Involvement of two glycoside hydrolase family 19 members in colony morphotype and virulence in <i>Flavobacterium columnare</i> -- Journal of Oceanology and Limnology,2017,35(6):1511-1523

	Cite this paper:
	ZHANG Xiaolin, LI Nan, QIN Ting, HUANG Bei, NIE Pin. Involvement of two glycoside hydrolase family 19 members in colony morphotype and virulence in Flavobacterium columnare[J]. Journal of Oceanology and Limnology, 2017, 35(6): 1511-1523

Involvement of two glycoside hydrolase family 19 members in colony morphotype and virulence in Flavobacterium columnare

ZHANG Xiaolin^1,2, LI Nan¹, QIN Ting¹, HUANG Bei³, NIE Pin¹

1 State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China;
2 University of Chinese Academy of Sciences, Beijing 100049, China;
3 College of Fisheries, Jimei University, Xiamen 361021, China

Abstract:

Flavobacterium columnare is the pathogenic agent of columnaris disease in aquaculture. Using a recently developed gene deletion strategy, two genes that encode the Glyco_hydro_19 domain (GH19 domain) containing proteins, ghd-1 and ghd-2, were deleted separately and together from the F. columnare G₄ wild type strain. Surprisingly, the single-, △ghd-1 and △ghd-2, and double-gene mutants, △ghd-1 △ghd-2, all had rhizoid and non-rhizoid colony morphotypes, which we named △ghd-1, △ghd-2, △ghd-1 △ghd-2, and N△ghd-1, N△ghd-2, and N△ghd-1 △ghd-2. However, chitin utilization was not detected in either these mutants or in the wild type. Instead, skimmed milk degradation was observed for the mutants and the wild type; the non-rhizoid strain N△ghd-2 exhibited higher degradation activity as revealed by the larger transparent circle on the skimmed milk plate. Using zebrafish as the model organism, we found that non-rhizoid mutants had higher LD₅₀ values and were less virulent because zebrafish infected with these survived longer. Transcriptome analysis between the non-rhizoid and rhizoid colony morphotypes of each mutant, i.e., N△ghd-1 versus (vs) △ghd-1, N△ghd-2 vs △ghd-2, and N△ghd-1 △ghd-2 vs △ghd-1 △ghd-2, revealed a large number of differentially expressed genes, among which 39 genes were common in three of the pairs compared. Although most of these genes encode hypothetical proteins, a few molecules such as phage tail protein, rhs element Vgr protein, thiol-activated cytolysin, and TonB-dependent outer membrane receptor precursor, expression of which was down-regulated in non-rhizoid mutants but up-regulated in rhizoid mutants, may play a role F. columnare virulence.

Key words: Flavobacterium columnare|GH19 domain|gene deletion|rhizoid colony|non-rhizoid colony

Received: 2016-06-03 Revised: 2016-07-14

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