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Volume 24 No. 3

Buffalo Bulletin (September 2005) Vol.24 No. 3

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LOCALLY INFILTRATIVE, NON-METASTASIZING TRANSITIONAL CELL CARCINOMALEADING TO DYSTOCIA, ANURIA AND CONSTIPATION IN AN INDIAN WATER

BUFFALO (BUBALUS BUBALIS)

Naresh Kumar Sood, Kuldip Gupta, Neeraj Sood, Mrigank Honeparkhe, Amarjit Singh

Department of Animal Reproduction, Gynecology and Obstetrics, Punjab Agricultural University,Ludhiana – 141 004, India.

ABSTRACT

A nine-year-old buffalo at full term ofpregnancy was presented to the PAU clinics withthe complaints of dystocia, anuria and constipation.Surgery was performed to relieve dystocia, but asthe condition of the animal deteriorated aftersurgery, the animal was euthanized. Necropsyrevealed thickening of the wall of the urinarybladder which was hard to cut and rough with fewirregular papilliform projections in the lumen.Microscopic examination indicated thickening ofwall of the urinary bladder due to urothelial hyper-plasia and hypertrophy. Neoplastic transitionalepithelial cells were seen infiltrating the laminapropria and occasionally to the tunica muscularisof the bladder. Marked lymphoid cell infiltrationwas observed in between the infiltrating tumourcells and also in the form of well-defined multifo-cal perivascular aggregates. No significant histo-pathological findings were detected in regional anddistant lymph nodes and other visceral organs, thus,ruling out the possibility of metastasis.

INTRODUCTIONTransitional cell carcinomas are rare in

bovines (Gupta, 1983). Only a few cases of transi-tional cell carcinoma have been reported previouslyin buffaloes (Gupta and Singh, 1975; Nanda andSharma, 1985) with limited or extensive metasta-sis. The present study deals with an unusual case oftransitional cell carcinoma associated with dysto-cia, anuria and constipation in an Indian waterbuffalo (Bubalus bubalis).

HISTORY AND CLINICALEXAMINATION

A nine-year-old she buffalo (Bubalus bubalis)in sixth parity and full term pregnancy was presentedto the University Clinics with the complaints ofdystocia (all previous parturitions normal), alongwith anuria and constipation. The animal looked dulland dehydrated. On per-rectal examination, a largehard irregular growth was palpated encroaching thepelvic brim. A caesarian section performed torelieve dystocia (unresponsive to conventionaltherapy), resulted in delivery of a female fetus.During surgery, a hard irregular tumour like growthinvolving urinary bladder, urethra and extending upto pelvic brim was noticed. Since the condition ofthe animal deteriorated progressively after surgeryand it developed an abrupt shooting pain, it washumanely euthanised with owner’s consent.

MATERIALS AND METHODSNecropsy of the animal was conducted and

detailed gross lesions were recorded. Tissue slices,one cm thick, from all the grossly affected andunaffected organs including lymph nodes werecollected in 10% neutral buffered formalin andprocessed for routine histopathology.

RESULTS AND DISCUSSIONAt necropsy, the wall of the urinary bladder

was found to be 7-8 cm thick, rough and hard to cut,


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and had a few irregular papilliform projections intothe narrow bladder lumen. The growth also appearedto involve urethra and cervix and extended into thepelvic brim. Both the kidneys were moderatelyenlarged, normal in colour and slightly soft intexture. Fecoliths were recovered from rectum ofthe animal. Incidentally, a sharp iron needlepiercing the reticular wall and diaphragm was alsolocated at necropsy. All the other visceral organsappeared grossly unaffected.

On histopathologic examination, the wall ofthe urinary bladder was found to be markedlythickened with areas of urothelial hyperplasia,hypertrophy and disruption. The sheets of neoplas-tic transitional epithelial cells infiltrated the laminapropria and occasionally to the tunica muscularisof the bladder (Figures 1 and 2). In places, thesurface epithelium was almost completely denuded.Marked lymphoid cell infiltrates, singly or in smallgroups, were also observed interspersed randomlybetween the infiltrating tumour cells and also in theform of well-defined multifocal perivascular aggre-gates (Figures 2 and 3). The thickening of the laminapropria of the bladder wall was due to markedfibrovascular response. In addition, the microvas-culature of the lamina propria was congested anddilated in places. The tunica muscularis revealedmultifocal areas of smooth muscle necrosissurrounded by extensive fibroplasia. An interestingfinding was the presence of skeletal muscle bundlesbetween the smooth muscles of the serosal muscu-laris layer (Figure 4). The wall of the urethra andcervix was also markedly thickened due to fibrosis,besides tumour cell infiltration in the pelvicurethra. Kidneys revealed acute diffuse tubularnecrosis and mild multifocal non-suppurative inter-stitial nephritis. In addition, focal myocardialdegeneration and hyalinization along with loss ofpurkinje fibres were also observed (Figure 5). Nosignificant histopathological findings were detectedin regional and distant lymph nodes and othervisceral organs, thus, ruling out the possibility ofmetastasis.

The present case and the earlier reports (Guptaand Singh, 1975; Nanda and Sharma, 1985) had a

common history that they affected old femalebuffaloes which is in contrary to the findings of nosuch sex predisposition in dogs (the most frequentlyaffected species). In the present case, although thetumour was infiltrating deep down to the tunicamuscularis of the bladder, yet no lymph node or vitalorgan metastasis was detected, which may be dueto the immune response generated by local lymphoidcell infiltration as tumour infiltrating lymphocyteshave been incriminated in checking the tumourspread and thereby, better prognosis in humancancers including transitional cell carcinomas(Underwood, 1974; Lipponan et al., 1992 and Eerolaet al., 2000). The presence of multifocal perivascu-lar lymphoid aggregates without presence of tumourcell hinted at the possibility of type IV hypersensi-tivity reaction in this case as such a response hasbeen recorded previously in cases of enzooticbovine hematuria including transitional cell carci-nomas in cattle (Krishna et al., 1991). Theextensive fibrosis in bladder wall may also beattributed to type IV hypersensitivity (Roitt et al.,1996). The genital and urinary tumours irrespectiveof their size have rarely been associated with dysto-cia in cattle although two such cases had beenreported in old buffaloes (Nanda and Sharma. 1985).The encroachment of the tumour onto the pelvicbrim along with thickening of cervix might have ledto obliteration of the birth canal and thereby dysto-cia, in the present case, as it might have failed todilate at the time of birth. The anuria in such cases(Nanda and Sharma, 1985) might be due toobliteration of neck of urethra by tumour compres-sion as well as atonicity of the bladder wall due tosmooth muscle necrosis and extensive bladder wallfibrosis, the findings in this case. The presence ofskeletal muscles between the smooth muscles of thebladder might be attempt at reinforcing the bladderwall strength. The constipation as observed in thepresent case might either be due to compression bytumour or an effect of anuric uremia. The degenera-tive changes and necrosis in the myocardium andpurkinje fibers as well as nephrotic changes in thekidneys might also be the offshoots of uremictoxemia. The sudden shooting pain, which necessi-tated immediate euthanasia, might have been due topenetration of sharp iron needle into the diaphragm


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at that time because of increased intra-abdominalpressure by foetal dystocia and ruminal atony byuremia. The present case appeared to be unique in

LEGENDS TO THE FIGURES

Figure 1. Hyperplasia, hypertrophy and disruption of overlying transitional epithelium of urinary bladder and infiltration of neoplastic transitional epithelial cells into the lamina propria and muscularis of the bladder. Marked lymphoid cell infiltration is also seen. (H&Ex75)

Figure 2. Disruption of transitional epithelium. The tumour cell nests are also seen in the lamina propriasurounded by lymphoid cells either singly or in small groups. (H&Ex150)

the sense that multiple clinical effects were associ-ated with transitional cell carcinoma and yet noevidence of metastasis.


