Solid state fermentation,storage and viability of Streptomyces similanensis 9X166 using agro-industrial substrates against Phytophthora palmivora-induced black rot disease in orchids

Streptomyces similanensis 9X166 is known to be an antagonist of the black rot pathogen of orchids, Phytophthora palmivora. In this study, we investigated the production of highly viable S. similanensis 9X166 cells by solid state fermentation using agro-industrial substrates, and the shelf life of a S. similanensis 9X166 dried solid. Rice bran was found to be the most appropriate raw material for production of both viable cells and β-1,3-glucanase. A medium containing 12?g of rice bran and coconut husks at a ratio of 10:2, supplemented with 10?mL of mineral salts produced the highest number of viable cells and greatest level of β-1,3-glucanase. Ammonium sulfate was the most suitable nitrogen source, and an initial moisture content of 65% and a temperature of 30°C were found to be optimal conditions for the production of viable cells and β-1,3-glucanase. Storing the dried fermented solid under non-vacuum conditions resulted in the highest cell viability. The specific rate of degradation on viability increased as the temperature increased to 37°C, according to the Arrhenius equation. There was no difference between the storage time estimated by the Arrhenius equation from the specific rate of degradation compared to the validated storage time of S. similanensis 9X166 dried solids when maintained at the ambient temperature in Thailand. At 60 days, the product retained 10⁶ CFU/g of S. similanensis 9X166 in dried solid, which was the minimal effective amount for 100% inhibition of P. palmivora in living orchids.     

Molecular phylogeny of symbiotic basidiomycetes of fungus-growing termites in Thailand and their relationship with the host

Termitomyces-related symbiotic basidiomycetes in the nests of fungus-growing termites (Macrotermitinae) of several genera in Thailand were cultivated and analyzed phylogenetically based on the DNA sequence of nuclear ribosomal RNA genes. The relationships of the symbiotic fungi with host termites and their locality were apparently complex, supporting intricate mechanisms for the termites to acquire the symbionts.     

Symbiotic Fungi Produce Laccases Potentially Involved in Phenol Degradation in Fungus Combs of Fungus-Growing Termites in Thailand

Fungus-growing termites efficiently decompose plant litter through their symbiotic relationship with basidiomycete fungi of the genus Termitomyces. Here, we investigated phenol-oxidizing enzymes in symbiotic fungi and fungus combs (a substrate used to cultivate symbiotic fungi) from termites belonging to the genera Macrotermes, Odontotermes, and Microtermes in Thailand, because these enzymes are potentially involved in the degradation of phenolic compounds during fungus comb aging. Laccase activity was detected in all the fungus combs examined as well as in the culture supernatants of isolated symbiotic fungi. Conversely, no peroxidase activity was detected in any of the fungus combs or the symbiotic fungal cultures. The laccase cDNA fragments were amplified directly from RNA extracted from fungus combs of five termite species and a fungal isolate using degenerate primers targeting conserved copper binding domains of basidiomycete laccases, resulting in a total of 13 putative laccase cDNA sequences being identified. The full-length sequences of the laccase cDNA and the corresponding gene, lcc1-2, were identified from the fungus comb of Macrotermes gilvus and a Termitomyces strain isolated from the same fungus comb, respectively. Partial purification of laccase from the fungus comb showed that the lcc1-2 gene product was a dominant laccase in the fungus comb. These findings indicate that the symbiotic fungus secretes laccase to the fungus comb. In addition to laccase, we report novel genes that showed a significant similarity with fungal laccases, but the gene product lacked laccase activity. Interestingly, these genes were highly expressed in symbiotic fungi of all the termite hosts examined.     

Symbiotic fungi produce laccases potentially involved in phenol degradation in fungus combs of fungus-growing termites in Thailand

Fungus-growing termites efficiently decompose plant litter through their symbiotic relationship with basidiomycete fungi of the genus Termitomyces. Here, we investigated phenol-oxidizing enzymes in symbiotic fungi and fungus combs (a substrate used to cultivate symbiotic fungi) from termites belonging to the genera Macrotermes, Odontotermes, and Microtermes in Thailand, because these enzymes are potentially involved in the degradation of phenolic compounds during fungus comb aging. Laccase activity was detected in all the fungus combs examined as well as in the culture supernatants of isolated symbiotic fungi. Conversely, no peroxidase activity was detected in any of the fungus combs or the symbiotic fungal cultures. The laccase cDNA fragments were amplified directly from RNA extracted from fungus combs of five termite species and a fungal isolate using degenerate primers targeting conserved copper binding domains of basidiomycete laccases, resulting in a total of 13 putative laccase cDNA sequences being identified. The full-length sequences of the laccase cDNA and the corresponding gene, lcc1-2, were identified from the fungus comb of Macrotermes gilvus and a Termitomyces strain isolated from the same fungus comb, respectively. Partial purification of laccase from the fungus comb showed that the lcc1-2 gene product was a dominant laccase in the fungus comb. These findings indicate that the symbiotic fungus secretes laccase to the fungus comb. In addition to laccase, we report novel genes that showed a significant similarity with fungal laccases, but the gene product lacked laccase activity. Interestingly, these genes were highly expressed in symbiotic fungi of all the termite hosts examined.