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Studies on Rumen Fluid Analysis in Ruminal Acidotic Goats

Tambe Satish Valmik K. Padmaja P. Nagaraj A. Gopala Reddy
Vol 7(9), 250-258
DOI- http://dx.doi.org/10.5455/ijlr.20170716012034

Thirty goats that were investigated for ruminal acidosis, with the history of accidental ingestion of carbohydrate rich diet, inappetence to anorexia, suspended rumination and clinical manifestations of distended rumen and diarrhoea were selected and were divided based on ruminal fluid pH and severity of ruminal acidosis into three different groups consisting of 10 goats in each. Ruminal fluid analysis was undertaken and compared against ten apparently healthy goats (group IV) from the same area with similar husbandry practices which served as healthy control. Basing on ruminal fluid pH, these goats were divided as mild acidosis (pH=5.5 - 6.5), moderate acidosis (pH = 4.5 - 5.5), and severe acidosis (pH = 4.0 - 4.5) group I, II and III respectively. Ruminal fluid parameters such as Methylene Blue Reduction Time (MBRT), Sedimentation activity time (SAT) and protozoal dead count significantly increased in group I. While, Ruminal acidotic goats had decreased rumen fluid parameters such as rumen fluid pH, protozoal motility, density, and iodophilic activity in group I. Rumen fluid color was grey in group I and milky grey in both group II and III, consistency was semisolid or slightly watery in group I and watery in both group II and III, odour was acidic in both group I and II and pungent acidic in group III of acidotic goats. Complete absence of protozoa in rumen fluid of both group II and III, and replacement of rumen fluid gram negative bacteria by gram positive bacteria in all acidotic goats. Based on study of all above parameters, the goats were confirmed as suffering from ruminal acidosis.


Keywords : Dietary Abnormalities Ruminal Acidosis Rumen Fluid pH MBRT SAT

 

Introduction

Goats were among the first animals to be domesticated. As indicated by the archaeological evidence, they have been associated with man in a symbiotic relationship for upto10, 000 years (Ensminger and Parker, 1986). India has the largest population of goats in the world. F.A.O. (2010) estimated that there are around 920 million goats in the world out of which 154 million goats are found in India and 150.7 million goats in China (Jindal, 2013). The erstwhile Andhra Pradesh contributes 9.629 million goats population according to animal husbandry annual report 2012-2013 (18th Livestock census, 2007). The goat is a multipurpose animal which provides meat, milk, fibers (pashmina, mohair and cashmere), skin, bones and manure. It is known as the ‘poor man’s cow’ in India and as ‘wet nurse of infants’ in Europe. Goat can be kept with little expense. Marginal or undulating lands unsuitable for other type of livestock may be used and inexpensive shelter will suffice. Of all the domestic animals, goats undoubtedly have the greatest range of adaptation. A goat thrives well on poor agricultural lands where they browse on incidental vegetations. Goat has ability to get acclimatized under diversified agro-climatic condition, unfastidious type in choosing of available forest and short generation period. Goat have a wide range of adaptive traits which enable them to survive and produce, including disease resistance, heat resistance, water tolerance, ability to cope with poor feed quality (Baker and Grey, 2004). Majority of health problems encountered in goats are rumen related. Rumen acidosis is abnormal fermentative disorder, a common aliment that was presented to peri-urban clinics. People living here rear goats in semi-intensive method are more susceptible to rumen lactic acidosis (Padmaja and Praveena, 2011).

Acidosis is also known as lactic acidosis, rumen acidosis or grain overload is a carbohydrate fermentation disorder of the rumen that can affect animals of all ages. As the name implies, acidosis results in acidic pH of rumen (normal being 6.2-6.8) which is caused by feeding of highly fermentable carbohydrates, feeding of low fiber diet, poor management practices or a combination of these. Degree of acidosis varies from seriousness, a slight drop in feed intake (mild) to death (severe). Acute form of the disease in ruminants is characterized by indigestion, rumen stasis, toxemia, in coordination, collapse and frequently death (Tufani et al., 2013). With acute acidosis, ruminal acidity and osmolality increase markedly as acids and glucose accumulate; these can damage the ruminal and intestinal wall, decrease blood pH, and cause dehydration that proves fatal. Laminitis, polioencephalomalacia and liver abscesses often accompany acidosis (Owens et al., 1998). Lactic acidosis can also cause chemical ruminitis, metabolic acidosis, lameness, hepatic abscessation, pneumonia and death if untreated (Lean and Wade, 2000). The goats being reared under free grazing system are more vulnerable for accidental excessive ingestion of carbohydrate-rich diets. Majority of goat population of peri-urban area, Rajendranagar and Mylardevpally are free grazing and therefore possibility of accidental ingestion of carbohydrate products is more.