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Figure 3. Infiltration of tumour cell nests into the tunica muscularis, fibroplasia and presence of scattered lymphoid cells as well as perivascular lymphoid cell aggregates. (H&E x 75)

Figure 4. Presence of several skeletal muscle bundles interspersed between the more abundant smoothmuscles in the tunica muscularis of the urinary bladder, serosal surface. (H&E x 100)


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Figure 5. Conspicuous myocardial degeneration and hyalinization of myofibrils along with necrosis ofpurkinje fibers charactrized by loss of their nuclei. (H&E x 150)

REFERENCES

Eerola, A.K., Y. Soini, and P. Paakko (2000). A highnumber of tumor-infiltrating lymphocytes areassociated with a small tumor size, low tumorstage, and a favourable prognosis in operatedsmall cell lung carcinoma. Clin. Cancer Res.,6(5): 1875-1881.

Gupta, P.P. (1983). Infiltrative transitional cellcarcinoma of urinary bladder in a buffalo.Indian Vet. J., 60(11): 934.

Gupta, P.P. and B. Singh (1985). Infiltrative transi-tional cell carcinoma of urinary bladder in abuffalo (Bubalus bubalis). Vet. Pathol., 12(5-6): 468-469.

Krishna, L., J. Vaid, and R.K. Dawra (1991).Enzootic bovine hematuriain cattle: II. Patho-morphological immunofluorescent and immuno-logical studies. Indian J. Vet. Pathol., 15(1):30-34.

Lipponen, P.K., M.J. Eskelinen., K. Jauhiainen.,E. Harju, and R. Terho(1992). Tumorinfiltrating lymphocytes as an independentprognositic factor in transitional cellbladder cancer. Europ. J. Cancer, 29: 69-75.

Nanda, A.S. and R.D. Sharma (1985). Dystociadue to urinary bladder carcinoma in twowater buffaloes (Bubalus bubalis)-clinicalcase report. Theriogenol., 24 (3): 327-329.

Roitt, I., J. Brostoff and D. Male (1996). Immu-nology (Mosby, London)

Underwood, J.C.E. (1974). Lymphoreticularinfiltration in human tumors: prognostic andbiological implications. Brit. J. Cancer, 30:537-548.


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STUDIES ON ETIOLOGY, SYMPTOMATOLOGY, DIAGNOSIS AND THERAPY INULCERATIVE THELITIS OF BUFFALOES IN ANDHRA PRADESH (INDIA)

M. Lokanadhamu, B. Sreedevi and T. Venkata Reddy

Department of Veterinary Epidemiology and Preventive Medicine, ANGRAU, College ofVeterinary Science, Tirupati – 517 502, Andhra Pradesh, India

ABSTRACT

Ulcerative thelitis is a disease affectingprimarily high-yielding primiparous graded Murrahmilch buffaloes and causing serious economic lossesto the farmers in coastal districts of Andhra Pradesh.The disease is characterized by acute inflammationof one or more teats with subsequent thickening,narrowing or closure of teat canal leading to incom-plete drainage of milk. The quality of milk appearsto be normal unlike in clinical mastitis. This isfollowed by ulceration, focal necrosis, andsloughing off the affected teat. Healing may bedelayed due to the trauma of milking and secondarybacterial infections. The treatment of the conditionwas not successful because the etiology of thedisease was unknown. Hence the present investiga-tion was undertaken to study the epidemiology,etiology and clinico-therapeutic aspects of thedisease. The milk samples collected from ulcerativethelitis cases were normal in colour and consistencyand were negative in all mastitis diagnostic tests.The milk samples subjected to cultural examinationdid not reveal any pathogenic bacterial (or) fungi.The milk samples filtered and inoculated intrader-mally in to rabbits produced mild erythematouslesions after 48 h. The haematological changes werestudied in ulcerative thelitis affected buffaloes andwere compared with healthy postpartum buffaloes.There was a slight increase in the neutrophil countand slight decrease in the lymphocyte countcompared to healthy animals. The difference wasstatistically significant (t-test) indicating that thecondition might be localized without systemicinvolvement. The serum protein profile wasstudied and there was a significant (t-test) increases

in total proteins and globulin when compared tohealthy buffaloes, but only a slight increase in thealbumin and albumin : globulin ratio (A : G) ratios,which were statistically not significant. Theseresults indicate that the condition might be of aninfectious origin. The probable viral etiology of thedisease was investigated using the vesicular fluidcollected from ulcerative thelitis affected gradedMurrah buffaloes. The material subjected to elec-tron microscopy revealed non-envelopedhexagonal viral particles (uranyl acetate stain) andenveloped hexagonal viral particles (phosphotung-stic acid stain). Based on the above findings, thera-peutic trial was conducted in 24 animals, which weregrouped into three groups of eight animals each. TheFirst group were treated with Acyclovir orally andtopically along with conventional therapy. Ahighest cure rate of 62.5% was obtained. In GroupII, in which animals were treated with Acyclovirtopically along with conventional therapy, a curerate of 37.5% was obtained. Group III animals weretreated with conventional therapy; a cure rate of12.5% only was recorded.

INTRODUCTION The farmers of Andhra Pradesh are showing

more inclination to rear buffaloes rather than cowsas the buffalo milk is more rewarding price wisebecause of high butter fat and solid non-fat content,selective preferential consumption of buffalo milkfor its wholesomeness, and other beliefs. So thegraded Murrah buffalo population is increasingenormously in the coastal districts of Andhra


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Pradesh. A peculiar condition affecting teats whichhas no resemblance to mastitis, namely, ulcerativethelitis has emerged as an obstacle to the milkproducing capacity of buffaloes in Andhra Pradesh.This condition was reported in primiparous gradedMurrah buffaloes and is characterized by acuteinflammation of one or more teats, ulceration,necrosis and sloughing off the affected teat. Treat-ment with antibiotics and other anti-mastitisremedials was found to be of no curative value.Hence, the present investigation was undertaken tostudy the probable etiology and the clinico-thera-peutic aspects of this ulcerative teat disease, namelyulcerative thelitis in graded Murrah buffaloes.

MATERIALS AND METHODS

Collection of SamplesSamples were collected from the typical

cases of ulcerative thelitis of graded Murrah buffa-loes in different coastal districts of Andhra Pradesh.Milk samples, blood smears, whole blood(uncoagulated), serum, vesicular fluid, affected teattissues were collected and preserved until used.

Testing of Milk SamplesDifferent mastitis diagnostic tests were

conducted as per the standard protocols.The somaticcell count of milk samples was carried outaccording to the method of Schalm et al., (1971)and electrical conductivity was measured using amanual meter (Milk Checker, Eisai, Tokyo, Japan).Later, the milk samples were subjected to bacterio-logical examination to check the possibility ofbacterial etiology of the disease. Bacteriologicalinvestigation was undertaken by transfer of a loopfulof the milk sample to tryptose broth and incubationat 37 °C overnight. Milk agar slants were preparedas per the procedure given by Christie and Koegh(1940). The broth culture was streaked on milk agarslants for primary isolation of pathogens. A tenta-tive identification of bacterial growth was done,based on colony morphology and gram’s staining.The pure cultures were identified up to genus level

using appropriate selective media (Mannitol saltagar, Mac Conkey lactose agar and blood agar) andbiochemical tests were conducted as per the Bergey’sManual of Systematic Bacteriology (Sneath Peteret al., 1986).

