Rumen Fluid Analysis

Oreta M. Samples, RVT, MPH, DHSc

KVMA Mid-America Veterinary Conference

INTRODUCTION

The analysis of rumen fluid is carried out to assess the function and activity of the ruminant forestomach
as well as the diagnosis of diseases of the forestomach. This is a task for which the veterinary technician
may be quite useful providing that they are familiar with ruminant anatomy as well as physiological
considerations regarding handling and testing of collected fluids.

DISCUSSION

Basic Anatomy and Physiology Refresher:

It is important to have a working knowledge of the ruminant digestive tract when attempting sampling of
rumen fluid. The ruminant digestive system is made up of four compartments: rumen, reticulum, omasum
and abomasum with each section being responsible for specific digestive functions. The “forestomach”
contains the rumen, reticulum and omasum while the abomasum is regarded as the “true stomach”. Table
1 shows the different functions of each of the four areas.

TABLE 1

Section Function
Rumen Mixing/churning digesta

Reticulum Mixing/distribution of anaerobic bacteria

Omasum Absorbs water and salts

Abomasum Secretes enzymes, etc. for digestive breakdown

Basic digestion within ruminants begins with the animal chewing up food that is subsequently mixed with
bicarbonate/phosphate enriched saliva. Once swallowed, the forestomach begins to contract mixing even
more buffers with the forage. At this point, the animal regurgitates their food and re-chews it. This is
commonly known as “chewing cud”, an action that allows for more saliva to be mixed with foodstuff
before being swallowed. The food then moves throughout the other three compartments for complete
digestion.

The rumen, the first compartment to contribute to digestion is itself an ecosystem populated with a variety
of unicellular organisms. These organisms are responsible for digestion of foodstuffs, utilization of
resulting sugars and acids, synthesis of vitamins and production of waste products such as methane. The
rumen fluid when collected and analyzed allows the clinician to assess the number of functional
anaerobes, assess the functional capability and monitor output of waste materials.

Collection:
The collection of rumen fluid is not a difficult procedure; however it must be undertaken in a humane
manner. Because of the nature of the collection method, it is impossible to render the process a strictly
sterile procedure; however it should be carried out as aseptically as possible. After all, the process is
being carried out in an anaerobic environment, in an attempt to retrieve living anaerobic bacteria for
analysis, therefore nothing should be introduced to the environment; (for this reason) one should avoid
excessive salivary contamination of the orogastric tube as well as other outside contaminants when
passing tube.

There are two methods of rumen fluid collection: orogastric collection and ruminocentesis. Ororuminal
collection is often favored when small amounts of fluid must be collected and may be attempted by a
veterinary technician due to the ease of the procedure. Ruminocentesis by contrast, yields larger amounts
of fluid and lacks the salivary contamination sometimes experienced with orogastric collection. The
negative aspect of ruminocentesis is the measure of difficulty in locating rumen and the chance of post
operative infection of the puncture site. Also, while a veterinary technician may attempt successful
ororuminal collection, a veterinarian should perform the act of ruminocentesis which is oftentimes
regarded as a surgical procedure. For the purpose of veterinary technician education, the remainder of this
presentation will focus on ororuminal collection and subsequent analysis.

Ororuminal Methodology:
This procedure is best accomplished with the cow firmly secured within a head gate, the head restrained
by halter and rope. For safety reasons, always ensure the animal is under positive control of restrainer. It
is helpful to have a dedicated restrainer in addition to the individual attempting collection.

Standing close to cow, wrap arm around top of cows’ neck using hand to open mouth; insert a Frick’s
Speculum into the mouth and directed slightly up and over the tongue to provide access into throat. See
Table 2 for the difference between a good passage and a bad passage of Frick’s Speculum.

TABLE 2
TABLE 2: PASSAGE: GOOD VERSUS BAD
GOOD PASSAGE BAD PASSAGE
Gurgling sounds heard Coughing
Smell of rumen fluid (sour-sweet) Air passage thru speculum is noted

Once the speculum is confirmed to be correctly placed, thread the orogastric tube through the Frick’s
Speculum, down the throat and into the rumen. In a manner similar to confirming placement of the
speculum, the tube is confirmed as being in the rumen by gurgling sounds which may be heard as well as
the scent of fermented rumen fluid (sour-sweetish) detected. Once the tube is confirmed to be in place,
there are two methods which may be used for collecting the fluids as denoted below:

COLLECTION METHOD # 1

The collection is begun by kinking the tube which traps fluid in the end of the tube by negative pressure.
At this point, someone should be standing by with a glass container readied to catch the fluid. Swiftly but
gently withdraw the tube through the speculum and empty the fluids into the waiting vessel. Frick’s
Speculum is likewise withdrawn.

COLLECTION METHOD # 2

After confirming the placement of the tube in the rumen, attach a dosing syringe to the tube and aspirate
to maximum ability, then kink tube and withdraw slowly and steadily; empty fluid immediately into
collection vessel. This may yield more fluid volume than the first method. Regardless of the method
which is used, beware – cows often sneeze/cough upon withdrawal of orogastric tube…on you.

Because of the frailty of the rumen bacteria, consider point of care (POC) testing for performing the pH
and Methylene Blue Reduction Test. The longer the fluid is out of the body, the less reliable the results
will be. Once collection is completed, the cow may be released, howver, before releasing cow from chute,
evaluate breathing, cow should not be coughing or breathing heavily. If satisfied release the cow.

