Publication of The College of Agricult UN IV E R S I T Y O F C A L 1 F O R N I A QUICK DECLINE OF CITRUS AS INFLUENCED BY TOP-ROOT RELATIONSHIPS W. P. BITTERS E. R. PARKER C ALI FOR N IA AGR I C U ITU RAl EXPERIMENT STATION BULLETIN 733 A THREAT . . . to the California Citrus Industry In 1939, an unfamiliar disorder affecting orange trees appeared in a small number of orchards in two areas of Los Angeles County. Soon after that, it was discovered in Orange County. The striking character of the symptoms on affected trees resulted in the name "quick decline." A 1940 survey of the Los Angeles County Agricultural Department revealed 146 affected trees in seven orchards. Subsequent findings were: 1941, 277 affected trees in 21 orchards; 1942, 791 trees in 35 orchards; 1943, 1,394 trees affected; 1944, 7,484 trees. By 1949, there were about 200,000 affected trees in Los Angeles County. The disease also spread over a large part of Orange County, and to many orchards in San Bernardino County. Some infected trees were found in west- ern Riverside County in 1949, and in southern Ventura County in 1951. An estimated half million trees have been affected as of 1952. In California there are approximately 250,000 acres of oranges and grapefruit, or about 22 million trees. Of these, it is estimated that from 40 to 50 per cent — 8% to IOV2 million trees — have top-root combinations that are susceptible and may eventu- ally be affected. Clearly, this disease may be expected to have a serious effect on the economy of the citrus industry in California and to cause major adjustments in it. The cover picture shows two Valencia trees on sour orange root. The one at the top has the "collapse" type of quick decline symptoms. The tree in the lower photograph shows the "gradual decline" type symptoms. (See page 8.) THE PROBLEM: Will the right top A PROGRESS REPORT on What Is Being Done about It Research on quick decline is a continuing project. This bulletin, the first of a series of progress reports, includes a history of the disease and studies of the tolerance of the commercially important scions to quick decline when those scions are budded on a wide range of species and varieties of rootstocks. Experiments on alterations in the top and root relationships of established trees are also described. Briefly, this is what research, to date, has contributed toward solution of the problem of quick decline: It is now known that quick decline is a virus disease, and that the virus is trans- mitted from tree to tree by an insect vector. Only certain budded combinations, and occasionally seedlings, are affected. Quick decline is probably identical with diseases long prevalent in South Africa and Java, with the "bud union decline" of Australia, and the "tristeza" disease of Argentina and Brazil. The disease has been particularly devastating to the citrus industry in the two South American countries because their orchards were planted chiefly with susceptible combinations. In rootstock experiments of limited scope, with plantings in 1945 and 1946, trees were subjected to natural infection as well as artificial inoculation with the quick decline virus. A more comprehensive experiment, with Valencia orange scions on 125 different rootstocks, was planted in 1948. Half of the trees in this second planting were subjected to inoculation by the spur-bud technique. Generally, present results of the two trials agree: the same combinations showed symptoms in both plantings. However, after-inoculation symptoms progressed more rapidly in the Baldwin Park plantings. (See table 1, page 12.) Inoculation by means of diseased buds generally caused a reduction in tree growth fie tight oof stock resistance to QUICK DECLINE? proportionate to the symptoms in evidence although there were exceptions. In some cases, the growth was reduced in trees which showed no other visual symptoms. Susceptibilities Trees on the various rootstocks were classed as tolerant, susceptible, or uncertain in their reaction to the disease. The following designations generally hold true as of August, 1950: Sweet orange scions are apparently susceptible to quick decline when budded on stocks of sour orange, shaddock, hybrids of shaddock x sweet orange, grapefruit, all lemons (except Rough lemon types), some mandarin hybrids (mostly tangelos), Citrus Ichangensis hybrids, Palestine sweet lime, and Citrus macroptera. Stem pitting has appeared on Morton citrange stock. Based on the effects of artificial inoculation, the reaction of