DETERMINATION OF FECAL ANALYTES

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: ANIMAL HEALTH
• Reporting Frequency: Annual
• Accession No.: 0204434
• Project Start Date: Oct 1, 2005
• Project End Date: Sep 30, 2010
• Program Code: [(N/A)]- (N/A)

Recipient Organization
PURDUE UNIVERSITY
(N/A)
WEST LAFAYETTE,IN 47907

Performing Department
VETERINARY CLINICAL SCIENCES

Non Technical Summary
Producers and veterinarians often can not determine the cause of diarrhea in large animal species. Because they must treat diarrhea symptomatically, many animals with non-infectious diseases are given antibiotics. This may lead to further disruption of normal gut function and increase the risk of antibiotic residue in animal-derived foods. The goals of this project are 1) to develop a means of determining fecal electrolyte concentrations by using specially designed probes to collect fecal ultrafiltrate, and 2) to use this technology in the field to characterize diarrhea in large animal species. We will measure fecal sodium, potassium, and chloride and show that they do not differ between fecal fluid and fecal ultrafiltrate. Normal fecal electrolyte concentrations and fecal osmotic gap values (FOG) will be calculated in a variety of large animal species under different management systems. It will then be determined if FOG can be used to differentiate osmotic from secretory diarrhea in a clinically relevant manner. It is expected that in instances of osmotic diarrhea fecal electrolyte concentrations will be low (high FOG) while in instances of secretory diarrhea fecal electrolyte concentrations will be high (low FOG). The potential impact of this research will be to allow producers and veterinarians to rapidly differentiate between fecal and osmotic diarrheas in large animal species. This is important because osmotic diarrheas generally do not require treatment with antibiotics.

Animal Health Component

70%

Research Effort Categories

Basic

20%

Applied

70%

Developmental

10%

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
305	3910	1020	30%
311	3910	1160	60%
315	3910	1160	10%

Knowledge Area
311 - Animal Diseases; 305 - Animal Physiological Processes; 315 - Animal Welfare/Well-Being and Protection;

Subject Of Investigation
3910 - Cross-commodity research--multiple animal species;

Field Of Science
1160 - Pathology; 1020 - Physiology;

Keywords

feces

fecal analysis

antibiotic residues

residue analysis

electrolytes

calves

diarrhea

gastrointestinal diseases

osmosis

secretion

animal diseases

animal pathology

animal physiology

chemical analysis

validation

sodium

potassium

chlorides

cows

swine

sheep

pathogenesis

Goals / Objectives
The overall objectives of this proposal are to validate a method for measuring fecal electrolyte concentrations in large animal species and then determine the accuracy of using fecal electrolyte concentrations to differentiate between osmotic and secretory diarrheas. The specific research objectives of this proposal are to: 1) Collect fecal ultrafiltrate using specially designed ultrafiltrate probes 2) Measure ultrafiltrate sodium, potassium, and chloride concentrations and determine if the electrolyte concentrations in fecal ultrafiltrate mirror total fecal values. 3) Use the measured electrolyte values along with the serum osmolarity to calculate the fecal osmotic gap (FOG). 4) Determine the normal FOG range in the cows and calves on various diets. 5)Measure fecal electrolytes in other large animal species: swine, ovine, and porcine models. 6) Measure FOG in calves with experimentally induced secretory and osmotic diarrhea, and determine if the FOG can be used to differentiate between the two. 7) Assess the ability of fecal ultrafiltrate electrolyte concentrations and FOG to characterize diarrheal type in large animal species with naturally occurring disease. 8) Assess if FOG determination provides insight into either the pathogenesis or response to treatment of large animal diarrheal diseases.

