A serine proteinase inhibitor from frog eggs with bacteriostatic activity

By Sephadex G-50 gel filtration, Resource Q anionic exchange and C4 reversed phase liquid high performance liquid chromatography, a proteinase inhibitor protein (Ranaserpin) was identified and purified from the eggs of the odour frog, Rana grahami. The protein displayed a single band adjacent to the molecular weight marker of 14.4 kDa analyzed by SDS-PAGE. The inhibitor protein homogeneity and its molecular weight were confirmed again by MALDI-TOF mass spectrometry analysis. The MALDI-TOF mass spectrum analysis gave this inhibitor protein an m/z of 14422.26 that was matched well with the result from SDS-PAGE. This protein is a serine proteinase inhibitor targeting multiple proteinases including trypsin, elastase, and subtilisin. Ranaserpin inhibited the proteolytic activities of trypsin, elastase, and subtilisin. It has an inhibitory constant (K-i) of 6.2 x 10(-8) M, 2.7 x 10(-7) M and 2.2 x 10(-8) M for trypsin, elastase, and subtilisin, respectively. This serine proteinase inhibitor exhibited bacteriostatic effect on Gram-positive bacteria Bacillus subtilis (ATCC 6633). It was suggested that ranaserpin might act as a defensive role in resistance to invasion of pests or pathogens. This is the first report of serine proteinase inhibitor and its direct defensive role from amphibian eggs. (C) 2007 Elsevier Inc. All rights reserved.

Melanoma cell growth inhibition by beta gamma-CAT, which is a novel non-lens betagamma-crystallin and trefoil factor complex from frog Bombina maxima skin

In vertebrates, non-lens beta gamma-crystallins are widely expressed in various tissues but their functions are unknown. The molecular mechanisms of trefoil factors, initiators of mucosal healing and being greatly involved in tumorigenesis, have remained

Acute toxicity of beta gamma-CAT, a naturally existing non-lens beta gamma-crystallin and trefoil factor complex from frog Bombina maxima skin secretions

In vertebrates, non-lens beta gamma-crystallins are widely expressed in various tissues, but their functions are unknown. The molecular mechanisms of trefoil factors, initiators of mucosal healing and being greatly involved in tumorigenesis, have remained

Frog skins keep redox homeostasis by antioxidant peptides with rapid radical scavenging ability

The question of how amphibians can protect themselves from reactive oxygen species when exposed to the sun in an oxygen-rich atmosphere is important and interesting, not only from an evolutionary viewpoint, but also as a primer for researchers interested in mammalian skin biology, in which such peptide systems for antioxidant defense are not well studied. The identification of an antioxidant peptide named antioxidin-RL from frog (Odorrana livida) skin in this report supports the idea that a peptide antioxidant system may be a widespread antioxidant strategy among amphibian skins. Its ability to eliminate most of the 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) radical tested within 2 s, which is much faster than the commercial antioxidant factor butylated hydroxytoluene, suggests that it has a potentially large impact on redox homeostasis in amphibian skins. Cys10 is proven to be responsible for its rapid radical scavenging function and tyrosines take part in the binding of antioxidin-RL to radicals according to our nuclear magnetic resonance assay. (C) 2010 Elsevier Inc. All rights reserved.

Peptidomics and genomics analysis of novel antimicrobial peptides from the frog, Rana nigrovittata

Much attention has been paid on amphibian peptides for their wide-ranging pharmacological properties, clinical potential, and gene-encoded origin. More than 300 antimicrobial peptides (AMPs) from amphibians have been studied. Peptidomics and genomics analysis combined with functional test including microorganism killing, histamine-releasing, and mast cell degranulation was used to investigate antimicrobial peptide diversity. Thirty-four novel AMPs from skin secretions of Rana nigrovittata were identified in current work, and they belong to 9 families, including 6 novel families. Other three families are classified into rugosin, gaegurin, and temporin family of amphibian AMP, respectively. These AMPs share highly conserved preproregions including signal peptides and spacer acidic peptides, while greatly diversified on mature peptides structures. In this work, peptidomics combined with genomics analysis was confirmed to be an effective way to identify amphibian AMPs, especially novel families. Some AMPs reported here will provide leading molecules for designing novel antimicrobial agents. (C) 2009 Elsevier Inc. All rights reserved

Five novel antimicrobial peptides from skin secretions of the frog, Amolops loloensis