sweet orange scions is uncertain on stocks of trifoliate orange, trifoliate orange hybrids, some mandarins, many mandarin hybrids, and some sweet oranges. Sweet orange scions appear to be tolerant on stocks of the acid and mandarin limes, some mandarins, on several mandarin hybrids, on some sweet oranges, on Rough lemon types, and on Citrus moi. In some cases, the determination of tolerance or susceptibility is clear. In others, not enough time has elapsed for prpper classification. Generally, the effects of artificial inoculation are apparent, but in some cases their duration is not yet known. Observations will be continued, to clarify results. Topworking Topworking susceptible sweet orange trees to lemons was generally successful even when some of the trees were showing symptoms of the disease at the time. It was found that all of the orange top should be removed in topworking. Topworking to grapefruit was unsatisfactory. Grapefruit/sweet orange/sour orange was found susceptible to quick decline. Inserting a tolerant interstock into susceptible trees did not prevent occurrence of the disease ; in fact, it seemed to hasten the development of symptoms. When the top was susceptible on a particular rootstock, the top remained suscep- tible when the species used for the root was inserted as an intermediate stock over a normally tolerant stock. Inarching Inarching of susceptible combinations with tolerant seedlings did not prevent prog- ress of the disease in infected trees. The growth of the inarches and the vigor of the tops were increased by various girdling procedures. A complete girdling, or an inverted "funnel" between the inarches, resulted in the greatest inarch growth. Rough lemon seedlings, when used for inarching, gave better results than did sweet orange seedlings. When inarched trees were heavily pruned, they formed a new top which was normal in appearance. The top volume of these trees five years after inarching and three years after top pruning did not exceed the size of young trees on tolerant roots planted at the time of inarching. It appears that the replanting of tolerant combinations is thus more practical than inarching. The following pages contain a detailed discussion of the experiments and the re- sults obtained, with supporting illustrations and tables. [4] QUICK DECLINE of CITRUS As Influenced by TOP- ROOT RELATIONSHIPS W. P. BITTERS E. R. PARKER Importance of Top-root Relationships to Quick Decline As early as 1944, Halma, Smoyer, and Schwalm {30) 2 observed that tops of sweet orange {Citrus sinensis Osb.) on sour orange {Citrus aurantium Linn.) roots appeared to be affected by quick decline, while sweet orange tops on sweet orange roots appeared to be resistant. The following year, these same workers published additional observations {31) suggesting that sweet orange trees on roots of trifoliate orange, grapefruit, and Rough lemon were also resistant combi- nations. They further reported, on the basis of limited observations, that lemon trees were not affected by the disease. In foreign countries, studies of citrus diseases that closely resemble quick de- cline {2, 25, 39, 54, 55, 57) have shown that sweet orange and some other top species are susceptible on other roots in addition to sour orange. The importance of top-root relationships in determining susceptibility is thus well established. Although a citrus variety, as a root- stock, may be tolerant to quick decline, it may still be undesirable since all root- 1 Paper No. 744, University of California Cit- rus Experiment Station, Riverside, California. 2 Italicized figures in parentheses indicate references to "Literature Cited," page 33. stocks do not have the same horticultural value. They vary in their resistance to diseases other than quick decline, their ability to grow as replants in old citrus soils, and their wide effects on scion vari- eties budded upon them. Many of the cit- rus species and varieties included in these trials have never previously been tested for their desirability as stocks. It is im- portant, therefore, that tolerant roots not have a deleterious effect on the longevity of the tree or an undesirable effect on fruit yields and fruit quality of the scion variety budded upon it. Relation of Quick Decline to Similar Diseases Elsewhere A disorder very similar to quick de- cline was first observed in South Africa as early as 1899 {16, 45). In 1891, sev- eral members of the Cape Department of Agriculture, recognizing the merits of sour orange rootstock in