The taxonomic status, distribution and conservation of the lowland anoa Bubalus depressicornis and mountain anoa Bubalus quarlesi

1. The anoas are two species of dwarf buffalo, the lowland anoa Bubalus depressicornis and mountain anoa Bubalus quarlesi that are endemic to the island of Sulawesi, Indonesia. The classification of the subgenus Anoa within Bubalus is upheld by assessment of recent genetic and morphological research. The classification of anoas into two species is still debated, but with the absence of significant opposing evidence, this position is adopted here. 2. Information about the distribution of the two species is presented that adds to but largely supports existing reports. However, it is still uncertain whether the two putative species are sympatric or parapatric in their distribution. A review of anoa distribution from historical reports and recent field data (1990s to 2002) highlights their decline throughout Sulawesi, especially in the southern and north-eastern peninsulas. The decline has been attributed to local hunting for meat and habitat loss. Most populations are rapidly becoming fragmented, suggesting that the conservation of viable populations may eventually require management of metapopulations. 3. There is an urgent requirement for conservation efforts to: (i) protect anoas from hunting; (ii) prevent habitat loss in key sites; (iii) complete genetic studies to better determine the number of anoa taxa and Management Units and assess their distribution; and (iv) determine the status of the remaining anoa populations.     

In vitro culture of the strobilar stage of Echinococcus granulosus from protoscoleces of human, camel, cattle, sheep and goat origin from Kenya and buffalo origin from India

Protoscoleces from human, camel, cattle, sheep, goat (all from Kenya) and buffalo (from India) hydatid cysts were cultured under identical conditions in vitro using the diphasic culture system of Smyth (1979b). Organisms from all sources grew and segmented in culture. Genital anlagen developed in all cultured worms but further genital differentiation occurred only in cultures of cattle (testes) and camel (testes and genital pore) material. The possible significance of these results is discussed in relation to the general epidemiology of hydatid disease and the potential infectivity of the different strains to man.     

Studies on activation and inhibition of cathepsin B from buffalo liver

Cathepsin B (EC 3.4.22.1) was purified from buffalo liver. The enzyme activity against-benzoyl-dl-arginine-naphthylamme (BANA) was substantially reduced by heat (above 37C) and by nondenaturing concentrations of urea (3 M) and guanidine hydrochloride (1 M). Cathepsin B was significantly activated by 1.5 mM EDTA alone. The activation of the enzyme was further enhanced in the presence of thiol compounds, e.g., cysteine thioglycolic acid, 2,3-dimercapto-1-propenol, and dithioerythritol (DTE). The minimum concentration of the thiol compound required for optimal activation of cathepsin B was found to be lowest (0.2 mM) for DTE. The BANA hydrolyzing activity of cathepsin B was substantially reduced by Cu²⁺ (20–200M) and Ca²⁺ (30–250 mM) as well as by thiol blocking reagents, e.g., iodoacetate, 5,5-dithiobis(2-nitro-benzoic acid) (DTNB), andp-hydroxymercuribenzoate (pHMB). The enzyme activity was completely abolished when the molar ratio of the reagent: cathepsin B was close to 1. The number of free sulfhydryl groups in cathepsin B was determined to be 2 by titration against DTNB and pHMB. Modification of one free thiol group of cathepsin B resulted in complete loss of BANA hydrolyzing activity.     

Divergence between the Anoas of Sulawesi and the Asiatic Water buffaloes, inferred from their complete amino acid sequences of hemoglobin ß chains

Four complete amino acid sequences of hemoglobin β chains were determined for the swamp and the river types of the Asiatic water buffalo (Bubalus bubalis) and two species of the subgenus Anoa in Bubalus; B. (A.) depressicornis (H. Smith, 1827), the lowland anoa, and B. (A.) quarlesi (Ouwens, 1910), the mountain anoa. The two types of the bubalis were identical in the 145 amino acid residues of the β chains and, compared to this sequence, the two residues were substituted in the depressicornis (β49Thr → Ser and 134Ala → Thr) and the five were in the quarlesi (β53Val → Ile, 74Met → Ile, 111Val → Ile, 115Arg → His and 134Ala → Thr). While both Anoa species diverged from the bubalis by the β134Ala → Thr, they differed from each other by the five substitutions. The Anoa species are endemic to Sulawesi of Indonesia. Their speciation and the present coexistence were discussed with reference to probable immigrations of two ancestral Anoa species to Sulawesi at so long interval that had caused a reproductive isolation between the two wild animals. The earlier immigrants were postulated to be ancestral to the quarlesi and the later ones to the depressicornis.     

Selection of buffalo bulls: Sexual behavior and its relationship to semen production and fertility

Forty-four buffalo bulls, used for artificial insemination, were studied to develop libido, mating ability and sexual behavior indices for selection purposes. For each index, 5 categories (i.e., excellent, very good, good, fair and poor) were established. The sexual behavior index was found to be more reliable than the libido and mating ability indices. Buffalo bulls in good to excellent categories were considered acceptable sires.

Reaction time, sexual aggressiveness, and scores of libido, mating ability and sexual behavior differed significantly among the various categories of the 3 indices. Libido significantly correlated with mating ability (r=0.89; P<0.001). Sexual behavior expressed significant relationship with age (r=0.41; P<0.01) and body weight (r=0.48; P<0.01), but was nonsignificant with the scrotal circumference (r=0.28; P>0.05) of buffalo bulls. However, these relationships were absent (P>0.05) in the acceptable sires. Semen production was correlated with sexual behavior in only the fair and poor categories of buffalo bulls (r=0.84; P<0.005). Sexual behavior had no relationship with the fertility rate of buffalo bulls (r=0.44; P>0.05). It is concluded that the sexual behavior index can be used successfully for the selection of buffalo bulls. Excellent- to good bulls should be used in an artificial breeding program if they qualify in the other selection indices.     

