A comparative study of culture dependent and independent techniques (metagenomics) of bacterial communities associated with Persian oak tree

During the last decade, about 20-30% of the Zagros forests have faced a problem called oak decline. The aim of present study was to investigate the bacterial diversity in the healthy and declined forest areas by comparing two methods culture-dependent and culture-independent methods.

Sampling was done from different tissue and soil of oak trees in the forests of Ilam province. In the culture-based method, using microbiological and molecular methods the bacteria in the samples were isolated and identified to the genus and species level. In order to study metagenomics in culture-independent method, total microbial DNA was extracted directly from the samples and prepared metagenomic library using conventional and index primer, the diversity of bacterial was examined using Illumina's Miseq platform and then the results of the two methods were compared.

Results

Based on the culture-based method, only 3 phylum were identified, while in the metagenomics method, 9416 OTUs were obtained, that were divided into 12 phylum. In both methods, 3 families of Bacillaceae, Enterobacteriacea and Xanthomonadaceae were observed as the dominant families. The species diversity within a sample (Alpha diversity) was higher in the rhizosphere sample and in the Eyvan and Gale jar areas than other samples. The species diversity between samples (Beta diversity) was in Eyvan, Gale jar, Chavar, Tangedalab and Arghavan appear more similar to each other than the sites samples. Bacterial community of stems and leaves were more similar and also bulk and rhizosphere were more similar in bacterial species composition.

In the metagenomics method, very few species are identified, while in the culture method, the probability of this is low. Also, in the culture-independent method, alpha and beta diversity can be studied more comprehensively. In the culture-based method, the bacterial population can be identified up to the species level, while in the culture-independent method, it is usually identified up to the genus level. Therefore, it is suggested to use a combination of two methods simultaneously in studies of microbial communities.