry of newly deposited Hg(II) into aquatic food webs, suggesting that the bulk of mercury incorporation in aquatic food webs would occur within hours following its deposition from the atmosphere.

Infectious bovine keratoconjunctivitis (IBK) is a common ocular disease in cattle, associated with a 6.8 to 13.6 kg decrease in weaning weight. Antibiotic therapy is available but it is unclear if pain mitigation as an adjunct therapy would reduce the weight loss associated with IBK. Before assessing the impact of pain mitigation therapies, it is first necessary to validate approaches to qualifying ocular pain. The objective of this study was to evaluate approaches to qualifying ocular pain in bovine calves (Bos taurus) with IBK. Our a priori assumption was that scarification or corneal ulcerations consistent with IBK are painful compared to normal eyes. To quantify this difference in pain, we assessed 4 tools: pressure algometry-mechanical nociceptive threshold (PA-MNT), corneal touch thresholds (CTT) obtained with the use of a Cochet-Bonnet aesthesiometer, and assessment for the presence of blepharospasm and photophobia as metrics for pain. Using a 1-eye randomized controlled challenge trial, 31 calves with healthy eyes were randomly allocated to treatment groups, and then a left or right eye was randomly assigned for corneal scarification and inoculation with Moraxella bovoculi or Moraxella bovis. A repeated measures analysis of variance was used for PA-MNT, with significance set at P < 0.05. A log (base 10) transformation was used to stabilize the variance, and Tukey's t tests were used to test differences between assessment days for each landmark. Calves had statistically significantly lower PA-MNT scores (which indicates more pain) the day after scarification relative to baseline measurements (4 d before scarification). For example, at 1 landmark the median PA-MNT (kg/force) prescarification was 4.82 (95% confidence interval [CI]: 3.92-5.93) and 3.43 (95% CI: 2.79-4.22) postscarification. These data suggest PA-MNT may be a tool for quantifying ocular pain in calves. No differences (P < 0.1) in PA-MNT scores between scarified and not-scarified eyes were detected for any landmark on any day. This result suggests that the pain response occurs over the entire face, not just the affected eye. Corneal ulcerations consistent with IBK were not associated with statistically significant differences in PA-MNT or CTT at eye or calf levels. Not surprisingly, scarified eyes were more likely to exhibit blepharospasm and photophobia compared to healthy eyes. Due to blepharospasm, the use of the Cochet-Bonnet to evaluate corneal sensitivity by CTT was of limited value.

Selection of study endpoints is one of the most important decisions in the design of effective clinical trials. Late mortality (e.g., 28 days) is an unambiguous endpoint, accepted by regulatory agencies, but it is viewed as problematic among researchers in the study of resuscitation for acute trauma injury with hemorrhagic shock. In February 2008, physicians, ethicists, statisticians, and research scientists from the military, academia, industry, the Federal Drug Administration, and the National Heart Lung and Blood Institute gathered to discuss the obstacles confronting the trauma community in their efforts to improve patient outcomes. The primary meeting objective was to generate preliminary suggestions for a series of follow-up meetings that will develop consensus guidelines for the design of large multicenter clinical trials. Twenty short presentations and discussions, summarized here, outlined the group's concerns and suggestions. Successful and failed, completed or ongoing, clinical studies provided insight as to endpoints that may be of value for future trauma and shock studies. In addition to the importance of appropriate endpoints in study design, other related topics were discussed, including trauma epidemiology, patient enrollment and inclusion criteria, community consultation and the difficulty of obtaining informed consent in acute trauma research, and the inclusion of quality of life in composite endpoints. The consensus was that more discussion was needed and that consideration of new endpoints for clinical trials in emergency trauma research was a worthwhile and necessary goal.

