Bubaline Species Research Articles

Buffalo anti-PDC-109 antibodies improve the semen quality profiles and in-vitro zona binding index and minimize the cryoinjury of sperm in cryopreserved buffalo semen

Objective: To optimize the concentration of PDC-109 protein in semen for higher cryopreservability and fertility by sequestration or neutralization of PDC-109 by its antibodies (anti-PDC-109 antibodies) in bubaline species. Methods: PDC-109 protein was purified by applying two-step chromatography procedures. Purified protein was injected in rabbits to raise antibodies. These raised anti-PDC-109 antibodies were used in neutralization or sequestration of PDC-109 in in-vitro model. Ejaculates were collected from buffaloes and splited for four groups. Group 1 received egg yolk Tris glycerol extender, without anti-PDC-109 antibodies, while group 2 to 4 received anti-PDC-109 antibodies 266 μg/mL, 80 μg/mL, and 26 μg/mL in Tris-fructosecitrate buffer, respectively. Semen quality parameters viz., forward progressive motility, viability, total morphological abnormality, acrosomal integrity, plasma membrane integrity, cryoinjury and in-vitro zona binding index were evaluated. Results: Semen quality parameters of neat semen were within the normal range of bubaline species. Sperm motility, livability, acrosomal integrity, plasma membrane integrity, and cholesterol content of sperm were decreased and total sperm abnormality was increased significantly in post-thaw semen compared to those in pre-freeze and fresh semen (P<0.05). Semen in group 2 had higher sperm motility, livability, acrosomal integrity, plasma membrane integrity, and cholesterol content of sperm and lower total sperm abnormality significantly compared to those in group 1, 3 and 4 at pre-freeze and post-thaw stages (P<0.05). Conclusions: Sequestration or neutralization of PDC-109 by its antibodies significantly improves pre-freeze, and post-thaw semen quality parameters and in-vitro zona binding index with simultaneously reducing cryoinjury or cryodamage in the sperm of bubaline species.

Molecular Characterization of TLR4 Gene of Swamp and Riverine Type of Water Buffaloes

Molecular characterization of TLR4 genes of swamp and riverine types of water buffaloes was conducted to determine unique genotypic characteristics specific to each type of water buffalo and provide baseline information for explaining differences in disease resistance between each type. The nucleotide sequence of the riverine TLR4 gene produced 2,526 bp, whereas the swamp TLR4 sequence produced 2,322 bp. Riverine and swamp buffalo TLR4 nucleotide sequences were 99% similar to that of B. bubalis sequence from GenBank. The translated amino acid sequence of TLR4 showed that the riverine buffalo TLR4 sequence had a higher percentage similarity to B. bubalis sequence as compared to the swamp buffalo TLR4 amino acid sequence. Phylogenetic analysis confirmed the degree of relationship between the bubaline species and showed the distinctness of each breed. The predicted functional domains identified were 13 LRRs and a single LRRCT. Transmembrane protein and globular cytoplasmic domain TIR1 were found to be conserved regions.

Single nucleotide polymorphism in STAT5A could not endorse variation in milk production traits in Indian bovine population.

The Signal Transducer and Activator of Transcription 5A (STAT5A) gene involved in activating the transcription of milk protein genes was predicted to be influencing milk production traits. The present study was undertaken to investigate the suitability of the polymorphism of STAT5A as a marker for milk traits in Ongole, crossbred cattle and Murrah buffaloes from Southern India. Blood samples (n = 502) for DNA isolation and milk samples (n = 222) from different genetic groups were collected from various farms. The gene variants upon polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) on the exon 7 region of STAT5A were subjected to GLM analysis to evaluate their association with milk production traits. The frequencies of C and T alleles at the STAT5A/AvaI locus were 0.98 and 0.02 (Jersey crossbred), 0.94 and 0.06 [Holstein-Friesian (HF) crossbred], 0.97 and 0.03 (Ongole). T allele was not observed in Murrah buffaloes. The least squares mean lactation milk yield of CC and CT genotypes of STAT5A were 2,096.90 ± 48.63 and 2,294.41 ± 215.85 kg in Jersey crossbred, 2,312.92 ± 91.01 and 2,392.82 ± 207.66 kg in HF crossbred and 528.40 ± 22.10 and 396.37 ± 76.17 kg in Ongole cattle, respectively. The milk fat content of the CC genotype was higher (P > 0.05) in Jersey crossbred cattle. The CT genotypes of Ongole and HF crossbred cattle recorded a higher fat per cent than the CC genotypes. Significant associations were not observed in support of STAT5A as a marker for milk production traits in either Ongole or crossbred cattle of indicine admixture and no reason could be found to consider this locus as universal markers for milk production traits in indicine cattle and buffaloes. Considering the monomorphic nature of the gene in buffaloes and their higher milk fat content as compared to bovine milk, much remains to be explored regarding the underlying differences across the bovine and the bubaline species.