Project Methods
Feces will be collected from 10 calves. Feces will be weighed, and equal amounts of deionized water added. The mixture will be homogenized and a three-fiber ultrafiltrate probe inserted into the fecal slurry. A total of 5 ml ultrafiltrate will be collected per sample. This will be analyzed for sodium, potassium, and chloride using a clinical chemistry analyzer. Fecal electrolytes will also be determined using standard methodology. Concentrations of each analyte will be compared between fecal fluid and fecal ultrafiltrate using a paired T-test. Osmolality of serum will be determined using an Advanced Instruments Model 3D3 Osmometer. FOG will be calculated on all samples using the formula: FOG= serum osmolality - 2X (fecal Na + fecal K) Feces and blood samples will then be collected from a number of large animal species in various management systems. (At least 10 animals per group will be sampled). Twenty one-day-old dairy bull calves will be used for the next study. Calves will randomly be assigned to either the secretory or osmotic diarrhea group. They will be housed in individual stalls with free access to water and receive 10% of their body weight per day in commercial milk replacer. Osmotic diarrhea will be induced by oral administration of sucrose. Ten grams of sucrose will be added to the milk replacer at each feeding. Secretory diarrhea will be induced by oral administration of white phenolphthalein. This agent was chosen as it has been proven to cause a secretory diarrhea without introducing the presence of a toxic or infectious agent. White phenolphthalein will be given at a dosage of 10 mg/kg orally BID. The treatments will be continued until diarrhea is detected. Ten ml blood will be collected by jugular venipuncture and the osmolarity of the resultant serum will be measured. Analysis of variance will be used to compare sodium, potassium, and chloride concentrations between the fecal fluid and fecal ultrafiltrate. It is anticipated that no difference in electrolyte concentration will be detected. A paired T-test will be used to determine if the FOG or any of the electrolyte values differ significantly between control values and the two treatment groups. In addition to phenolphthalein, E.coli heat-stabile enterotoxin will be given to calves in order to induce a more severe secretory diarrhea. If FOG can be demonstrated as a useful way to differentiate diarrhea types experimentally, fecal and serum samples will be collected from veterinary patients with diarrheal disease. After the case is complete, either by death of the animal or discharge, the case record will be abstracted to determine if the exact etiology of the diarrhea was determined. It will then be determined if the FOG value or any single electrolyte concentration correlates with a specific type of diarrhea in animals with naturally occurring disease. Calculated FOG values will also be compared to outcome of the case, length of hospital stay, presence of complications, and presenting complaint. From this we will be able to determine if FOG is predictive of clinical course or outcome in hospitalized patients.

Progress 10/01/09 to 09/30/10

Outputs
OUTPUTS: There are no new outputs to report. There was a malfunction to the freezer where all samples were being stored. They thawed for an unknown period of time and could not be analyzed. PARTICIPANTS: Dr. Janice Sojka - Principal Investigator TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
The preliminary material has been presented to students in large animal medicine block courses.

Publications

• No publications reported this period

Progress 10/01/05 to 09/30/10

Outputs
OUTPUTS: The work in this project was terminated early before results were obtained. The primary community to which the information was disseminated was to students in the school of veterinary medicine at Purdue University. They received information on GI disease and diarrhea in several didactic and small group educational experiences. PARTICIPANTS: In addition to Dr. Sojka, the following people worked on this project: Dr. Elsa Janle, Department of Foods and Nutrition Dr. Laurie Gallatin, Veterinary Clinical Sciences Department. TARGET AUDIENCES: Veterinarians PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Because the work was terminated before all the results were obtained, there were no lasting outcomes to the project.

Publications

• No publications reported this period

Progress 10/01/08 to 09/30/09

Outputs
OUTPUTS: The output during this calendar year was the development of a new ultrafiltrate probe system that can be used to extract cell-free extracts of fecal slurry. PARTICIPANTS: Dr. Janice Sojka worked on this project. TARGET AUDIENCES: The target audience for this effort is primarily veterinarians, particularly food animal veterinarians. Secondary targets are animal producers, veterinary students, and physiologists. PROJECT MODIFICATIONS: There have been no major changes to the approach. However, development of a functional probe has been much more problematic and time consuming than originally believed.

Impacts
The impact of the probe development is that we can now begin measuring fecal electrolyte concentrations on fecal samples using standard chemistry analyzers with no additional post-collection processing.