foreign coun- tries {38) , advocated that citrus varieties in South Africa be budded on sour orange stock. Of the trees so budded, only those with lemon tops remained healthy after a few years; the others be- came sickly and died. The failure of trees on sour orange roots was investigated by a Commission of Inquiry of the Cape De- partment of Agriculture, in 1904 (4), THE AUTHORS: W. P. Bitters is Associate Horticulturist in the Experiment Station, Riverside. E. R. Parker was Horticulturist in the Experiment Station, Riverside. (Deceased. [5] which found that most of the healthy trees in South Africa had been budded on Rough lemon (C. limon hybrid) stock. The failure of sour orange stock in South Africa was originally attributed to "in- compatibility." A malady resulting in the same general symptoms was next observed in Java, about 1928, by Toxopeus {55). By the use of reciprocal grafts of sweet orange, sour orange, and Japanese citron (Citrus reticulata hybrid) , he showed that the use of a tolerant intermediate stock neither prevented nor delayed the decline. As long as the top was sweet orange and the stock or interstock was sour orange, the combination failed. Toxopeus further showed that the parallel insertion of both sweet orange and Japanese citron tops into sour orange roots results in death of the plants. Inarching sweet orange trees with sour orange or Japanese citron seed- lings resulted only in the failure of the trees with sour orange inarches. Toxo- peus suggested that the sweet orange top produced some substance that was toxic to the sour root. About 1931, a similar disease appeared in Argentina (5). The trouble was re- ported in Brazil in 1937 (5). Moreira (42) referred to it as the "tristeza" dis- ease (tristeza, in Portuguese, means mel- ancholy or sadness), and described its very rapid spread. Bitancourt (6) pro- posed that an infectious agent was in- volved as the cause of the disease in Brazil. Orange, grapefruit, and mandarin scions on sour orange root declined, but sour orange seedlings or lemons budded on sour orange roots remained healthy. Moreira (42) reported that sweet orange tops on Thornton tangelo stock were af- fected by the disease, but that the Samp- son tangelo was completely tolerant when used with sweet orange tops. However, Marloth (39), in studies made at Nels- pruit, Transvaal, reported the behavior of Sampson tangelo root budded to sweet orange to be similar to that of sour orange. Bertelli and Bertelli (3) reported the presence of tristeza in Uruguay in 1940. It was suspected in Venezuela in 1949 (11). The same, or a very similar, disease was first noticed in Australia, about 1941 (10, 34), where it has been called "bud- union decline." It is also found in New Zealand (34) , on the Gold Coast of West Africa (24, 32), where it is prevalent on lime trees, and in Ceylon (47, 48). It is also believed to be in China (53, and un- published information from Dr. Lin Kung-Hsiang, Professor of Plant Pathol- ogy, Lingnan University, Canton, China), Mexico, and other countries. A disorder resembling quick decline has recently been reported in Louisiana (8). In 1952 it was reported from Hawaii (23) and also from Florida (9) . Evidence has been presented recently, by McClean (36, 37) , that the condition known as "stem- pitting" of grapefruit in South Africa is probably a different symptom manifesta- tion of the disease that causes the failure of sweet orange trees on sour orange roots. The observations of Grant, Costa, and Moreira (14, 28, 29) on tristeza, and of Wallace and Drake (56) on quick decline substantiate this fact. Nature and Transmission of Quick Decline and Similar Diseases Although several theories as to the cause of quick decline and of similar, if not identical, diseases elsewhere were ad- vanced and investigated (12, 22), the possibility that these diseases would prove to be of virus origin appeared most probable (5) . This theory was thoroughly analyzed by Webber (57), in 1943, for the diseases then prevalent in South Af- rica and in Brazil. Subsequent research has proved the virus nature of the dis- ease. Schneider (22) investigated the anatomical relationships in trees affected with quick decline. He found a necrosis of the sieve tubes, just below the bud union, which appeared to precede any other symptom, and pointed out that a [6] similar necrosis was caused by a virus in the case of cherry trees affected with the "buckskin" disease (46, 49). The simi- larity of the pathological anatomy of quick decline