Intra-dermal Testing of Whey in RabbitsTwo ml of milk from each sample was taken

into sterile screw-capped vials. Microbial rennet wasadded at a final concentration of 0.25 g per kg ofmilk, the vials were centrifuged, and the whey wasseparated. The flank region of healthy, adult NewZealand white rabbit was neatly depilated a daybefore inoculation. The area was cleaned and disin-fected. Whey sample was injected intradermallyalong with negative controls, leaving a distance oftwo cm between two sites, by using a 27-gaugehypodermic needle with a tuberculin syringe.Sequential changes on the skin after inoculation wererecorded every day for two weeks.

HaematologyAll the haematological estimations were

done on the day of blood collection. Totalleucocyte count was calculated as per the standardprotocols. Blood smears were prepared on cleangrease-free slides for estimation of differentialleucocyte count (Feldman et al., 2000). Total serumproteins, albumin, globulin and A/G ratio were esti-mated by adopting modified Biuret and Dumasmethod using a diagnostic reagent kit supplied bySpan Diagnostics Ltd., Ludhiana (Surat), India. Thevalues were expressed in g/100 ml of serum.

HistopathologySkin biopsies from ulcerative thelitis

affected buffaloes were taken from two cases at thestage of scab formation and were examined. Scabtissues were fixed immediately in 10% neutralbuffered formaline and processed by routineconventional histo-pathological procedures. Fivemillimicron thickness sections were made fromparaffin embedded blocks and the sections werestained with haematoxylin and eosin (Rweyemamu,et al., 1969).


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Electron microscopic StudiesThe electron microscopy of suspected

samples was conducted as per the method describedby Gibbs et al. (1970b). A drop of undiluted vesicu-lar fluid was applied to a lightly carboned celloidin- coated copper grid. When the suspension hadpartially dried, the grid was washed by touching itthree times to the surface of a drop of distilled water.A small drop of 2 % potassium phosphotungstatesolution (PTA) at pH 7 was than applied to the gridand after ten seconds excess stain was removed asdescribed earlier. The grid was then allowed to dry.The grids thus prepared were examined immediatelyunder an electron microscope (Hitachi model H-7500) at Ruska Labs Rajendra Nagar, Hyderabadand electronmicrographs were taken. The grids weredouble-stained with saturated uranyl acetate (UA)in methanol and then with 4% lead citrate air driedand examined under electron microscope.

The scab tissues fixed in 3% glutaraldehydewere post fixed in 1% aqueous osmium tetraoxidefor 2 h and rinsed with 3 changes of distilled waterfor 10 minutes. Dehydration of the sample was doneusing ascending grades of acetone followed byinfiltration and embedding in resin media (spurr).Ultra thin sections of 50-70 nm were made withLeica ultra cut UCT - GA-D / E - 1/ 00 and wereplaced in copper mesh grids which were stained withsaturated uranyl acetate for 0.5 hour followed bycounter staining with 4% lead citrate. The grids wereair dried and observed under TEM, Hitachi H 7500,transmission electron microscope.

Therapeutic TrailIn the present study, a total of 24 clinical

ulcerative thelitis cases of graded Murrah buffaloeswere selected based on the clinical history and theseanimals were allocated at random into three groupsof eight animals each. Individual groups was treatedseparately with different combination of drugs forcomparing the therapeutic efficacy of different drugstill clinical recovery (Table 1). The clinical obser-vations were recorded every day up to 15 days.

Statistical AnalysisThe data was subjected to statistical analy-

sis (Snedecor and Cochran 1994).

RESULTS AND DISCUSSION

SymptomatologyThe important clinical signs noticed in the

cases of ulcerative thelitis include acute inflamma-tion of one or more teats with subsequentthickening, narrowing (or) closure of the teat canalleading to incomplete drainage of milk. There wasincrease in the size of the affected teats withglistening appearance followed by peeling off(Figure 1) and ulcer formation (Figure 2) whilemilking.

In rare cases, formation of vesicles(Figure 3) was seen and upon rupture serous fluidexude from the dermis resulting in ulcer formation.This may be followed by the commencement of thenecrotic changes in the teat, which eventually maylead to either partial or complete sloughing of theaffected teat (Figure 4). But, in some cases scabformation may be seen over the ulcers followed byhealing of the lesions (Turner et al., 1976).

Similar clinical signs were reported bymany workers in ulcerative mammillitis of cows(Gibbs et al., 1970a; Turner et al., 1976, Chauhanet al., 1989 and Janett et al., 2000) and in buffaloes(Sharma et al., 1998; Malleswara Rao et al., 2003).As primiparous animals were most commonlyaffected with this condition, there is lot of impacton milk production due to loss of teats, resulting ingreat economic loss to the farmers.

Examination of milk samples in ulcerativethelitis

Milk samples collected from animals affectedwith ulcerative thelitis revealed no changes in colourand consistency, which are common in clinicalmastitis. These samples, were subjected todifferent mastitis diagnostic tests viz., CMT, BTBtest, BCP test, WST test and chloride test, and found


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to be negative. The pH of the milk samples wasrecorded and was within the normal range of 6.5 to6.7. The values of electrical conductivity and thesomatic cell count were found to be normal.

Cultural examination of milk samplesThe bacterial etiology of the disease was ruled

out based on the results of cultural tests of thesuspected milk samples. In 38% of the samples, nogrowth was observed. Staphylococcus were isolatedin 50% and gram negative bacteria were isolated in12% of the samples. Later, coagulase andhaemolysin tests were conducted for the staphylo-coccus isolates, and IMViC tests were conductedfor gram negative isolates. No pathologicallysignificant bacteria could be identified in thesesamples and none of them revealed any fungalgrowth. This indicates that the bacteria isolated fromthe milk of the affected quarter could be the naturalinhabitants of udder and teat or environmentalcontaminants. These results are in accordance withthe findings of Ramaswamy et al. (2001) andMalleswara Rao et al. (2003).

Intradermal testing of whey in rabbitsThe bacteria free filtrate of whey obtained from

the milk of affected teats was used for studying thepathogenecity of the etiological agent by givingintradermal inoculation in to rabbit. A total of 26samples were tested out of which 18 samplesproduced mild erythematous lesions 48 h afterinoculation, and these persisted for another two days(Figure 5). Eight whey samples collected fromhealthy postpartum graded Murrah buffaloes did notproduce any lesions.

These results are in accordance with thefindings of Rweyemamu et al. (1968) and Chauhanet al. (1989). This experimental reproduction of thelesions in the rabbit gives strong evidence that theetiological agent of ulcerative thelitis could be anagent of infectious origin. According to the reportsof Deas and Johnston (1966); Pepper et al. (1966)and Castrucci et al. (1972) rabbits serve as excel-lent laboratory animals for conducting experimentsfor BHM isolates from cows whereas mice wererefractory.

Haematological changes in ulcerativethelitis cases

The total leucocyte count (Tables 2 and 3) ofulcerative thelitis affected buffaloes was found tobe higher (10,383.33 c/mm) than that of healthybuffaloes (9118.75 c/mm). However, the increasein TLC was found to be statistically non-significant(t-test). This indicates that the condition might belocalized affecting only teats without systemicinvolvement. Sharma et al. (1998) reported thesimilar results.

The differential leucocyte count of the bloodsamples (Tables 2 and 3) revealed the averageneutrophil count as 35.88 ± 1.19% and 41.50 ±1.62% in healthy postpartum and ulcerative thelitisaffected buffaloes, respectively. The lymphocytecount was 61.5 ± 1.5% and 56.06 ± 1.61% in healthyand affected buffaloes, respectively. The monocyte,eosinophil and basophil count of affected andhealthy buffaloes was with in the normal range(t-test). The slight increase in the neutrophil countmay be attributable to the secondary bacterialinfections of the teat lesions (t-test). Letchworth andCarmichael (1984) reported similar results statingthat no major changes in the hemogram were notedin BHV-2 inoculated cows.

