Effects of pH and the plasma or serum concentrations of total calcium, chloride, magnesium, l-lactate, and albumin on the plasma ionized calcium concentration in calves.

BACKGROUND: The plasma ionized calcium concentration (cCa2+ ) represents the biologically active form of calcium and is the preferred method for evaluating calcium status in animals. Different pH-corrective equations have been developed for human plasma, but the validity of the equations for bovine plasma is unknown. HYPOTHESIS: We hypothesized that pH-corrective equations for bovine plasma would be similar to those used for human plasma; cCa2+ was dependent on the plasma concentrations of total calcium (cTCa), chloride (cCl), L-lactate (cLactate), and albumin (cAlbumin); and the in vitro and in vivo cCa2+ -pH relationships would differ. ANIMALS: Ten healthy calves (in vitro study), 1426 critically ill calves. METHODS: The in vitro plasma log10 (cCa2+ )-pH relationship was determined by CO2 tonometry of 465 plasma samples. Plasma cCl was altered by equivolume dilution of plasma with 3 electrolyte solutions of different cCl. The in vivo plasma cCa2+ -pH relationship was investigated and validated using clinicopathologic data extracted from the medical records of 950 (model development) and 476 (model validation) critically ill calves. RESULTS: pH-corrective equations for bovine plasma were similar to those used for human plasma. Plasma cCa2+ increased in vitro with increases in plasma cCl. Plasma cCa2+ in critically ill calves was associated with plasma cTCa, blood pH, plasma cCl, serum cMg, and cL-lactate (R2 = 0.69) but not plasma cAlbumin. CONCLUSIONS AND CLINICAL IMPORTANCE: Calculation of cCa2+ from cTCa in calf plasma or serum requires adjustment for at least pH and cCl when 1 or both are outside the reference range.

Diagnosis and Treatment of Undifferentiated and Infectious Acute Diarrhea in the Adult Horse.

Acute, infectious, diarrhea in adult horses is a major cause of morbidity and is associated with numerous complications. Common causes include salmonellosis, clostridiosis, Coronavirus, and infection with Neorickettsia risticii (Potomac horse fever). Treatment is empirical and supportive until results of specific diagnostic tests are available. Supportive care is aimed at restoring hydration, correcting electrolyte imbalances, and limiting the systemic inflammatory response. The mainstays of therapy are intravenous fluid therapy, electrolyte supplementation where necessary, nonsteroidal anti-inflammatory agents, and nutritional support. Specific therapies include colloid oncotic support, antibiotics, hyperimmune plasma, polymyxin B, pentoxifylline, probiotics, binding agents, gastroprotectants, laminitis prevention, and coagulation prophylaxis.

Changes in the equine fecal microbiota associated with the use of systemic antimicrobial drugs.

BACKGROUND: The intestinal tract is a rich and complex environment and its microbiota has been shown to have an important role in health and disease in the host. Several factors can cause disruption of the normal intestinal microbiota, including antimicrobial therapy, which is an important cause of diarrhea in horses. This study aimed to characterize changes in the fecal bacterial populations of healthy horses associated with the administration of frequently used antimicrobial drugs. RESULTS: Twenty-four adult mares were assigned to receive procaine penicillin intramuscularly (IM), ceftiofur sodium IM, trimethoprim sulfadiazine (TMS) orally or to a control group. Treatment was given for 5 consecutive days and fecal samples were collected before drug administration (Day 1), at the end of treatment (Days 5), and on Days 14 and 30 of the trial. High throughput sequencing of the V4 region of the 16S rRNA gene was performed using an Illumina MiSeq sequencer. Significant changes of population structure and community membership were observed after the use of all drugs. TMS caused the most marked changes on fecal microbiota even at higher taxonomic levels including a significant decrease of richness and diversity. Those changes were mainly due to a drastic decrease of Verrucomicrobia, specifically the "5 genus incertae sedis". Changes in structure and membership caused by antimicrobial administration were specific for each drug and may be predictable. Twenty-five days after the end of treatment, bacterial profiles were more similar to pre-treatment patterns indicating a recovery from changes caused by antimicrobial administration, but differences were still evident, especially regarding community membership. CONCLUSIONS: The use of systemic antimicrobials leads to changes in the intestinal microbiota, with different and specific responses to different antimicrobials. All antimicrobials tested here had some impact on the microbiota, but TMS significantly reduced bacterial species richness and diversity and had the greatest apparent impact on population structure, specifically targeting members of the Verrucomicrobia phylum.