and of tristeza-affected orange trees was later established by Schneider (51) and Schneider, Bitan- court, and Rosetti (52). McAlpin et al. (34) indicated similar results with bud- union decline in Australia. The virus nature of quick decline was first demonstrated by Fawcett and Wal- lace in 1946 (20) . These workers showed that transmission of quick decline could be effected by inoculating healthy trees by insertion of buds from trees showing symptoms of the disease, and concluded that bacteria or fungi were not involved (20, 22). Subsequently, Meneghini (41) succeeded in transmitting tristeza by means of an insect vector, and concluded that this disease also was caused by a virus. Bennett and Costa (1, 2) trans- mitted tristeza by infected buds and the vector. Oberholzer (44), in 1947, re- ported the tissue transmission of the dis- ease present in the Union of South Africa. He also showed that susceptible combinations (for example, sweet orange tops on sour orange roots, both derived from seedlings) remained healthy when grown in a greenhouse free of insect pests. The virus causing the disease is apparently not seed transmitted (2, 44) . In 1948, Fawcett and Wallace (21) demonstrated that sweet orange tops on sweet orange roots are symptomless car- riers of the quick decline virus. Bennett and Costa (2) were able to transmit the virus of tristeza from symptomless sources, such as sweet orange on Rang- pur lime, Rough lemon on its own roots, "cravo" tangerine on its own roots, and an unidentified citrus on its own roots. Perhaps any variety of citrus that does not show obvious symptoms of the dis- ease when infected may serve as a host and act as a carrier. Budded trees have shown the most severe symptoms. Milder symptoms have been observed on some inoculated seedlings (2, 27), but the condition of such seedlings later im- proved. It is now recognized (56, 28, 29) that certain citrus seedlings are good indicator plants for the presence of the virus. Infection in nature and the resultant rapid spread of these diseases have been attributed to insect vectors. Meneghini (41), in 1946, and subsequently Bennett and Costa (2) demonstrated that tristeza virus could be transmitted by a species of black aphid, Aphis citricidus (Kirk.). This aphid has not been found in the United States. McAlpin et al. (34) in- timated that either red scale (Aonidiella sp.) or the black aphid was a vector for bud-union decline. In 1949 (35) he con- cluded that the citrus aphid (Aphis citricidus Kirk.) acts as a vector, but presented no data. This aphid has also been shown to transmit the causal virus of the disease on limes on the Gold Coast (32), stem pitting of grapefruit in South Africa (36, 37), and stem pitting in Brazil (14). In California, Dickson and his coworkers showed that Aphis gossypii Glover is a vector of the quick decline virus (17, 18). Wallace and Drake (56) have demonstrated that this aphis is also capable of transmitting the quick decline virus and causing stem pit- ting symptoms on West Indian lime seed- lings. Thus the importance of insect car- riers of the causal viruses of these dis- eases is well established. It has not always been conceded that quick decline, tristeza, and other "tris- teza-type" disorders are identical. How- ever, as a result of the recent investiga- tions of Hughes and Lister (32), Mc- Clean (36, 37), Costa, et al. (14), and Wallace and Drake (56), the evidence now appears conclusive that these dis- eases are caused by the same virus or by closely related strains. This conclusion was reached on the basis of the marked similarity of symptoms on budded trees and on Mexican lime seedlings, and the transmission of the causal virus of all [7] these diseases (except quick decline) by Aphis citricidus. This species has not been found in the United States. The slower rate of spread of the quick decline disease in California may be partly due to a low efficiency in virus transmission by Aphis gossypii and the local effects of climate upon the geographical distribu- tion of that vector. Recent evidence (26) also indicates that there are different strains of the tris- teza virus, and that they differ in viru- lence. A difference in virulence might reasonably account for some differences in various top-root responses (25, 27) under similar environmental conditions. Top-root responses may also possibly vary because of wide differences in en- vironmental conditions. Various citrus varieties are used as tops and roots in different countries. Apparent