The internal transcribed spacer regions (ITS1 and ITS2) of the
RNA gene repeat from Phytophthora species were amplified
using the polymerase chain reaction and sequenced. Sequences from P.
cambivora, P. cinnamomi, P. citricola, P. cryptogea,
P. fragariae var. fragariae, P. fragariae var.
rubi, P. megasperma var. megasperma and P.
nicotianae were compared with published
sequences and phylogenetic trees were produced. The resultant grouping
species generally agreed with groupings established
using classical morphological criteria, primarily sporangial morphology.
Amongst species with non-papillate sporangia two clades
were evident, one consisting of P. fragariae, P. cambivora
P. cinnamomi and the other of P. megasperma, P. drechsleri
and P. cryptogea. The latter three were placed in the tree between
the non-papillate groups and the papillate and semi-papillate groups
which formed three distinct clades. One group comprised P. citricola,
P. citrophthora and P. capsici, another P. megakarya
P. palmivora and a third P. pseudotsugae, P. cactorum,
P. idaei, P. nicotianae and P. infestans. More
P. megasperma, P. drechsleri
and P. cryptogea will need to be examined to settle more precisely
the relationship of these species to the others. PCR amplification
of ITS sequences using freeze-thawed mycelial scrapings from pure cultures
growing on agar followed by digestion with restriction
enzymes is a quick and easy way to compare and identify isolates without
the need for laborious DNA extraction procedures. With
improved technology, rapid automatic sequencing of PCR-amplified ITS regions
is now possible and yields detailed information of
relationships within the genus as well as allowing the design of
species-specific PCR primers for diagnostic purposes.