Effect of Incorporation of Additives in Tris‐Based Egg Yolk Extender on Buffalo (Bubalus bubalis) Sperm Tyrosine Phosphorylation During Cryopreservation

Phosphorylation of tyrosine residues on sperm protein is a known indicator of capacitation and a major intracellular signalling event. There is evidence that sperm cryopreservation promotes tyrosine phosphorylation and is associated with reduced fertility of spermatozoa. Under this study, cryoprotective role of different additives namely taurine, trehalose, catalase and 4‐bromophenacyl bromide on buffalo sperm quality was evaluated. Buffalo semen was cryopreserved in tris‐based egg yolk extender supplemented with additives like taurine (50 mm ) or trehalose (100 mm ) or 4‐bromophenacyl bromide (200 μm ) or catalase (100 U/ml) and used for assessment of levels of tyrosine phosphorylation in frozen‐thawed spermatozoa. The results obtained were compared with the level of protein tyrosine phosphorylation of semen cryopreserved in tris‐based egg yolk extender without additives. Proteins were extracted from a total number of nine ejaculates from three individual buffalo bulls chosen at random and analysed for tyrosine phospho‐proteins using SDS–PAGE followed by immunoblotting. Monoclonal anti‐phosphotyrosine antibody (Clone pT‐154) was used as primary antibody followed by treatment with HRP‐conjugated secondary antibody. Signals were detected on X‐ray film using chemiluminescence. Nine proteins (p20, p30, p32, p38, p49, p56, p59, p72 and p86) were found to be tyrosine phosphorylated in cryopreserved spermatozoa. Supplementation of additives significantly (p<0.05) reduced the level of protein tyrosine phosphorylation in spermatozoa. Moreover, this study showed improved (p<0.05) post‐thaw motility, viability and membrane integrity of spermatozoa on addition of these additives. The results obtained clearly indicate reduced level of capacitation like changes on supplementation of additives in terms of protein tyrosine phosphorylation.     

转基因植物疫苗研究进展

众多研究表明,病毒的抗原决定簇及细菌毒素亚单位均能在转基因植物中成功表达,并保持良好的免疫原性.与现有的疫苗生产体系相比,植物生产疫苗具有安全、经济、稳定、高效等优势.本文概述了利用不同受体植物生产疫苗的研究现状、免疫原性评价及免疫原基因的优化表达策略.     

Neuromuscular pedicle graft for restoration of arytenoid abductor function in dogs with experimentally induced laryngeal hemiplegia

Left laryngeal hemiplegia was induced by resection of the left recurrent laryngeal nerve in 12 dogs. A neuromuscular pedicle graft formed from the first cervical nerve and sternothyroideus muscle was transplanted after 1 week to the denervated cricoarytenoideus dorsalis muscle in 8 dogs. The remaining 4 dogs served as controls. Left arytenoid abduction was blindly evaluated by laryngoscopy with video photography at time 0, at 1 week, and at 19 weeks in all dogs. At 19 weeks, biopsy specimens of the left cricoarytenoideus dorsalis muscle and the neuromuscular pedicle were taken from 4 of the treatment dogs, and biopsy specimens of the left cricoarytenoideus dorsalis muscle were taken from the 4 control dogs. All biopsy specimens were blindly evaluated by histologic and histochemical examination. At 36 to 44 weeks, the remaining 4 treatment dogs, from which biopsy specimens had not been taken, were reevaluated by use of laryngoscopy with video photography. Complications and difficulties encountered during surgery included hemorrhage in the area of the cricoarytenoideus dorsalis muscle, location of a branch of the first cervical nerve that was long enough to prevent tension at the graft site, orientation of the muscle pedicle in the cricoarytenoideus dorsalis muscle without the use of an operating microscope, and preservation of the terminal portion of the first cervical nerve while forming the neuromuscular pedicle. Results of the arytenoid movement evaluations revealed improvement in arytenoid abductor function in the treatment group, compared with that in the control group at 19 weeks. Arytenoid abduction in the treatment group at this time, however, was still significantly decreased (P less than 0.05), compared with presurgical movement evaluations.(ABSTRACT TRUNCATED AT 250 WORDS)     

Traditional uses of medicinal plants for dermatological healthcare management practices by the Tharu tribal community of Uttar Pradesh, India