conflicts in the susceptibility of specific combinations may be due to any one, or a combination of, the above factors. SYMPTOMS OF QUICK DECLINE Symptoms of quick decline may be di- vided into two types — with one, trees show a relatively gradual decline; and with the other, trees show collapse or abrupt wilting (cover picture) . Although the latter type is the most striking in ap- pearance, and justifies the name quick decline, it is the less common. Gradual Decline. The first visual symptom of the more gradual decline of old orchard trees is the suppression of the normal seasonal flushes of growth. This lack of growth is usually accompa- nied by leaf discoloration. The leaves on affected trees lose their dark green luster and become dull and olive-green to yel- low-green in color. Early in the spring or late in the fall the leaves usually be- come somewhat yellow and display some yellowing of the midrib and the lateral veins. The new leaves on affected trees are small, leathery, and stand somewhat erect. After leaf discoloration begins, the leaves are shed gradually and the branches begin to die back from the tips. As the limbs become defoliated, weak multiple shoots develop from dormant buds, but there are no strong leaders. Such trees may survive for several years. They are incapable of bearing a com- mercial crop and are generally promptly removed from the orchards. Collapse. When affected trees col- lapse, an abrupt wilting of the leaves occurs within a period of a few days. This may or may not be preceded by initial symptoms similar to those that characterize trees affected with the grad- ual decline. The dried leaves and fruits generally remain attached to the twigs for several weeks or longer. Although such trees at first appear to be dead, they frequently develop new shoots from buds on the large limbs or trunk. The new growth consists of weak multiple shoots, and is not vigorous. Such trees then enter a stage of partial recovery, sometimes referred to as an equilibrium stage. Light crops of inferior quality fruit may then be obtained. The trees may again collapse during periods of soil moisture stress. Diseased trees have a tendency to blos- som very heavily, and the bloom may occur in off seasons. The number of fruits set is larger than normal, and there is a tendency for the fruits to be borne in clusters. The orange fruits color prema- turely, and appear more conspicuous. Heavy or off-season bloom and abnor- mally heavy crops of prematurely colored fruit are sometimes the first visible indi- cations of the disease. Among the earliest symptoms that can be observed on affected trees is the dying of the feeder roots, followed by the death and sloughing of the bark of the smaller lateral roots (22). These effects precede or accompany the appearance of top symptoms. The injury to the root system follows the necrosis of the sieve tubes of [8] the rootstock below the bud union. These tubes are blocked {50), thus preventing movement of carbohydrates to the root system. As the reserve supply of carbo- hydrates in the roots is utilized, starch disappears — first in the feeder roots {19, 22). When these have been entirely de- pleted of starch, they soon decompose {22) . Some functioning phloem usually persists or is formed- however, and slight root activity generally occurs. The rate and extent of the destruction of the root system are associated with the degree of the severity of the top symptoms. Application of the starch test (I-KI) to root tissues as an aid in the diag- nosis of quick decline and of tristeza has been a subject of much discussion. In 1945, Fawcett {19) found that trees visi- bly affected by quick decline showed no starch in the wood or bark of live roots. This phenomenon was observed as an outgrowth of previous work by Bitan- court (7) on trees affected with tristeza disease in Brazil. In quick decline, the disappearance of starch from the wood or bark of live roots at their extremities, and frequently from the wood of larger live roots, may be detected prior to, or coincident with, the appearance of top symptoms. In the earliest stages of the disease, the amount of starch varies in- versely according to the degree of severity of the top symptoms. However, Bitan- court (7) reported that tristeza top symp- toms may appear before the disappear- ance of starch from the roots, and this is also said to be true of trees suffering from "chlorosis" in Ceylon {48). It should be emphasized that the starch test is merely an aid