Morphological Characterization Of Rumen Protozoa Isolated From Sweet Sorghum Hydrolysis Using Carabao (Bubalus bubalis Linn) Rumen Fluid


  • Perla Florendo PCC AT CLSU



Rumen protozoa, Philippine Carabao, Hydrolysis, Bioethanol, Bagasse, Feeds


This study was conducted to characterize the morphology of rumen protozoa isolated in the batch type hydrolysis of sweet sorghum using 1% fresh carabao rumen fluid with duration of 0, 3, 6, 9, 12 and 15 days of the novel process hydrolysis. Hydrolysis was conducted using 5% chopped sweet sorghum bagasse. Nitrogen content was augmented with 0.35 g urea per 750 ml effective volume with low speed agitation of fermentation bottle at least 20 minutes twice daily and incubated at room temperature. Characterization was limited to microscopic evaluation of the morphological features of protozoa that was stained with methylene blue formalin saline solution. Protozoon population of 2.61 x 10 4 cfu/ml at initial period had significantly lowered down at the 15 days hydrolysis to 0.62 x 104 cfu/ml. Evaluation of morphology for the types of protozoa at different durations showed the predominance of species types with different orientations of caudal spines, shapes of the macromolecules, size of the body, and presence of adoral ciliary zones were features of species types from genus Eodinium Entodinium of the family Ophryoscolecidae. Evaluation of the hydrolysis condition showed that duration had significant effect on the pH (P>0.05). Sweet sorghum hydrolysis has initial pH 7 that significantly declined to pH 5 at 15   days duration. The population of the protozoa in the rumen fluid hydrolysis was significantly affected by duration of the hydrolysis (P>0.05). Entodinium were isolated at all durations of the acidic hydrolysis of sweet sorghum bagasse using carabao rumen fluid. In conclusion, morphological characteristics of the rumen protozoan shows how diverse are the composition of rumen protozoa in the carabao rumen fluid hydrolysis. The presence of the rumen protozoa is a justification of the novel carabao rumen fluid hydrolysis conversion of lignocelluloses in sweet sorghum bagasse into soluble carbohydrates for bioethanol production. The information is vital for animal feed utilization and bioethanol production optimization.


ABE, M., IRIKI, T., TOBE, N. and I. SHIBUIH. (1981). Sequestration of Holotrich protozoa in the reticule-rumen of cattle. Applied and Environmental Microbiology 41: 758-765.

ABENES, F. and P. FLORENDO. (2008) Bacteria in water buffalos rumen may help produce cellulose biofuels.,

ABOU AKKADAA, R. & H. HOWARDB. (1960). The biochemistry of rumen protozoa.3. The carbohydrate metabolism of Entodinium. Biochem. J. 76, 445.

BAUCHOP, T. and T. CLARKER. (1976). Attachment of the ciliate Epidinium crawley to plant fragments in sheep rumen. Applied and Environmental Microbiology 32: 417-422.

COLEMAN, G. S. (1979). The role of rumen protozoa in the metabolism of ruminants given tropical feeds. Tropical Animal Production, 3:199-213.

COLEMAN, G.S.(1998) Rumen Ciliates Protozoa.Advance in Parasitology.Vol 18:121-173.

DEHORITY, B. A. (1993). Laboratory manual for classification and morphology of rumen ciliate protozoa. The rumen microbial ecosystem. CRC Press, Boca Raton 593.1’7—dc 20.

DEHORITY, B.A. and R.S. MATTOWS. (1978). Diurnal changes and effect of ration on concentrations of the rumen ciliate Charonventriculi. Applied and Environmental Microbiology 36: 953-958.

DEHORITY, B.A. and P.A. TIRABASSO. (1989). Factors affecting the migration and sequestration of rumen protozoa in the family Isotrichidae. Journal of General Microbiology 135: 539-548.

DEMEYERD, I. (1981). Rumen microbes and digestion of plant cell walls. Agriculture and Environment 6: 295-337.