The aim of present study was to explore and document medicinal plants used for the traditional dermatological healthcare management practices by the the Tharu tribal community of Uttar Pradesh. The study was conducted during 2000–2004. Information was gathered from 230 informants residing in 46 villages in Terai region of Indo-Nepal boarder using questionnaires; oral interviews and group discussions. Total 92 medicinal plant species were cited for the preparation of 113 crude drug formulations. Voucher specimens of cited plant species were collected and identified as belonging to 82 genera and 49 families. Thirty-nine medicinal plant species were reported for the first time for dermatological healthcare problems from India. The dermatological healthcare problems managed were cut and wounds, ringworm, leprosy, eczema, scabies, leucoderma, boils, carbuncles, pimples, skin blemishes, spots, eruption, and burns etc. The most commonly and popularly used medicinal plant species for management of dermatological healthcare problems in the study area were Curcuma longa L., Azadirachta indica A. Juss and Melia azedarach L. It is concluded that dermatological healthcare management practice in the study area depends largely on wildly growing medicinal plant species. There is an urgent need to properly conserve the medicinal plant species growing in this area for human welfare. There is also need for further phytopharmacological studies to provide scientific explanation for the usages of 57 medicinal plant species for which to the best of our knowledge phytopharmacological literatures are not available.     

Embryonic and larval developments of brackish water catfish,Mystus gulio (Hamilton and Buchanan, 1822) induced with human chorionic gonadotropin and consequent larval rearing

Mystus gulio, the long whiskers catfish, is a popular food fish and potential candidate species for aquaculture in Sundarban area of India and Bangladesh. Recently, catch of this species has declined due to overfishing and various ecological changes. In the present study, mature fish was induced to spawn in captivity through intramuscular injection of human chorionic gonadotropin at the doses of 10 IU/g to female and 5 IU/g to male. Photomicrographs of all developmental stages of live embryo and larvae were documented with the aid of a light microscope. Results demonstrated that morula, blastula, gastrula and neurula and organogenesis ended at 1:30‐, 3:00‐, 5:30‐, 7:30‐ and 17:15‐hour post‐spawning (hps) respectively. Heartbeat and muscular contraction of embryo commenced at 8:30 and 11.15 hps respectively. Hatching of embryo started after an incubation period of 17:30 hour at an ambient temperature of 29 ± 1°C. The newly hatched larvae measured 2.17 ± 0.29 mm in total length with a yolk volume of 0.165 ± 0.03 mm³ started feeding 36 hr after hatching. The present study, on induced breeding and chronological development of M. gulio embryo, will have significant implications on conservation and seed production for aquaculture.     

Effects of biofloc under different carbon sources and protein levels on water quality,growth performance and immune responses in black tiger shrimp Penaeus monodon (Fabricius, 1978)

A 75‐day 2 × 3 factorial experiment was conducted to evaluate the effect of two levels of dietary protein (32 and 40%) and two different carbon sources (rice flour–R and molasses‐M), and without carbohydrate (control–C) in black tiger shrimp Penaeus monodon juveniles (3.37 ± 0.04 g) at 100 nos. m^?3 in 100 L fibre reinforced plastic tanks. Biofloc volume and total suspended solid were higher in molasses added groups (32 + M and 40 + M) followed by rice flour (32 + R and 40 + R) and controls (32C and 40C). Molasses and rice flour addition significantly reduced (P < 0.01) the total ammonia–N compared to controls. The highest Vibrio, Bacillus and Lactobacillus counts were recorded in 40 + M, 32 + M and 32 + R respectively. Among the treatments, significantly higher (P < 0.01) final body weight was recorded in 40 + R (8.5 ± 0.3 g), 40 + M (7.8 ± 0.3 g) and 32 + R (7.5 ± 0.4 g) compared to control groups, 32C (6.1 ± 0.3 g), 40C (6.4 ± 0.3 g) and molasses added group, 32 + M (5.7 ± 0.4 g). Rice flour supplementation significantly increased (P < 0.01) the total haemocyte count (×10⁶ cells mL^?1) in 32 + R (45.7 ± 3.7) and 40 + R (44.3 ± 3.1) compared to controls, 32C (27.3 ± 3.4) and 40C (25.8 ± 0.9). Similarly, higher superoxide dismutase, catalase, serum protein and glucose were recorded in the rice flour added groups, 40 + R followed by 32 + R. Among the treatments, the highest level of prophenoloxidase (OD 490 nm, 0.3 ± 0.0) and survival after challenge with Vibrio harveyi (55.6%) was observed in 32 + R. The study elucidates that rice flour addition produces optimum level of biofloc with better growth and immune responses compared to molasses and control. Furthermore, rice flour addition at 32% protein level could replace 40% protein feed.     

