The UC Riverside Citrus Variety Improvement Program (CVIP), the forerunner of the Citrus Clonal Protection Program (CCPP), began indexing candidate varieties in 1958. At that time, the full range of indicator plants that is utilized today was not known. In the early 1960s, the usefulness of ‘Dweet’ tangor as an indicator for Citrus Concave Gum Virus as well as other psorosis-like viruses was demonstrated. Consequently, starting in 1963 over 150 varieties not previously indexed on ‘Dweet’ were indexed on that indicator.
One of the results of this re-indexing was that a ‘Cleopatra’ mandarin (CRC 270, which had been indexed as VI 92) produced a leaf mottle resembling but distinct from that of psorosis or concave gum. The source tree showed no evidence of decline due to this virus, although it did show twig die-back, produced very small fruits, and was not vigorous. The trunk did not exhibit any discoloration or pitting. Because there were other selections of ‘Cleopatra’ available in the CVIP, this particular selection, which had been introduced from Florida in 1914, was eliminated from the program. Types of citrus other than ‘Dweet’ produced no symptoms but could act as carriers. This presumptive virus did not provide any protection against psorosis-like viruses and so was considered a distinct virus. Because this virus produced symptoms only in ‘Dweet’, it was named ‘Dweet Mottle Virus’ (Roistacher and Blue, 1968).
Dweet mottle virus remained a rather obscure virus. It was not observed to produce any losses in economic situations and was not reported to occur in commercial production. In fact, it was detected in the CCPP indexing program only one time after 1963. This was in a mandarin type introduced from New Zealand in the late 1990s.
In the mid 1980s, the Spanish group at the Instituto Valenciano de Investigaciones Agrarias (IVIA) reported a graft-transmissible disease that caused a bud-union incompatibility between ‘Nagami’ kumquat and ‘Troyer’ citrange (Navarro et al, 1984). The ‘Nagami’ in question (SRA-153) had been introduced from the Station des Recherches Agrumicoles in San Giuliano, Corsica. In addition to the incompatibility, the presumptive virus caused
vein-clearing in sweet orange and some other indicators and stem pitting in citron. After shoot-tip grafting, some of the plants produced were compatible with ‘Troyer’ and did not cause veinclearing but did pit the citrons, suggesting that there was more than one virus involved.
A later report (Galipienso et al, 2000) demonstrated that this virus caused bud-union creasing with ‘Nules’ clementine and ‘Eureka’ lemon on ‘Troyer’, whereas the same was not observed with ‘Pineapple’ sweet and ‘Marsh’ grapefruit on ‘Troyer’. The budunion problems were similar to those caused by Citrus tatterleaf virus. However, the pathogen did not act like CTLV in symptom expression in indicators or in mechanical transmissibility in herbaceous hosts. This report further strengthened the evidence that more than one virus was involved. All sources of the virus used in the reported experiments produced a chlorotic blotching in ‘Dweet’ tangor and stem pitting in citron. However, the bud union crease and vein-clearing in ‘Pineapple’ sweet orange were not observed in some shoot-tip grafted plants or from ‘Marsh’ grapefruit or ‘Pineapple’ sweet orange pre-inoculated with tissue from SRA-153 ’Nagami’.
The same group partially purified and characterized the apparent causal agent, and gave it the candidate name Citrus leaf blotch virus (Galipienso et al, 2001; Vives et al, 2001, 2002). These papers also indicated that CLBV was detected in trees in Spain and introductions from Japan and Florida. They also reported the development of probes usable for RTPCR as well as other molecular detection methodologies (Galipienso et al, 2004).