While investigating antimicrobial peptide diversity of Amolops loloensis, five novel antimicrobial peptides belonging to two families were identified from skin secretions of this frog. The first family including two members is esculentin-2-AL (esculentin-2-ALa and -ALb): the second family including three members is temporin-AL (temporin-ALd to -ALf). The family of esculentin-2-AL is composed of 37 amino acid residues (aa); the family of temporin-AL is composed of 16, 13 and 10 aa, respectively. All of these antimicrobial peptides showed antimicrobial activities against tested microorganisms. cDNAs encoding precursors of esculentin-2-ALs and temporin-ALs were cloned from the skin cDNA library of A. loloensis. All the precursors share similar overall structures. There is a typical prohormone processing signal (Lys-Arg) located between the acidic propiece and the mature peptide. The antimicrobial peptide family of esculentin-2 is firstly reported in the genus of Amolops. Combined with previous reports, a total of four antimicrobial peptide families have been identified from the genus of Amolops; three of them are also found in the genus of Rana. These results suggest the possible evolutionary connection between the genera Amolops and Rana. (C) 2009 Elsevier Inc. All rights reserved.

Feasibility of bi-culture of mud crab (Scylla serrata) and shrimp (Penaeus monodon)

A study on the feasibility of bi-culture of mud crab (Scylla serrata) and shrimp (Penaeus monodon) in brackishwater earthen ponds (0.1 ha each) was carried out for a period of five months (March-August). Nursed shrimp juvenile (ABL:· 3.36±0.23 em and ABW: 0.26±0.04 g) and crab juvenile (ACL: 2.61±0.22 cm, ACW: 4.63±0.11 cm and ABW: 43±2.64 g) were stocked following the experimental design of shrimp 2/m2 (Treatment-1), shrimp 2/m2 and mud crab l/m2 (Treatment-2) and shrimp 2/m2 and mud crab 0.5/m2 (Treatment-3). Crabs were fed with chopped trash tilapia @ 10~5%, while shrimp were fed with Saudi-Bangla shrimp feed @ 3~5% of biomass twice daily. Significantly (p<0.05) higher specific growth rate (SGR) of shrimp and mud crab was 1.86% (g/day) in T2 and 0.83% (g/day) in T3, respectively. The survival of shrimp and mud crab also varied significantly (p<0.05) with a higher mean value of74.63% in Tl and 51.04% in T3, respectively. The production of shrimp (424.09 kg/ha) was significantly (p<0.05) higher in Tl and that of mud crab (568.80 kg/ha) in T2. Significantly (p<0.05) highest total production of 871.29 kg!ha was in T2 followed by 708.52 kg/ha in T3 and 424.09 kg/ha in Tl. The results indicate that mud crab can be cultured at a stocking rate of 1/m2 together with shrimp at 2/m2 •

Seed production and culture techniques of Genetically Improved Farmed Tilapia (GIFT) in brackishwater environment

Effects of different levels of salinity on survival, growth and gonadal development of Genetically Improved Farmed Tilapia (GIFT) were studied under laboratory conditions in glass aquarium, for a period of ten weeks. The initial individual size of the GIFT was 20.23±4.45 and the salinity levels tested were 0, 5, 10, 15 and 20 ppt. The highest survival of 87.5% was found in 0 ppt and the lowest 60.5% in 20 ppt. Though the survival decreased progressively with increased salinity, there were no significant differences (P>0.05) among 0, 5, and 10 ppt. Similar to what has been observed in survival, the specific growth rate (SGR %/day) also decreased as of 1.30, 1.24, 1.08, 0.90 and 0.71, respectively, with the increased salinity of 0, 5, 10, 15 and 20 ppt. The gonadal development was highest in 0 ppt with a GSI value of 3.75 and lowest of 2.01 in 20 ppt. In the second experiment, gonadal development and seed production performance of GIFT in brackishwater condition were investigated for a period of three months. Each of the three fine meshed hapas of 20 square meters made from nylon net was placed in a freshwater (0 ppt) and in a brackish water (10-15 ppt) pond of the Brackishwater Station (BS). GIFT of 65 g average weight from a single cohort were stocked into three hapas at a rate of 2 per m. The male vs female ratio was 1:3. The development of gonad was faster with the higher gonadosomatic index (GSI %) of 3.85 % in freshwater condition than that of 2.73 % in brackish water. Within three months of the study period, a total of 70,510 and 44,250 GIFT fry were produced respectively, in freshwater and brackishwater conditions. Finally under third experiment, a participatory on-farm trial was carried out to evaluate the production performance of GIFT in monoculture and in polyculture with silver barb in coastal freshwater pond conditions. Nine ponds were selected for three treatment combinations of GIFT monoculture (T1), GIFT and silver barb polyculture (T2), and silver barb monoculture (T3). The ponds have been stocked in April, 05 at a density of 25,000 fry per ha. Fishes were fed with rice bran at the rate of 6% bw per day. In one month culture period, GIFT attained an average weight of 16.27 g in monoculture and 17.23 g in polyculture, against an average stocking weight of 0.37 g. Silver barb reached an average weight of 16.62 g in polyculture with GIFT and 10.01 g in monoculture, against an average stocking weight of 3.79 g.

