Volume 60 - Tomato Genetics Cooperative - University of Florida

University with 16s rRNA primers,CLi.po.F/O12c and PCR fragments sequenced using
protocols reported by Secor et al. (2009). A PCR fragment of about 1,100 bp was
obtained and sequenced in both directions. After correction by visual proofreading, a
966-nt region (contact D.P. Maxwell for sequence) was submitted to a BLAST analysis
at the National Center for Biotechnology Information data base; and a 100% nucleotide
identity was obtained with the 16s-23s rRNA intergenic spacer region for „Candidatus
Liberibacter solanacearum’ from potato from Guatemala (FJ395205, Secor et al., 2009)
and tomato from Sinaloa, Mexico (FJ957897, Munyaneza et al., 2009b). These
sequences also had a 100% nucleotide identity with „Ca. L. solanacearum‟ from tomato
in New Zealand (EU834130, Liefting et al., 2009a, 2009b), from bell pepper collected in
Sinaloa, Mexico (FJ957896, Munyaneza et al., 2009a) and „Candidatus Liberibacter
psyllaurous‟ (synonym „Ca. L. solanacearum‟, see discussion in Secor et al., 2009)
from Zebra chip symptomatic potatoes in California (FJ498802, Crosslin and Bester,
2009). In Central America, Rehman et al. (2010) report the widespread occurrence of
„Ca. L. solanacerum‟ (GQ926922) and its potato psyllid vector in potato fields in
Honduras. Pair wise comparison of this sequence from „Ca. L. solanacearum‟ from
potato with the Guatemalan „Ca. L. solanacearum’ sequence from tomato showed that
there was a difference of two SNP between the sequences (678 nt), and this might
indicate a different geographic origin of these two pathogens.
Subsequently to samples collected in December 2009, samples were collected
from tomatoes with either typical begomovirus symptoms and/or purple/yellowing
symptoms on younger leaves in the Department of El Progresso in March 2010. The
samples were prepared for transport to the University of California-Davis using AgDia
absorption strips. This is a method to capture total nucleic acids from plant tissue in
which sap is prepared from the target tissue, applied to an absorbent matrix on the end
of a plastic 'stick', and allowed to dry prior to transport. DNA extracts were prepared
from the 12 samples and tested for the presence of Liberobacter and begomovirus
infection by PCR. 'Ca. Liberibacter sp.' was detected in 4 of 12 samples, and these
samples showed symptoms of stunted and distorted growth; older leaves were yellow
and brittle and younger leaves were upcurled with yellowing and vein purpling.
Begomovirus infection was detected in all 12 samples.
Additionally, 49 tomato samples were collected from March to June, 2010 and
assayed for Liberibacter sp. using PCR primers CLi.po.F/O12c (Secor et al., 2009).
The expected size fragment for Liberibacter sp. was obtained from 17 samples and no
fragments were obtained with the other 32 samples. Positive PCR samples were
collected from the following Departments: Sacatepéquez, Guatemala and Baja
Verapaz,. These PCR fragments will be sequenced for more definitive identification.
Universal PCR primers for phytoplasma (Smart et al., 1996) were used with the
DNA samples from tomato collected in Guatemala and all were negative. Because of
the purple top symptoms, phytoplasma were originally suspected as being present.
In the last two years, there has been considerable effort devoted to
understanding the etiology of Zebra Chip (see Secor et al., 2009) and the new
Candidatus Liberibacter sp. associated with solanaceous plants (see Liefting et al.,
2009a, 2009b). These efforts plus the report here indicate that this unculturable
bacterium transmitted by the tomato/potato psyllid will result in a serious disease for
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