Clinical utility of serum biochemical variables for predicting acid-base balance in critically ill horses.

BACKGROUND: Profiles from serum biochemical analyzers include the concentration of strong electrolytes (including l-lactate), total carbon dioxide (tCO2 ), and total protein. These variables are associated with changes in acid-base balance. Application of physicochemical principles may allow predicting acid-base balance from serum biochemistry without measuring whole blood pH and pCO2 . OBJECTIVES: The purpose of the study was to determine if the acid-base status of critically ill horses could be accurately predicted using variables included in standard serum biochemical profiles. METHODS: Two jugular venous blood samples were prospectively obtained from critically ill horses and foals. Samples were analyzed using a whole blood gas and pH analyzer (BG) and a serum biochemistry multi analyzer system (AMAS). Linear regression, Deming regression, and Bland-Altman plots were used for method comparison and P < .05 was considered significant. RESULTS: Values from 70 horses and foals for Na, K, Cl, and total protein concentrations, and consequently the calculated variables used for acid base interpretation, were different between the AMAS and BG analyzer. Using physicochemical principles, BG results accurately predicted pH, whereas the AMAS results did not when a fixed value for pCO2 was used. CONCLUSIONS: Measurement of pCO2 is required in critically ill horses for accurate prediction of whole blood pH. Differences in the measured values of Na and Cl concentration exist when measured in serum by the AMAS and in whole blood or plasma by BG, indicating that the accurate prediction of whole blood pH is analyzer-dependent. Application of physicochemical principles to plasma or serum provides a practical method to evaluate analyzer accuracy.

False-Positive Clostridium difficile in Negative-Control Reactions Peak and Then Decrease with Repetitive Refrigeration of Immunoassay.

Aberrant false-positive reactions in negative-controls during ELISA testing for Clostridium difficile indicated the potential for false-diagnoses. Experiments with 96-well products showed a maximum peak of false-positive immunoassay reactions with the provided negative-control reagents after 5 refrigeration-to-room temperature cycles (P < 0.001), decreasing thereafter with additional refrigeration cycles. Because repetitive refrigeration causes a peak of false-positives, the use of single negative-controls per ELISA run might be insufficient to monitor aberrant preanalytical false-positives if immunoassays are subject to repetitive refrigeration.

Clinicopathologic variables associated with hypokalemia in lactating dairy cows with abomasal displacement or volvulus.

OBJECTIVE: To identify potential mechanisms for hypokalemia in dairy cows with left-displaced abomasum (LDA), right-displaced abomasum (RDA), or abomasal volvulus (AV). DESIGN: Retrospective analysis of clinicopathologic data from 2 convenience samples of cows. SAMPLE: 112 lactating dairy cows with AV (group 1); 1,332 lactating dairy cows (group 2) with LDA (n = 1,160) or RDA or AV (172). PROCEDURES: Data were analyzed via Spearman ρ and multivariate stepwise regression. RESULTS: 78 of 112 (70%) group 1 cows were hypokalemic (mean serum potassium concentration, 3.5 mEq/L; reference range, 3.9 to 5.8 mEq/L). For group 1 cows, serum chloride concentration had the strongest positive association with serum potassium concentration, and serum potassium concentration was negatively associated with plasma bicarbonate and serum glucose, creatinine, and urea concentrations. Six hundred thirty-six of 1,160 (55%) of group 2 cows with LDA were hypokalemic (mean serum potassium concentration, 3.7 mEq/L). Ninety-two of 172 (53%) group 2 cows with RDA or AV were hypokalemic (mean serum potassium concentration, 3.8 mEq/L). For group 2 cows, serum chloride concentration had the strongest positive association with serum potassium concentration, and serum potassium concentration was negatively associated with indices of feed intake (serum bilirubin concentration) and hydration status. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested hypokalemia was associated with hypochloremia, alkalemia, low feed intake with high amount of milk produced, hypovolemia, and hyperglycemia in lactating dairy cows. Treatment of hypokalemia should include surgical correction of abomasal displacement, increased dietary potassium intake via dietary dry matter intake or oral administration of KCl, and correction of hypochloremia, alkalemia, metabolic alkalosis, and dehydration.