The total serum protein content was 6.07 ±0.13 g/dl and 7.07 ± 0.22 g/dl in healthy postpartumand ulcerative thelitis affected buffaloes, respec-tively (Tables 4 and 5). There was a significantincrease in the total serum protein content inulcerative thelitis (t-test).

The total globulin content was 3.02 ± 0.08g/dl and 3.96 ± 0.26 g/dl in healthy and affectedbuffaloes, respectively (Tables 4 and 5). There wassignificant increase in the globulin content andnon-significant decrease in the A:G ratio (t-test).These results indicate that the condition might beof an infectious origin. These observations are inconcurrence with the results of Satija et al. (1979)and Canfield et al. (1984). Letchworth andCarmichael (1984) reported that there was anincrease in antibody production 7 days of postinfection in BHV-2 inoculated cows.


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Histo-pathologyHisto-pathology of the scab tissue revealed

congestion (Figure 6), hyperkeratinization, thinningand fracture of stratum corneum, intraepidermalvesiculation and presence of moderate cellularinfiltration mainly of lymphocytes (Figure 7) inepidermis. Extensive calcification (Figure 8) withintracellular edema in dermis was noticed in onecase. Several authors have recorded similar obser-vations in the histopathology of teat tissues fromnatural and experimental cases of BHM of cattle(Martin et al. 1969; Rweyemamu et al., 1969; Gibbset al., 1970a and Gibbs and Collings 1972). Similarmicroscopic changes were recorded in BHM ofbuffaloes by Sharma et al. (1998). These observa-tions suggest that the present condition might becaused by a viral infection.

Histo-pathology of rabbit skin after intra-dermal inoculation of suspected whey samplerevealed the formation of syncytia (Figure 9),intercellular oedema, areas of congestion and mildinfiltration of cells. Chauhan et al. (1989) observedsimilar histo-pathological changes in rabbit skin afterexperimental inoculation of material along withCowdry type A inclusion bodies.

Electron Microscopy of the suspectedsamples from ulcerative thelitis

The transmission electron microscopy (TEM)of the vesicular fluid was studied with two differentstaining techniques at Ruska Labs, Rajendra Nagar,Hyderabad, and this revealed naked and enveloped(Figure10) hexagonal viral particles.

Figure 1. Ulcerative thelitis of graded Murrahbuffalo showing epidermal peeling of the affectedteat skin

Figure 2. Ulcerative thelitis of graded Murrahbuffalo showing typical ulceration on the affectedteat

Figure 3. Ulcerative thelitis of gradedMurrah buffalo showing vesical informationon the skin of affeected teat

Figure 4. Ulcerative thelitis of graded Murrahbuffalo showing necrosis of the affected teat


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Gibbs et al. (1970a) and Weaver et al.(1972) also demonstrated bovine herpes mammilli-tis virus by electronmicroscopy of the vesicularfluids collected from bovine ulcerative mammillitiscases. Several authors reported that electron micros-copy can be used as a tool for the diagnosis ofbovine herpes mammillitis cases in cows and buffa-loes (Martin et al., 1969; Rweyemamu et al., 1969;Gibbs et al., 1970a; Gibbs et al., 1972; Sharma etal., 1998 and Malleswara Rao et al., 2003).

Though the virus was demonstrated in thevesicular fluid, the scab tissue from the affected teatserves as the best material for conducting electronmicroscopy. Hence, in the present study a portionof scab from affected teat was subjected to electronmicroscopy. The tissue sections showed thepresence of viral particles in the cytoplasm (Figure11) dispersed irregularly with two limiting

membranes. In the cytoplasm, viral particles oc-curred free and were in groups of two or more. Vi-ral particles were also found in vacuoles. Apoptoticchanges were seen in the nucleus (Figure 12). Simi-lar findings were observed in cows by Martin et al.(1969) and in buffaloes by Sharma et al. (1998).

Treatment of ulcerative thelitisThe treatment of ulcerative thelitis with

conventional therapy was not successful. As theresults of the present investigation suggestedprobable viral etiology of the condition, the effi-cacy of Acyclovir, a known anti-herpes drug, wastested along with an antibiotic and symptomatictherapy. A total number of 24 cases were dividedinto three groups of eight animals each and weretested with different combinations of drugs.

Figure 5. Intradermal inoculation of suspectedwhey samples into rabbit, showing developmentof mild erythematous reaction

Figure 6. Histo-pathology of the teat scab-Notesevere congestion (H&E x 280)

Figure 7. Histo-patology of the teat scab-Notehyperkeratinization thinning and fracture ofstratum corneum with cellular infiltration (H&Ex 280 )

Figure 8. Histo-pathology of the teat scab-Noteextensive calcification with oedema (H&E x 280)


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Table 1. Dosage regimens of therapeutics in ulcerative theilitis

S. No. Group Therapeutic Formulation Dose rate Route No. of Durationregiment (drug used mg/ kg buffaloes ofcombination) (ml) body weight treated treatment

(day)

1. I Injection 15 5 IM 8 5Enrofloxacin (10% w/v)(Floxidin)Inj. Diclofnac 20 1 IMsodium (Zobid)Inj. Avil 10 0.5 IMPheniraminemaleateOint. Acivir 5% w/v Topical 10Tab Acivir Tab 800 Oral 5

DT x 3Tablets TID

Lysine as a Topical 10Tablets

2. II Injection 15 5 IM 8 5Enrofloxacin (10% w/v)(Floxidin)Inj. Diclofenac 20 1 IMsodium (Zobid)maleateInj. Avil 10 0.5 IMPheniramineOint. Acivir 5% w/v Topical 10

3. III Injection 15 5 IM 8 5Enrofloxacin (10% w/v)(Floxidin)Inj. Diclofenace 20 1 IMsodium (Zobid)Inj. Avil 10 0.5 IMPheniraminemaleate


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Group I buffaloes were treated with Acyclovirorally and topically along with conventional drugs,and a 62.5% cure rate was obtained in 12 days aftertherapy. Lysine was applied as a topical paste onthe affected teats in bovine herpes mammillitis casesas it appears to be an effective agent for reductionof occurrence, severity and healing time for recur-rent herpes simplex infections in humans. Herpessimplex was found to be sensitive to the ratiobetween lysine and arginine. High levels ofarginine seem to initiate reactivation of herpessimplex infections in humans and lysine supplemen-tation shifts the ratio away from arginine andtoward lysine (Griffith et al., 1987).

In Group II animals, which were treatedwith topical application of Acyclovir cream alongwith conventional drugs, a cure rate of 37.5% wasobtained within 26 days after therapy.

In Group III, animals were treated with onlyconventional drugs and a 12.5% cure rate obtainedafter 35 days (Table 6).

The results of therapeutic trial wereanalysed to ascertain the effective therapeuticregimen for ulcerative thelitis. The regimen followedin Group I was more efficacious than Group II andGroup III, and that in Group II was more efficaciousthan that in Group III. Statistical analysis of theresults revealed significant differences betweenGroup I and Groups II and III and also between groupII and group III (t-test).

Watson (1989) reported the efficacy ofAcyclovir and other antiviral drugs in the treatmentof bovine herpes mammillitis (BHV-2) infections.

Based on the findings of the present study,it can be suggested that early treatment of teatlesions with oral and topical application of Acycloviralong with conventional therapy can restore the milkyield after 12 days in the clinical cases of ulcerativethelitis.