in the diagnosis of quick decline — used only to determine if the movement of the food supply has been interrupted. But this reaction could be caused by any type of girdling injury, such as gopher- ing and gummosis, as well as by quick decline. Stem Pitting. The stem pitting as- pect of quick decline is another phase of the symptom complex. Stem pitting symptoms may or may not accompany typical quick decline symptoms. The term stem pitting is somewhat misleading since it implies that the disorder occurs wjr Fig. 1— Left: close-up of a healthy Morton citrange stock, showing its typical smooth, symmetrical nature. Center: close-up of a Morton citrange stock affected by stem pitting, showing the resulting grooves and asymmetrical nature of the trunk. Right: Morton citrange stock affected by stem pit- ting which did not extend across the bud union. Note pits and grooves in the wood and corre- sponding peglike growth on inner surface of rolled-back bark. [9] only on the stem. However, stem pitting is an anatomical abnormality which may occur on the twigs, branches, or trunk of citrus seedlings or budded trees, and on the latter it may also be found on the stock as a result of the plant's being in- fected with the quick decline virus. The top alone may show pitting in some com- binations, the root alone in some, and both top and root in others. Stem pitting is characterized by the in- ability of the cambium to function nor- mally, at least in localized areas. Portions of the xylem (wood) fail to develop, or do not develop normally. This results in numerous small pits or depressions in the wood. In contrast, small masses of tissue develop in place of some of the normal phloem (bark) elements so that, corres- ponding to the sunken, undeveloped areas in the wood, are small, elevated, peglike growths on the inner surface of the bark (fig. 1, right) . In advanced stages the pits tend to anastamose (connect), and the resulting striations, or grooves, tend to follow a twisted course parallel to the grain of the wood. The grooving may be more severe in some areas than in others, and may result in an asymmetrical shape of the stock (fig. 1, center). This symp- tom of quick decline first appeared in the field in 1952 and was more fully investi- gated in October of that year. Results are reported elsewhere. OBJECTIVES AND PROCEDURE The first purpose of the present study (of which this bulletin is a progress re- port) is to determine the tolerance or susceptibility to quick decline of the com- mercially important varieties of citrus when grown on a large number of species and varieties of rootstocks. The second purpose considers the feasibility of alter- ing existing top-root relationships of sus- ceptible orchard trees. Such alterations include top working (substituting a toler- ant top), inarching (substituting a tol- erant root), and bridgegrafting (substi- tuting a tolerant intermediate between top and root). Rootstock Trials The rootstock investigations were un- dertaken in 1945 at a time when only sweet orange tops on sour orange roots were known to be susceptible to quick decline. Reports from other countries had indicated that other susceptible combina- tions would be found. Two sets of experi- ments were begun at that time. Rootstock Trials Planted 1 945 and 1946. For these experiments, 620 trees of various top-root combinations were obtained from commercial nurseries or from nurseries of the Citrus Experiment Station. They were planted as replants in the orchards of nine cooperators in the quick-decline area of the Azusa-Covina district (Los Angeles County). Trees on 10 rootstock were included. All of these trees were exposed to natural infection by quick decline. However, the trees in one orchard were also inoculated, in the spring of 1947, with buds from diseased trees, in a manner described by Wallace and Fawcett (20). The Baldwin Park and Associated Plantings. In the spring of 1945, seeds of 125 types of citrus and citrus relatives were gathered from the variety orchards at the University of California Citrus Ex- periment Station, Riverside, from similar orchards at the University of California at Los Angeles, or from other sources. 