Examination of Rumen Fluid (PDF)

‫بسم هللا الرحمن الرحيم‬ BSU University Vet Medicine Physiology depart practical part Examination of rumen fluid Dr. salah hosny Assistant lecturer at fuc.vet.med.BSU.univ. Animal physiology specialist Master degree in vetrinar...

‫بسم هللا الرحمن الرحيم‬ BSU University Vet Medicine Physiology depart practical part Examination of rumen fluid Dr. salah hosny Assistant lecturer at fuc.vet.med.BSU.univ. Animal physiology specialist Master degree in vetrinaryscience.Topics. 1.Indication. 2.Methods of collection. 3.Methods of examination. 1.Indication. 1.Diagnosis of rumen disease. 2.Therapeutic purposes. 2.Methods of collection. 1.Needle puncture of the rumen : -Helps eliminate salivary contamination. 2.Oral or nasal passage of stomach tube : -Helps avoid peritoneal contamination. -Discard first 100 ml to reduce salivary contamination. 3.Manual method from slaughtered animal. 3.Methods of examination. 1.Physical examination : -Color. -Consistency. -Odor. 2.Chemical examination : -PH. -Sedimentation activity test. -Methylene blue reduction test ( redox potential ). -Nitrate reduction test ( NRT ). -Cellulose digestion test. -Glucose fermentation test. -Rumen fluid chloride. -Other tests. 3.Examination for protozoa ( fauna ). 4.Examination for bacteria 5.Examination for fungi  1.Physical examination. -Color : Color Causes  Normal Yellow / brown Corn silage/straw diet color varies Normal Olive / brown Concentrate based ration depending on the Green Pasture-based diet nature of feed Milky gray grain overfeeding (lactic acidosis) prolonged ruminal stasis and/or Abnormal Dark greenish or brown decomposition/putrefaction of rumen contents. Gray with clots of milk Abomasal reflux (calves) 1.Physical examination. -Consistency : Consistency Causes Normal -Slightly viscous Inactive bacteria and protozoa, Watery Starvation Abnormal Frothy (Bubbles) Bloat  1.Physical examination. -Odor : Odour Causes Normal Aromatic Ammonia smell Urea toxicity Moldy rotting Protein putrefaction Abnormal grain overfeeding (lactic acidosis) Acidic/ sour  2.Chemical examination. -PH :  Immediately after sampling.  Using PH paper or PH meter. pH Causes Normal 6–7 5.0-5.6 Subacute rumen acidosis  5.2 Lactic acidosi Abnormal 7.5 – 8.5 -Simple indigestion - Starvation 8.0 – 10 -Urea poisoning 7.5 Putrefation / protein decomposition  2.Chemical examination. -Sedimentation activity test :  Immediately after sampling.  Provide a rapid evaluation of microfloral activity.  Put sample of rumen fluid in test tube and let to stand.  Measure time for sedimentation of fine particles and flotation of coarse particles. Sedimentation time Causes Normal 4 - 8 minutes Rumen acidosis. Rapid sedimentation Simple indigestion. with no floatation Starvation Abnormal Inactive microflora from indigestible roughages No sedimentation Frothy bloat. or floatation Vagal indigestion.  2.Chemical examination. -Methylene blue reduction test(redox potential):  Measure metabolic activity of ruminal flora.  20 ml of rumen fluid + 1 ml of 0.03 % MB in TT.  Let stand at RT.  Measure time needed for color change. Methylene blue Causes reduction time Normal 5 -6 minutes Active rumen microbes Abnormal up to 15 minutes Diminished microfloral activity e.g., indigestible roughage, rumen acidosis.  2.Chemical examination. -Cellulose digestion test :  10 ml rumen fluid + 0.3 ml of 16 % glucose in TT.  Suspend a thread of pure cellulose or cotton thread tied by a glass bead to the lower end to immerse in the rumen fluid.  Tightly close the tube , incubate at body temp 39 c.  Record time for the thread to be broken. Cellulose digestion time Causes Fully active rumen fluid, Normal 48-54 hours resulting in digestion of cotton thread Abnormal Not broken within normal time Inactive rumen fluid.  2.Chemical examination. -Glucose fermentation test :  Indirectly measure the ability of microflora to break down glucose through measuring the volume of gas formed.  