Background: Human polymerized hemoglobin (PolyHeme, Northfield Laboratories) is a universally compatible oxygen carrier developed to treat life-threatening anemia. This multicenter phase III trial was the first US study to assess survival of patients resuscitated with a hemoglobin-based oxygen carrier starting at the scene of injury., Study Design: Injured patients with a systolic blood pressure

Background: Acute chest syndrome (ACS) is a complication of sickle cell disease that can cause significant morbidity. Transfusion therapy has been shown to significantly increase oxygenation in patients with ACS and RBC exchange is considered the standard of care in patients at high risk of respiratory failure., Case Report: A patient with ACS and several high-risk features, including thrombocytopenia, profound anemia, bilateral pulmonary infiltrates, staphylococcal sepsis, and pulmonary embolism is presented. The patient refused transfusion on religious grounds and received 12 units of human polymerized Hb solution (poly SFH-P injection, PolyHeme, Northfield Laboratories) over the course of 13 days. The patient's respiratory status improved and she was discharged home without receiving RBC transfusions., Conclusion: This is the first reported case that describes the use of PolyHeme in a patient with sickle cell disease, ACS, and sepsis. This therapy is thought to have been lifesaving for this patient.

Background: Human polymerized hemoglobin (PolyHeme, Northfield Laboratories, Evanston, IL) is a universally compatible, immediately available, disease-free, oxygen-carrying resuscitative fluid being developed as a red cell substitute for use in urgent blood loss. PolyHeme should be particularly useful when red cells may be temporarily unavailable. This article assesses survival at life-threatening RBC hemoglobin concentration ([Hb]) in massively bleeding patients who do not receive red cells., Study Design: There were 171 patients who received rapid infusion of 1 to 20 units (1,000 g, 10 L) of PolyHeme in lieu of red cells as initial oxygen-carrying replacement in trauma and urgent surgery. The protocol simulated the unavailability of red cells, and the progressive fall in RBC [Hb] in bleeding patients was quantified. Thirty-day mortality was compared with a historical control group of 300 surgical patients who refused red cells on religious grounds., Results: A total of 171 patients received rapid infusion of 1 to 2 units (n = 45), 3 to 4 units (n = 45), 5 to 9 units (n = 47), or 10 to 20 units (n = 34) of PolyHeme. Forty patients had a nadir RBC [Hb] < or = 3 g/dL (mean, 1.5 +/- 0.7 g/dL). But total [Hb] was adequately maintained (mean, 6.8 +/- 1.2 g/dL) because of plasma [Hb] added by PolyHeme. The 30-day mortality was 25.0% (10/40 patients) compared with 64.5% (20/31 patients) in historical control patients at these RBC [Hb] levels., Conclusions: PolyHeme increases survival at life-threatening RBC [Hb] by maintaining total [Hb] in the absence of red cell transfusion. PolyHeme should be useful in the early treatment of urgent blood loss and resolve the dilemma of unavailability of red cells.

Background: Guidelines for allogeneic transfusion emphasize minimizing use to avoid transmission of serious illness. However, there is little information on the risks associated from withholding transfusion., Study Design and Methods: A retrospective cohort study of patients who declined RBC transfusions for religious reasons was performed. This analysis was restricted to consecutive patients > or = 18 years old, who underwent surgery in the operating room from 1981 to 1994 and had a postoperative Hb count of 8 g per dL or less. The primary outcome was defined as any inhospital death occurring within 30 days of the surgery. Secondary outcome was 30-day mortality or in-hospital 30-day morbidity. Morbidity was defined as myocardial infarction, arrhythmia, congestive heart failure, or infection., Results: Of 2083 eligible patients, 300 had postoperative Hb counts of 8 g per dL or less. The study population was predominantly female (70.3%) with a mean age of 57 years (SD, +/- 17.7). In patients with a postoperative Hb level of 7.1 to 8.0, 0 died (upper 95% CI, 3.7%), and 9.4 percent (95% CI, 4.4-17.0%) had a morbid event. In patients with a postoperative Hb level of 4.1 to 5.0, 34.4 percent (95% CI, 18.6-53.2%) died and 57.7 percent (95% CI, 36.9-76.6%) had a morbid event or died. After adjusting for age, cardiovascular disease, and Acute Physiology and Chronic Health Evaluation II score, the odds of death in patients with a postoperative Hb level of < or = 8 g per dL increased 2.5 times (95% CI, 1.9-3.2) for each gram decrease in Hb level., Conclusions: The risk of death was low in patients with postoperative Hb levels of 7.1 to 8.0 g per dL, although morbidity occurred in 9.4 percent. As postoperative blood counts fall the risk of mortality and/or morbidity rises and becomes extremely high below 5 to 6 g per dL.