Cleavage Rate of in vitro Matured Oocytes Fertilized with Conventional Semen in Buffaloes

Background: In vitro embryo production in buffaloes has gained much importance in this current scenario due to ever increasing population and high demand of milk and meat. Slaughter house derived bubaline ovaries are a cheap and abundant source of cumulus oocyte complexes.Methods: Oocytes from the buffalo ovarian follicles were recovered by aspiration technique as it facilitates quick recovery. Total 155 ovaries were used in the present study. Surface follicles were measured using vernier calliper and categorized into three groups viz. less than 3 mm, 3-5 mm and greater than 5 mm based on follicular diameter and oocytes were processed for IVM, IVF and IVC using conventional non sorted semen.Result: Overall percentage of small, medium and large follicles in the ovaries were recorded as 16.29 ± 0.94%, 8.14±0.60%, 5.35 ± 0.76%, respectively. Overall recovery rate of COCs was 38%. The percentage of these oocytes were 16.74% (A), 15.25% (B), 25.26% (C), 18.33% (D) and 29.87% (E) respectively. Maturation rate of oocytes were 81.96 ± 2.70%. Fertilization rate was 74.98 ± 3.87%, Cleavage rate % was 40.84±2.51% and Blastocyst percentage was 21.57±1.75% respectively. Application of in vitro embryo production technique using slaughter house ovaries can salvage the genetic potential of bubaline species.

Hematology and biochemistry of buffalo (Bubalus bubalis): influence of sex and age on reference values.

This research aimed to obtain a hematological and biochemical profile of buffalos (Bubalus bubalis) bred in a region of the Brazilian Legal Amazon. A total of 73 animals of mixed races, divided into three groups, were studied: group 1 (G1) comprised animals up to 11 months old; in group 2 (G2), subjects were 12 to 23 months old; and, in group 3 (G3), they were 24 months old and older. We performed the hematological analysis manually, and, for the biochemistry, we used biochemical analyzers. Age of the animals has influenced the monocyte count. The counting of these cells was higher (p > 0.05) in younger animals (G1). There was no difference (p > 0.05) in hematological parameters concerning sex. In regard to biochemistry, we found that age influenced results for albumin and urea. Animals from G1 had higher albumin values when compared to G2 and G3. Serum urea values were higher in animals from G3. Regarding the serum mineral parameters, there were no significant results when the different age groups and both sexes were compared. Thus, the hematological and biochemical values obtained can work as a reference for the bubaline species of animals bred in the studied region, under the same management and breeding conditions. Sex and age of the animals are necessary for the interpretation of the tests.

Identification of important miRNAs in bubaline mammary gland at heifer stage - An in-silico approach

The present study was undertaken to identify the microRNAs (miRNAs) expressed in the mammary tissue of a bubaline heifer. Small RNAs were isolated from the mammary gland tissue and enriched for miRNA fraction. The linker-ligated small RNAs were reverse transcribed to synthesize cDNA and amplified by PCR. The PCR products were ligated to the pGEM-T Easy vector; cloned into DH5 alpha cells and sequenced. Sequencing of 40 clones, randomly chosen from this library, produced 115 concatemerized short sequences. The short sequences were checked for their matches with the help of refseq_rna database, EST database (NCBI) and Ensembl. The analysis was performed for more than 90% identity with miRNAs across different species in miRBase. Alignment of putative small RNA sequences with the bovine genome was attempted in GenBank (NCBI) and Ensembl using BLAST. The small RNA sequences with a partial matches within the bovine genome and/or with flanking sequences (upstream or downstream) were analyzed for hairpin structures using the Mfold web server. Reverse complements were also assessed for the homology search. miRNA sequences showing only a partial match with already reported sequences were considered as a putative bubaline miRNAs. Six developmentally important putative miRNA precursors were identified from this study using cloning and sequencing followed by the Bioinformatics approach. This study will help in the elucidation of pathways involving miRNAs in bubaline species at the heifer stage.