Genetic studies of water buffalo blood markers. I. Red cell acid phosphatase,albumin, catalase,red cell α-esterase-3, group-specific component,and protease inhibitor

We have developed the methodologies for typing and family studies to establish the modes of inheritance of water buffalo red cell acid phosphatase (Acp), protease inhibitor (Pi), and group-specific component (Gc) on isoelectric focusing and albumin (Alb), red cell -esterase-3 (Est-3), and catalase (Cat) on polyacrylamide gel electrophoresis. Family studies showed that Pi, Gc, Alb, and Cat are coded by autosomal genes with two codominant alleles, while Est-3 is autosomal with two codominant alleles and a recessive null allele and Acp exhibits three codominant alleles.This project was funded by the Australian Centre for International Agricultural Research through Grant PN 8364 and the Malaysian programme for Intensification of Research in Priority Areas through Grant IRPA 1-07-05-057.     

Age-related changes in body weight,scrotal size and plasma testosterone levels in buffalo bulls (Bubalus bubalis)

Body weight, testicle size and peripheral testosterone concentrations were measured in 35 water buffalo bulls at 5, 15, 17, 21, 25 and 38 months of age. These parameters were studied in all animals during the same month (October), so the changes due to age were independent of changes in photoperiod and temperature. Body weight increased linearly with age. The testicular size measured in terms of scrotal circumference as related to age, showed a curvilinear increase; the average rate of testicular growth was maximum between 15 and 25 months. Plasma testosterone levels were low between 5 and 21 months. A significant rise in plasma concentration of testosterone was observed at 25 months reaching peak levels at 38 months. The mean age of sexually mature bulls at the time of first ejaculation of semen with motile sperm, was 24.9±0.9 months (n=9). It has been concluded that in the Nili-Ravi buffalo bulls the sexually quiescent period (prepubertal) extends up to 15 months of age and sexual maturation as indicated by the presence of motile sperm in the ejaculate is attained at about 25 months.     

Changes in profiles of serum sex steroids of male buffaloes from birth to maturity

Age-related changes in testosterone, progesterone and estradiol 17-beta were determined by radioimmunoassay (RIA) in the serum of 155 male buffalo calves of varying ages. The calves were classified into 17 age groups. The mean weight of calves increased from 33.6 +/- 9.6 kg at one week of age to 531 +/- 41.4 kg at 42 months. The testosterone levels were less than 100 pg/ml from birth until 15 months of age, followed by peak concentrations of 422 +/- 79 pg/ml at 24 to 30 months and 793 +/- 193 pg/ml at 42 to 48 months (corresponding to puberty and maturity, respectively). The progesterone levels were higher in newly born calves and mature bulls. Otherwise, the levels continued to be low throughout the period of growth and development. Estradiol 17-beta was significantly higher in postnatal calves up to two months of age. The testosterone revealed a positive correlation with weight and age while E2 17-beta showed a negative correlation with age. These results do not support a direct role of peripheral progesterone and estradiol 17-beta in the onset of puberty and sexual maturity of buffalo bulls.     

Polymorphism of mitochondrial DNA (mtDNA) in cattle and buffaloes

Mitochondrial DNA (mtDNA) from two breeds of cattle, viz., [Hariana (Bos indicus), Holstein (Bos taurus)] and Indian water buffalo (Bubalis bubalus), was analyzed using 13 restriction endonucleases which recognized an average of about 40 six-base sites. Polymorphism among cattle was detected with six of these enzymes. The two Holstein differed at six sites, whereas the Hariana breed (Bos indicus) did not show any site polymorphism. Surprisingly, the Hariana type differed by only one site from one of the Holstein types. The total size of buffalo mtDNA was estimated to be 16.4 kb. Polymorphism within the Murrah buffalo breed was observed with respect to aBglI site. Scarcely any of the restriction fragments of buffalo mtDNA matched those of cattle mtDNA.     

Caffeic acid improves microscopic sperm parameters and antioxidant status of buffalo (Bubalus bubalis) bull semen following freeze-thawing process

This study investigates the effects of cryopreservation and supplementation of buffalo's semen with Caffeic acid. It studies the effects of different Caffeic acid concentrations on cryopreservation capacity of the buffalo and evaluates their influence on various sperm parameters like motility, viability, progressive motility, sperm plasma membrane integrity, and antioxidant status. Twenty-four semen samples were collected with an artificial vaginal from three adult water buffalos. The semen samples were evaluated and the qualified ejaculates were separated and were diluted in a Tris-based extender. The resulting samples were classified into 5 groups: No antioxidant (control), Control sham (NaOH), Caffeic acid 50 μM, Caffeic acid 100 μM, and Caffeic acid 200 μM. The semen samples encountered cryodamage and the quality was deteriorating during the cryopreservation (P < 0.05). The semen evaluation after thawing showed that the groups of samples receiving 100 μM Caffeic acid had higher viability, total motility, and lower abnormal sperm and better linearity (LIN), curvilinear velocity (VCL), straight-line velocity (VSL) and path velocity and higher intact plasma membrane (P < 0.05) compared to other groups. It is notable that adding 100 μM Caffeic acid to freezing extenders enhances the CAT, GPx, SOD, and GSH and also ameliorates total antioxidant capacity of spermatozoa after thawing. It is notable that the addition of 100 μM Caffeic acid decreases the amount of Malondialdehyde. These reactions lead us to conclude that 100 μM Caffeic acid enhances the semen quality of water buffalo after thawing.