CONCLUSIONIn the present study, the examination of milk

and blood samples collected from the cases of ul-cerative thelitis revealed no evidence of bacterialor fungal etiology. Electron microscopic studies ofscab tissue and vesicular fluid clearly showed thepresence of hexagonal herpes virus like particles.With probable similarity to BHV-2 of mammillitisin cows. These findings were further strengthenedby the efficacy of the anti herpes therapy in curingthe condition. It is suggested that herpes antiviraldrugs like acyclovir may be tried as the choice oftreatment in very early stage of ulcerative thelitis ofbuffaloes along with concurrent antibacterial andsymptomatic therapy for minimising teat damage.Based on the observations in the present study, it isconcluded that the disease might be due to bovineherpes virus-2. However, further studies are to beundertaken using a large number of samples andisolation and molecular characterization of theetiological agent is to be carried out for evolvingappropriate control measures for the disease.

Figure 9. Histo-pathology of rabbit skin showingformation of syncytia in epidermis (H&E x 280)

Figure10. Electron microscopy of vesicular fluidfrom ulcerative thelitis showing hexagonal viralparticles with capsomeres (UA stain; x 100,000)


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Table 2 . Total leucocyte count and differential leucocyte count of healthy post parturient buffaloes

Sample No. TLC Differential Leucocyte countthousand /cubic mm N(%) L(%) M(%) E(%) B(%)

1 10800 35 64 1 0 02 6250 41 54 3 2 03 10500 34 63 2 1 04 10000 33 66 1 0 05 6900 32 64 4 0 06 11900 38 58 4 0 07 7850 40 58 2 0 08 8750 34 65 3 1 0Mean 9118.75 35.88 61.50 2.50 0.50 0SD 2005.16 3.36 4.28 1.20 0.76 0SE 708.93 1.19 1.51 0.42 0.27 0

Figure11. Electron microscopy of skin biobsy ofteat from ulcerative thelitis showing viral particlesin the cytoplasm (x 51600)

Figure 12. Electron microscopy of skin biobsy ofteat from ulcerative thelitis showing apoptoticchangesin the nucleus (x10320)


65

Table 3 . Total leucocyte count and differential leucocyte count of ulcerative thelitis cases ofbuffaloes

Sample No. TLC Differential Leucocyte countthousand / cubic mm N (%) L (%) M (%) E (%) B (%)

1 11,500 36 60 3 1 02 12,800 36 62 2 0 03 12,000 47 50 3 0 04 13,500 46 50 4 0 05 12,600 38 61 0 1 06 11,800 33 64 3 0 07 13,000 39 58 3 0 08 13,800 38 60 2 0 09 7,700 60 38 2 0 010 6,600 55 43 2 0 011 8700 40 58 2 0 012 9600 40 59 1 0 013 9900 44 55 1 0 014 9800 41 56 2 1 015 7,500 38 58 4 0 016 12,600 36 61 3 0 017 6,650 42 55 2 1 018 6,850 38 61 1 0 0MEAN 10,383.33 41.50 56.06 2.22 0.22 0.00SD 2544.20 6.88 6.84 1.06 0.43 0.00SE 599.67 1.62 1.61 0.25 0.10 0

t-test of healthy and ulcerative thelitis cases of buffaloes

Particular Total Neutrophils* Lymphocytes* Monocytes Eosinophils Basophils Leucocyte count

Healthy UT Healthy UT Healthy UT Healthy UT Healthy UT Healthy UT

Mean 9118.75 10383.33 35.88 41.50 61.50 56.06 2.50 2.22 0.50 0.22 0 0Variance 4020669 6472941 11.27 47.32 18.29 46.76 1.43 1.12 0.57 0.18 - -Observations 8 18 8 18 8 18 8 18 8 18 8 18t Stat -1.24 -2.18 2.07 0.59 1.20 -P (T< = t) on 0.11 0.02 0.02 0.28 0.12 -t (critical) or 1.71 1.71 1.71 1.71 1.71 -P (T< = t) tw 0.23 0.04 0.05 0.56 0.24 -t Critical tw 2.06 2.06 2.06 2.06 2.06 -

* SignificantUT– Ulcerative thelitis


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Table 4. Total serum protein (g/dl), serum albumin (g/dl), serum globulin (g/dl) and A to G ratio-values of healthy post parturient buffaloes

Sample No. Total proteins Albumin Globulin A/G ratio

1 5.76 2.98 2.78 0.0712 6.65 3.23 3.42 0.933 6.11 2.85 3.16 0.944 6.16 3 3.16 0.945 5.5 2.27 2.78 0.976 5.94 3.04 2.9 1.047 5.87 2.82 3.05 0.928 6.53 3.62 2.94 1.23MEAN 6.07 2.98 3.02 0.99SD 0.38 0.38 0.22 0.13SE 0.14 0.14 0.08 0.05

Table 5. Total serum protein (g/dl), serum albumin (g/dl), serum globulin (g/dl) and A to G ratio-values of buffaloes affected with ulcerative theilitis

Sample No. Total Protein Albumin Globulin A/G ratio

1 6.35 3.03 3.32 0.912 6.68 2.2 4.48 0.493 7.6 2.59 4.47 0.574 6.58 2.84 5.01 0.755 8.47 2.78 5.68 0.496 8.87 2.96 5.91 0.507 8.94 2.84 6.1 0.478 6.54 3.83 2.71 1.419 8.02 4.29 3.73 1.1510 7.56 3.88 3.68 1.0511 6.67 2.74 3.89 0.7012 5.85 3.05 2.8 1.0813 6.51 2.6 3.91 0.6714 6.68 4.11 2.57 1.5915 6.56 3.65 2.91 1.2516 6.37 2.71 3.66 0.7417 6.59 3.24 3.35 0.9718 6.35 3.23 3.12 1.04MEAN 7.07 3.14 3.96 0.87SD 0.94 0.58 1.10 0.35SE 0.22 0.14 0.26 0.08

t-test of healthy and Ulcerative thelitis cases of buffaloes


67

Particular Total Protein* Albumin Globulin* A/G ratioHealthy UT Healthy UT Healthy UT Healthy UT

Mean 6.07 7.07 2.98 3.14 3.02 3.96 0.99 0.87Variance 0.15 0.88 0.15 0.34 0.05 1.21 0.02 0.12Observations 8 18 8 18 8 18 8 18t Stat 2.89 0.74 2.36 -0.91P (T< = t) on 0.00 0.23 0.01 0.19t (critical) or 1.71 1.71 1.71 1.71P (T< = t) tw 0.01 0.47 0.03 0.37t Critical tw 2.06 2.06 2.06 2.06

* SignificantUT– Ulcerative thelitis

Table 6 . Comparative efficacy of therapeutic trial in ulcerative thelitis

Sl. No. Number of Number Days required Percentageanimals treated of cured for curing

Group I 8 5 12 62.5*Group II 8 3 26 37.5*Group III 8 1 35 12.5*

* t-test significant within the groups


68

REFERENCES

Canfield, P.J., F.J. Best., A.J. Fairburn, J. Purdieand M. Gilham (1984). Normal haematologicaland biochemical values for the swamp buffalo(Bubalis bubalis) in Australia. AustralianVeterinary Journal, 61: 89-93.

Castrucci, G., B. Pedini, V. Cilli and G. Arancia(1972). Characterization of a viral agentresembling bovine herpes mammillitis virus.Veterinary Record, 90: 325-335.

Chauhan, H.V.S., M.K. Gupta, G.J. Jha., R.S.Pandey, K.K. Singh, R.P. Sinha and A. Prasad(1989). Studies on the bovine herpes mammil-litis (BHM) in cattle. Indian VeterinaryJournal, 66: 106-109.