Expanded clays bipillared with [Al(13)O(4)(OH)(24)(H(2)O)(12)](7+) ions and with hexameric Cu complexes such as M[(&mgr;-OH)Cu(&mgr;-OCH(2)CH(2)NEt(2))](6)(ClO(4))(3), or with M[(&mgr;-OH)Cu(&mgr;-OCH(2)CH(2)NEt(2))](6)(PF(6))(3) where M = Fe, Al, Ga, form microporous materials whose stability and microporosity depend mainly on the identity of the hexamer central metal atom. In fact, a general decrease in thermal stability, interlamellar heights, surface areas, and pore volumes was noted when, in the (Cu,M) hexamer, M changed from gallium to aluminum to iron. Mossbauer results have indicated that only Fe(3+) in octahedral coordination is present in the iron-containing bi-PILC samples (bi-PILC = bipillared interlayered clays). It is believed that metals such as Fe(3+) and Cu(2+) can interact with the interlamellar Keggin ions thereby decreasing the stability of the alumina pillars. In contrast, the intermediate Al(13)-PILC structure is least affected when the more stable Cr complex is used. Bi-PILC materials containing 2.7-3.4% Cr stable to 500 degrees C have been obtained. The low polarity of the chosen solvent (acetonitrile) appears to inhibit the back-exchange of the intermediate PILC's Keggin ions with the hexameric Cu complexes. Elemental analysis together with XRD results suggests that the primary intercalation pathway was diffusion or ion exchange when Cr[(&mgr;-OCH(3))(&mgr;-OCH(2)CH(2)NEt(2))CuCl](3) or M[(&mgr;-OH)Cu(&mgr;-OCH(2)CH(2)NEt(2))](6)(ClO(4))(3), respectively, was used. In all preparations, bi-PILC were produced containing complexes that suffered ligand losses during the synthesis reaction. Molecular scale AFM images have shown that these complexes can be found also outside the clay interlamellar space.

The surfaces of both stretched and unstretched silk threads from the cobweb weaver, Latrodectus hesperus (Black Widow) have been examined by atomic force microscopy (AFM). AFM images of cobweb scaffolding threads show both unordered and highly ordered regions. Two types of fibers within the threads were observed: thicker (approximately 300 nm in diameter) fibers oriented parallel to the thread axis and thinner (10-100 nm) fibrils oriented across the thread axis. While regions which lacked parallel fibers or fibrils were observed on threads at all strain values, the probability of observing fibers and/or fibrils increased with strain. High-resolution AFM images show that with increasing strain, both mean fiber and fibril diameters decrease and that fibrils align themselves more closely with the thread axis. The observation of fibers and fibrils within the cobweb threads has implications for current models of the secondary and tertiary structure and organization of spider silk.

