ICRAF's first ten years

This article is about the institutional development of the International Council for Research in Agroforestry. It follows the Council's history from the initial ideas in the IDRC-sponsored study Trees, Food and People, its formalization in 1977, the early years up to 1980, when the Council was searching for an identity, the expansive years 1981–85, when in-house capability was built up and the foundation laid of the agroforestry discipline, to the present phase, from 1986, when major field research programmes are being initiated. The main emphasis is on the strategies and priorities behind the programmes in the different phases. A summary is given of the Council's development and growth in terms of funding and staffing. In the final section, some thoughts are expressed about ICRAF in the 1990s.Director, ICRAF; Forester     

Comparative Expression Analysis of Gametogenesis‐Associated Genes in Foetal and Adult Bubaline (Bubalus bubalis) Ovaries and Testes

This study was conducted to identify and analyse the expression of gametogenesis‐associated genes and proteins in foetal and adult buffalo gonads of both the sexes. Relative quantification of the genes was determined by qPCR and Western blotting. Immunohistochemistry was also performed for various gametogenesis‐associated proteins in foetal and adult gonads of both the sexes. We observed significantly (p < 0.05) increased expression of primordial germ cell‐specific, meiotic as well as genes associated with oocyte maturation and development in foetal ovaries as compared to the adult ones. However, significantly (p < 0.05) increased expression of proteins associated with oocyte maturation like GDF9 and ZP4 was found in adult ovaries, indicating temporal regulation of mRNA translation during oogenesis. Meiotic genes showed significantly (p < 0.05) increased expression in adult testes as compared to foetal testes and ovaries, indicating onset of meiosis at a later stage in spermatogenesis. In general, the expression of primordial germ cell‐associated as well as meiotic genes was higher in adult testes, indicating the increased biological activity in the organ. Immunohistochemistry revealed localized expression of gametogenesis‐associated proteins in ovarian follicles and seminiferous tubules of testes, while the surrounding somatic tissues were devoid of these proteins. The study gives an understanding of the sequential and temporal events of gene expression as well as mRNA translation during male and female gametogenesis. It could also be concluded that follicles and seminiferous tubules are the functional units of the female and male gonads, respectively, and their function could be enhanced by appropriate chemical and genetic intervention of the somatic tissue immediately surrounding them. This assumes importance in the context that buffalo attains sexual maturity at an older age of 2–3 years and have smaller ovaries with lesser number of primordial follicles in comparison with cattle, which is suggested to be the main reason of their poor breeding performance.     

Pigmentation, carotenoids, lipid peroxides and lipid composition of skin of red porgy (Pagrus pagrus) fed diets supplemented with different astaxanthin sources

A feeding experiment was carried out to determine the efficiency of different commercial sources, chemical forms and levels, of dietary astaxanthin, to appropriately pigment the red porgy (Pagrus pagrus) skin. According to this, total carotenoid content, profiles and chemical forms present in the skin were determined. In order to establish the potential for antioxidant protecting role of astaxanthin supplemented diets, peroxide levels and lipid composition of skin were also determined.

Red porgy alevins were fed six dietary treatments in triplicate; a basal diet (B) without carotenoids; two diets (N25 and N50) formulated to supply either 25 or 50 mg kg^− 1 of an esterified source of astaxanthin (Haematococcus pluvialis, NatuRose™); two diets (CP25 and CP50) with either 25 or 50 mg kg^− 1 of unesterified astaxanthin (Carophyll® Pink); and a positive control diet (B + S) proved as a successful pigmenting-diet in previous experiences (B + S, 88% basal diet:12% frozen shrimp) [Cejas, J., Almansa, E., Tejera, N., Jerez, S., Bolaños, A., Lorenzo, A., 2003. Effect of dietary supplementation with shrimp on skin pigmentation and lipid composition of red porgy (P. pagrus) alevins. Aquaculture 218, 457–469].