10ml of rumen fluid ± 0.5 ml of 16 % glucose solutsion.  Place at fermentation saccharometer and keep at 39 c.  Read result after 30 min and 60 min.  N.B : normal rumen fluid containing active microflora will ferment the glucose and result in gas formation. Rate of gas Causes fermentation Normal 1-2 ml gas/hr Active microflora Inactive microflora No or decreased gas Abnormal e.g., decomposition, rumen alkalosis or acidosis. Increased gas Bloat. 2.Chemical examination. -Nitrate reduction test ( NRT ) :  Measure activity of ruminal flora that degrade and synthesize nitrogen compounds.  10 ml of sieved ruminal fluid into each of 3 TT.  Add 0.2 , 0.5 and 0.7 ml of 0.025 % pot nitrate to the three TT.  Put the 3 TT in water bath at 39 c.  Every 5 min 1 drop of each tube is placed in a small ceramic plate.  To each drop add : - 2 drops of reagent 1 ( 2 ml sulphanilic acid in 30 % acetic acid to make 200 ml ). - 2 drops of reagent 2 ( 0.6 ml alpha-naphthylamine ± 16 ml conc acetic acid ± 140 ml DW ). - Observe change in color. Interpretation :  Samples that contain nitrates are colored red.  Rumen fluid of cattle fed a mixed ration will not change color after 5-10 min in tube 1 and 20 min in tube 2 and 30 min in tube 3.  Reduction is more rapid when cattle are fed green fodder or have ruminal decomposition or bloat.  Reduction is more slower when adeficient ration is fed or when the animal lacks appetite. 2.Chemical examination. -Rumen fluid chloride :  Measured in supernatant of a centrifuged sample.  Determined by portable chloride meter.  Normal rumen fluid chloride conc is less than 30 mEq\L.  Elevated rumen fluid chloride conc indicates : - Reflux of abomasal content into the rumen. - Supplementation of diet with salt. 2.Chemical examination. -Some other tests :  For evaluation of activity of the rumen microflora :   Measurement of titratible acidity.  Measurement of volatile fatty acids.  Measurement of lactic acid.  Measurement of ammonia.  Measurement of concentration of sodium, potassium.  Measurement of activity of enzymes 3 ) Examination for protozoa (fauna) : I ) Qualitative examination : 1 ) Motility : Technique: A drop of fresh rumen fluid is examined using the low power objective on a warm side at about 30°C. 3 ) Examination for protozoa (fauna) I ) Qualitative examination 1 ) Motility Interpretation: - Motility of rumen microfauna is judged as follows: Judgement on motility of rumen Causes microfauna +++ Highly motile and very crowded. Normal ++ Motile and crowded. + Sluggish motility and low number. Abnormal Rumen No or sporadic alive acidosis 0 microflora 3 ) Examination for protozoa (fauna) 2 ) Quantitative examination - Technique: ????? - Interpretation:  In cases of rumen function disturbances large species of microfauna disappear firstly, followed by medium and finally the small species.  The entire microfauna dies during severe digestive disorders, particularly when pH falls below 5.0.  In recent cases of moderate digestive disorders there is an increase in the proportion of dead to live protozoa.  4 ) Rumen microflora (Bacteria)  The rumen microflora is comprised of a large number of species with a variety of morphological forms, depending chiefly on the composition of the ration. - Cattle fed on high fibre diet have lower microbial activity. - Cattle fed on roughage have a high proportion of large bacterial forms. - Cattle fed on mixed rations should have mixed-size bacterial forms. - Cattle fed on high-grain diets should have relatively a uniform proportion of small bacterial forms. 4 ) Rumen microflora (Bacteria)  Rumen fluid from normal healthy cattle should have only a small percentage of Gram-positive bacteria.  A high proportion of Gram-positive bacteria (sometimes no Gram-negative bacteria at all) is indicative of lactic acidosis.  Absence of Gram-negative bacteria in the rumen sample is an abnormal finding.  3 ) Rumen Fungi  Present as yeasts e.g. Candida sp.  Sometimes present in exceptional numbers during rumen acidosis