Optimal stocking density for culturing tropical soil-dwelling earthworm, pontoscolex corethrurus

The present study was carried out to determine the optimal stocking density for culturing tropical soil dwelling earthworm, Pontoscolex corethrurus. F1 generation earthworms were cultured in four different stocking densities of 1, 4, 7 and 10 worms per vessel, corresponding to field densities of 50, 200, 350 and 500 individuals per m2. Earthworms were kept under laboratory conditions (25±2°C and 25% moisture) for the 14 weeks study period. The results showed that at higher earthworm densities (>350 individuals per m2), the earthworm growth was slower and sexual maturation was delayed as compared with their counterparts in lower stocking density. With the high survival rate and parthenogenetic reproduction mode, P. corethrurus could potentially be used as tropical soil rehabilitation agent

Bacillus thuringiensis entomotoxicity activity in wastewater sludge-culture medium towards Bactrocera dorsalis and their histopathological assessment

This study investigates the production of biopesticide based on Bacillus thuringiensis activity in culture media supplemented with semi-solid wastewater sludge as one of the raw ingredient. A series of testing using mixture of sludge and source of protein as B. thuringiensis growth media were carried out and selection of media was based on viable spore count. The entomotoxicity test of B. thuringiensis was carried out against larvae of fruit fly using diet incorporation method. Further impact of entomotoxicity was observed based on histology deformities on columnar epithelial cell and goblet cell of the midgut. A mixture of sludge with 60% wheat bran produced up to 1.64 × 1010 CFU/mL of viable spore count within 10 days of incubation. Based on entomotoxicity test, incorporation of 12 mL of semi-solid wastewater sludge-culture media into fruit fly artificial diet caused the highest fruit fly mortality at 64.8%. The value of semi-solid wastewater sludge-culture media concentration for LC50 was determined at 8.43%. Effect of entomotoxicity can be seen started from 3rd instar larvae where histopathological studies showed that up to 10% of columnar epithelial cells in the intestine were swollen and severe reduction of goblet cell’s size. Thus, it decreases the survivality of the fruit fly larvae. The present study indicated that semi-solid wastewater sludge has the potential to enhance B. thuringiensis entomotoxicity activity

Bacillus thuringiensis entomotoxicity activity in wastewater sludge-culture medium towards Bactrocera dorsalis and their histopathological assessment

This study investigates the production of biopesticide based on Bacillus thuringiensis activity in culture media supplemented with semi-solid wastewater sludge as one of the raw ingredient. A series of testing using mixture of sludge and source of protein as B. thuringiensis growth media were carried out and selection of media was based on viable spore count. The entomotoxicity test of B. thuringiensis was carried out against larvae of fruit fly using diet incorporation method. Further impact of entomotoxicity was observed based on histology deformities on columnar epithelial cell and goblet cell of the midgut. A mixture of sludge with 60% wheat bran produced up to 1.64 × 10 10CFU/mL of viable spore count within 10 days of incubation. Based on entomotoxicity test, incorporation of 12 mL of semi-solid wastewater sludge-culture media into fruit fly artificial diet caused the highest fruit fly mortality at 64.8%. The value of semi-solid wastewater sludge-culture media concentration for LC50 was determined at 8.43%. Effect of entomotoxicity can be seen started from 3rd instar larvae where histopathological studies showed that up to 10% of columnar epithelial cells in the intestine were swollen and severe reduction of goblet cell’s size. Thus, it decreases the survivality of the fruit fly larvae. The present study indicated that semi-solid wastewater sludge has the potential to enhance B. thuringiensis entomotoxicity activity

Optimal Stocking Density for Culturing Tropical Soil-dwelling Earthworm, Pontoscolex corethrurus (Kepadatan Optimum untuk Pengkulturan Cacing Tanah Tropika, Pontoscolex corethrurus)

ABSTRACT The present study was carried out to determine the optimal stocking density for culturing tropica

Variation in mycorrhizal specificity for in vitro symbiotic seed germination of grammatophyllum speciosumBlume

Grammatophyllum seeds are minute and lack endosperm. As with their other orchids counterpart, the seeds are dependent on mycorrhizal fungi for seed germination in nature. The ability to uptake nutrients from substrate is assisted by preferable fungal symbionts. Seeds of Grammatophyllum speciosum Blume. were used to determine the specificity of its fungus relationship using fungi isolated from roots of G. speciosum, G. stapeliiflorum and G. scriptum. A total of 31 different species of fungus was isolated and inoculated onto G. speciosum seed on Oat Meal Agar (OMA). The result obtained from the test demonstrated that seed germination rates were best when co-cultured with Fusarium sp. number 3 isolated from G. speciosum. An increment in 63.3% was measured when compared to the seed’s original size. The seed can also germinate when inoculated with fungus isolated from different species, implying that G. speciosum is a generalist in its association with fungal symbionts

