Kusavadee Sangdee

Selection of entomopathogenic fungus for biological control of chili anthracnose disease caused by Colletotrichum spp.

The literature on the entomopathogenic fungi in the genus Cordyceps describing its use in the agricultural area as a biocontrol agent is limited. In this study, a total of 47 isolates of entomopathogenic fungi were isolated from dead cicada nymphs obtained from various locations in the northeast of Thailand. These isolates were primarily screened for antagonistic activity to inhibit the mycelial growth of one isolate of Colletotrichum gloeosporioides and one isolate of C. capsici. The screen revealed that five isolates of entomopathogenic fungi showed good inhibitory effects on the fungal mycelial growth and were chosen for further confirmation of their antagonistic effects against five isolates of C. gloeosporioides and five isolates of C. capsici by the dual culture method. After investigation, the isolate Cod-NB1302 had the best inhibitory effect. Moreover, the mycelium extract and culture filtrate of isolate Cod-NB1302 also had inhibitory effects on the mycelial growth and conidial germination of all isolates of plant pathogenic Colletotrichum spp. under in vitro conditions. Interestingly, the mycelium extract and culture filtrate effectively reduced the size of the disease lesion and disease severity on chili fruits after inoculation with the plant pathogenic fungi. However, the mycelium extract exhibited greater antifungal activity than the culture filtrate. Finally, the isolate Cod-NB1302 was identified as Ophiocordyceps sobolifera based on the sequence of three ribosomal nuclear DNA genes and two protein-coding genes. These findings suggest that the isolate Cod-NB1302 is a potential candidate, with antagonistic activity, for use as a source of antifungal agents to control anthracnose disease caused by the plant pathogenic Colletotrichum spp.

Development of SCAR primers based on a repetitive DNA fingerprint for Escherichia coli detection

The present study aimed to use enterobacterial repetitive intergenic consensus (ERIC) fingerprints to design SCAR primers for the detection of Escherichia coli. The E. coli strains were isolated from various water sources. The primary presumptive identification of E. coli was achieved using MacConkey agar. Nineteen isolates were selected and confirmed to be E. coli strains based on seven biochemical characteristics. ERIC-PCR with ERIC 1R and ERIC 2 primers were used to generate DNA fingerprints. ERIC-PCR DNA profiles showed variant DNA profiles among the tested E. coli strains and distinguished all E. coli strains from the other tested bacterial strains. A 350 bp band that predominated in five E. coli strains was used for the development of the species-specific SCAR primers EC-F1 and EC-R1. The primers showed good specificity for E. coli, with the exception of a single false positive reaction with Sh. flexneri DMST 4423. The primers were able to detect 50 pg and 100 CFU/ml of genomic DNA and cells of E. coli, respectively.

Evaluation of antigen preparation methods for polyclonal antibody production against Streptomyces spp.

Aim: To determine the optimum antigen preparation method for producing specific polyclonal antibody specific for Streptomyces species. Study Design: Experimental study. Place and Duration of Study: Faculty of Science, Mahasarakham University, Mahasarakham Province, Thailand, between May 2011 and December 2011. Methodology: Two Streptomyces isolates were used for antisera production. The sonication method was chosen for antigen preparation. Antigen suspensions were emulsified with incomplete Freund’s adjuvant and 1 ml was injected into rabbit thigh muscle for the first, second and third immunization. The fourth and fifth immunizations were injected intravenously. Antibody titer, detection limit and specificity were measured using indirect-ELISA. Streptomyces antigen mixed with soil was investigated. Results: The 5 min sonication method gave a higher protein than other test methods, so this protocol was chosen for all subsequent work. The sonicated Streptomyces antigen was

Investigation of antibacterial and anti-cancer activities of Streptomyces sp SRF1 culture filtrate

Purpose : To evaluate the antibacterial activity and cytotoxic effects of Streptomyces sp. SRF1 culture filtrate extract against breast cancer cell line. Methods : The activity of the extract against Gram-positive and Gram-negative bacteria was initially screened by an agar-well diffusion method. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values were measured by broth microdilution method. Time-kill assays were also performed, and extract-induced morphological and ultrastructural changes to bacterial cells were investigated. Sulforhodamine B (SRB) assay was performed to determine the cytotoxicity of the extract against the human breast cancer cell line, MCF-7. Results: Antibacterial activity by the extract was detected against four strains of Gram-positive pathogens including one strain of methicillin-susceptible Staphylococcus aureus (MSSA) and 3 strains of methicillin-resistant Staphylococcus aureus (MRSA) - with low MIC and MBC values. This activity was bactericidal after 6 h exposure. Morphological alterations were detected on the cell surface of both MSSA and MRSA. The extract also inhibited MCF-7 cell growth with half-maximal concentration (IC50) of 211.67 ± 33.95 μg/mL in 72 h. Conclusions: Streptomyces sp . SRF1 culture filtrate extract exhibits potent antibacterial and anticancer activities and thus, represents a potential source of antibacterial and anticancer drugs. Keywords : Antibacterial activity, Anti-breast cancer, Staphylococcus aureus , Streptomyces sp. SRF1

Disease suppressive activity of extracts from entomopathogenic fungus Ophiocordyceps sobolifera against chili anthracnose fungi Colletotrichum spp. in a pot experiment

This study evaluated the efficacy of the extracts of Ophiocordyceps sobolifera isolate Cod-NB1302 for the biological control of chili anthracnose disease caused by Colletotrichum capsici and C. gloeosporioides under pot conditions. Among the extracts, mycelial extract treatments provide the best reduction in disease severity. Interestingly, two bioactive constituents, adenosine and cordytropolone, from the mycelial extract, inhibited growth of the fungal pathogens. Moreover, these bioactive compounds had a synergistic effect against the fungal pathogens in a pot experiment. These results confirmed the disease suppressive activity of the mycelial extract.

Colony Characteristics, Nucleoside Analog Profiles, and Genetic Variations of Medicinal Fungus Polycephalomyces nipponicus (Ascomycetes) Isolates from Northeast Thailand

The entomopathogenic fungus Polycephalomyces nipponicus is known to have activity against human pathogenic bacteria and the malaria pathogen; however, information about its genetic variation is limited. In this study, cicada nymphs infected with entomopathogenic fungi were collected from various locations in the northeast of Thailand. Internal transcribed spacer sequencing was used to identify the fungal pathogen P. nipponicus. A total of 36 isolates of P. nipponicus from 6 provinces were investigated for variations in fungal morphology, nucleoside analog content, and genetics. The results showed that colony morphology varied depending on the strain of the tested fungi, without influence from its geographic origin. A similar finding was observed with regard to the production of nucleoside analog content. Interestingly, the important bioactive compound adenosine was detected in the mycelial extract of all 36 isolates. This indicates that P. nipponicus could possibly be used as a source of potential therapeutic bioactive compounds. In addition, random amplified polymorphic DNA analysis, as supported by the Nei Index and Shannon Index values, showed high genetic variation within and between the populations. These findings represent what is, to our knowledge, the first information on the colony morphology, adenosine analog profile, and genetic variation of P. nipponicus.