Molecular characterization of a mosaic locus in the genome of 'CandidatusLiberibacter asiaticus'

Abstract

BackgroundHuanglongbing (HLB) is a highly destructive disease of citrus production worldwide. 'Candidatus Liberibacter asiaticus', an unculturable alpha proteobacterium, is a putative pathogen of HLB. Information about the biology and strain diversity of 'Ca. L. asiaticus' is currently limited, inhibiting the scope of HLB research and control.ResultsA genomic region (CLIBASIA_05640 to CLIBASIA_05650) of 'Ca. L. asiaticus' showing hyper-sequence variation or locus mosaicism was identified and investigated using 262 bacterial strains (188 from China and 74 from Florida). Based on the characteristic electrophoretic profiles of PCR amplicons generated by a specific primer set, eight electrophoretic types (E-types) were identified, six E-types (A, B, C, D, E, and F) in China and four E-types (A, C, G, and H) in Florida. The 'Ca. L. asiaticus' strains from China consisted predominately of E-type A (71.3%) and E-type B (19.7%). In contrast, the 'Ca. L. asiaticus' strains from Florida was predominated by E-type G (82.4%). Diversity of 'Ca. L. asiaticus' in China was also evidenced. Strains from the high altitude Yunnan Province consisted of five E-types with E-type B being the majority (62.8%), whereas strains from the low altitude coastal Guangdong Province consisted of only two E-types with E-type A as the majority (97.0%). Sequence analyses revealed that variation of DNA amplicons was due to insertion/deletion events at CLIBASIA_05650 and the downstream intergenic region.ConclusionsThis study demonstrated the genomic mosaicism of 'Ca. L. asiaticus' resulted from active DNA insertion/deletion activities. Analyses of strain variation depicted the significant inter- and intra-continent diversity of 'Ca. L. asiaticus'.

Population Structures of ‘Candidatus Liberibacter asiaticus’ in Southern China

“…It remains unclear what caused the multiple amplicon phenomenon. It could be due to co-infection/existence of multiple genotypes (5,11,18), but others, such as nonoptimized PCR amplifications, cannot be excluded. In fact, amplification of plant/insect symbiont DNA could be explained by occasional nonoptimized PCR events.…”

“…L. asiaticus' showing locus mosaicism was recently characterized and the 'Ca. L. asiaticus' isolates in China could be divided into high and low altitude groups (18). Chen et al (2) analyzed the tandem repeat number (TRN) variation at a genomic locus (CLIBASIA_01645) of 'Ca.…”

Detection and Characterization of Miniature Inverted-Repeat Transposable Elements in "Candidatus Liberibacter asiaticus"

“…A previously published procedure was followed for PCR (29). Briefly, a reaction mixture (25 l) included 2 l of template DNA, 0.3 l of Taq DNA polymerase at 5 U/l (TaKaRa Bio Inc., Shiga, Japan), 0.4 l of each forward and reverse primer (10 M), and 2.5 l of 2.5 mM deoxynucleoside triphosphate.…”

“…Primers were designed using Primer 3 software (32). As an extension of a genomic diversity research (29), primer set LapPF1-f/LapPF1-r (5=-GCCACTTTGGGGTA GCAGTA-3=/5=-AAAACTTTCGTCACGGCTTT-3=) were found to yield multiple amplicons from some "Ca. Liberibacter asiaticus" isolates ( Fig.…”

“…In the past few years, there have been several reports of DNA polymorphisms in "Ca. Liberibacter asiaticus" populations using single primer sets (24,(26)(27)(28)(29). By analyzing two Chinese "Ca.…”