[논문]Enterobacter cloacae, an Emerging Plant-Pathogenic Bacterium Affecting Chili Pepper Seedlings

Garcia-Gonzalez, Tanahiri; Saenz-Hidalgo, Hilda Karina; Silva-Rojas, Hilda Victoria; Morales-Nieto, Carlos; Vancheva, Taca; Koebnik, Ralf; Avila-Quezada, Graciela Dolores

doi:10.5423/ppj.oa.06.2017.0128

Enterobacter cloacae, an Emerging Plant-Pathogenic Bacterium Affecting Chili Pepper Seedlings 원문보기

The plant pathology journal, v.34 no.1, 2018년, pp.1 - 10

Garcia-Gonzalez, Tanahiri (Universidad Autonoma de Chihuahua, Chihuahua, Campus 1) , Saenz-Hidalgo, Hilda Karina (Universidad Autonoma de Chihuahua, Chihuahua, Campus 1) , Silva-Rojas, Hilda Victoria (Colegio de Postgraduados, Campus Montecillo) , Morales-Nieto, Carlos (Universidad Autonoma de Chihuahua, Facultad de Zootecnia y Ecologia) , Vancheva, Taca (UMR Interactions Plantes Microorganismes Environnement (IPME), IRD-Cirad-Universite Montpellier) , Koebnik, Ralf (UMR Interactions Plantes Microorganismes Environnement (IPME), IRD-Cirad-Universite Montpellier) , Avila-Quezada, Graciela Dolores (Universidad Autonoma de Chihuahua, Facultad de Zootecnia y Ecologia)

Abstract ▼ AI-Helper

A previously unreported bacterial disease on chili pepper (Capsicum annuum L.) seedlings affecting as many as 4% of seedlings was observed in greenhouses in Chihuahua, Mexico (Delicias and Meoqui counties). Initial lesions appeared as irregular small spots on leaves and brown necrosis at margins tips were observed. Later, the spots became necrotic with a chlorotic halo. Advanced disease was associated with defoliation. A Gram negative, rod-shaped bacterium was isolated from diseased chili pepper seedlings. Three inoculation methods revealed that isolated strains produce foliage symptoms, similar to those observed in naturally infected seedlings. Pathogenic strains that caused symptoms in inoculated seedlings were re-isolated and identified to fulfill koch's postulate. Polyphasic approaches for identification including biochemical assays (API 20E and 50CH), carbon source utilization profiling (Biolog) and 16S rDNA, hsp60 and rpoB sequence analysis were done. Enterobacter cloacae was identified as the causal agent of this outbreak on chili pepper seedlings.

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가설 설정

(A) Initial symptoms in jalapeño pepper seedlings showing water-soaked in the middle of the leaves. (B) Chili pepper leaves with irregular brown spots and slightly yellowing. (C) Yellowing on the leaves with darkening in the affected area, apical necrosis is observed in some leaves.

제안 방법

Biochemical identification was conducted using API 20E and API 50CH strips (BioMerieux Inc., Durham, NC, USA), according to the manufacturer’s instructions.
In the third one, the bacterial suspension was injected with a needle-less 1-ml syringe, pressing the syringe on the lower side of three leaves per seedling. Four seedlings were inoculated per bacterial strain and SDW treatment controls, and each experiment was conducted two times.
, 2008). MUSCLE (v3.7; Mill Valley, CA, USA) configured for highest accuracy (MUSCLE with default settings) was used to align the sequences. Regions containing gaps or poorly aligned were deleted with Gblocks (v0.
The aim of this work was to characterize and identify the causative agents responsible for a new disease on chili pepper (Capsicum annuum) that is affecting as many as 4% of seedlings grown in commercial greenhouses in northern, Mexico.

대상 데이터

Sequence comparisons were performed on the websites of the European Bioinformatics Institute (http://www.ebi.ac.uk/services). DNA sequences were aligned using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970).

이론/모형

uk/services). DNA sequences were aligned using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970). For multiple sequence alignments, the MUSCLE algorithm was used (Edgar, 2004).
DNA sequences were aligned using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970). For multiple sequence alignments, the MUSCLE algorithm was used (Edgar, 2004).
Genomic DNA was extracted by a modified hexadecyl trimethyl ammonium bromide (CTAB) method (Wang et al., 2010). One loop of 48 h-old bacterial culture grown on NA plates were lysed.
Internal branch reliability was estimated by the aLRTtest (SH-Like). The phylogenetic tree graphic and edition was performed with TreeDyn (v198.3; Montpellier, France).
91b; Castresana Lab, Barcelona, Spain). The phylogenetic tree was rebuilt on the PhyML program (v3.0; Oregon University, USA) with maximum likelihood, assuming the number of invariant sites and 4 gamma-distributed rate categories, to account for rate heterogeneity across sites, the HKY85 substitution model was used. The gamma shape parameter was calculated from the data.

성능/효과

3). Sequences of our isolates had 99% identity (1 mismatch) to sequences from E. cloacae strains P101 and EcWSU1 (accession numbers CP006580 and CP002886). A comparison with the hsp60 fragments used by others revealed that our sequences were 100% identical to E.
cloacae, portions of the 16S rRNA genes were amplified by PCR and sequenced. The obtained 16S rRNA gene fragments were identical to portions of the 16S rRNA genes of several E. cloacae strains (e.g., GenBank accession numbers KT328458, KR061390, KF516245, CP006580).

후속연구

cloacae. More research is needed to understand the epidemiology of this new disease and to develop management strategies to reduce its expansion.

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