All fish fed carotenoid supplemented diets displayed a pink-coloured skin after 4 months of feeding in contrast to the greyish appearance displayed by fish fed the basal diet not supplemented with carotenoids (B). Furthermore, astaxanthin diesters were the major carotenoid in the skin of pink fish. A second carotenoid, tentatively identified as tunaxanthin diester, was also detected. The best results in terms of skin natural reddish hue, total carotenoid and astaxanthin contents were found by using the esterified forms of dietary astaxanthin (N25, N50 and B + S). Interestingly, the lowest levels of lipid peroxides were found in the fish fed these three treatments. However, no effect of treatment on lipid composition was found. In conclusion, red porgy alevins are able to efficiently utilise dietary natural or synthetic astaxanthin, and deposit this pigment in its esterified form to acquire an acceptable pink-coloured skin compared to that of the wild fish.     

Field evaluation of live chilling with CO2 on stunning Atlantic salmon (Salmo salar) and the subsequent effect on quality

Atlantic salmon were slaughtered in three ways on a commercial slaughter line: (1) killed by a percussive stun after crowding; (2) killed by percussive stun after crowding, pumping and live chilling; (3) killed by exsanguination after crowding, pumping and live chilling. The live‐chilled fish were exposed to seawater (2°C) saturated with carbon dioxide (pH 5.5–5.7) for 40 min. The fish were calm after live chilling, but not unconscious, as eye rolling was observed in all individuals. Subsequent exsanguination of the unstunned fish resulted in death. Both rapid live chilling and the subsequent exsanguination appeared stressful to the fish, as a large and rapid pH drop coupled with earlier onset of rigor mortis, indicative of high muscle activity during the process were observed. The muscle core temperature during ice storage showed that live chilling only has an effect on carcass temperature during the first 6 h post mortem. After 6 h, no significant differences in temperature were detected between live‐chilled and traditionally ice‐chilled fish. We conclude that commercial use of live chilling in combination with high levels of CO₂ does not stun Atlantic salmon. Live chilling followed by exsanguination of the unstunned fish appears to be highly stressful and should be avoided.     

The effect of slaughtering procedures on blood spotting in rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar)

Rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar) were submitted for various slaughter and bleeding procedures to see what effect this would have on blood drainage of the muscles. Results show that the bleeding method is of less importance, while it is the timing that is important. No significant difference in bloodspotting was observed between fish that were bled live by a gill cut or percussive killed and bled by gutting. Most of the drainage of blood in the fish muscle seems to occur within the first hours postmortem, so rigor mortis is of little importance. The visual appearance of the fillet was influenced by number and size of the bloodstains. Colour measurements with Hunter L*, a*, b* did not reveal this. We conclude that a gill cut is not necessarily to obtain bleeding, so the industry can omit this phase and go directly to gutting.     

Efficacy of piperazine dihydrochloride against Ascaris suum and Oesophagostomum species in naturally infected pigs

A controlled trial was performed to evaluate the efficacy of piperazine dihydrochloride in a new granular formulation (Ascarex D) against naturally occurring infections with Ascaris suum, Oesophagostomum dentatum and O quadrispinulatum. Treatment effects were estimated on the basis of parasites recoverable from the intestinal contents. Given orally at 200 mg per kg body weight the compound showed an efficacy of 99 to 100 per cent against A suum and the nodular worms. Egg excretion of the respective species was reduced by 98 per cent and 100 per cent six days after treatment. No adverse reactions were observed after the treatment.     

Oxygen Concentration and Cysteamine Supplementation During In vitro Production of Buffalo (Bubalus bubalis) Embryos Affect mRNA Expression of BCL‐2, BCL‐XL,MCL‐1, BAX and BID