Christie and Koegh (1940). Practical medicalmicrobiology. Mackie, and Cartney, eds.Volume 2, 13th ed. Churchill Livingstone NewYork. p. 312-313.

Deas, D.W. and W.S. Johnston (1966). An outbreakof an ulcerative skin condition of the udderand teats of dairy cattle in the east of Scot-land. Veterinary Record, 78: 828-829.

Feldman, B.F., J.G. Zinkl and N.C. Jain (2000).Schalm’s Veterinary Hematology. 5th ed.Published by Lippincott Williams and Wilkins,New York.

Gibbs, E.P.J. and D.F. Collings (1972). Observationson bovine herpes mammillitis (BHM) virusinfections of heavily pregnant heifers andyoung calves. Veterinary Record, 90: 66-68.

Gibbs, E.P.J., R.H. Johnson and A.D. Osborne(1970a). The differential diagnosis of viral skininfections of bovine teat. Veterinary Record,87: 602-609.

Gibbs, E.P.J., R.H. Johnson and C.A. Voyle (1970b).Differential diagnosis of viral infections of thebovine teat skin by electron microscopy.Journal of Comparative Pathology, 80: 455-463.

Gibbs, E.P.J., R.H. Johnson and A.D. Osborne(1972). Field observations on the epidemi-ology of bovine herpes mammillitis. VeterinaryRecord, 91: 395-401.

Griffith, R.S., D.E. Walsh., K.H. Myrmel, R.W.Thompson and A. Behforooz (1987). Successof L-Lysine therapy in frequently recurrentherpes simplex infection. Treatment andProphylaxis. Dermatologica, 175 (4) : 183-190.

Janett, F., N. Stauber., E. Schraner., H. Stocker andR. Thun (2000). Bovine herpes mammillitisclinical symptoms and serological course.Schweiz Arch Tierheilkd, 142: 375-380.

Letchworth, G.J. and L.E.Carmichael (1984). Localtissue temperature a critical factor in the patho-genesis of bovine herpes virus-2. Infection andImmunity 43: 1072-1079.

Malleswara Rao, U.V.N., B. Sreedevi and T. VenkataReddy (2003). Studies on ulcerative mammil-litis of buffaloes in Andhra Pradesh (India).Buffalo Bulletin, 22 (4) : 80-90.

Martin, W.B., Z. Helem James., I.M. Lauder., MaxMurray and H.M. Pirie (1969). Pathogensis ofbovine mammillitis virus infection in cattle.American Journal of Veterinary Research, 30:2151-2166.

Pepper, T.A., L.P. Stafford., R.H. Johnson and A.D.Osborne (1966). Bovine ulcerative mammil-litis caused by a herpes virus. VeterinaryRecord, 78: 569-570.


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Ramaswamy, V., N. Rajendran., D. Chandrasekran.,J. Johnson Rajeshwar and K. Sukumar (2001).Studies on the etiology and effect of obstruc-tive thelitis in buffaloes. Final report of IndianCouncil of Agricultural Research (ICAR),Veterinary College and Research Institute,Namakkal.

Rweyemamu, M.M., R.H. Johnson and M.R. McCrea (1968). Bovine herpes mammillitis III.Observations on experimental infection.British Veterinary Journal, 124: 317-324.

Rweyemamu, M.M., A.D. Osborne and R.H.Johnson (1969). Observations on the histopa-thology of bovine herpes mammillitis.Research in Veterinary Science, 10: 203-207.

Satija, K.C., S. Rajpal, R. Pandey and V.K. Sharma(1979). Electrophoresis of buffalo (Bosbubalis) serum proteins including immunoglo-bulins. Infectious Immunology, 24: 567-570.

Schalm, O.W., E.J. Caroll and N.C. Jain (1971).Bovine mastitis. Lea and Febigee Philadelphiap.94-127.

Sharma, S., K.B. Singh, M.S. Oberoi and N. Sood(1998). Studies on the occurrence of bovineherpes mammillits in buffaloes. BuffaloBulletin, 17:79-81.

Sneath Peter, H.A., S. Mair Nicholas, M. ElisabethSharp and G. Holt John (1986). Bergey’smanual of systematic bacteriology Volume2.

Snedecor, G.W. and W.G. Cochran (1994). Statisti-cal methods 8th ed. Oxford and IBH publica-tion. Co . Pvt. Ltd., Calcutta.

Turner, A.J., I.R. Morgan, W.E. Sykes and W.A.Nicholls (1976). Bovine herpes mammillitisof dairy cattle in Victoria. Australian Veteri-nary Journal, 52: 170-173.

Watson, C.A. (1989). Study of selected antiviralcompounds on bovine herpes mammillitisvirus (BHV-2). European Society for Veteri-nary Virology Liege, Belgium, p. 6-7.

Weaver, L.D., R.W. Dellers and A.H. Dardiri (1972)Bovine herpes mammillitis in New York.Journal of American Veterinary MedicalAssociation, 160: 1643-1644.


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RESEARCH ABSTRACTS

HEALTH AND DISEASES

M.A. Javaid, M.A. Saeed, M. Aleem, I.N. Bashir.Department of Parasitology, Faculty ofVeterinary Science, University of Veterinaryand Animal Sciences, Lahore-54000, Pakistan.Effect of gentamicin and enrofloxacin onuterine characteristics, hematology, cervi-cal mucus and pregnancy rate in postpar-tum endometritic nili-ravi buffaloes. BuffaloJournal (2004), 3:279-288.

The objective of the present study was todetermine if Gentamicin (GENT, 10%, @ 4 mg/kgbody weight, n=20) or Enrofloxacin (ENRO, 10%,@ 5 mg/kg body weight, n=20) I/M for 3 consecu-tive days is effective against the endometritis.Twenty postpartum, untreated, healthy buffaloeswere kept as control (CON). The uterine charac-teristics and blood samples were examined before(day 0) and after treatment (day 4). Mucus sampleswere examined for fern pattern at first estrus,before and after the treatment. Both the treatmentsdid not differ significantly (P>0.05) amongst them-selves; however, they had significantly (P<0.05)beneficial effect on cervical diameter, location ofuterus, texture of uterine wall and symmetry ofuterine horns. Similarly, in GENT and ENRObuffaloes, the estrus mucus characteristics and bloodparameters i.e., erythrocyte sedimentation rate(ESR), hemoglobin (Hb) concentration, total eryth-rocyte count (TEC), total leukocyte count (TLC) anddifferential leukocyte count (DLC) were signifi-cantly (P<0.05) improved. After treatment, estruswas observed in 45% (9/20) of GENT and 55% (11/20) of ENRO buffaloes. The pregnancy rate was60% (3/5) in GENT and 50% (3/6) in ENRO buffa-loes. This was comparable to pregnancy rate in CONbuffaloes (62%, 8/13). Is is concluded that bothGentamicin and Enrofloxacin are equally effectivefor the treatment of endometritis.

S.S. Sidhu, Kulbir Singh, Prahlad Singh. Depart-ment of Animal Reproduction, Gynaecologyand Obstertrics, College of Veterinary Science,Punjab Agricultural University, Ludhiana-141004 (Punjab), India. Bacteriology andcomparative efficacy of LPS and CST basedantibiotics in puerperal metritis in buffa-loes (Bubalus Bubalis). Buffalo Journal(2004), 3:271-278.