Passive immunity in tiger shrimp (Penaeus monodon) fed with monoclonal antibody to white spot syndrome virus

A study was conducted on the stability of monoclonal antibody (MAb) in the hepatopancreas and hemolymph of Penaeus monodon and its effect on protection against white spot syndrome virus (WSSV) upon challenge. MAb C-5 raised against WSSV was purified and coated onto a commercial shrimp feed at dosages of 5, 10 and 15 mg/kg feed. The feed was fed to P. monodon and stability of the MAb in hepatopancreas and hemolymph was determined by immunodot and Western blot. Immunodot results indicated the presence of MAb for 2 h post-feeding in hepatopancreas and hemolymph which was dose-dependent. MAb was also detected in hemolymph by Western blot up to 1 h post-feeding. Shrimp fed with MAb were challenged with WSSV by oral and injection methods. In shrimp fed with 15 mg antibody/kg feed (0.45 μg MAb/g shrimp/day) WSSV infection significantly delayed both in oral and injection challenges with a survival of 65 and 70 % (p < 0.05), respectively, during 15 days post-challenge. MAb was stable in shrimp for passive immunization against WSSV and could be a potential tool for prophylaxis against the virus.     

Effects of dietary protein on growth performance,nutrient utilization,digestive enzymes and physiological status of grey mullet,Mugil cephalus L. fingerlings reared in inland saline water

A 60 days feeding trial was conducted to illustrate the effect of graded levels of protein on the growth and metabolic enzymes of grey mullet (Mugil cephalus L.) fingerlings reared in inland saline water (ISW). Six isoenergetic (16 MJ/kg) and isolipidic (60 g/kg) diets containing 240, 260, 280, 300, 320 and 340 g crude protein (CP)/kg diet were formulated and fed to triplicate. Weight gain %, specific growth rate, protein utilizing efficiency, feed efficiency and RNA:DNA ratio were significantly higher (p < .05) in 320 and 300 g CP/kg diets. Fish fed with 240 g CP/kg diet showed significantly higher (p < .05) feed intake, whole‐body lipid content, hepatosomatic index value and liver glycogen content. Transaminase enzymes and malate dehydrogenase activities were elevated in fish fed 340 g CP/kg diet. Protease activity increased with increasing dietary CP level, but amylase activities showed an inverse relationship. No significant (p > .05) variations were observed for lactate dehydrogenase, oxidative stress enzymes, blood parameters and serum osmolality among all the treatment groups, but red blood cell count increases with increasing dietary CP levels. Based on the results, feeding dietary protein level of 300 g CP/kg is economically viable for rearing of grey mullet in ISW.     

Identification and pathogenicity study of emerging fish pathogens Acinetobacter junii and Acinetobacter pittii recovered from a disease outbreak in Labeo catla (Hamilton, 1822) and Hypophthalmichthys molitrix (Valenciennes, 1844) of freshwater wetland in West Bengal,India

The disease outbreaks in aquaculture system of wetlands are the major cause of fish mortality. Among various bacterial septicaemic diseases, fish mortality caused by Acinetobacter spp. is recently reported in different fish species. Fish disease outbreak was investigated in a wetland of West Bengal, India to identify the aetiological factors involved. The moribund fish were examined and subjected to bacterial isolation. Two bacterial causative agents were identified as Acinetobacter junii and Acinetobacter pittii by biochemical characterization and 16S rRNA gene amplification. Both the isolates were oxidase‐negative, nitrate‐negative, catalase‐positive and indole‐negative. The molecular identification using 16S rRNA gene sequencing and phylogenetic tree analysis further confirmed the two Acinetobacter spp. with 97%–99% similarity. The antibiotic resistance patterns of these two bacteria revealed that both of them were resistant to β‐lactam, cefalexin, cephalothin, amoxyclav, cefuroxime, cefadroxil, clindamycin, vancomycin and penicillin. In addition, A. pittii was also resistant to other antibiotics of cephams group such as ceftazidime and cefotaxime. In the challenge experiment, both A. junii and A. pittii were found to be pathogenic with LD₅₀ of 1.24 × 10⁵ and 1.88 × 10⁷ cfu/fish respectively. Histopathological examination of gill, liver and kidney revealed prominent changes supporting bacterial septicaemia. The investigation reports for the first time on concurrent infection by A. junii and multidrug‐resistant (MDR)‐A. pittii as emerging fish pathogens to cause severe mortality in Labeo catla and Hypophthalmichthys molitrix in a freshwater wetland.