3 The seeds were germinated in deep flats in a greenhouse at Riverside. About 35 of the resulting seedlings of each variety were transplanted to a nursery in Corona in the fall of 1945. Both Riverside and Corona were free of known cases of quick 3 Seeds were furnished by E. Mortensen, Uni- versity of Texas Agricultural Experiment Sta- tion, Winter Haven, Texas; F. E. Gardner, U. S. Department of Agriculture, Orlando, Florida; and by commercial growers. [10] decline until 1949. The various seedlings were budded with a uniform strain of Valencia sweet orange in the spring of 1947. In addition, 35 seedlings each of sweet orange, sour orange, and Rough lemon were double-worked to all possible combinations of those three species. About 3,500 trees were balled from this nursery in May of 1948 and planted in the following manner: 2,112 trees of Valencia on 125 different rootstocks were placed at Baldwin Park in June, 1948. In addition, there were 93 of the double- worked trees mentioned above. Each com- bination was planted in single-row plots of 2, 4, or 6 trees each, in a randomized block experiment with four replications. One half of the trees (the odd-numbered in the rows) was inoculated in August, 1948. The other half (the even-numbered in the rows) was not inoculated. Each inoculated tree was budded high on the trunk with one bud from each of three diseased trees. Six trees were used as sources of inoculum, and buds from each source tree were used in two of the ran- domized blocks. The inoculation buds were held in place with strips of gummed tape. The "take" of these buds was poor, and the inoculation process was repeated in the same manner in October, 1948. Rubber budding strips were used for wraps. A good "take" was obtained in the second budding attempt. The inocu- lation technique of inserting three tissue grafts is considered to have resulted in the introduction of large quantities of inoculum. The check trees were not budded. Twelve hundred trees from the Corona nursery were planted in the orchards of 10 cooperators, half of which were out- side any known quick-decline area. None of these trees was inoculated, but those in the decline area were exposed to natural infection. Some plantings were on old citrus soil. These are expected to be long-term plantings, and it is hoped that they will furnish much of the infor- mation in regard to the horticultural value of those stocks shown to be tolerant to quick decline as a result of the inocu- lation trials. Results are not reported here. Trees with other tops and roots were planted in 1949, 1950, 1951, and 1952. Still others are being grown and will be planted later. Alteration of Top-Root Relation- ships of Orchard Trees in Topwork- ing. Eighteen trees of Valencia sweet orange on sour orange root, showing varying stages of quick decline, were en- tirely or partly topworked in 1945, by means of cleft grafts, to Marsh seedless grapefruit at the E. B. Griffith orchard near Covina. Twenty-eight similar trees were partly or entirely topworked to Eureka lemons at the J. B. Stair grove in the Glendora area. Both orchards are in Los Angeles County. The grafts were in- serted in the primary scaffold or in the secondary limbs of the trees. All sweet orange growth was subsequently removed. Inarching. The substitution of tolerant roots for the susceptible roots of budded orchard trees was attempted by this pro- cedure. Inarches were established in 1945 and 1946 in the groves of 10 cooperators in the Los Angeles County quick-decline area. A total of 255 Valencia trees on sour orange root, of different ages and in vary- ing stages of quick decline, were inarched with either sweet orange or Rough lemon seedlings. Generally, four inarches were inserted per tree. Some four- and five- year-old trees received only two inarches per tree. The trees were not cut back at the time of inarching, but some of them, later on, were severely pruned and treated in other ways. Suckers were al- lowed to grow on the inarches for the first year in order to encourage inarch growth. Additional unaffected trees, bordering on the quick-decline zone, were inarched in 1949. The trees are Valencia sweet orange on sour orange roots. Several spe- cies of citrus and citrus relatives were used for inarching. Results of the 1949 inarching are not yet available. [in Bridgegrafting. Bridgegrafting over the bud union with grafts from a tolerant intermediate seedling was attempted on a small scale. In 1945 a total of 257 bridgegrafts was placed in 70 trees in orchards of seven cooperators. Rough lemon was used as the source of supply for the grafting wood. The number of bridgegrafts per tree varied with the size of the tree, some of the larger trees having six to eight. A high percentage of the bridgegrafts failed to become established. RESULTS Rootstock Experiments 