Bacteriology and treatment of puerperalmetritis was investigated in 25 buffaloes (15normal calving and 10 suffering from puerperalmetritis) at the Dairy Farm and Veterinary Clinicsof Punjab Agricultural University, Ludhiana.Normal calving buffaloes (n=15) were divided intothree sub groups: Group A (n=5, treated with 40 mlsterile normal saline, intrauterine, once only), GroupB (n=5, treated with 250 µg E. coli LPS dissolvedin 40 ml normal saline as single intrauterine in-fusion) and Group C (n=5, given intramuscular in-jection of antibiotic based on culture and sensitiv-ity test – CST, for 7 days). Clinical group (n=10)was further divided into two sub groups D (n=5,given LPS as in Group B) and E (n=5, given antibi-otics as in Group C). Bacterial load in sub groupsA, B and C increased significantly (P<0.05) fromday 0 to 3 (0.0054±0.0054x103 to 0.221±0.054x103,0.006±0.006x103 to 0.516±0.0132x103 and0.0086±0.0086x103 to 0.0992±0.0098x103 CFU perml respectively). After day 3, in Group A, decreasein bacterial load was significant (P<0.05), whereas,in Groups B and C the reduction in bacterial loadon day 3 was non-significant. Bacterial load inclinical Group D was much higher than in normalcalving buffaloes. Group D indicated a significant(P<0.05) decline in bacterial load from day 0(76.42±18.98x103 CFU per ml) to day 3(6.58±3.43x103 CFU per ml). In clinical Group Ethe significant (P<0.05) decline in bacterial load wasevident from day 3 (40.98±16.99x103 CFU per ml)


71

to day 7 (18.34±8.26x103 CFU per ml). Reductionin bacterial load on days other than specified wasnon-significant in both the clinical Groups D and E.Clinical group buffaloes revealed mixed infectionswith predominance of Coliform, Pseudomonas, andCorynebacterium (Actinomyces) organisms, besidessome incidental microbes (Staphylococci, Strepto-cocci and Bacillus). Pseudomonas and Corynebac-terium persisted till later stages of infection inclinical group. It was consluded that the therapyinitiated immediately at parturition in normallycalving and puerperal metritic buffaloes enhancedexpulsion of infection from the uterus. The therapyinitiated with LPS was more effective than antibi-otics for early clearing of uterine infections.

J.S. Soodan, S.S. Randhawa, C.L. Arora. PunjabAgricultural University, Ludhiana, Punjab 141004, India. Alteration in macro and microminerals in experimentally inducedhypocuprosis in buffalo calves. IndianJournal of Animal Sciences (2005), 75(1):8-10.

Hypocuprosis was achieved in 13 buffalocalves by administering varying doses of ammoniummolybdate. The mean plasma and hair copperlevels were 1.15 plus or minus 0.10 and 8.28 plusor minus 0.38 ppm respectively. Mo inducedhypocuprosis resulted in a significant increase inplasma Cu concentration (1.43 plus or minus 0.18PPM) associated with decline in hair (6.02 plus orminus 0.22 ppm) and liver (22.59 plus or minus10.01 ppm) Cu levels. In addition, increased Fe,phosphorus and decreased zinc concentrations werealso observed. The Mo concentration in the plasma,hair and liver revealed a significant increase through-out the period of induction. Ca, Mg, Mn, K and Sdid not show any significant alteration.

J.S. Soodan, S.S. Randhawa, N.K. Sood. PunjabAgricultural University, Ludhiana, Punjab 141004, India. Radiological and histopathologi-cal alterations in bones in molybdenum-induced hypocuprosis in buffalo calves.Indian Journal of Animal Sciences (2005),75(1):14-16.

Hypocuprosis was achieved in 13 buffalocalves by administering a 20% solution of ammo-nium molybdate in water at 15 mg Mo/kg bodyweight for 3 days, followed by 7.5 mg Mo/kg bodyweight for 4 days and then 5 mg Mo/kg body weightfor another 38 days. The clinical findings observedin the bones were stiffness limbs with enlargementof joints adjacent to long bone. Radiographically,marked widening of cartilaginous growth plate ofmetacarpal and slight lipping was recorded, whilehistopathological analysis of the bones revealedosteoporotic changes in the bony spicules along withirregular and disruptive development of growthplates at costochondra junctions.

Ishwari Prasad Dhakal. Veterinary Medicine,Veterinary Teaching Hospital, Institute ofAgriculture and Animal Science (IAAS)Rampur Chitwan, Nepal. Normal somatic cellcount and subclinical mastitis in murrahbuffaloes. Buffalo Journal (2004), 3: 261-270.

This study was conducted to investigate thenormal somatic cell count and to define subclinicalmastitis in Murrah buffaloes. Data were collectedfrom sixty clinically normal buffaloes stationed atfive farms of Chitwan Nepal and Buffalo ResearchCenter, Hissar, India. Somatic cell count wasmeasured using the Newmann –Lampert stainingtechnique. The upper limit of somatic cell count was


72

determined ≥200,000/ml of milk based on the mean± 2sd of a total somatic cell count. Abnormal dataof the somatic cell count was repeatedly removedwhich lie beyond the values of more than m + 2sduntil all the data come to lie within (m + 2sd).Averages of somatic cell count of right front andright hind quarters were significantly higher thanleft front and left hind quarters. Subclinical masti-tis was diagnosed on the basis of samples withsomatic cell counts ≥200,000/ml with positivebacterial cultures. Subclinical mastitis was foundin 21.7% buffaloes and 8% of the quarter foremilksamples. Neutrophil counts were significantly higherin subclinical mastitis milk.

Ajit Kumar, K.D.R. Prasad, R.R. Kumar. Birsa Ag-ricultural University, Ranchi, Jharkhand 834006, India. Impact of helminthic diseasecontrol on the draught power capability(DPC) in working bullocks and buffaloes.Indian Journal of Animal Sciences (2005),75(2): 224 -225.

The apparent reduction in the health anddraught power capability (DPC) of workingbullocks and buffaloes affected naturally withparamphistomiosis and common GI nematodiosiswas observed to be improved within a fortnightfollowing control of the diseases by the respectiveparasite control package. The DPC was tentativelyestimated by taking observations with respect toploughing agricultural fields in village farmingconditions by the animals.

Raj Sukhbir Singh, B.K. Bansal. Department ofClinical Veterinary Medicine, Ethics andJurisprudence, Punjab Agricultural Univer-sity, Ludhiana-141 004, Punjab, India. Varia-tion in selected components of milk amongdifferent milk fractions and ITS relevanceto diagnosis of mastitis in buffaloes. BuffaloJournal (2004), 3:213-224.

Milk composition pertaining to somatic cellcount (SCC), electrical conductivity (EC), lactoseand pH was compared in three quarter milkfractions (foremilk, mid-milk and strippings) andone udder composite milk from healthy andmastitic buffaloes. A total of 225 quarters from 57lactating buffaloes were studied. All the componentsexcept pH showed a significant variation over themilk fraction and udder health. In general, EC andlactose decreased while SCC increased in strippingsas compared to that in foremilk. The levels of varia-tion over the milk fractions were different for healthyand mastitic quarters. The increase in SCC over themilking was much higher in specific mastitisquarters than that in healthy quarters. Mid-milkcontained significantly lower SCC than that offoremilk in healthy but not in mastitic quarters. ThepH of foremilk and strippings did not differ signifi-cantly, neither in healthy nor in mastitic quarters.The decrease in the EC and lactose content of milkin strippings from that of foremilk was observed tobe more in healthy quarters than in mastiticquarters. The difference between foremilk and mid-milk for SCC, EC, lactose was significant in healthyquarters only. Mastitis at its all levels i.e. specific,specific, non-specific and latent resulted in asignificant alteration in one or more of theparameters studied. The comparison of variousparameters in udder composite milk samplesrevealed significance of effect for milk SCC andlactose but not for EC and pH. The evaluation ofselected parameters in diagnosis of mastitis withrespect to milk fraction showed that highest levelof discrimination was obtained for EC in strippings(66.0%) and for lactose inforemilk fraction(85.65%).