Background: Human polymerized hemoglobin (PolyHeme) is a universally compatible, disease-free, oxygen-carrying resuscitative fluid. This is the first prospective, randomized trial to compare directly the therapeutic benefit of PolyHeme with that of allogeneic red blood cells (RBCs) in the treatment of acute blood loss., Study Design: Forty-four trauma patients (33 male, 11 female) aged 19-75 years with an average Injury Severity Score (ISS) score of 21+/-10 were randomized to receive red cells (n = 23) or up to 6 U (300 g) of PolyHeme (n = 21) as their initial blood replacement after trauma and during emergent operations., Results: There were no serious or unexpected adverse events related to PolyHeme. The PolyHeme infusion of 4.4+/-2.0 units (mean +/- SD) resulted in a plasma [Hb] of 3.9+/-1.3 g/dL, which accounted for 40% of the total circulating [Hb]. There was no difference in total [Hb] between the groups before infusion (10.4+/-2.3 g/dL control vs. 9.4+/-1.9 g/dL experimental). At end-infusion the experimental RBC [Hb] fell to 5.8+/-2.8 g/dL vs. 10.6+/-1.8 g/dL (p < 0.05) in the control, although the total [Hb] was not different between the groups or from pre-infusion. The total number of allogeneic red cell transfusions for the control and experimental groups was 10.4+/-4.2 units vs. 6.8+/-3.9 units (p < 0.05) through day 1, and 11.3+/-4.1 units vs. 7.8 +/-4.2 units (p = 0.06) through day 3., Conclusions: PolyHeme is safe in acute blood loss, maintains total [Hb] in lieu of red cells despite the marked fall in RBC [Hb], and reduces the use of allogeneic blood. PolyHeme appears to be a clinically useful blood substitute.

A practice parameter has been developed to assist physicians in the therapeutic use of red blood cell transfusions. The developers of this parameter used the best available information from the medical literature, as well as clinical experience and the extensive reality testing required by the College of American Pathologists for approval. In acute anemia, a fall in hemoglobin values below 6 g/dL or a rapid blood volume loss of more than 30% to 40% requires red blood cell transfusions in most patients. However, tissue oxygenation provides a better indication of physiologic need in situations where invasive monitoring provides this information. When these data are not available, heart rate and blood pressure measurements and the nature of bleeding (active, controlled, uncontrolled) supplement the hemoglobin value in guiding the transfusion decision. In sickle cell disease and thalassemias, red blood cells are transfused to prevent acute or chronic complications. Red blood cell transfusions are used in chronic anemias unresponsive to pharmacologic agents based on the patient's symptoms. Guidelines must be altered for neonates who require an increase in hematocrit to above 0.30 to 0.35 when respiratory distress is present. Indications for red blood cell transfusion for the pregnant or postpartum patient are similar to those for the nonpregnant patient. Risks of transfusion, particularly transmissible disease and incompatibility, remain but have been reduced. Thus, red blood cell transfusion continues to be a powerful therapeutic tool when used judiciously and carries less risk than in the recent past.

Although the efficacy of hemoglobin-based oxygen carriers was established more than 60 years ago, all prior clinical trials have demonstrated significant toxicity characterized by renal dysfunction, gastrointestinal distress, and systemic vasoconstriction. The mechanisms of these toxicities now appear to be understood. Tetrameric forms of the hemoglobin molecule extravasate from the circulation and interact with endothelium-derived relaxing factor, leading to unopposed vasoconstriction. Although numerous efforts are under way to chemically modify the native tetramer, it is likely that all tetrameric forms of the hemoglobin molecule will continue to extravasate. We have focused on developing a polymerized form of hemoglobin that is virtually free of unreacted tetramer. The development and characterization of this polymerized pyridoxylated hemoglobin solution (Poly SFH-P) is described. Clinical trials have been completed successfully in volunteers and are now under way to assess the safety and efficacy of Poly SFH-P as a clinically useful red blood cell substitute for treatment of acute blood loss in the setting of trauma and surgery.