Exogenous cholesterol prevents cryocapacitation-like changes, membrane fluidity, and enhances in vitro fertility in bubaline spermatozoa.

A study was conducted to determine the optimum dosage of the exogenous cholesterol-loaded cyclodextrins (CLC) to get maximum cryoprotection for bubaline spermatozoa. In the present study, 120×106 spermatozoa were incubated in 2, 3 and 4mg of CLC as grouped as Gr II, III and IV, respectively, and sperm progressive motility, intracellular Ca2+ , capacitation status by protein tyrosine phosphorylation (PTP) assay and zona binding per cent (ZBP) and cleavage rate (CR) of the cryopreserved buffalo spermatozoa by in vitro fertility assay were assessed in comparison with an untreated control group (Gr I). Results revealed that there was a significant (p<.05) linear decrease in percentage of sperm population with higher intracellular Ca2+ and percentage of sperm population with medium or high capacitated by PTP in CLC treated from 2 to 3mg and then increased to 4mg/120×106 spermatozoa whereas sperm progressive motility, percentage of sperm population with low capacitated, ZBP and CR were increased significantly (p<.05) in sperm population treated from 2 to 3mg CLC and then decreased to 4mg/120×106 spermatozoa. The study has clearly indicated that CLC at 3mg/120×106 spermatozoa has maximum beneficial effects in protection of sperm progressive motility, membrane fluidity (low intracellular Ca2+ ); prevention of cryocapacitation (low capacitation pattern in immunolocalization) and enhancement of in vitro ZBP and CR. Post-thaw motility of the CLC-treated sperm has shown positively significant (p<.05) correlation with sperm population with low intracellular Ca2+ , low capacitated sperm population, ZBP and CR, whereas it was negatively (p<.05) correlated with sperm population with high intracellular Ca2+ , medium or high capacitated sperm. The present study has revealed for the first time that incubation of spermatozoa with CLC of higher dose (>3mg/120×106 spermatozoa) had adverse effects on sperm cryopreservation, although incubation of sperm with 3mg/120 million prior to processing had minimised the freezing-thawing-associated damages in bubaline species.

Anti-Chlamydia abortus and Anti-Mycobacterium avium subsp. paratuberculosis Antibodies in Buffaloes in the State of Pernambuco, Brazil

Background: The occurrence of economic losses in buffaloes may be related to reproductive problems such as chlamydiosis caused by the bacteria Chlamydia abortus considered as a zoonotic agent; and digestive problems highlighting the infection by Mycobacterium avium subsp. paratuberculosis (Map), responsible for paratuberculosis. There is a little information about these diseases in buffaloes, therefore the aim of this study was to determine the occurrence of anti-Chlamydia abortus and anti- Mycobacterium avium subsp. paratuberculosis (Map) antibodies in water buffaloes in the state of Pernambuco, Brazil.Materials, Methods &amp; Results: The 262 bubaline sera belonging to the serum bank of the Infectious Diseases Laboratory (LIDIC) of the Federal Rural University of Pernambuco (UFRPE) were analyzed. The samples were from nine properties distributed in the municipalities of Agreste and Zona da Mata of the state of Pernambuco. For the detection of anti-Chlamydia abortus and anti-Map antibodies was used a technique of Enzyme immunoabsorption Assay (ELISA) of the IDEXX® by following the manufacturer's instructions. Regarding the detection of anti-Chlamydia abortus antibodies, it was observed that 47.70% (125/262) of the samples were positive. All properties showed at least one positive animal for the investigation of anti-Chlamydia abortus antibodies. It was also verified the occurrence of 7.25% of suspected animals for the investigation of anti-C. abortus antibodies. No positives animals were observed for Map in the bubaline analyzed.Discussion: The occurrence of anti-C. abortus antibodies in buffaloes in the region can be explained by the fact that properties with buffalo breeding has sanitary management that allows the contact between animals, thus increasing the risk of transmission of the agent. In addition, the variation found from 35% to 68.75% may be associated with divergences to the type of management and breeding system used in each property. The percentage of suspected animals may suggest that the number of positive animals is higher or that there were non-specific reactions with other species of Chlamydia, but it is not possible to determine the seroconversion without the accomplishment of paired serology. Because it is a zoonosis, C. abortus may be a risk to the health of the population involved, since the transmission of the bacteria to humans can occur by contact with secretions and excretions of these animals. No positive animals were found for the occurrence of anti-Map antibodies, however, there are reports of the infection identifying animals with clinical signs or properties with a history of the disease, and this may be related to the differences in each property in relation to the history and sanitary management. Moreover, the ELISA test may not be sensitive to the agent depending on the phase of the infection, because if the response is predominantly cellular, the number of antibodies is diminished, making the serological diagnosis difficult. The use of more sensitive tests for the bubaline species can also favor the diagnosis of the infection. Although the occurrence of anti-Map antibodies in the analyzed samples was not detected, it is important to carry out routine epidemiological studies, since the disease has already been registered in water buffaloes in the state of Pernambuco. Considering the occurrence of anti-Chlamydia abortus antibodies it is suggested to conduct studies in order to isolate the agent and verify its real importance in the reproductive disorders of the bubaline species, since this agent causes reproductive losses and has a zoonotic character.

