Citrograph May-June 2010 - Citrus Research Board

grafted bud for further indexing. This
method is effective against most citrus
viruses; however, is not effective against
viroids, and its effect on citrus tatter leaf
virus has been reported to be variable.
All imports received by CCPP originating
outside the U.S. are routinely subjected
to thermaltherapy as a precaution.
Shoot-tip-micrografting (STG) is the
other form of disease clean-up therapy
employed by the CCPP. Some pathogens,
particularly the citrus viroids (e.g.
exocortis, cachexia), are difficult or impossible
to eliminate by thermaltherapy
and are much more readily eliminated by
STG. STG is a procedure where several
new growth tips slightly less than 1cm in
length are taken from one of the original
infected import propagations. Under a
dissecting microscope, an apical meristem
of about 0.15 mm, barely visible
to the naked eye, is removed from the
infected growth tip and grafted onto a
seedling grown in vitro. If small enough
when removed from the growth tip, the
apical meristem is not yet developed
enough to contain the pathogen, and
therefore the disease will not be present
in the micro-grafted propagation. STG
propagations are returned to glass tubes
and placed under light in a culture chamber.
When the scion of the micrografted
propagation reaches about 2 cm, it is regrafted
onto a clean rough lemon seedling
and moved to the greenhouse. STG
is effective against all graft-transmissible
agents including viroids (Fig. 6).
The two therapy techniques – thermotherapy
and shoot-tip-grafting – are
complimentary, providing a flexible system
for pathogen elimination bypassing
the limitations (thermotherapy ineffective
for viroids) and/or practical restrictions
(STG long growth-regeneration
time, specialized equipment, in vitro
contaminants, and possible rootstockscion
incompatibilities) that one method
may have.
Following any therapy procedure, all
propagations produced during therapy
must go through indexing again to determine
their disease status. If the subsequent
indexing indicates that a disease is
still present, then the plant material must
be subjected again to therapy. This cycle
of therapy and testing continues until all
tests are negative. When propagation of
a budline tests negative in a pre-indexing
following therapy, it may then enter the
full scale VI index (see section B) (Fig.2,
4). If a budline is shown to be free of
Fig.6. Shoot-tip-grafting. Preparation of shoot tip from the source plant (A i) and
excision of apical meristem (2-3 leaf primordia barely visible with naked eye) (A
ii). Preparation of rootstock seedling, seed germination in vitro (B i) and removal
of portion of shoot and root growth (B ii). Necessary tools (scalpels, scissors,
and forceps) for shoot-tip-grafting (C i) and dissecting microscope (C ii). Inverted
T-cut on rootstock seedling under the dissecting microscope and placement of
the apical meristem in contact with cambial tissue ((D I and ii). In vitro growth
of shoot-tip grafted plants in liquid media (E i) and apical meristem growth after
shoot-tip-grafting (E ii).
known diseases in the VI Index, it is
then considered ready for release from
quarantine (Fig.2, 5) (see section D).
D. Quarantine release
When an introduced budline has
tested negative for all known budtransmissible
diseases in the VI index,
the CCPP then applies for its release
from both state and federal quarantine.
The CCPP must first obtain release from
CDFA by outlining the testing procedures
and test results. Once released by
the State of California, an application
for federal quarantine release is sent to
USDA/APHIS containing the testing
information and a copy of the letter of
approval by the State of California for
release from quarantine. The distribution
of citrus material that has been released
from quarantine is also a highly regulated
and carefully executed procedure
that involves close interaction between
CDFA, CCPP, and citrus nurserymen
and growers (Fig. 2, 5).
The above described introductory
procedure and release from quarantine
is available to private entities that
wish to import patented or other proprietary
varieties into California. The
propagator/owner signs an agreement
with UCR for the recovery of the cost
of the testing and therapy procedures
(currently set at $10,000), and when
the variety is released from quarantine
it is delivered to the owner and is not
maintained in the CCPP Foundation
Blocks (see section E).
E. Maintenance
The Lindcove Foundation and Evaluation
Block
The newly introduced varieties that
have been pathogen tested and found
“clean” and released from state and
federal quarantine are propagated at
Rubidoux on suitable rootstocks for field
planting in the Foundation and Evaluation
Block. The CCPP Foundation and
Evaluation Block is located at the University
of California Lindcove Research
and Extension Center near Exeter in the
San Joaquin Valley of California. This is a
field planting of about 20 acres and now
contains over 1,200 trees and over 300
different scion and rootstock varieties.
The Foundation and Evaluation
Block is planted on fumigated soil and
has a wide planting distance between
rows and trees to allow for better visual
evaluation of each tree. Each tree of
the Foundation and Evaluation Block
is examined several times each year
by CCPP and interested University
and industry people for horticultural
trueness-to-type, fruit quality, freedom
from budsports and chimeras, spontaneous
genetic disorders, and symptoms
of disease. Until 2007, each tree was
annually re-indexed biologically and by
ELISA for tristeza and was tested up to
three more times by ELISA during the
May/June 2010 Citrograph 31