Mining of unexplored habitats for novel chitinases—chiA as a helper gene proxy in metagenomics

Abstract

The main objective of this study was to assess the abundance and diversity of chitin-degrading microbial communities in ten terrestrial and aquatic habitats in order to provide guidance to the subsequent exploration of such environments for novel chitinolytic enzymes. A combined protocol which encompassed (1) classical overall enzymatic assays, (2) chiA gene abundance measurement by qPCR, (3) chiA gene pyrosequencing, and (4) chiA gene-based PCR-DGGE was used. The chiA gene pyrosequencing is unprecedented, as it is the first massive parallel sequencing of this gene. The data obtained showed the existence across habitats of core bacterial communities responsible for chitin assimilation irrespective of ecosystem origin. Conversely, there were habitat-specific differences. In addition, a suite of sequences were obtained that are as yet unregistered in the chitinase database. In terms of chiA gene abundance and diversity, typical low-abundance/diversity versus high-abundance/diversity habitats was distinguished. From the combined data, we selected chitin-amended agricultural soil, the rhizosphere of the Arctic plant Oxyria digyna and the freshwater sponge Ephydatia fluviatilis as the most promising habitats for subsequent bioexploration. Thus, the screening strategy used is proposed as a guide for further metagenomics-based exploration of the selected habitats.Electronic supplementary materialThe online version of this article (doi:10.1007/s00253-012-4057-5) contains supplementary material, which is available to authorized users.

Effects of Trichoderma viride chitinases on the peritrophic matrix of Lepidoptera

“…Firstly, we would like to confirm the detrimental effects of T. viride chitinases on relevant insect pests. Secondly, we will characterise and test other fungal chitinases produced by isolated fungi or we will search novel enzymes into yet‐to‐be explored uncultivable microorganisms by metagenomic approaches …”

Chitin Amendment Increases Soil Suppressiveness toward Plant Pathogens and Modulates the Actinobacterial and Oxalobacteraceal Communities in an Experimental Agricultural Field

“…Thus, the genus Streptomyces, along with Oxalicibacterium (in the presence of fungi), might play key roles in disease-suppressive soils as being part of the amendment-reactive microbiota (5,19,20,54). A further lead, obtained from chiAbased Roche 454 deep pyrosequencing analysis of the June-10 chitin-amended field and soil from a microcosm experiment (17,21), recently indicated an increase in the abundance of Oxalobacteraceae-like sequences. In the present study, the relative abundances of both groups, Actinobacteria and Oxalobacteraceae, were positively correlated with chitin amendment over experimental time.…”

“…The gels were prepared using an optimized denaturant gradient for each type of gene (Table 1) (18,21,(32)(33)(34)(35). DGGE profiles were obtained using 16 h of electrophoresis at an optimized voltage in 0.5ϫ Tris-acetate-EDTA (TAE) buffer at 60°C.…”

“…The investigation of oxalobacteraceal communities was supported by previous findings in our laboratory of oxalobacteraceal dominance based on 454 pyrosequencing of chitinolytic bacteria. The chiA-gene-based analyses showed that particular oxalobacteraceae may be important colonizers of chitin-amended soils and rhizosphere, as well as other underexplored habitats (21). Additionally, recent reports (22,23) of the genome sequences of the plant-growth-promoting Herbaspirillum sp.…”

A broad pH range and processive chitinase from a metagenome library

“…The presence and diversity of chitinases have been investigated in some metagenome libraries from diverse environments such as soil (11), aquatic habitats (12) and extreme habitats (13) using PCR or sequencing approaches. However, only a few works went further on the characterization of found chitinases.…”