This study examined the effects of O₂ concentration (5% vs 20%) during in vitro maturation (IVM), fertilization (IVF) and culture (IVC) or supplementation of IVM and IVC media with cysteamine (50 and 100 μm , respectively; IVM, IVF and IVC carried out in 20% O₂), on blastocyst rate and relative mRNA abundance of some apoptosis‐related genes measured by real‐time qPCR in immature and in vitro‐matured buffalo oocytes and in embryos at 2‐, 4‐, 8‐ to 16‐cell, morula and blastocyst stages. The blastocyst rate was significantly higher (p < 0.05) while the percentage of TUNEL‐positive cells was significantly lower (p < 0.05) under 5% O₂ than that under 20% O₂. The mRNA expression of anti‐apoptotic genes BCL‐2 and MCL‐1 was significantly higher (p < 0.05) and that of pro‐apoptotic genes BAX and BID was lower (p < 0.05) under 5% O₂ than that under 20% O₂ concentration at many embryonic stages. Following cysteamine supplementation, the blastocyst rate and the relative mRNA abundance of BCL‐XL and MCL‐1 was significantly higher (p < 0.05) and that of BAX but not BID was lower (p < 0.05) at many stages of embryonic development, although it did not affect the percentage of TUNEL positive cells in the blastocysts significantly. The mRNA expression pattern of these genes during embryonic development was different in 5% vs 20% O₂ groups and in cysteamine supplemented vs controls. At the 8‐ to 16‐cell stage, where developmental block occurs in buffalo, the relative mRNA abundance of BCL‐2 and MCL‐1 was highest under 5% O₂ concentration and that of BAX and BID was highest (p < 0.05) under 20% O₂ concentration. These results suggest that one of the mechanisms through which beneficial effects of low O₂ concentration and cysteamine supplementation are mediated during in vitro embryo production is through an increase in the expression of anti‐apoptotic and a decrease in the expression of pro‐apoptotic genes.     

Ecological and landscape effects on genetic distance in an assemblage of headwater fishes

Environmental divergence along hierarchically structured longitudinal gradients may constitute barriers to gene flow in river networks for headwater specialised species. While known, this phenomenon has not been well studied, especially with regard to degree of headwater specialisation. We examined six headwater species that differ in habitat specialisation to assess whether patterns of differentiation vary according to geographic or environmental distance. We also identified regional environmental or anthropogenically induced fragmentation effects by comparing within‐drainage patterns of genetic distance across replicate watersheds. We used a comparative modelling framework to determine whether isolation by distance or isolation by resistance of large river habitats was a better predictor of genetic distance across species. The influence of reservoir presence and regional network characteristics that may influence the hydrology and size of large river habitats were also assessed. Resistance effects from large rivers were closely related to headwater specialisation, with increased specialisation leading to increased resistance and loss of drainagewide population connectivity. These results affirm that dendritic networks naturally fragment headwater specialised species. Further isolation from anthropogenic fragmentation was detected in two of the six drainages, indicating interactions with system‐specific conditions. Landscape variables related to the hydrology of large rivers also affected genetic distance in predicted ways, supporting the importance of large rivers in genetically structuring headwater species in drainage networks.     

Expression and localization of angiopoietin family in corpus luteum during different stages of oestrous cycle and modulatory role of angiopoietins on steroidogenesis,angiogenesis and survivability of cultured buffalo luteal cells

The objective of this study was to document the expression and localization of angiopoietin (ANGPT) family members comprising of angiopoietin (ANGPT1 and ANGPT2), and their receptors (Tie1 and Tie2) in buffalo corpus luteum (CL) obtained from different stages of the oestrous cycle, and the modulatory role of ANGPT1 and ANGPT2 alone or in combinations on progesterone (P₄) secretion and mRNA expression of phosphotidylinositide‐3kinase‐protein kinase B (PI3K‐AKT), phosphoinositide‐dependent kinase (PDK), protein kinase B (AKT), Bcl2 associated death promoter (BAD), caspase 3 and von willebrand factor (vWF) in luteal cells obtained from midluteal phase (MLP) of oestrous cycle in buffalo. Real‐time RT‐PCR (qPCR), Western blot and immunohistochemistry were applied to investigate mRNA expression, protein expression and localization of examined factors whereas, the P₄ secretion was assessed by RIA. The mRNA and protein expression of ANGPT1 and Tie2 was maximum (p < .05) in mid luteal phase (MLP) of oestrous cycle. The ANGPT2 mRNA and protein expression was maximum (p < .05) in early luteal phase, decreased in MLP and again increased in late luteal phase of oestrous cycle. ANGPT family members were localized in luteal cells and endothelial cells with a stage specific immunoreactivity. P₄ secretion was highest (p < .05) with 100 ng/ml at 72 hr when luteal cells were treated with either protein alone. The mRNA expression of PDK, AKT and vWF was highest (p < .05) and BAD along with caspase 3 were lowest (p < .05) at 100 ng/ml at 72 hr of incubation period, when cultured luteal cells were treated with either protein alone or in combination. To conclude, our study explores the steroidogenic potential of angiopoietins to promote P₄ secretion, luteal cell survival and angiogenesis through an autocrine and paracrine actions in buffalo CL.