73

I.R. Akbar, A. Shakoor, A. Iqbal, S.U. Rahman, Q.Azeez, U. Waheed. Department of ClinicalMedicine and Surgery, University of Agricul-ture, Faisalabad-38040, Pakistan. Evaluationof the effects of commercial homoeopathicand allopathic preparations on the somaticcell counts and milk composition of Nili-Ravi buffaloes. International Journal ofAgriculture and Biology (2005), 7(1): 94-96.

This study involved 20 subclinically mastiticquarters of 10 Nili-Ravi buffaloes. The animals weredivided into two groups: 10 quarters from six buf-faloes (group A) and 10 quarters from four buffa-loes (group B). Sampling was done before and afterseven days of treatment with either homeopathic(Mastcare injection) or allopathic (Spectrazolintramammary tube+Tribrissen injection) commer-cial preparations. Determinations were made fordirect and indirect microscopic somatic cell counts(SCC) by standard techniques followed by analysisof milk composition (fat, protein, acidity, solidsnot-fat and total solids). Statistical analysis revealedno significant differences in milk compositionbetween groups A and B, while the differences weresignificant before and after the treatments. It wasinferred from the study that the efficacy of bothpreparations was similar. However, the homeopathictreatment had an edge being more economical.

MANAGEMENT AND PRODUCTION

K.S. Pajai, N.P. Dakshinkar. Department of Medi-cine, Nagpur Veterinary College, Nagpur -440006, India. Effect of various treatments onbiochemical parameters and milkproduction in subclinically ketotic buffa-loes. Indian Veterinary Journal (2005),82(1):86-87.

This study was carried out to determine theeffect of several treatments on the blood chemistryand milk production of subclinically ketotic buffa-loes. 10 buffaloes having hypogalactia and positivereaction to Rothera’s test were divided into 2 groups.Group A was subjected to treatment with long-acting insulin along with 20% dextrose, followedby treatment with Rickelvit (calcium levulinate +vitamin D3 + vitamin B12). Group B was treatedintramuscularly with prednisolone at 10 ml/animalalong with 20% dextrose at 1.5 ml/kg body weight,followed by Rickelvit given at 30 ml and Calborolat 200 ml. Biochemical parameters were estimatedand milk production was recorded before and aftertreatment. In group A, treatment with insulin anddextrose decreased the average milk yield, whereasproduction increased in group B after treatment withRickelvit. Plasma glucose decreased after treatmentwith insulin and dextrose and could not be restoredeven after treatment with Rickelvit. Serum calciumlevels decreased insignificantly after insulin anddextrose treatment, but increased by 0.11 mg%after treatment with Rickelvit. Increase in choles-terol, total protein and albumin levels was observedafter treatment with insulin and dextrose. In groupB, treatment with prednisolone and dextroseincreased the average milk yield. This increasedfurther after treatment with Rickelvit and Calborol.

Plasma glucose and calcium levels increased aftertreatment with prednisolone, dextrose, Rickelvit andCalborol. On the other hand, serum cholesteroldecreased after treatment with prednisolone anddextrose. However, the cholesterol level returnedto pretreatment values after treatment with Rickelvitand Calborol.


74

PHYSIOLOGYP. Secchiari, G. Campanile, M. Mele, F. Zicarelli,

A. Serra, M. del Viva, L. Amante, J.F.Hocquette, S. Gigli. DAGA, Sezione ScienzeZootecniche, Universia di Pisa, via delBorghetto, 80, Pisa, Italy. Fatty acid compo-sition and CLA content of milk fat fromItalian buffalo. Indicators of Milk and BeefQuality (2005), 339-343.

Aim of the present work was to characterizethe fatty acid composition and CLA content of milkfrom milk from buffalo fed hay or fresh forage. Milksamples from 88 lactating buffalo belonging to sevenherd located in South Italy were collected. Twosamplings have been performed: the first whenanimals fed a ration included green forage and thesecond when fresh forage was substituted by hay ina total mixed ration (TMR) diet. Milk samples wereanalysed for fatty acid composition. Results showedthat when buffalo cows are fed fresh foragepercentages of medium-chain fatty acids and satu-rated fatty acids significantly decreased in milk fat,while that of monounsaturated fatty acids and poly-unsaturated fatty acids increased. The conjugatedlinoleic acid average content (CLA) of milk wasenhanced by the inclusion of fresh forage in the TMRdiet (0.66 vs. 0.46), in a similar way to whatreported for dairy cattle. A wide individual varia-tion of milk CLA content may be observed, whenanimals were submitted to asimilar dietary regimen.The slope of the relationship between rumenic acid(RA, cis9, trans11 CLA) and vaccenic acid (VA,trans11, 18.1) was comparable to that reported fordairy cattle (RA=0.12+0.23 (VA+RA); p<0.01;R2=0.89).

Virendra Singh, P. Amarpal Kinjavdekar, H. P.Aithal, K. Pratap. Indian Veterinary ResearchInstitute, Izatnagar, Uttar Pradesh 243 122,India. Haematobiochemical changesfollowing epidural administration ofxylazine and ketamine in buffalo calves.Indian Journal of Animal Sciences 2005, 75(2): 178-181.

The haematobiochemical effects of epiduralketamine, xylazine and their combination in buffa-loes were evaluated to examine the safety of thesedrugs. 12 clinically healthy male buffaloes weredivided into 3 groups of 4 animals each. In animalsof groups A, B and C, xylazine (0.05 mg/kg),ketamine (2 mg/kg) and a combination of xylazine(0.05 mg/kg) and ketamine (2 mg/kg), respectively,were administered at the first intercoccygeal epidu-ral space. Blood samples were collected for theestimation of haemoglobin (Hb), packed cellvolume (PVC), total leukocyte count (TLC), plasmaglucose, total proteins, albumin, globulin, A:Gratio, urea nitrogen, creatinine and gamma-glutamyltransferase (GGT) up to 48 h after injec-tion of the drug(s). Reduction in Hb, PCV and TLCwas observed from 30 to 120 min in animalsbelonging to all groups. A significanthyperglycaemia was recorded in all the groups. Thiswas slightly longer in group C. A significant(P<0.05) decrease in plasma proteins was recordedin animals of group A. In animals of groups B andC, an insignificant change in the total plasmaproteins was recorded. An insignificant decrease inplasma albumin and an increase in plasma globulinwere recorded in all groups. A transient but signifi-cant (P<0.05) increase in urea nitrogen, creatinineand GGT was recorded in all the groups. Thevalues for these parameters, however, remainedwithin the normal reference range. Since the changesin various haematobiochemical parameters wereonly transient and reversible, these drugs could beconsidered safe for epidural analgesia in buffaloes.However, the combination of xylazine and ketaminewas better than xylazine alone.


75

CONTENTS

BUFFALO BULLETINIBIC, KASETSART UNIVERSITY, P.O. BOX 1084

BAGKOK 10903, THAILANDURL : http//ibic.lib.ku.ac.thE-mail : [email protected]

Tel.: 66-2-9428616Fax : 66-2-5611369

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Locally imfitrative, non-metastasizing transtional cell carcinoma leadingto dystocia,anuria and constipation in an Indian water buffalo (Bubalus bubalis)

Naresh Kumar Sood, Kuldip Gupta, Neerai Sood, Mrigank Honeparkhe,and Amarjit Singh.......................................................................................................... 51

Studies on etiology, symptomatology, diagnosis and therapy in ulcerative thelitisof buffaloes in Andhra Pradesh (India)

M. Lokanadhamu, B. Sreedevi and T. Venkata Reddy..................................................... 56

RESEARCH ABSTRACTS........................................................................................... 70

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