Seasonal and Ageing-Depending Changes of Aquaporins 1 and 9 Expression in the Genital Tract of Buffalo Bulls (Bubalus bubalis).

The presence of Aquaporins 1 (AQP1) and 9 (AQP9), integral membrane water channels that facilitate rapid passive movement of water and solutes, was immunohistochemically detected in the excurrent ducts collected from sexually mature buffalo bulls of proven fertility during the mating (late autumn-winter) and non-mating (late spring to the beginning of autumn) seasons. Furthermore, the research was performed also on the epididymal cauda of a senile buffalo bull with inactive testis. Aquaporins 1 and 9 were immunolocalized at distinct levels. In the efferent ducts, AQP1 immunoreactivity was strongly evidenced at the apical surface of the non-ciliated cells and weakly along the basal membrane of the epithelial cells. The latter reactivity disappeared during the non-mating season. No AQP1 immunoreactivity was detected in the epithelium of epididymis and vas deferens, whereas AQP1 was expressed in the smooth muscle layer of the vas deferens. Aquaporin 1 was present in the blood vessels and in small nerve bundles all along the genital tract. The supranuclear zone of the epididymal principal cells was AQP9 immunoreactive, limited to the corpus and cauda regions, and vas deferens. The samples collected in the two reproductive seasons showed a weaker AQP9 immunoreactivity during the non-mating season. A typical AQP9 immunoreactivity was noticed in the old buffalo examined. The tested AQP molecules showed a different expression pattern in comparison with laboratory mammals, primates, equine, dog and cat. In addition, seasonal differences were noticed which are possibly useful in regard to the comprehension of the morphophysiology of reproduction in the bubaline species, which are still a matter of debate.

Molecular comparison of Slc11a1 and Slc11a2 genes of swamp- and riverine-type water buffaloes.

Solute-linked carrier 11a and 11a2 (Slc) have been associated with disease resistance and/or susceptibility across animal species. These genes have an important mechanism in the regulation against intracellular infection. This study analysed the genetic characteristic of Slc 11a and 11a2 in swamp-type and riverine-type water buffaloes to understand their immunological distinction. Characterization of Slc11a1 and Slc11a2 genes from swamp- and riverine-type water buffaloes was carried out by molecular cloning, sequencing and phylogenetic analysis. The cloned cDNA of Slc11a1 and Slc11a2 contained an open reading frame of 1647 and 1723 nucleotides, encoding 549 and 574 amino acids, respectively. Nucleotide sequence homology of both Slc11a1 and Slc11a2 had 99% in swamp and riverine type, which gives almost identical polypeptide. However, Slc11a1 and Slc11a2 have substitutions of 5 and 1 amino acid residues, correspondingly. These substitutions suggest as a potential gene markers for resistance and/or susceptibility to intracellular infection. Furthermore, phylogenetic analysis confirmed the degree of relationship between the bubaline species and justifies the distinctness of each breed by the bootstrap value generated.

