Transcriptome Profiling Reveals the EanI/R Quorum Sensing Regulon in Pantoea Ananatis LMG 2665T.

Siphathele Sibanda,Collins Tanui,Teresa Coutinho,Lucy Moleleki,Stanford Kwenda,Divine Shyntum

doi:10.3390/genes9030148

Abstract

Pantoea ananatis LMG 2665T synthesizes and utilizes acyl homoserine lactones (AHLs) for signalling. The complete set of genes regulated by the EanI/R quorum sensing (QS) system in this strain is still not fully known. In this study, RNA-sequencing (RNA-seq) was used to identify the EanI/R regulon in LMG 2665T. Pairwise comparisons of LMG 2665T in the absence of AHLs (Optical density (OD)600 = 0.2) and in the presence of AHLs (OD600 = 0.5) were performed. Additionally, pairwise comparisons of LMG 2665T and its QS mutant at OD600 = 0.5 were undertaken. In total, 608 genes were differentially expressed between LMG 2665T at OD600 = 0.5 versus the same strain at OD600 = 0.2 and 701 genes were differentially expressed between LMG 2665T versus its QS mutant at OD600 = 0.5. A total of 196 genes were commonly differentially expressed between the two approaches. These constituted approximately 4.5% of the whole transcriptome under the experimental conditions used in this study. The RNA-seq data was validated by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). Genes found to be regulated by EanI/R QS were those coding for redox sensing, metabolism, flagella formation, flagella dependent motility, cell adhesion, biofilm formation, regulators, transport, chemotaxis, methyl accepting proteins, membrane proteins, cell wall synthesis, stress response and a large number of hypothetical proteins. The results of this study give insight into the genes that are regulated by the EanI/R system in LMG 2665T. Functional characterization of the QS regulated genes in LMG 2665T could assist in the formulation of control strategies for this plant pathogen.

Highlights

Pantoea ananatis is an ubiquitous, Gram-negative, yellow pigmented, rod shaped bacterium that has been described as an “emerging pathogen” of global importance [1]
The bacterial strains used in this study are P. ananatis LMG 2665T (virulent natural isolate from pineapple obtained from Belgian co-ordinated collections of microorganisms (BCCM)/LMG, University of Gent, Gent, Belgium), LMG 2665T ean∆I/R (P. ananatis LMG 2665T ∆ resistant to kanamycin) [5], Chromobacterium violaceum 026
The sampling points were determined by detection of acyl homoserine lactones (AHLs) using the CV026 assay as described

Summary

Introduction

Pantoea ananatis is an ubiquitous, Gram-negative, yellow pigmented, rod shaped bacterium that has been described as an “emerging pathogen” of global importance [1]. This bacterium infects plants and is capable of surviving in different environments [1]. This pathogen causes disease symptoms in a number of economically important plants such as Eucalyptus [1,2], staple food crops, namely maize, rice and sorghum [3] as well as cash crops, for example, honeydew melons, pineapples and onions [1]. Unlike many other plant pathogens, P. ananatis does not possess Type 2 Secretion System (T2SS) and Type 3 Secretion System (T3SS) [9].

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