Comparison of the fecal microbiota of healthy horses and horses with colitis by high throughput sequencing of the V3-V5 region of the 16S rRNA gene.

The intestinal tract houses one of the richest and most complex microbial populations on the planet, and plays a critical role in health and a wide range of diseases. Limited studies using new sequencing technologies in horses are available. The objective of this study was to characterize the fecal microbiome of healthy horses and to compare the fecal microbiome of healthy horses to that of horses with undifferentiated colitis. A total of 195,748 sequences obtained from 6 healthy horses and 10 horses affected by undifferentiated colitis were analyzed. Firmicutes predominated (68%) among healthy horses followed by Bacteroidetes (14%) and Proteobacteria (10%). In contrast, Bacteroidetes (40%) was the most abundant phylum among horses with colitis, followed by Firmicutes (30%) and Proteobacteria (18%). Healthy horses had a significantly higher relative abundance of Actinobacteria and Spirochaetes while horses with colitis had significantly more Fusobacteria. Members of the Clostridia class were more abundant in healthy horses. Members of the Lachnospiraceae family were the most frequently shared among healthy individuals. The species richness reported here indicates the complexity of the equine intestinal microbiome. The predominance of Clostridia demonstrates the importance of this group of bacteria in healthy horses. The marked differences in the microbiome between healthy horses and horses with colitis indicate that colitis may be a disease of gut dysbiosis, rather than one that occurs simply through overgrowth of an individual pathogen.

Application of strong ion difference theory to urine and the relationship between urine pH and net acid excretion in cattle.

OBJECTIVE: To develop an equation expressing urine pH in terms of independent variables, derive an equation relating urine pH to net acid excretion (NAE), and apply this new knowledge to determine the role that monitoring urine pH should play when diets with low cationanion difference are fed to dairy cattle. ANIMALS: 11 Holstein-Friesian cows. PROCEDURES: A physicochemical strong ion approach was used to develop a general electroneutrality equation for urine that involved urine pH and strong ion difference (SID [difference between strong cation and strong anion concentrations]), PCO(2), the concentration of ammonium ([NH(4)(+)]) and phosphate ([PO(4)]), and 3 constants. The general electroneutrality equation was simplified for use in bovine urine and applied to 321 data points from 11 cows fed different diets. RESULTS: Urine pH was dependent on 4 independent variables (urine SID, [NH(4)(+)], PCO(2), and [PO(4)]) and 3 constants. The simplified electroneutrality equation for bovine urine was pH approximately {pK(1)' - log(10)(S PCO(2))} + log(10)([K(+)] + [Na(+)] + [Mg(2+)] + [Ca(2+)] + [NH(4)(+)] - [Cl(-)] - [SO(4)(2-)]). The relationship between urine pH and NAE (in mEq/L) for cattle fed different diets was pH = 6.12 + log(10)(-NAE + [NH(4)(+)] + 2.6). CONCLUSIONS AND CLINICAL RELEVANCE: A change in urine SID, [NH(4)(+)], PCO(2), or [PO(4)] independently and directly led to a change in urine pH. Urinary [K(+)] had the greatest effect on urine pH in cattle, with high urine [K(+)] resulting in alkaline urine and low urine [K(+)] resulting in acidic urine. Urine pH provided an accurate assessment of NAE in cattle when pH was > 6.3.

Equine renal tubular disorders.

Renal tubular disorders have been sporadically reported in horses. Only three types of tubular defects have been recognized: (1) nephrogenic diabetes insipidus, attributable to unresponsiveness of the renal tubules to antidiuretic hormone; (2) distal renal tubular acidosis (RTA; type I); and (3) proximal RTA (type II). The following review focuses on RTA and nephrogenic diabetes insipidus.

Congenital colonic malformation ("short colon") in a 4-month-old standardbred foal.

During exploratory laparotomy of a foal with colic, a congenital abnormally developed large colon was identified incidentally. Long-term follow-up showed that the colt was more prone to gas-colic with diet and exercise changes than were other horses, due possibly to the short colon.

Natural and experimental infection of neonatal calves with Clostridium difficile.