Publications

• No publications reported this period

Progress 10/01/06 to 09/30/07

Outputs
OUTPUTS: Results have been disseminated to large animal and food animal practitioners via one on one discussion and consultation. TARGET AUDIENCES: Target audience is large animal, mixed practice, and food animal veterinary practitioners.

Impacts
Development of a means to collect fecal ultrafiltrate resulted in achieving the anticipated outcomes in this study.

Publications

• No publications reported this period

Progress 10/01/05 to 09/30/06

Outputs
Producers and veterinarians often can not determine the cause of diarrhea in large animal species. Because they must treat diarrhea symptomatically, many animals with non-infectious diseases are given antibiotics. This may lead to further disruption of normal gut function and increase the risk of antibiotic residue in animal-derived foods. The goals of this project are 1) to develop a means of determining fecal electrolyte concentrations by using specially designed probes to collect fecal ultrafiltrate, and 2) to use this technology in the field to characterize diarrhea in large animal species. To date, the sample collection is complete and data analysis has begun. The serum osmolality has been determined on each calf at every testing point. The average serum osmolality was 283 +/- 7.3 mOsm. There was no difference between control and treatment groups in this regard. This was not unexpected as the calves were not ill or appreciably dehydrated during the trial period. The fecal ultrafiltrate has been prepared, and will be analyzed for electrolyte content in the coming months. All data analysis should be complete by 3/2007.

Impacts
The long term impact of this research will be to reduce antibiotic use in calves and other food producing animals. This is because once the FOG system is validated, producers will have a means to rapidly differentiate diarrhea into either secretory or osmotic types. If an osmotic type was diagnosed (thought to be the most common type) the producer would know that no health benefit would be expected by antibiotic administration. Management factors could be reviewed if an osmotic diarrhea were found, and potential viral or parastitic etiologies could be investigated

Publications

• No publications reported this period

Progress 10/01/04 to 09/30/05

Outputs
The work to date is in the preliminary stages. To investigate the effect of osmotic and secretory diarrhea on fecal electrolytes, the following experiment was performed: 8 1-3 week-old Holstein bull calves were used in this portion of the experiment. They were fed colostrum till 24 hours of age, then commercial calf replacer. Blood and feces were collected for control samples. All samples at all time points were frozen at -80 degrees within 3 hours of collection. Osmotic diarrhea was induced by oral administration of sucrose (Domino Table Sugar). Five hundred grams of sucrose was added to the milk replacer at each feeding. The sucrose was given until the calf developed diarrhea as assessed by the presence of semi-liquid or liquid feces. This occurred within 12 hours of sucrose administration in all instances. Feces and serum were collected for analysis. Once samples were collected, the treatment was discontinued. All calves regained normal fecal consistency within 24 hours of cessation of sucrose. To induce a secretory diarrhea, phenolphthalein was given at a dosage of 25 mg/kg orally BID. It was given until diarrhea was detected, at which point feces and serum were collected and the treatment immediately stopped. Phenolphthalien-induced diarrhea was extremely transitory in nature, and even high doses of this substance had little effect. This was a cross over design; each calf was given both sucrose and phenolphthalein in a randomized fashion. Although not originally designed as a cross over, the calves developed diarrhea so quickly after treatment, and then had it resolve so rapidly after treatments were discontinued that it was felt that this was a valid experimental design. A minimum of a 48 hour wash out period was used between treatments. At the end of the final collection period, the calves were humanely euthanized. All samples are currently frozen awaiting analysis.

Impacts
The long term impact of this research will be to reduce antibiotic use in calves and other food producing animals. This is because once the FOG system is validated, producers will have a means to rapidly differentiate diarrhea into either secretory or osmotic types. If an osmotic type was diagnosed (thought to be the most common type) the producer would know that no health benefit would be expected by antibiotic administration. Management factors could be reviewed if an osmotic diarrhea were found, and potential viral or parastitic etiologies could be investigated.

Publications

• No publications reported this period