Materials and Methods

Goats brought to the Teaching Veterinary Clinical Complex, Campus Veterinary Hospital, and Veterinary Ambulatory Clinic, Mylardevpally, College of Veterinary Science, Rajendranagar, Hyderabad with the history of dietary abnormalities, excessive ingestion of carbohydrate-rich diet viz., wheat and wheat flour, roti, chapatti, boiled rice, vegetables like potatoes, banana and ceremonial wastes were particularly included in the present study. Detailed clinical examination was carried out for each clinical case. The cases showing the symptoms of diarrhoea, abdominal distension, anorexia, distension of rumen, and diarrhoea were selected and screened for ruminal acidosis basing on ruminal fluid pH. Ruminal fluid was collected by following aseptic precaution from left paralumbar fossa (Ruminocentesis) as per procedure described by Rosenberger (1979). Thirty goats were found suffering with ruminal acidosis and basing on ruminal fluid pH they were divided into three different groups viz. mild, moderate and severe with rumen fluid pH as 5.5 – 6.5, 4.5 – 5.5 and 4.0 – 4.5 respectively consisting of 10 goats in each. Color, consistency and odour of rumen fluid were recorded as per Rosenberger (1979). The color of the rumen fluid was recorded immediately after collection by visual inspection of the sample. Although the normal color of rumen fluid varies from pure green to yellowish brown (depending on feed), the change in color towards slight milky to milky green or milky greyish was specifically assessed in ruminal acidotic goats.

The consistency of rumen liquor of healthy goats is slightly viscous. The alterations in consistency of rumen liquor varying from normal to less viscous or watery foamy or to slimy pulp were recorded in affected goats. Normal odour of rumen fluid is aromatic because of the gases produced by fermentation. The characteristic change in odour from sour (acidic) to pungent-acidic was recorded in goats suffered from ruminal acidosis. The pH of ruminal fluid was measured immediately after collection without much exposure to atmospheric air using indicator paper (Himedia Laboratory, Mumbai). Ruminal fluid pH ≤ 5.5 was the criteria to differentiate acidemic goats. The severity of ruminal acidosis was assessed by recording rumen fluid pH in affected goats and grouped as mild, moderate and severe with rumen fluid pH as 5.5 – 6.5, 4.5 – 5.5 and 4.0 – 4.5 respectively (Chakrabarti, 2006). The MBRT of rumen fluid was measured by following procedure described by Dirksen (1965) and Rosenberger (1979). Theiodophilic activity of protozoa in rumen fluid was estimated and graded as per method given by Misra et al. (1972). Microbial examination of rumen fluid was performedSedimentation activity time (SAT) or Stratification of rumen fluid was noted as per the procedure described by Nicholas and Penn (1958). The protozoal motility of rumen fluid was estimated based on procedure described by Misra and Singh (1974). The Protozoal density / protozoal concentration in rumen fluid were estimated as per the method described by Misra et al. (1972).

A drop of fresh rumen fluid was placed on a clean glass slide and was covered with a cover glass. The live and dead protozoal proportion (ratio of live: dead) examined under low power field of microscope based on viability of protozoa. An air-dried smear of ruminal fluid was stained by Gram’s Method and observed under oil immersion of the microscope. Proliferation of Gram +ve cocci and rods at the expense of Gram –ve was specifically recorded as per procedure described by Coles (1974). The statistical analysis of the data was subjected to one way ANOVA using statistical package for social sciences (SPSS) version 15. Differences between means were tested using Duncan’s multiple comparison test and significance was set at 5 percent (P<0.05). The values were represented as Mean ± Standard Error.

Results and Discussion

Goats of mixed breed, age and sex were presented to Teaching Veterinary Clinical Complex, Campus Veterinary Hospital, Ragendranagar and Veterinary Ambulatory Clinic, Mylardevpally, College of Veterinary Science, Rajendranagar, Hyderabad for period of 9 months from September 2012 to July 2013 with history of dietary abnormalities, excessive ingestion of carbohydrate-rich diet viz., wheat and wheat flour, roti, chapatti, boiled rice, vegetables like potatoes, banana and ceremonial wastes were particularly included in the present study. Detailed clinical examination was carried out for each clinical case. The cases showing the symptoms of diarrhoea, abdominal distension, anorexia, distension of rumen, and diarrhoea were selected and screened for ruminal acidosis basing on ruminal fluid pH. The present study was carried out to ascertain the ruminal fluid alterations in ruminal acidotic goats and compared with healthy goats. The results obtained are presented as follows.