Production of Bioactive Compounds by Bacillus subtilis against Sclerotium rolfsii

This study aims to investigate the characteristic of bioactive compound produced by Bacillus subtilis against Sclerotium rolfsii and the influence of additive supplements on the antagonistic activity of B. subtilis. The fact that B. subtilis produced an antifungal substance which has inhibitory effect on wide range of fungi, including S. rolfsii, is well known. To learn the effect of pH, temperature and light condition on the production of antifungal compound, B. subtilis was inoculated in Potato Dextrose Broth at various initial pH, temperatures and light conditions, respectively. This antagonist was found to produce antifungal compound that stable at 80C with 58.3 % inhibition on S. rolfsii. The activity was constant within a wide range of pH (3–11). However, treatment with pH11 lead to higher antifungal activity (31.57 % inhibition) and it was also found to produce substance that can endure dark condition (46.24 % inhibition) with fungicidal effect on S. rolfsii. A series of experiments also been carried out to enhance the antifungal production by supplementing different carbon source preparation into bacterial liquid culture. B. subtilis were grown in minimal medium containing 1 % of oil palm root, Ganoderma lucidum or chitin, respectively prior to bioassay. Crude culture from oil palm root supplemented culture shown significantly reduction in S. rolfsii growth compared to other carbon source crude culture or the antagonism alone, suggesting that this approach may provide improved biocontrol efficiency

The Evidence of Non n-glycan Linked Mannose in Exochitinase 42kDa, from Trichoderma harzianum BIO10671 Glycosylation

Chitinase 42 kDa produced by Trichoderma harzianum has been proven as a prime compound to be excreted onto the hyphae of the pathogen causing localised cell wall lysis at the point of interaction. Later it will initiate the process of the host cell becomes empty of cytoplasm, disintegrates and shows a rapid collapse. This study investigates the existence of N-glycan linked mannose in chitinase 42 kDa produced by the Malaysian T. harzianum strain BIO10671. The chitinase 42 kDa from T. harzianum BIO10671 was initially purified using anion exchange chromatography prior to a series of experiments such as immunoblotting against the chitinase 42 kDa antibody, lectin staining for detecting any terminal linked mannose, and galactofuranose detection to determine the presence of galatofuranase components in glycoproteins. The enzyme purification harvested about 12-fold of chitinase 42 kDa from T. harzianum BIO10671 with strong indication of the chitinase 42 kDa presence on SDS-Page. This was confirmed by immunoblotting with a strong response around 42 kDa after overnight incubation in chitinase 42 kDa antibody suggesting that the gene for chitinase 42 kDa was greatly expressed in this strain. There are no intervation of galatofuranose on any of the terminal mannose in chitinase 42 kDa as shown by negative results on samples treated with or without endoglycosidase-H and lectin staining. Therefore, it can be concludeed that glycosylation occurred in the chitinase 42 kDa from T. harzianum 42 kDa was not in the form of N-glycan linked mannose as expected