Objective: Limited cardiac reserve, secondary to coronary disease, may be associated with end organ morbidity. In this study, we investigate the significance of anemia in the pathogenesis of this phenomenon., Design: Nonrandomized controlled animal trial., Settings: Animal laboratory in a university hospital., Subject: Anesthetized dogs. INTERVENTIONS/MEASUREMENTS: Fourteen anesthetized dogs underwent isovolemic hemodilution with 6% hetastarch from a baseline hematocrit of 40 to 20%. Radioactive microspheres were used to evaluate regional blood flow and cardiac index. Systemic oxygen delivery, consumption, serum lactate, and systemic vascular resistance were recorded during each experiment. Arterial venous oxygen difference was determined from arterial and mixed venous blood. Seven dogs had an iatrogenic critical stenosis of their left anterior descending coronary artery (experimental group); seven dogs did not (control)., Main Results: Only in the control animals, the cardiac index increased by 35% with hemodilution to 20%. Systemic oxygen delivery decreased in both the control and the experimental animals. Systemic oxygen consumption and lactate levels were unchanged in both groups. In the renal cortex, spleen, distal colon, ileum, gallbladder, and stomach body, regional O2 delivery was significantly decreased with hemodilution to 20% in both groups. This finding was also observed in the left ventricle and cervical spinal cord in the experimental group. In addition, regional O2 delivery was reduced in the spleen, distal colon, and gallbladder with hemodilution to only 30%. Regional blood flow in the stomach body, gallbladder, ileum, renal cortex, and distal colon, in both groups, and the spleen in the control group was unchanged from baseline with hemodilution to 20%. However, regional blood flow under all other circumstances (control or experimental) was significantly increased with hemodilution to 20% with the exception of the spleen, which showed significant regional blood flow decrease in the experimental group only., Conclusions: These data suggest that with limited cardiac reserve, anemia may compromise aerobic splanchnic circulation. These observations may further our understanding of the pathogenesis of cholecystitis, gastric stress ulcers, ileal endotoxin translocation, and ischemic colitis in critically ill patients with coronary artery disease.

Although the efficacy of hemoglobin-based oxygen carriers was established more than 60 years ago, all prior clinical trials have demonstrated significant toxicity characterized by renal dysfunction, gastrointestinal distress, and systemic vasoconstriction. The mechanisms of these toxicities now appear to be understood. Tetrameric forms of the hemoglobin molecule extravasate from the circulation and interact with endothelial derived relaxing factor, leading to unopposed vasoconstriction. Although numerous efforts are underway to chemically modify the native tetramer, it is likely that all tetrameric forms of the hemoglobin molecule will continue to extravasate. We have focused on developing a polymerized form of hemoglobin that is virtually free of unreacted tetramer. The development and characterization of this polymerized pyridoxylated hemoglobin solution (Poly SFH-P) is described. Clinical trials have been completed successfully in volunteers, and are now underway to assess the safety and efficacy of Poly SFH-P as a clinically useful red cell substitute in the treatment of acute blood loss in the setting of trauma and surgery.

The risks inherent in the use of homologous blood products have increased efforts toward identifying alternatives to transfusion. We have previously shown that the administration of recombinant human erythropoietin (rhEpo) enhances the erythropoietic response to acute blood loss. Recombinant human interleukin-3 (rh-IL-3) is a hematopoietic growth factor that has been shown to act synergistically with rhEpo in accelerating erythropoiesis in vitro. The purpose of this study in a primate model was to determine if the administration of rhIL-3 in combination with rhEpo could augment the erythropoietic response to acute blood loss more than rhEpo therapy alone. Twenty-four adult male baboons were randomized into four groups. The induction of acute normovolemic anemia to a hematocrit of 20% was accomplished via exchange-transfusion with 6% hetastarch. The groups were then treated for 7 consecutive days with the following growth factors: group I (n = 7), no growth factors; group II (n = 5), rhIL-3 alone (100 micrograms/kg/d); group III (n = 6), rhEpo alone (1000 U/kg/d); group IV (n = 6), rhEpo (1000 U/kg/d) plus rhIL-3 (100 micrograms/kg/d). All animals received folate, vitamin B12, and intravenous iron-dextran immediately following the exchange-transfusion. Response to therapy was monitored for 35 days. There were no adverse reactions following growth factor administration. The analysis of erythropoietic rates between study days 1 through 11, as determined via linear regression analysis, revealed that hematocrits increased significantly faster in the groups receiving rhEpo compared to controls. The administration of rhIL-3, however, did not increase the rate of erythropoiesis when compared to controls, nor did it augment response when added to the rhEpo regimen. The results of this study demonstrate that the administration of rhIL-3 alone had no significant effect on erythropoiesis in this setting of acute blood loss. Further, despite promising in vitro data, rhIL-3 provided no additional stimulation of erythropoiesis in animals receiving rhEpo. Nevertheless, the study confirms that the pharmacologic acceleration of erythropoiesis by rhEpo alone remains an attractive alternative to homologous transfusion.

