Hand-made Cloning: A Guide for Cloning Water Buffaloes.

The present review is a detailed discussion on comparable benefits of hand-made cloning (HMC) technique than micro-manipulation based conventional cloning and developed in the author’s laboratory. Hand-made cloning technique does not require micromanipulators, because the manipulations required for both enucleation and nucleus transfer are performed by hand. The HMC technique includes manual bisection of zona-free oocytes and the simultaneous fusion of the somatic cell with two cytoplasts to produce a cloned embryo. The benefits of HMC include low setup costs for limited equipment, no requirement of highly trained expertise and in vitro effciency comparable to traditional somatic cell nuclear transfer technology. Embryos produced by HMC can be cryopreserved and capable of producing live births. The HMC technique is now applied to different species and can be used in large scale nuclear transfer programs.

Increased capacitation of buffalo sperm by heparin as confirmed by electron microscopy and in vitro fertilization.

Capacitation of buffalo sperm was evaluated by induced acrosome reaction (AR) upon the exposure of 10 mM Ca2+. Culture of sperm for 8 hr in BO medium supplemented with 10 micrograms/ml heparin significantly (P < 0.01) increased the percentage of AR and confirmed by transmission electron microscopy. Vesiculization of outer acrosomal membrane and plasma membrane was observed significantly higher (P < 0.01) following 8 hr of sperm culture with heparin. Culture of sperm with heparin also increased rate of fertilization of in vitro matured oocytes and their subsequent development up to morula/blastocyst stage (P < 0.01). The study demonstrates that capacitation of buffalo sperm by heparin required at least 8 hr exposure of sperm to heparin for maximum acrosome reaction.

In vitro maturation and fertilization of goat oocytes.

Goat oocytes were isolated from 3-5 mm diam. follicles. The oocytes with compact cumulus mass were matured and fertilized in vitro. Three different media, viz. modified Krebs-Ringer bicarbonate, Dulbecco's and Ham's F-12 with three different additives (bovine serum albumin, BSA; follicle stimulating hormone, FSH and fetal calf serum, FCS) were tested. The three basal media gave almost similar results with Ham's F-12 being slightly better. Addition of BSA (10 mg/ml) increased the rates of maturation and penetration. FSH + BSA (2.5 micrograms/ml + 10 mg/ml) further enhanced the rates while FCS (10%) proved to be even more effective. In modified Krebs-Ringer bicarbonate and Dulbecco media with additives FCS + BSA, around 60% oocytes matured to metaphase II of which 53% were penetrated by capacitated goat spermatozoa while in F-12 medium 70% reached metaphase II and 63% were penetrated. Ham's F-12 medium with additives FCS + BSA was slightly better for maturation and penetration of goat oocytes in comparison to two other media tested.

Enlarging pulmonary histoplasmoma associated with pulmonary tuberculosis.

Histoplasmosis is not a frequently reported infection from India. Similarity of its clinical and radiological features with pulmonary tuberculosis, may cause a difficult diagnostic dilemma when the two occur together. We report a case who initially presented with cavitary lesions over right lung responding well to anti tuberculosis chemotherapy but subsequently a small opacity present over the left upper zone showed a relentless increase till it attained a massive size few years later. Operative removal and histopathological examination of the mass confirmed it to be histoplasmoma. She became sputum AFB smear positive and culture negative for a short while during this period and this is explained on the basis of previous reports.

In vitro capacitation of goat spermatozoa.

Goat epididymal and ejaculated spermatozoa were incubated in Krebs-Ringer bicarbonate buffer containing pyruvate and lactate as energy source. A 3 hr incubation for epididymal and 4 hr for ejaculated spermatozoa was required for the capacitation and acrosome reaction to take place. Calcium is an essential requirement which was needed for motility maintenance/activation and for the initiation of acrosome reaction. A 2-fold increase in cAMP content was measured over 3 hr period of incubation of goat epididymal spermatozoa which was not seen when calcium ions were either omitted or chelated with EGTA. There is thus a definite involvement of Ca2+ ions and cAMP in capacitation and acrosome reaction of goat spermatozoa.

Necrobiotic pulmonary nodule leading to pyopneumothorax in a case of rheumatoid arthritis--a rare clinical presentation.

A 49-year-old male patient, an active case of sero positive rheumatoid arthritis involving multiple joints with bilateral necrobiotic pulmonary nodules is presented. Initial symptoms were of progressively increasing dyspnoea. The nodule in the right lung subsequently cavitated leading to pyopneumothorax. The diagnosis of rheumatoid lung was confirmed by post-mortem.

Osler-Rendu-Weber disease presenting as recurrent haemoptysis.

A young adult male with a short history of recurrent haemoptysis due to vascular malformation of right lung is presented. Lesion was not visible in the chest radiograph, nor on bronchography. CT-scan of thorax, however, showed a bunch of circinate opacities in the upper lobe of the right lung. Histopathological examination of the resected lung tissue confirmed the diagnosis.

Malignant mesothelioma in the young (with prosthetic aortic valve an unusual association).

A case of malignant mesothelioma (MM) in a 15 year old boy is reported. He had a prosthetic aortic valve implanted in 1980 at our centre for rheumatic aortic regurgitation. Fever, weight loss and chest pain as initial symptoms suggested possibility of prosthetic valve endocarditis. Dysphagia as another initial symptom was interesting and caused further diagnostic dilemma. Role of Computerised Tomography (CT) scan in diagnosis of MM is highlighted.