Clostridium difficile toxins were associated with calf diarrhea in a recent retrospective study; however, no causal relationship has been prospectively investigated. This infection study tested whether the oral inoculation of neonatal calves with a toxigenic strain of C. difficile (PCR-ribotype 077) results in enteric disease. Fourteen 6-24 h old male colostrums-fed Holstein calves, received either three doses of C. difficile (1.4 x 10(8) +/- 3.5 x 10(8) cfu) (n = 8) or sterile culture broth (n = 6). Calves were euthanized on day 6 or after the onset of diarrhea, whichever came first. Fecal and intestinal samples were blindly cultured for C. difficile, and tested for its toxin A/B (C. difficile TOX A/B II ELISA, Techlab). PCR-ribotyping was used to compare inoculated and recovered isolates. Diarrhea was observed in all control calves and 3/8 of inoculated calves (p = 0.03), but it did not occur in calves that tested positive for C. difficile toxins. Fecal toxins were identified only from two controls. PCR-ribotyping confirmed the presence of C. difficile PCR-ribotype 077 in samples of all inoculated calves, but not from controls. The identification of five other PCR-ribotypes in 3/8 (37.5%) and 2/6 (33.3%) of inoculated and control calves, respectively, indicated early natural infection (< or = 24h of age). Five of 14 cecal samples had C. difficile (p = 0.01). In conclusion, the oral administration of C. difficile PCR-ribotype 077 to neonatal calves resulted in fecal/intestinal colonization but not in detection of toxins, or signs of enteric disease. Further studies are required to investigate the clinical relevance of C. difficile in calves.

Potential role of Clostridium difficile as a cause of duodenitis-proximal jejunitis in horses.

Duodenitis-proximal jejunitis (DPJ) is an idiopathic condition in the horse characterized by inflammation and oedema of the duodenum and proximal jejunum. Clinical signs include colic, ileus, depression, fluid accumulation in the small intestine and stomach, and endotoxaemia. The objective of this study was to investigate prospectively the role of Clostridium difficile in this idiopathic disease. Nasogastric reflux from 10 consecutive cases with DPJ and 16 consecutive horses with other causes of nasogastric reflux was cultured for C. difficile, other Clostridium spp., and Salmonella. Toxigenic strains of C. difficile were isolated from 10/10 (100%) of horses with DPJ and 1/16 controls (P<0.0001). No other known pathogenic clostridia were isolated from either group. Results of this study suggest that C. difficile might be an important cause of this syndrome.

Experimental determination of net protein charge, [A]tot, and Ka of nonvolatile buffers in bird plasma.

The quantitative mechanistic acid-base approach to clinical assessment of acid-base status requires species-specific values for [A]tot (the total concentration of nonvolatile buffers in plasma) and Ka (the effective dissociation constant for weak acids in plasma). The aim of this study was to determine [A]tot and Ka values for plasma in domestic pigeons. Plasma from 12 healthy commercial domestic pigeons was tonometered with 20% CO2 at 37 degrees C. Plasma pH, Pco2, and plasma concentrations of strong cations (Na, K, Ca), strong anions (Cl, L-lactate), and nonvolatile buffer ions (total protein, albumin, phosphate) were measured over a pH range of 6.8-7.7. Strong ion difference (SID) (SID5=Na+K+Ca-Cl-lactate) was used to calculate [A]tot and Ka from the measured pH and Pco2 and SID5. Mean (+/-SD) values for bird plasma were as follows: [A]tot=7.76+/-2.15 mmol/l (equivalent to 0.32 mmol/g of total protein, 0.51 mmol/g of albumin, 0.23 mmol/g of total solids); Ka=2.15+/-1.15x10(-7); and pKa=6.67. The net protein charge at normal pH (7.43) was estimated to be 6 meq/l; this value indicates that pigeon plasma has a much lower anion gap value than mammals after adjusting for high mean L-lactate concentrations induced by restraint during blood sampling. This finding indicates that plasma proteins in pigeons have a much lower net anion charge than mammalian plasma protein. An incidental finding was that total protein concentration measured by a multianalyzer system was consistently lower than the value for total solids measured by refractometer.

Clostridium difficile PCR ribotypes in calves, Canada.