We assessed limit to cardiac compensation during isovolemic hemodilution (HD) in 14 anesthetized dogs. Radioactive microspheres were used to evaluate myocardial blood flow (MBF) and its transmural distribution (endo/epi). Myocardial O2 consumption (MVO2) and percent lactate extraction were determined. Coronary vasodilator reserve was assessed from reactive hyperemic responses. Dogs were divided into group 1, with intact left anterior descending coronary artery (LAD), and group 2, with critical stenosis of LAD. Measurements were obtained at baseline and during graded HD (Hespan) until cardiac failure (CF). CF occurred at lower hematocrit in group 1 compared with group 2 (9 +/- 1 vs. 17 +/- 1%). In group 1, MBF increased during HD to maintain MVO2 constant; increases in MBF were transmurally uniform until CF, when decreased endo/epi and lactate production suggested subendocardial ischemia. Coronary vasodilator reserve decreased progressively during HD and was absent at CF. In group 2, stenotic LAD demonstrated constant MBF (resulting in decreased MVO2) during HD. At CF, these responses along with reduced endo/epi and lactate production indicated local myocardial ischemia. We conclude that 1) with normal coronary circulation, cardiac function was well maintained over a wide range of hematocrits because increases in MBF were transmurally uniform and sufficient to maintain myocardial oxygenation: CF occurred during extreme HD when MBF became maldistributed, resulting in subendocardial ischemia; 2) critical coronary stenosis impaired coronary vascular adjustment to HD and reduced significantly tolerance of left ventricle to HD; and 3) present findings underscore the importance of recruitment of coronary vasodilator reserve in preserving total and regional myocardial oxygenation during HD.

This article defines a rational approach to the treatment of hemorrhagic shock. All patients that are hypovolemic following hemorrhage require fluid resuscitation. Some patients require red cell restoration and very few require correction of any clotting deficiencies. A physiologic approach to these problems will lead to optimal patient care in these circumstances.

Stimulation of erythropoiesis is an attractive alternative to the risks of homologous transfusion. The availability of rHuEPO has made erythropoietic acceleration possible. The use of rHuEPO perioperatively and in preoperative autologous donation will likely find firm indications. Combinations of present methods of autologous blood use with the various rHuEPO regimens will be the best methods of minimizing perioperative homologous blood exposure.

We have described whole body oxygen (O2) extraction ratio (ER) as a reliable indicator of transfusion need in acute normovolemic anemia. In normal hearts, myocardial lactate production (-LACT), indicating anaerobic metabolism, does not occur until the ER greater than 50% and Hct less than 10%. It is not known if the ER is valid in the setting of limited coronary vascular reserve. This study assesses the effect of a critical left anterior descending (LAD) coronary stenosis on the compensation to acute blood loss anemia. Adult dogs were anesthetized, paralyzed, and mechanically ventilated. A critical LAD stenosis was created in seven animals (STEN). There were seven controls (CON). Animals underwent isovolemic exchange transfusion with 6% HES until cardiac failure (CF). Catheters were placed in the aorta, pulmonary artery, and anterior interventricular coronary vein. Cardiac failure occurred at Hct = 8.6% +/- 0.4% in the CON and 17.0% +/- 0.5% in the STEN animals. Cardiac output increased in the CON, but not in the STEN animals. Blood flow in the LAD increased in the CON but not the STEN animals. -LACT began in the CON and STEN animals at Hct less than 20% and coincided with an ER greater than 50% in both groups. We conclude that CF occurs at a higher hematocrit with a critical LAD stenosis. The whole body ER greater than 50% remains a valid indicator of myocardial metabolism in anemia in the presence of limited coronary vascular reserve. The ER may be a useful guide to transfusion therapy.