Localization of IGF proteins in various stages of ovarian follicular development and modulatory role of IGF-I on granulosa cell steroid production in water buffalo (Bubalus bubalis)

The present study aimed to determine the expression of insulin like growth factor (IGF) genes in the bubaline ovarian follicles and modulatory role of IGF-I on progesterone production from granulosa cells (GC) of pre-ovulatory follicle in vitro. According to size, follicles were classified into four groups: GI (small), GII (medium), GIII (large) and GIV (preovulatory). All IGF genes were expressed in both GC and theca interna (TI) cells. The relative expression of IGF-I and IGF receptor I (IGFR-I) genes increased with follicle size and was greatest in the pre-ovulatory follicle (P<0.05). Expression of IGF-II and IGFR-II genes was minimal in GC but was readily detected in TI cells. In TI cells, the gene expression was greater in medium and large as compared to small and pre-ovulatory follicles. The expression of all binding protein (IGFBP) genes was detected in both GC and TI cells. Expression of IGFBP-3 gene increased with follicle size and was greatest in pre-ovulatory follicles (P<0.05). The expression of IGFBP-2 and IGFBP-4 was less in pre-ovulatory follicles but expression of IGFBP-5 and IGFBP-6 genes were greater at this stage. The GC culture was conducted for three time durations and with three doses of IGF-I. Expression of steroidogenic genes (StAR, CYP11A1, HSD3B) and progesterone concentration were increased in a dose and time dependent fashion. The present study, therefore, provided evidence of an autocrine/paracrine role of IGFs in follicular development and a stimulatory role of IGF1 in steroid production in GC of preovulatory follicles in the bubaline species.

Functional Characterization of Bubaline Recombinant-Cathelicidin

Cathelicidin proteins are new candidates for drug designing, as their peptides exhibit antimicrobial activity against bacteria, viruses, and fungi. Bubaline species are well known for their tolerance to environmental stress and disease with the innate immune system playing a pivotal role in meeting these functions. Functional characterization of cathelicidin antimicrobial peptides of bubaline origin could give us an insight into how bubaline species is more tolerant to diseases than their other bovidae counterparts. In the present study a novel cathelcidin peptide of bubaline origin was characterized by cloning and in vitro expressing the recombinant peptide. Adult dairy bubaline peripheral blood was collected aseptically from the jugular vein in 0.5 M EDTA. The total RNA was extracted and reverse transcribed to cDNA. The sequence encoding the mature peptide portion of cathelicidin was cloned in a E. coli system using pProExHta vector and confirmed by custom sequencing. The positive clones were induced at room temperature with 1 mM IPTG to express recombinant cathelicidin which was confirmed by Western and Dot Blotting. The antimicrobial activity of the recombinant cathelicidin was tested against gram positive cocci Staphylococcus aureus. The bacteria produced small zones of inhibition around discs of 5 and 10 μg recombinant cathelicidin while no inhibitory zone could be detected around 2 μg recombinant cathelicidin.

Produção de interleucina-1beta e severidade da mastite pós-inoculação de Staphylococcus aureus na glândula mamária de bovinos e bubalinos

O presente trabalho objetivou avaliar, em vacas e em búfalas submetidas à mastite induzida por inoculação de Staphylococcus aureus, a concentração da citocina pró-inflamatória interleucina-1β (IL-1β), a contagem de células somáticas (CCS) e a correlação destas com alguns parâmetros da resposta local e sistêmica à inflamação. Os animais tiveram uma glândula mamária inoculada e o processo inflamatório foi monitorado pela cultura bacteriológica do leite, CCS, quantificação da IL-1β na secreção láctea, avaliação da aparência/consistência da glândula, aparência da secreção láctea (resposta localizada à inflamação) e aferição da temperatura retal (resposta sistêmica à inflamação). Houve elevação nos níveis de IL-1β, na CCS e resposta localizada e sistêmica à inflamação, tanto na espécie bovina como na bubalina. A cinética da produção da citocina foi diferente nas duas espécies (P&lt;0,05), sendo que as búfalas apresentaram elevação mais rápida, porém com níveis menos elevados, quando comparadas às vacas. As duas espécies alcançaram contagens máximas semelhantes (P&gt;0,05) de CS/mL de leite, com concentrações diferentes (P&lt;0,05) de IL-1β/mL de leite. Os parâmetros utilizados para verificar a resposta localizada à inflamação demonstraram escores médios mais elevados na espécie bovina. Correlação positiva entre a concentração da IL-1β no leite, CCS e parâmetros utilizados para avaliar a severidade da mastite foi verificada somente na espécie bovina. Os resultados evidenciaram que a cinética de produção da IL-1β foi diferente nas espécies bovina e bubalina e demonstraram que as búfalas desenvolveram um processo inflamatório mais brando, com restabelecimento mais rápido dos parâmetros utilizados para avaliar a severidade da mastite