We investigated Clostridium difficile in calves and the similarity between bovine and human C. difficile PCR ribotypes by conducting a case-control study of calves from 102 dairy farms in Canada. Fecal samples from 144 calves with diarrhea and 134 control calves were cultured for C. difficile and tested with an ELISA for C. difficile toxins A and B. C. difficile was isolated from 31 of 278 calves: 11 (7.6%) of 144 with diarrhea and 20 (14.9%) of 134 controls (p = 0.009). Toxins were detected in calf feces from 58 (56.8%) of 102 farms, 57 (39.6%) of 144 calves with diarrhea, and 28 (20.9%) of 134 controls (p = 0.0002). PCR ribotyping of 31 isolates showed 8 distinct patterns; 7 have been identified in humans, 2 of which have been associated with outbreaks of severe disease (PCR types 017 and 027). C. difficile may be associated with calf diarrhea, and cattle may be reservoirs of C. difficile for humans.

Experimental determination of net protein charge and A(tot) and K(a) of nonvolatile buffers in canine plasma.

Acid-base abnormalities frequently are present in sick dogs. The mechanism for an acid-base disturbance can be determined with the simplified strong ion approach, which requires accurate values for the total concentration of plasma nonvolatile buffers (A(tot)) and the effective dissociation constant for plasma weak acids (K(a)). The aims of this study were to experimentally determine A(tot) and K(a) values for canine plasma. Plasma was harvested from 10 healthy dogs; the concentrations of quantitatively important strong ions (Na+, K+, Ca2+, Mg2+, Cl-, L-lactate) and nonvolatile buffer ions (total protein, albumin, phosphate) were determined; and the plasma was tonometered with CO2 at 37 degrees C. Strong ion difference (SID) was calculated from the measured strong ion concentrations, and nonlinear regression was used to estimate values for A(tot) and K(a), which were validated with data from an in vitro and in vivo study. Mean (+/- SD) values for canine plasma were A(tot) = (17.4 +/- 8.6) mM (equivalent to 0.273 mmol/g of total protein or 0.469 mmol/g of albumin); K(a) = (0.17 +/- 0.11) x 10(-7); pK(a) = 7.77. The calculated SID for normal canine plasma (pH = 7.40; P(CO2) = 37 mm Hg; [total protein] = 64 g/L) was 27 mEq/L. The net protein charge for normal canine plasma was 0.25 mEq/g of total protein or 0.42 mEq/g of albumin. Application of the experimentally determined values for A(tot), K(a), and net protein charge should improve understanding of the mechanism for complex acid-base disturbances in dogs.

Use of a quantitative strong ion approach to determine the mechanism for acid-base abnormalities in sick calves with or without diarrhea.

Acid-base abnormalities are frequently present in sick calves. The mechanism for an acid-base disturbance can be characterized using the strong ion approach, which requires accurate values for the total concentration of plasma nonvolatile buffers (A(tot)) and the effective dissociation constant for plasma weak acids (K(a)). The aims of this study were to experimentally determine A(tot), K(a), and net protein charge values for calf plasma and to apply these values quantitatively to data from sick calves to determine underlying mechanisms for the observed acid-base disturbance. Plasma was harvested from 9 healthy Holstein-Friesian calves and concentrations of quantitatively important strong ions (Na+, K+, Ca2+, Mg2+, Cl-, L-lactate) and nonvolatile buffer ions (total protein, albumin, phosphate) were determined. Plasma was tonometered with CO2 at 37 degrees C, and plasma P(CO2) and pH measured over a range of 15-159 mm Hg and 6.93-7.79, respectively. Strong ion difference (SID) was calculated from the measured strong ion concentrations, and nonlinear regression was used to estimate values for A(tot) and K(a) from the measured pH and P(CO2) and calculated SID. The estimated A(tot) and K(a) values were then validated using data from 2 in vivo studies. Mean (+/- SD) values for calf plasma were A(tot) = 0.343 mmol/g of total protein or 0.622 mmol/g of albumin; K(a) = (0.84 +/- 0.41) x 10(-7); pK(a) = 7.08. The net protein charge of calf plasma was 10.5 mEq/L, equivalent to 0.19 mEq/g of total protein or 0.34 mEq/g of albumin. Application of the strong ion approach to acid-base disturbances in 231 sick calves with or without diarrhea indicated that acidemia was due predominantly to a strong ion acidosis in response to hyponatremia accompanied by normochloremia or hyperchloremia and the presence of unidentified strong anions. These results confirm current recommendations that treatment of acidemia in sick calves with or without diarrhea should focus on intravenous or PO administration of a fluid containing sodium and a high effective SID.