In the present study, 30 clinical cases of ruminal acidosis were selected, based on rumen fluid pH, the goats were divided in three different groups as group I (mild ruminal acidosis), group II (moderate ruminal acidosis) and group III (severe ruminal acidosis) consisting of ten goats in each. Ten apparently healthy goats served as healthy control for the comparative study. In the present study on clinical cases of ruminal acidosis occurred due to the accidental ingestion of large amount of wheat grain, stale chapattis and roti, stale rice, mangoes, banana, vegetables waste, ceremonial waste, vegetables market waste, hotel waste, fed with excessive amount of jowar grains, corn, wheat flour and let-out for grazing in their surroundings for few hours. The cases were presented to clinic 12-24 hrs after accidental ingestion of excessive quantity of carbohydrates rich diet.

Rumen Fluid Examination

In the present study, ruminal fluid parameters physical like viz. color, consistency, odour and pH of rumen fluid, micro-biochemical like viz. protozoal motility, density, live and dead count, MBRT, iodophilic activity of protozoa, SAT, and gram’s staining of rumen fluid were investigated in cases of ruminal acidotic goats and compared with healthy goats.

Physical Examination of Rumen Fluid

Color of Rumen Fluid

The color of rumen fluid was noticed in affected and healthy control goats by visualization. The color of rumen fluid in group I was grey and milky grey in both group II and group III as compared with group IV was greenish (seven goats) to yellowish-brown (three goats).The physical parameters like, the color of rumen fluid in group I was grey and it was milky grey in both group II and III as compared to healthy goats, it was yellowish-brown to greenish. These findings are in accordance with Padmaja and Praveena (2011), Gupta et al. (2012), Karale (2012) and Rahima et al. (2012).

Consistency of Rumen Fluid

The consistencyof rumen contents in acidotic goats was slightly watery (six goats) and semi-solid (four goats) in group I, while watery in both group II and III as compared against group IV was viscous (six goats) and thick (four goats). The consistency could be attributable to increased lactic acid concentration and osmolality of rumen fluid resulting in withdrawal of extra-cellular fluid in to rumen recorded (Slyter, 1976). These findings corroborate with that of Choudhary et al. (2011), Gupta et al. (2012), Karale (2012), and Rahima et al. (2012).

Odour of Rumen Fluid

The odour of the rumen fluid in group I and II was acidic and in group III was pungent-acidic and in group IV, it was aromatic. The change of odour of rumen fluid might be due to excess putrefaction / fermentation of carbohydrates rich diet by proliferated gram-positive organisms, which was correlated with the earlier observations recorded Padmaja and Praveena (2011), Gupta et al. (2012), Karale (2012), and Rahima et al. (2012).

Ruminal Fluid pH

The pH of ruminal fluid in all the ruminal lactic acidotic cases was significantly (p<0.05) lower than the normal range of healthy goats (7.05 ± 0.12). The mean values of rumen fluid pH were obtained 5.60 ± 0.07, 5.10 ± 0.07 and 4.0 ± 0.17 for group I, II, and III respectively. It was observed that the ruminal fluid pH in all the ruminal lactic acidotic cases was significantly (p<0.05) lower than the normal range of healthy goats. Ingestion of large amount of carbohydrates rich diet leads to change in the microbial population in the rumen. The changes in the microbial species inhabiting the rumen normal fermentation were accompanied by a change in the type of fermentation, becoming a lactic fermentation. This had an impact on the pH values of rumen liquid, which decreased in proportion to increased amount of lactate reported by Nikolov (1998). The similar finding were documented by Padmaja and Praveena (2011), Gupta et al.(2012), Karale (2012),Rahima et al. (2012) Tufani et al. (2013) and Ullah et al. (2013).

Protozoal Activity

Protozoal Motility

The protozoal motility of rumen fluid was noted in group I + (slow sluggish, rotatory movements), and was nil (no motility) in group II and III as against group IV was +++ (swirls, rapid or vigorous movements). The micro-biochemical parameters like, protozoal motility of rumen fluid in group I was +/0 (sluggish/no motility) and was nil in group II and III, because of complete absence of protozoa as compared to group IV which had +++ (rapid vigorous motility). These findings are in agreement with the observations of Gupta et al. (2012), Karale (2012), Rahima et al. (2012) and Ullah et al. (2013).

Protozoal Density

The protozoal density of rumen fluid in group I was 1-10 protozoa per low power field graded as + (few), and in both group II and III were nil (0) protozoa per low power field was graded as – (none). Whereas, in group IV the protozoal density were more than 30 protozoa per low power field graded as +++ (abundant). The rumen normal protozoa might be destroyed due to changed rumen eco-system i.e. increase lactate and lactate producing gram positive bacteria, which increased abnormal fermentation in rumen. Similar observations were recorded by Darwin et al.(2007), Chaudhary et al. (2009), Selvaraj et al. (2009), and Karale (2012).

