Production and Characterization of Protease from Serratia marcescens

Extracellular protease produced by Serratia marcescens isolated from the gut of termite (Centrocestus formosanus) was investigated in this study. The effect of temperature, pH, carbon and nitrogen sources on protease production was examined. The extracellular protease was separated from the culture supernatant of the organism through precipitation with ammonium sulfate. The protease was purified by Sephadex G-150 gel filtration and diethylaminoethanol (DEAE) - Sephadex A-50 ion exchange chromatography. The effect of temperature, pH, and salts on the proteolytic activity of the enzyme was also investigated. Optimal protease production was obtained after about 12-hour incubation. The optimum temperature and pH for protease production was found to be 40°C and pH 7, respectively. Protease production was highest when rice bran and casein were used as the carbon and nitrogen sources, respectively. The native molecular weight of the protease was estimated to be 56 kDA. The optimal temperature for hydrolysis of casein was 50°C and the optimal pH was 8. The protease was found to be inhibited by ethylenediaminetetraacetic acid (EDTA), NaCl, CaCl2, FeCl2 and HgCl2. Proteolytic activity was however enhanced by KCl. The maximum velocity Vmax and Km values were 40 mg/min/mL and 0.91 mg/mL, respectively

Dry flower disease of Macadamia in Australia caused by Neopestalotiopsis macadamiae sp. nov. and Pestalotiopsis macadamiae sp. nov

Incidence of dry flower disease of macadamia (Macadamia integrifolia), expressed as blight of the flowers, necrosis and dieback of the rachis, is increasing in Australia. In the 2012/13 production season, incidence of dry flower disease resulted in 10% to 30% yield loss in the affected orchards. Etiology of the disease has not been established. This study was established to characterise the disease and identify the causal pathogen. A survey of the major macadamia producing regions in Australia revealed dry flower disease symptoms, regardless of cultivar or location at all stages of raceme development. Based on colony and conidial morphology, the majority (41%) of fungal isolates obtained from tissue samples were identified as Pestalotiopsis and Neopestalotiopsis spp. The phylogeny of the combined partial sequence of the internal transcribed spacer, beta-tubulin and translation elongation factor 1-alpha gene loci, segregated the isolates into two well supported clades, independent of location or part of the inflorescence affected. Further morphological examination supported the establishment of two new species, which are formally described as Neopestalotiopsis macadamiae sp. nov. and Pestalotiopsis macadamiae sp. nov. Using spore suspensions of isolates of both species, Koch?s postulates were fulfilled on three macadamia cultivars at all stages of raceme development. To our knowledge, this is the first report of species of Neopestalotiopsis and Pestalotiopsis as causal agents of inflorescence disease in macadamia

Genome-Wide Association Study for Abscission Failure of Fruit Pericarps (Stick-Tights) in Wild Macadamia Germplasm

Macadamia pericarps that fail to abscise (‘stick-tights’) are an important trait to select against in breeding as they can harbour pests and diseases. Traditional macadamia breeding cycles are lengthy and expensive due to long juvenilities and large tree sizes. Thus, genome-wide association studies (GWAS) are an important investigative tool to identify candidate trait-linked markers to enable potential reductions in evaluation and selection cycles via marker-assisted selection (MAS) in young seedlings. This study assessed 199 wild macadamia germplasm accessions for stick-tight prevalence across two years. As the number of stick-tights per tree is limited by the number of nuts per tree, we conducted association analyses to identify SNPs linked with the number of stick-tights per tree, and examined whether such SNPs were also associated with, and thus confounded with, the number of nuts per tree. We also assessed associations with the proportion of stick-tights per total number of nuts. Thirty-two SNPs were associated with at least one of the stick-tight traits in one year (p < 0.001). Of all such SNPs, only one was associated with the number of nuts per tree (p < 0.001), indicating that most associations were not confounded with yield