Chitin amendment increases soil suppressiveness toward plant pathogens and modulates the actinobacterial and oxalobacteraceal communities in an experimental agricultural field

Abstract

A long-term experiment on the effect of chitin addition to soil on the suppression of soilborne pathogens was set up and monitored for 8 years in an experimental field, Vredepeel, The Netherlands. Chitinous matter obtained from shrimps was added to soil top layers on two different occasions, and the suppressiveness of soil toward Verticillium dahliae, as well as plant-pathogenic nematodes, was assessed, in addition to analyses of the abundances and community structures of members of the soil microbiota. The data revealed that chitin amendment had raised the suppressiveness of soil, in particular toward Verticillium dahliae, 9 months after the (second) treatment, extending to 2 years following treatment. Moreover, major effects of the added chitin on the soil microbial communities were detected. First, shifts in both the abundances and structures of the chitin-treated soil microbial communities, both of total soil bacteria and fungi, were found. In addition, the abundances and structures of soil actinobacteria and the Oxalobacteraceae were affected by chitin. At the functional gene level, the abundance of specific (family-18 glycoside hydrolase) chitinase genes carried by the soil bacteria also revealed upshifts as a result of the added chitin. The effects of chitin noted for the Oxalobacteraceae were specifically related to significant upshifts in the abundances of the species Duganella violaceinigra and Massilia plicata. These effects of chitin persisted over the time of the experiment.

Fig 1

Effects of chitin amendment on the soilborne pathogen Verticillium dahliae (A) or Pratylenchidae (nematodes) (B) over sampling points and crops. Averages of triplicate measurements with standard error bars are given; significant values (P < 0.05) are indicated with “∗” (see also reference 26). MS, number of microsclerotia; n, number of individuals.

Fig 2

Relative abundance of total bacteria (A), total fungi (B), Oxalobacteraceae (C), or Actinobacteria (D) in unamended and chitin-amended soil. Error bars represents standard errors of the means (geometric) for three replicates, and sampling points indicated with “∗” were significantly different (comparison of chitin-amended with unamended soils, P < 0.05).

Fig 3

Clustering of DGGE profiles based on UPGMA and Jaccard correlation coefficient. Shown are total bacteria (A), total fungi (B), Oxalobacteraceae (C), or Actinobacteria communities (D) in unamended and amended soils. Sampling time points of unamended soils, Dec-09, June-10, Nov-10, March-11, and April-12, are referred to as Dec, June, Nov, March, and April. Sampling time points of chitin-amended soils, Dec-09, June-10, Nov-10, March-11, and April-12, are referred as Dec+, June+, Nov+, March+, and April+. Sampling points indicated with “∗” were significantly different (P < 0.05).

Fig 4

Comparison of 16S rRNA gene sequences of the oxalobacteraceal community. The stacked column graph shows relative distribution of different bacterial species based on BLASTN analysis. Average relative abundance from three replicates as the ratio between sequence type abundance and total number of sequences in the group is shown.

Fig 5

Relative abundances of bacterial chitinolytic communities as assessed on the basis of the chiA gene. Sampling points indicated with “∗” were significantly different between chitin-amended and unamended soils (P < 0.05).