FLORENDO, P. DC. S.P. BANGIT, F.L. MAMUAD, E. C. ATABAY AND L.C. CRUZ (2017). Cryopreservation of Bubalus bubalis L Rumen Bacteria: Effect on Viability and Efficiency of Conversion of Crop Residues into Soluble Sugars for Biofuel Production. International Journal of Agricultural Technology 13(7.1):1131-1145.

FLORENDO, P. DC. SHARMA-SHIVAPPA, R. and FELLNER, V. (2018). Cattle rumen microorganisms hydrolysis for switchgrass saccharification, volatile fatty acids and methane production. International Journal of Agricultural Technology 14(1):31-43.

FLORENDO, P. DC. S.P. BANGIT, F.L. MAMUAD, E. C. ATABAY AND L.C. CRUZ (2022). Microbial Identification and Population Succession In the Novel Carabao Hydrolysis Pretreatment of Crops Residues Lignocelluloses Intended for Cellulose Ethanol Production. CLSU International J. Science and Technology

FRANZOLIN, R. and B.A. DEHORITY, (1996). Effect of prolonged high-concentrate feeding on rumen protozoa concentrations. Journal of Animal Science 74: 2803-2809.

HUNGATE, R.E., REICH, L.J., and S.R. PRIN. (1971). Parameters of rumen fermentation in a continuously fed sheep: evidence of a microbial rumination pool. Applied Microbiology 22:1 104-1 11 3.

IPCC (2007). Climate Change (2007): The Physical Science Basis. Contribution of Working Group I to the Fourth Assessment Report of the Intergovernmental Panel on Climate Change. Edited by S. SOLOMON, D. QIN, M. MANNING, M. MARQUIS, K.AVERYT, M. M. B.TIGNOR, H. L. MILLER, Jr & Z. CHEN. Cambridge: Cambridge University Press.

JOHNSON, K.A. and D.E. JOHNSON (1995).Methane emissions from cattle. J Animal Science 73, 2483–2492.

LIN, Y. and S. TANAKA. (2006). Ethanol fermentation from biomass resources: current state and prospects. Appl. Microbial. Biotechnol. 69(6): 627-642.

NAGARAJA, T.G. and E.C. TITGEMEYER (2007). Rumen acidosis in beef cattle: the current microbiological and nutritional outlook. Journal of Dairy Science 90: 17-38.

ORPIN, C.G. and J. LETCHERA. (1978). Some factors controlling the attachment of the Rumen holotrich protozoa Isotrichia intestinalis and I Prostomato plant particles in vitro. Journal of General Microbiology 106: 33-40.

OWENS, F.N., SECRIST, D.S. and W.J. HILL. (1998). Acidosis in cattle: a review. Journal of Animal Science, 76: 27-286.

MOSSEL, D., CORRY, A. and CORRY, J.E., STRUIJK, C.B. and R. M. BAIRD. (1995). Essentials of

the microbiology of foods: a textbook for advanced studies. Chichester (England): John Wiley and Sons. 699

POTTER E.L. and B.A. DEHORIY. (1973) Subsequent Daily Digestibility in the Ovine Effects of Dietary Change or Rumen Inoculation upon J Anim Sci, 37:1408- 1413.

PUNIAB, S., LEIBHOLZ, J. and J. FAICHNEGY (1987). The role of rumen protozoa in the utilization of paspalum (Paspulum dilututum) hay by cattle. British Journal of Nutrition 57: 395- 406.

PURSER, D.B. (1961).A diurnal cycle for holotrich protozoa of the rumen. Nature, London190: 831- 832.

USHIDA, K. (2011). Symbiotic Methanogens and Rumen Ciliates. Microbiology Monographs, 19: 25- 34.

TOMPKIN, RB (1963). The establishment of the Rumen Protozoa, Entodinium IN VITRO. The Ohio State University, P h .D.

WELLER, R.A. and F. PILGRIMA. (1974). Passage of protozoa and volatile fatty acids from the rumen of the sheep and from a continuous in vitro fermentation system. British Journal of Nutrition 32: 341-351.

WILLIAMS A.G and G. S. COLEMAN. (1997). The Rumen in The Rumen Microbial Ecosystem, 3: 1-3.