Economic analysis of carp based culture technologies in selected areas of Bangladesh

Six carp based culture technologies such as, carp-pangas, carp polyculture, carp-golda, pangas monoculture, golda monoculture and nursery have been selected to determine the cost and returns of respective technologies in Bangladesh. The sample farmers selected for these technologies were 55, 100, 65, 50, 51 and 55 respectively and thus the total sample size stood at 376. The study covered 7 districts of Bangladesh, namely, Mymensingh, Bogra, Noakhali, Comilla, Jessore, Khulna and Bagerhat. Both primary and secondary data were used for this study. It was found that farmers used a good number of feeds for the selected technologies and they maintained no standard doses for them. Remarkable differences were found among the prices of different feeds and other inputs used for different technologies in different locations. Prices of all inputs were found to be increasing and this increase was more in recent years compared to previous years. Though all the technologies were found to be profitable, the feed situation was not satisfactory. Except rice polish all the local feeds showed deficit in supply to meet the national demand for the country. If this situation persists and no proper measures are taken to secure the local feed supply, the present development of supplementary feed-based aquaculture would be fully dependent on imported feeds and would not be sustainable in future. This study strongly suggests the corresponding authority to handle the matter with proper attention considering its significant livelihood impact on the economy of the country.

Culture practices of freshwater giant prawn in some selected areas of Mymensingh

A study was conducted to examine the culture practices of freshwater giant prawn, Macrobrachium rosenbergii in some selected areas of Mymensingh. Based on a sample of 100 farmers from three different upazila, namely Phulpur, Gouripur and Ishwargonj in Mymensingh district, 94% of farmers cultured prawn with fish in their pond. Only 6% of farmers cultured prawn, fish and dike crops for higher economic return. The culture period is typically nine months; hatchery produced post-larvae were stocked from May to June and harvested from November to January. Per hectare production of prawn, 375 kg/yr was very low because the farmers followed simple culture method. Most of the farmers made a profit of Tk. 68,403/ha/yr and the major costs incurred were for purchasing prawn seed and feed. The culture of prawn in pond system is technically possible under different conditions though expansion of small-scale prawn farming mainly depends on reduction of production costs. Future targets could be integration of pond prawn culture with other agricultural activities especially dike cropping and rice production in the monsoon.

Shifting rice farming to fish culture in some selected areas of Mymensingh, Bangladesh: the process, conflicts and impacts

The evolutionary process of converting low-lying paddy fields into fish farms and its impact on agrarian communities in some selected areas of Mymensingh district were studied. This study was conducted through participatory rural appraisal (PRA) covering 12 villages from each of selected upazillas viz. Fulpur and Haluaghat of Mymensing [sic] district. A total of 12 PRA sessions were conducted where 90 farmers participated during 29 July to 26 August 2004. It is seen that the use of low-lying paddy fields was mostly confined to Broadcast Aman (B. Aman) rice production until 1960s. With the introduction of modern rice farming technology, the farmers started to produce Boro rice in Rabi season and B. Aman rice in Kharif season. With the passage of time, aquaculture technologies have been evolved and the farmers realized that fish farming is more profitable than rice cultivation, and then they started to utilize their paddy fields for alternate rice-fish farming and rice-cum-fish farming. Now a days, aquaculture based crop production system is in practice in more than 25% of the low-lying paddy fields. Conversion of rice fields in to fish ponds has brought up a change in the livelihood patterns of the rural farmers. The areas where the farmers involved themselves in the new production systems were fingerling collection, transportation and marketing of fry and fingerlings. During 1960s to 1970s, a few people used to culture fish in the permanent ponds for their own consumption, the species produced were rohu, catla, mrigal, ghainna, long whiskered catfish, freshwater shark (boal), snake head (shol) etc. Small fishes like climbing perch, stinging catfish, walking catfish, barb, minnows etc. were available in the rice fields during monsoon season. In 1980s to mid 1990s, some rice fields were converted into fish ponds and the people started to produce fish for commercial purposes. When rice-fish farming became profitable, a large number of people started converting their rice fields in to rice-fish culture ponds. Culture of some exotic fishes like silver carp, tilapia, grass carp, silver barb etc. also started in the paddy fields. Higher income from fish farming contributed positively in improving the housing, sanitation and education system in the study areas. It is seen that the medium and medium high lands were only used for alternate rice fish farming. The net income was high in any fish based cropping system that motivated the farmers to introduce fish based cropping system in the low-lying inland areas. As a result, the regional as well as communal income disparities occurred. However, the extraction of ground water became common during the dry period as the water was used for both rice and fish farming. Mass conversion of paddy fields into rice-fish culture ponds caused water logging in the study areas. In most cases, the participated farmers mentioned that they could be easily benefited by producing fish with T. Aman or only fish during the monsoon season. They agreed that this was an impressive technology to them and they could generate employment opportunities throughout the year. Finally, the social, economic and technical problems which are acting as constraints to rapid expansion of fish production system were reported from the interviewee.