First report of Bipolaris cactivora causing brown leaf spot in rice in Malaysia

In March 2013 to February 2017, brown spot symptoms on the leaves of rice plants were observed during a survey conducted in 23 granary fields throughout Peninsular Malaysia in Kedah, Penang, Perak, and Selangor states. In Malaysia, leaf spot symptoms can be seen on the rice seedling up to mature rice plant. Symptoms of the leaf spot include brown necrotic spots, which are irregular and measure between 2 and 10 mm wide. Small pieces (5 mm) of necrotic tissues from the spots were surface sterilized for 1 min in 0.5% NaOCl, washed twice with sterile distilled water, and plated onto peptone pentachloronitrobenzene agar. Plates were incubated at 28°C under 12-h light/dark for 7 days. The fungal colonies were purified using the hyphal-tip technique. A total of 152 fungal isolates were obtained, and the conidial morphology matched that of Curvularia and Bipolaris species, members of the Pleosporaceae family (Brecht et al. 2007; Garibaldi et al. 2014). Primers ITS1/ITS4 and GPDH-F/R were used to amplify and subsequently sequence the internal transcribed spacer (ITS1-5.8S-ITS2) region (White et al. 1990) and glycerol-3-phosphate dehydrogenase (gpd) gene, respectively. The gpd primer (GPDH-F, GTATCGTCTTCCGCAATG; GPDH-R, GAGGCGTTGGAGAGCAC) was designed using the genome of Curvularia victoriae FI3 that was obtained from the JGI website (https://genome.jgi.doe.gov/Cocvi1/Cocvi1.home.html). Bipolaris cactivora was successfully identified based on morphological characters and gene sequencing. Other species including B. sorghicola, B. papendorfii, B. sorokiniana, Curvularia aeria, C. affinis, C. eragrostidis, C. geniculata, C. hawaiiensis, and C. lunata were also successfully isolated and identified. In general, B. cactivora produced dark olive to black mycelia, floccose with black pigmentation on potato dextrose agar (PDA), and formed four to seven septate conidia, mostly straight and solitary with average size 56.9 × 13.4 µm. A phylogenetic tree was constructed using a neighbor-joining method, showing isolates B. cactivora A1022, K1282, K1283, and K1284 were grouped into the same clade of B. cactivora (ITS/gpd, HM598679/HM598682, Tarnowski et al. 2010). To examine virulence of B. cactivora, pathogenicity tests were performed three times by inoculating 5-week-old rice plants (variety MR211) with isolate B. cactivora K1283. Ten plants were inoculated by placing a conidial suspension (4 × 104 conidia/ml) from 8-day-old culture colonies, and 10 control plants were inoculated with distilled water. After 10 days, all inoculated leaves showed leaf spot at the inoculation points. Average length of necrosis formed by B. cactivora K1283 was 3.0 mm. B. cactivora K1283 was successfully reisolated from the necrotic tissues observed in symptomatic plants. No symptoms were observed in the control plants, and B. cactivora was not isolated from the controls. B. cactivora has been reported in Florida (U.S.A.) and Italy, where it occurs on bermudagrass and monstrose apple cactus, respectively (Brecht et al. 2007; Garibaldi et al. 2014). This study shows B. cactivora to be highly virulent on rice leaves, and to our knowledge this is the first report of B. cactivora causing leaf spot disease on rice

Comparison of plant nutrient contents in vermicompost from selected plant residues

In this experiment, earthworm, Eudrilus eugeniae was fed with different plant residues: grass clippings, sago waste and rice straw. These organic wastes were also left to decompose naturally as the control. Analysis on samples vermicompost showed that humic acid content was highest in rice straw, followed by grass clippings compared to those of sago wastes. Total phosphorus (P) was lower in vermicomposts as compared to the controls, however, highest extractable P was found in rice straw vermicompost (0.33 ± 0.00 g/kg). Sago waste vermicompost (22.79 ± 0.01 g/kg) contained highest total calcuim (Ca) compared to the grass clippings (1.39 ± 0.01 g/kg) and rice straw (8.22 ± 0.06 g/kg) vermicompost. Significant positive correlations between nutrient contents in raw plant residues and vermicompost were observed for total nitrogen (N) (r = 0.779), potassium (K) (r = 0.998) and Ca (r = 0.997). The study showed that nutrient in initial wastes material affects the nutrient contents of vermicompost. This research suggested that among the plant residues studied, vermicomposting of rice straw produced vermicompost with the highest plant extractable nutrient contents.Keywords: Earthworm, vermicomposting, rice straw, grass clippings, sago wastes, organic wastes, humic acidsAfrican Journal of Biotechnology Vol. 12(17), pp. 2207-221

Molecular characterization and pathogenicity of Streptococcus agalactiae serotypes Ia ST7 and III ST283 isolated from cultured red hybrid tilapia in Malaysia

Streptococcus agalactiae is one of the most important pathogens infecting tilapia worldwide. In this study, we determined the serotype, sequence type (ST), virulence gene profile and pathogenicity of S. agalactiae isolated from cultured fish. A collection of 256 S. agalactiae that were previously isolated from outbreaks of streptococcosis in red hybrid tilapia in Peninsular Malaysia were studied. The genotype and ST of the isolates were determined by molecular serotyping of the capsular polysaccharide (cps) gene clusters, and multi-locus sequence typing (MLST). The virulence gene profiles were constructed using m-PCR for 14 standard virulence genes with published primers. Two serotypes, with different associated STs, were identified in this study: serotypes Ia ST7 and III ST283. Serotype Ia ST7 lacks lmb, scpB, pavA, fbsB, cyl, bca, cspA and bac genes, which were present in serotype III ST283. The LD50–240h of S. agalactiae in red hybrid tilapia for serotypes Ia ST7 and III ST283 were 8.7 × 103 CFU/mL and 6.3 × 103 CFU/mL, respectively. The histopathological findings were consistent with meningitis, necrosis of hepatocytes, coagulative tubular necrosis and hypocellular of the spleen. Histopathological scores revealed significant differences (p < 0.05) between the two serotypes in all lesions in the brain, liver (except hepato-necrosis) and spleen, but not the kidney. This study revealed that S. agalactiae serotypes Ia ST7 and III ST283 currently exist in cultured tilapia in Malaysia. This molecular epidemiology study could be useful for the future development of cross protective vaccines against streptococcosis in the region