LABOATORY TESTING:

Physical characteristics such a color, odor and consistency should be noted immediately upon collection.
The physical characteristics are directly influenced by the animal’s diet and general digestive state of
health. Table 3 illustrates some possible causes for various differences in these physical characteristics.
Because diet oftentimes contributes to the color of the rumen fluid, it is a good idea to get feeding history
during talks with owner. Also, technicians should familiarize themselves with the smell and appearance of
normal rumen fluid as to the untrained eye, normal fluid may appear abnormal.

TABLE 3
TABLE 3: Physical Characteristics Evaluation
Color Evaluation
Yellow/Brown Corn silage/straw diet
Brown/Olive Concentrate diet
Green Pasture Diet
Milky gray/Brown Lactic Acidosis

Odor Evaluation

Aromatic Normal
Acidic/sour Lactic Acidosis
Rotting Rumen Putrification/Infection

Consistency Evaluation

Excess Viscosity High saliva content

Watery, few particles Anorexic
Bubbles Bloat

The pH should be measured immediately after collection with the most reliable results being obtained
from samples collected 4-8 hours after consumption of a fresh total mixed ration. Consider point of care
testing for this result for accuracy. Technicians should document for further evaluation the time of last
meal as well as the diet which the animal is currently eating for later use by the diagnostician. Table 4
below provides a listing of the pH values one might expect to see and possible interpretations.

TABLE 4

TABLE 4: pH Values

pH Interpretation

8 and above Saliva contamination, putrification

7-8 Reduced feed intake
6-7 Normal pH of cattle
5 – 5.6 High grain diet or pasture fed/early
Lactic Acidosis
5.5 Lactic Acidosis

Methylene Blue Reduction tests may be done to assess the number of functional anaerobic bacteria
available within the rumen. Prepare a test tube by adding .5 mL of a .03% solution of Methylene Blue
stain. Add 10 mL of fresh rumen fluid and set a timer. Always add rumen fluid to the dye; never reverse
the sequence as the results will not be accurate. Note the amount of time it takes for the rumen solution to
clear of color. Table 5 indicates the time necessary to assess functionality of present bacteria.

Table 5
TABLE 5: METHYLENE BLUE REDUCTION TEST

Time Interpretation

2-6 minutes Adequate bacteria are present

10+ minutes Inadequate bacteria are present

The Chloride concentration test is often of interest in cases of possible pylorus obstruction/blockage.
Because elevated levels of chloride can cause reflux of the abomasum (a type of internal vomiting, not to
be confused with regurgitation) it may be necessary to measure the amount of chloride present in rumen
fluid. Because this requires determination using a chloride meter, samples are generally sent overnight to
diagnostic laboratory facilities for results.

Protozoan presence in rumen fluid is measured through microscopic examination of one drop of fluid on a
slide. Protozoa are counted that as they appear in one field and should be moving (motile). To assess their
energy levels add a drop of Lugol’s Iodine; healthy protozoa will be stained uniformly while those with
decreased energy supplies will not. Protozoan inhabitants of the rumen are found in smaller numbers than
bacteria however because they are significantly larger than bacterial organisms, their members may
occupy an equal amount of volumetric space as bacteria. The types of protozoa are also more specific
with most being anaerobic ciliates – most notably holotrichous (cilia covers entire organisms’ surface)
and entodiniomorphid (devoid of cilia except for over the adoral zone).

Bacterial identification is generally accomplished through Gram Staining exercises of rumen fluid.
Although Gram positive bacteria may be seen, the fluid predominantly will be populated with Gram
negative bacteria of a heterogeneous nature unless there is a case of lactic acidosis which renders a larger,
more uniform population of gram positive bacteria. It is also expected to see bacteria which are both
motile and non-motile within the sample. Note that the longer the fluid is outside of the animals’ body,
the less motile the bacteria will be. Avoid cold shock that may occur if container of fluid is sat on cool
surface or refrigerated. Bacterial identification should be done soon after collection for best results as
these fermenters of carbohydrates are subject to death in an aerobic environment.

CONCLUSION

Ruminant species by the nature of their anatomy provide unique challenges both in the practice of
effective nutritional husbandry as well as diagnostically. The simplicity of diagnosis and treatment may
be greatly enhanced by a technician’s possession of knowledge and skills to aid in collection as well as
laboratory testing of rumen fluid. For those who enjoy the challenges of working with large animal
species, the ability to not only safely collect but accurately test rumen fluid makes the technician a
valuable asset to the large animal clinician.

REFERENCES

Anderson, D. &. (2009). Current Veterinary Therapy: Food Animal Practice, 5th Ed. St. Louis, MO:
Sauders-Elsevier.

Bassert, J. &. (2010). McCurnin's Clinical Textbook for Veterinary Technicians, 7th Ed. St. Louis, MO:
Saunders-Elsevier.

Bowen, R. (2009, November). Rumen Physiology and Rumination. Retrieved from Colorado State
University: http://www.vivo.colosate.edu/hbooks/pathphys/digestion/herbivores/rumination.html

Chuba, L. (2009). Rumen Microbiology and Fermentation. In L. Chuba, Animal Nutrition Handbook (pp.
55-79). Auburn, AL: Self-published.

Hall, J. &. (2009). Nutrition and Feeding of the Cow-Calf Herd: Digestive System of the Cow. Retrieved
from Virginia Cooperative Extension: http://pubs.ext.vt.edu/400/400-010/400-010_pdf.pdf

N.A. (2012). Answers.com. Retrieved from Answers.com:

http://www.answers.com/topic/ruminant#ixzz1mwTVXRDP

Reece, W. (2000). Functional Anatomy and Physiology of Domestic Animals, 3rd Ed. Philidelphia, PA:
Lippincott, Williams and Wilkins.

Rockett, J. &. (2007). Veterinary Clinical Procedures in Large Animal Practice. Clifton Park, NY:
Delmar-Cengage.