Luís Navarro of IVIA, in conversation with one of the authors (RRK) and Chet Roistacher in 2001, revealed that CLBV might be similar to Dweet mottle virus, based upon the symptom expression in ‘Dweet’. Consequently, we sent him tissue of the Dweet mottle positives maintained at the CCPP. This allowed the Spanish group to compare DMV and CLBV. They recently reported that the symptom expression of the two putative viruses is somewhat different: CLBV from SRA-153 induced bud-union crease of ‘Nules’ on ‘Troyer’, vein-clearing in ‘Pineapple’ sweet, chlorotic blotching in ‘Dweet’, and stem-pitting in ‘Etrog’, whereas DMV induced only the chlorotic blotching in ‘Dweet’ and stempitting in ‘Etrog’. Furthermore, they reported that the nucleotide identity between CLBV and the two California sources of DMV was over 96 %. They interpret these results as indicating that at the least DMV and CLBV are closely related. Dweet mottle may be caused by CLBV, with another virus being present in SRA-153 ‘Nagami’ causing the bud-union crease and vein-clearing (Vives et al, 2004).
The overall status of CLBV in California is unknown at this time. The recent report of seed transmission of CLBV (Guerri et al, 2004) makes it a concern for the citrus nursery industry. It is possible that more attention will need to be paid to the phytosanitary status of seed source trees than in the past. It should be noted that recently Citrus Variegated Chlorosis was also reported to be seedtransmitted (Li et al, 2003). In addition, there are anecdotal indications that certain individuals in Spain have alleged that CLBV was introduced into Spain in C-35 seeds from California. Furthermore, the recently reported bud-union problems between ‘Fukumoto’ and ‘Beck’ navels and certain citrange rootstocks resemble the bud-union problem associated with CLBV (as well as CTLV).
Consequently, we have recently assayed all rootstock varieties, kumquats, and ‘Fukumoto’ navels in the CCPP Foundation Block at Lindcove Research and Extension Center utilizing RT-PCR (Galipienso et al, 2004) with our local DMV positives and a CLBV positive from Florida DPI (received via RF Lee) used as positive controls. Whereas the positives consistently produced a positive result from the RT-PCR, none of the FB trees did so. We have also tested all trees maintained in the Repository Protected Collection (the other source of clean citrus propagative material in California) in the same manner and to this point have detected no positives. If any positives are detected either at CCPP or NCGRCD, they will be re-sanitized.
The fact that all C-35 in California is derived from trees in the FB means that, given that the FB trees are apparently free of CLBV, other C-35 seed source trees should also be free of CLBV. However, ‘Troyer’ was introduced to California before indexing began, and it is possible that there are some seed-source ‘Troyer’ that do not derive from the FB trees. Therefore, clean FB ‘Troyer’ would not necessarily mean that all commercial ‘Troyer’ are also clean. A similar situation exists with the navels of interest. All ‘Fukumoto’ in California derive from the FB trees and freedom of these trees from CLBV would also mean that other trees are also free. However, ‘Beck’ navels have never passed through the CCPP and are not maintained in the FB, so nothing can be conjectured about ‘Beck’ at this point.
The close identity of CLBV and DMV has probably prevented CLBV from becoming introduced to California. All introductions of new citrus germplasm are indexed into ‘Dweet’ tangor (among other indicators). This would detect CLBV, which gives a reaction in ‘Dweet’ tangor, even if the actual identity of the virus was not known at the time of the index. Any apparent positives of this sort, even if misidentified, would have been eliminated by thermal therapy or shoot-tip grafting before release. Thus, probably CLBV and/or DMV are probably not present in California or, if present, have very low incidence.
It should be noted that our experience with CLBV is just beginning. Conversations with L Navarro and J Guerri of IVIA during the recent 2004 meeting of the International Organization of Citrus Virologists suggest that detection of CLBV is not always straightforward. The Spanish researchers told us that CLBV appears to be distributed irregularly in the trees. Detection is variable even in small greenhouse trees, and sometimes leaves from the same tree give variable results. We are thus continuing to assess the reliability of the RT-PCR test under our conditions. We are also currently observing the reaction of our DMV positives in indicators other than ‘Dweet’ in order to assess whether the reaction under our conditions is the same as that reported from Spain. If any growers or extension personnel have questions or concerns regarding DMV or CLBV, we invite them to contact us.
(Robert Krueger is the Curator at the USDA-ARS Citrus Germplasm Repository, Riverside; John Bash is a Staff Research Associate at U.C. Riverside; Richard Lee is the Director of the USDA-ARS Germplasm Repository, Riverside)
