Caracterização da comunidade bacteriana endofítica de citros por isolamento, PCR específico e DGGE

Abstract

Resumo -O objetivo deste trabalho foi caracterizar a comunidade bacteriana endofítica de plantas assintomáticas (escapes) e afetadas pela clorose variegada dos citros (CVC) por meio de isolamento em meio de cultura, técnica de gradiente desnaturante em gel de eletroforese (DGGE) e detecção de Methylobacterium mesophilicum e Xyllela fastidiosa por meio de PCR específico, para estudar esta comunidade e sua relação com a ocorrência da CVC. A análise da comunidade bacteriana via DGGE permitiu a detecção de X. fastidiosa, bem como Klebsiella sp. e Acinetobacter sp. como endófitos de citros. Foram observados também Curtobacterium sp., Pseudomonas sp., Enterobacter sp. e Bacillus spp. Utilizando primers específicos, Methylobacterium mesophilicum e X. fastidiosa também foram observadas, reforçando hipóteses de que estas bactérias podem estar interagindo no interior da planta hospedeira.Termos para indexação: Citrus sinensis, Xylella fastidiosa, Methylobacterium, bactérias endofíticas. Characterization of the endophytic bacterial community from citrus by isolation, specific PCR and DGGEAbstract -The aim of this work was to characterize endophytic bacterial community of assintomatic (escape) and Citrus Variegated Chlorosis (CVC)-affected citrus plants using isolation in culture medium, denaturing gradient gel electrophoresis (DGGE) technique and Methylobacterium mesophilicum as well as Xylella fastidiosa specific PCR, allowing to assess this community and its interactions with CVC. The study of bacterial community by DGGE analysis allowed the detection of X. fastidiosa, as well as Klebsiella sp. e Acinetobacter sp., which were not detected previously. Curtobacterium sp., Pseudomonas sp., Enterobacter sp. and Bacillus spp. were also observed as endophyte in citrus plants. Using specific primers Methylobacterium mesophilicum and X. fastidiosa were observed, reinforcing that these bacteria could interact inside the host plant.

The diversity of citrus endophytic bacteria and their interactions with Xylella fastidiosa and host plants

“…Furthermore, the frequency of endophytic bacteria in healthy, escape, and CVC-affected Citrus sinensis plants was studied using cultivation as well as cultivation-independent methods (Araújo et al ., 2002). Bacteria from the genus Methylobacterium were the most frequently isolated endophytes from CVC-symptomatic citrus plants ( C. sinensis ) (Araújo et al ., 2002; Lacava et al ., 2004, 2006a,b); however, Araújo et al . (2002) observed that M. extorquens was only isolated from CVC-affected plants while M. mesophilicum was isolated from healthy plants, indicating that specific Methylobacterium species may be related to the citrus plant physiological condition.…”

“…Several reports highlighted the relationships among bacterial populations and suggest that CVC symptoms in citrus plants could be influenced by the population balance among Methylobacterium spp., Curtobacterium flaccumfaciens and X. fastidiosa (Araújo et al ., 2002; Lacava et al ., 2004, 2006b, 2007a; Ferreira Filho et al ., 2012). Understanding the relationship among endophytic bacteria within sweet orange trees and X. fastidiosa may lead to strategies to control CVC using endophytic bacteria (Ferreira Filho et al ., 2012) or inducing physiological shifts in both the host plant and microbial community that result in more balanced and stable interactions.…”

Detection of siderophores in endophytic bacteria Methylobacterium spp. associated with Xylella fastidiosa subsp. pauca

“…The genus Methylobacterium, which occupies the same ecological niche as Xfp, was the most frequently isolated endophytic bacterium from CVC-symptomatic citrus plants (Citrus sinensis) (Araújo et al, 2002;Lacava et al, 2004Lacava et al, , 2006aLacava et al, , 2006b. Recently, an interaction between Methylobacterium species and Xfp was strongly indicated (Araújo et al, 2002;Lacava et al, 2004).…”

Molecular phylogeny and biotechnological potential of bacterial endophytes associated with Malpighia emarginata

“…Its positive activity was also reported in an assay for growth enhancement in plants (Sessitsch et al, 2004). Lacava et al (2006) characterized the endophytic bacterial community of citrus and investigated its relationship with the occurrence of citrus variegated chlorosis (CVC). In asymptomatic plants, the endophyte community has been reported to be composed of the genera Klebsiella sp, Acinetobacter sp, Curtobacterium sp, Pseudomonas sp, Enterobacter sp, Bacillus spp, and Methylobacterium mesophilicum, whereas CVC-affected plants are preferential hosts of Methylobacterium spp.…”

“…In fact, Arnold and Herre (2003) noted that these factors affect the occurrence of endophytes. After isolation, many endophytes could not be cultivated, perhaps due to their need of culture media or specific conditions for growth (Lacava et al, 2006).…”