Adoption, productivity and profitability of rice-fish and fingerling culture in three selected districts of Bangladesh

The study aimed to investigate the productivity and profitability of rice-fish farming. It also determined the factors affecting rice-fish and fingerling production in Bangladesh. Among the total respondents about 55% were involved in rice-fish farming, 37% in pond fish culture and rest 9% in fish nursery. Integrated rice-fish farming contributed about 83% of the annual farm income in Comilla while it was about 20% in both Kishoregonj and Chandpur districts. Among the different farming systems integrated rice-fish culture and rearing fingerlings in the nursery pond were more profitable at the farm level. Farmers earned a net return of Tk. 49,714/ha from integrated fish culture with boro rice while the net return of boro rice-alone was only Tk. 11,179/ha. This indicated that farmer could earn 218% higher net return from integrated rice-fish farming than that of boro rice only farming. Moreover, integrated rice-fish culture also reduced pesticides cost to about 77% and weeding cost to about 51% in boro season. Fingerling nursing earned highest net returns among the three technologies.

Seed production and culture techniques of climbing perch, Anabas testudineus

Six treatments each with 12 replications designed to optimize the dose of inducing agent PG to achieve fertilization and hatching success of climbing perch, Anabas testudineus were tested. The females were given single injection of 7-12 mg PG/kg body weight and the males were given 4 mg PG/kg body weight. Fertilization and hatching rate varied from 67±4.55% to 66±3.0% and 59±4.88% to 57±6.21% for the doses of 10, 11 and 12 mg PG/kg of body weight, respectively. The hormone dose had significant (P<0.05) effect on fertilization and hatching. Six mini shallow cisterns (570 cm x 105 cm) were used to investigate the efficacy of zooplankton and Artemia nauplii as feed for spawn rearing. Three-day old spawns were stocked in six mini shallow cisterns at a stocking density of 100 individuals/L of water. Two treatments each with three replications were used to develop culture technique of the climbing perch. In case of treatment-1, the spawns were fed with Artemia nauplii three times daily, while in treatment-2, zooplankton were used as feed in the same manner as in treatment-1. After 14 days of rearing, mean final weight of the fry of treatments-1 and 2 were 95.55±6.71 and 57.69±5.40 mg, respectively. In treatment-1, spawn fed with Artemia nauplii showed significantly (P<0.05) higher mean weight than the spawn fed with zooplankton (treatment 2).

On farm trial of freshwater pearl culture

On farm preliminary trial of freshwater pearl culture was done through 20 entrepreneurs in Boilor and Sutiakhali villages of Mymensingh district during 2004. A group of 20 enthusiastic women were selected and trained on the art of mantle tissue dissection, operation for mantle tissue implantation and preparation of ponds for pearl culture. A total of 200 juvenile freshwater mussel, Lamellidens marginalis, were collected from the wild and were used for mantle issue operation. The operated mussels were then transferred to farmer's pond and were subjected to observational trial. Length and weight of each of the test mussels were recorded before hanging them at a depth of 40 cm in net bags (3 mussels/net bag) in ponds at the rate of 24,700 mussels/ha of pond area. Ponds were routinely fertilized with organic and inorganic fertilizers thorough out the mussel rearing period. Water temperature, pH, plankton density and soil organic matter were monitored fortnightly. Growth of pearl is yet to be monitored through sacrifice of the mussels but X-ray photography of a few mussels indicated the initiation of pearl formation in most of them.

Utilization of frog waste

Commercial frog waste samples have been converted into meals by cooking at 0.7 kg/sq. cm for 30 min, draining off the stick water and drying the press cake either in the sun, tunnel dryer under controlled conditions or hot air oven. Yield of the meal varied between 18.6 to 21.5% of the fresh frog waste. Chemical analyses of the meals have shown that the meals prepared from frog waste conform to standards prescribed for fish meal and livestock feed and can therefore be used for supplementation of poultry/animal feed.