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

Abstract

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.

Keywords: 16S rDNA; Capsicum annuum L.; bacterium; hsp60; rpoB.

Fig. 1

Symptoms observed in chili (Capsicum annuum L.) pepper during seedlings production in commercial greenhouses in Mexico. (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. (D) Eventually, seedlings got completely defoliated after 60 days.

Fig. 2

Pathogenicity test in healthy jalapeño pepper seedlings from 50 days-old. (A) Apical necrosis observed in jalapeño pepper, 23 days after inoculation by injection method. (B) Foliar water-soaked necrosis with chlorotic halo. (C) Reduction on the foliar lamina. (D) Defoliated seedlings from 90 days-old.

Fig. 3

Neighbor-joining tree after analyzing 264 nucleotides of the hsp60 gene 429 of the 14 isolates and reference strains of the genus E. cloacae from Hoffmann and Roggenkamp (2003).

Fig. 4

Neighbor-joining tree after analyzing 545 nucleotides of the rpoB gene and strains of the genus E. cloacae from Miyoshi-Akiyama et al (2013).