Characterization of CTLA-4, PD-1 and PDL-1 of swamp and riverine type water buffaloes

Characterization of CTLA-4, PD-1 and PDL-1 genes from swamp and riverine type water buffaloes was done by molecular cloning, sequencing and phylogenetic analysis. The cloned cDNA of CTLA-4, PD-1 and PDL-1 contained an open reading frame of 666, 849 and 870 nucleotides, encoding a polypeptide of 221, 282 and 298 amino acids, respectively. Nucleotide sequence homology of both CTLA-4 and PDL-1 had 99.8% in swamp and riverine type, which gives the identical polypeptide. Meanwhile, PD-1 genes of swamp and riverine type water buffaloes had 99.2% of homology in nucleotide sequence, which has substitution of two amino acid residues. The hexapeptide motif, phosphatidylinositol 3'-kinase and potential glycosylation sites were conserved within the tribe Bovinae. Phylogenetic analysis confirmed the degree of relationship between the bubaline species and justify the distinctness of each breeds by the bootstrap value generated.

Molecular cloning, sequencing and phylogenetic analysis of inflammatory cytokines of swamp type buffalo contrasting with other bubaline breeds

The current research concerned in the cloning, sequencing and phylogenetic analysis of inflammatory cytokine (IL-1alpha, IL-1beta, IL-6 and TNF-alpha) genes from swamp buffalo and two bubaline breeds, CB (cross between swamp and riverine type buffalo) and the Bulgarian Murrah buffalo. Multiple sequence comparison showed a high homology between the bubaline breeds, which ranged from 99.3% to 100.0% similarity, whereas from 98.6% to 99.0% compared to cattle. The phylogenetic analysis had confirmed and justified the degree of relationship between these bubaline species and their distinctness to each other by the bootstrap value (%) generated. These findings were discussed with particular attention to the diversity of the inflammatory cytokine proteins within closely related species. The result of this study concluded that a small difference in the cytokine structures might be the reason behind or has a contributory factor on the previous reports about the existence of disease resistance. However, in-depth study is necessary to further qualify these findings.

A set of cattle microsatellite DNA markers for genome analysis of riverine buffalo (Bubalus bubalis).

One hundred and eight microsatellite primer pairs, originally identified from cattle, were evaluated for their applicability in buffalo. Eighty-one primer pairs (75%) amplified discrete products, and of these, 61 pairs (56%) gave polymorphic band patterns on a panel of 25 buffaloes. The mean number of alleles per polymorphic marker was 4.50 +/- 0.20, and the mean heterozygosity per polymorphic marker was 0.66 +/- 0.02. Successful genotyping of buffaloes using cattle specific primers suggests that the latter can be a valuable resource for genome analysis in bubaline species.

External Characters and Karyotypes of the Captive Tamaraws, Bubalus (B.) mindorensis, at the Gene Pool in the Island of Mindoro, Philippines

Measurement values of sixteen external characters and karyotypes were described for one female and five males of the adult captive tamaraws, Bubalus (B.) mindorensis endemic to the island of Mindoro. Principal component analyses among bubaline species, using nine or ten of the characters as variables, derived more than 95% of total variance on the first component axis, in which the tamaraw was located almost at the midpoint of two distant bubaline groups : the dwarf water buffaloes of Sulawesi (Anoa depressicornis and A. quarlesi) and the Asiatic water buffaloes (the swamp and the river types of Bubalus bubalis) . All the six tamaraws had 46 chromosomes in diploid form (2N) and 58 as the fundamental number (FN) including the sex chromosomes. In contrast, all the bubaline species of Syncerus, Bubalus and Anoa were reported to have FN=60 with exception of the swamp type of the Asiatic buffalo (FN=58 but 2N 48) . The X chromosome of the tamaraws was the largest acrocentric, and the Y was also acrocentric, probably among the smallest acrocentrics. The karyotype of the tamaraws differs from that of the swamp type buffalo both by one additional pair of metacentrics and the two decreased pairs of acrocentrics in the autosomes.