Live and Dead Protozoal Proportion in Rumen Fluid

In group I, the mean live and dead protozoal counts were 10.50 ± 0.86 and 89.50 ± 0.86 respectively, and in group II and III, the mean live and dead protozoal counts were absent as compared against group IV which were 89.20 ± 0.59 and 10.80 ± 0.59 respectively. There was significant difference (p<0.05) in live and dead protozoal count in group I as compared to healthy goats and group II and III, complete absence of protozoa was observed. The findings were correlated with that of Selvaraj et al. (2009), but the complete absence of protozoa was reported by earlier workers Darwin et al. (2007), Radostits et al. (2007), Choudhary et al. (2011), Rahima et al. (2012) and Ullah et al. (2013).

Iodophilic Activity

The iodophilic activity was + in group I and in group II and III, nil (0) because protozoa did not find in those two groups as compared to group IV was +++/++++. These findings are agreement with Misra et al. (1972), Kasaralikar et al. (2007) and Karale (2012).

Methylene Blue Reduction Time (in minutes)

The mean MBRT of ruminal fluid in ruminal acidotic goats was 11.65 ± 0.37, 24.90 ± 0.82 and 33.10 ± 0.87 min for group I, II and III, respectively. While in group IV, it was 2.75 ± 0.17.The mean MBRT of rumen liquor was increased significantly (P < 0.05) in ruminal acidotic goats as compared against healthy goats. The more time required for reduction of methylene blue dye in rumen fluid might be due to inactivity or death of normal micro- flora and fauna (Vasu and Nagarajan, 1985). These findings corroborate with that of earlier researchers Vasu and Nagarajan (1985), Basak et al. (1993), Darwin et al. (2007), Kasaralikar et al. (2007), and Karale (2012).

Sedimentation Activity Time (in minutes)

The mean SAT values of ruminal fluid in group I was 66.20 ± 1.44, nil (0) in both group II and III. No sedimentation activity was recorded in group II and III. The mean SAT was increased significantly (P < 0.05) in ruminal acidotic goats as compared to the healthy goats (5.20 ± 0.35). No sedimentation activity was recorded in group II and III might be due to absence of normal microflora in rumen lquior. Prolongation of SAT and quick fermentation of particulate matter might be due to gross microbial inactivity as per the findings of Nicholas and Penn (1958), Rosenberger (1979). These findings agreed with earlier reports of Misra et al. (1972), Basak et al.(1993), Kasaralikar et al. (2007) and Karale (2012).

Table 1: Mean values of rumen fluid parameters in acidotic and healthy goats

S. No. Parameters Ruminal Acidotic Goats Group IV (Healthy Goats)
Group I (mild) Group II (moderate) Group III (severe)
Physical Parameters of Rumen Fluid
1 Color Grey Milky grey Milky grey Greenish (7) and yellowish-brown (3)
2 Consistency Slightly watery (6) and semisolid (4) Watery Watery Thick (4) and Viscous (6)
3 Odour Acidic Acidic Pungent acidic Aromatic
4 pH 5.6* ± 0.07 5.1* ± 0.07 4.0* ± 0.17 7.05 ± 0.12
Micro-biochemical Parameters of Rumen Fluid
1 MBRT (min) 11.65* ± 0.37 24.90* ± 0.82 33.10**±0.87 2.75 ± 0.17
2 SAT (min) 66.20* ± 1.44 Nil Nil 5.20 ± 0.35
3 Protozoal motility +/Nil Nil Nil +++
4 Protozoal density +/Nil Nil Nil +++
5 Live protozoal proportion 10.50* ± 0.86 Nil Nil 89.20 ± 0.59
Dead protozoal proportion 89.50* ± 0.86 Nil Nil 10.80 ± 0.59
6 Iodophilic activity + Nil Nil +++/++++
7 Gram’s staining Gram positive Gram positive Gram positive Gram negative

*Significant at (p<0.05)

Bacterial Staining Method (Gram’s Staining)

In all the group of ruminal acidotic goats, there was predominance of Gram positive cocci and rod-shaped bacteria as compared to healthy goats which had Gram negative cocco-bacilli. The findings are in accordance with Chaudhary et al. (2009), Padmaja and Praveena (2011), Gupta et al. (2012) and Karale (2012).

Conclusion

Thirty goats were presented with the history of dietary abnormalities, excessive ingestion of carbohydrate-rich diet. The cases were showing the symptoms of diarrhoea, abdominal distension, anorexia, distension of rumen, and diarrhoea were selected and screened for ruminal acidosis basing on ruminal fluid pH which was less than normal as compared with ten healthy goats. It was concluded that the goats were confirmed as suffering from ruminal acidosis and rumen fluid analysis was undertaken.

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