1945-1946 Plantings. The appear- ance of quick-decline symptoms on trees of this series of experiments up to Febru- ary, 1950, is shown in Table 1. Of the trees with sweet orange tops, those on bittersweet roots were first visibly af- fected by quick decline, and developed the most severe symptoms. Those on sour orange roots were next to show the effects Table 1 — Symptom Indications of Quick Decline in Various Top-root Combinations in Preliminary Rootstock Trials Planted 1945 and 1946 (Last observation, February, 1950) No. of trees 45 67 39 4 56 52 136 48 34 39 7 9 18 6 6 3 3 6 3 3 3 3 10 Scion Intermediate Stock Symptoms* Sweet orange f Sweet orange f Sweet orange f Sweet orange f Sweet orange f Sweet orange § Sweet orange || Sweet orange Sweet orange | Sweet orange f Grapefruit t Lemon** Lemon** Sweet orange f Grapefruit! Grapefruit 1 Grapefruit! Lemon** Lemon** Lemon** Sour orange Sour orange Sour orange Bittersweet Sour orange | Grapefruit Pink shaddock Sampson tangelo Sweet orange Rough lemon Troyer citrange Morton citrange King mandarin Sour orange Troyer citrange Morton citrange Sweet orange Sweet orange Sour orange Bittersweet Sweet orange Sour orange Bittersweet Sour orange Bittersweet Sweet orange + + + + + + + + ♦ Sour orange Sour orange Sweet orange Sweet orange Sour orange Sweet orange f Sweet orange f Sweet orange Sweet orange * + = diseased; — = healthy. t Valencia variety. X Keen sour orange and unknown sour orange varieties. § Valencia, Bedmar, (Verna), and Viciedo varieties. || Valencia and Washington Navel varieties. \ Marsh variety. ** Eureka and Lisbon varieties. [12] of the disease. While the bittersweet is considered a variety of sour orange, the two differ both morphologically (58) and chemically {40). Sweet orange trees on grapefruit roots have shown mild symp- toms from time to time. As of 1952, how- ever, no mature commercial trees of sweet orange on grapefruit roots show symp- toms of quick decline, although some have been exposed to natural infection for 10 years. It is expected, however, that such trees will eventually be affected by the disease. In the 1945-1946 plantings, sweet orange trees on pink shaddock roots showed the first symptoms of decline in 1949. In the 1945-1946 plantings, sweet orange tops on roots of sweet orange, Rough lemon, Trover citrange, and on King mandarin did not show apparent symptoms. Certain of the artificially in- oculated trees of some of these combina- tions are, however, showing evidence of the disease in the Baldwin Park experi- ments. The Morton citrange stocks budded to Valencia orange show severe stem pitting symptoms in 1952. The health and vigor of the top was not ap- parently affected at that time. Two-year-old trees of grapefruit on sour orange roots inoculated with dis- eased buds in 1947 showed no leaf symp- toms by 1950. These observations were not in accord with some later inocula- tions. The susceptibility of this combina- tion is also indicated as a result of the topworking experiments discussed on page 28. Lemon scions on Troyer and Morton citranges have been healthy in so far as quick decline is concerned. Lemon tops are known to be tolerant to quick decline, even on sour orange roots. The experiments with the use of inter- stocks with various tops and roots indi- cate that the introduction of susceptible interstocks into otherwise satisfactory combinations has resulted in quick de- cline. Thus, trees of sweet orange/sour orange/sweet orange (fig. 2) were found to be susceptible. Likewise, grapefruit/' sour orange/sweet orange, and grape- fruit/sweet orange/sour orange trees have declined. Lemon tops, however, Fig. 2.— Left: five-year-old tree of sweet orange/sour orange/sweet orange, inoculated and show- ing symptoms of quick decline. Right: five-year-old tree of sour orange/sweet orange/sour orange, inoculated but healthy. [13] were healthy on these combinations of stocks and interstocks. Sour orange tops were also found toler- ant with sweet orange, sour orange, and bittersweet roots and interstocks (fig. 2). Sour orange seedlings in other trials grew satisfactorily. The development of quick decline symptoms on the trees in the 1945-1946 plantings has been slow, and the symp- toms have been mild. This may have re- sulted from the fact that many of the trees grew slowly, probably because they were frequently planted in orchards that were due for removal and had poor care. On such trees, symptom expression is initially less pronounced than on nor- mally vigorous trees. Baldwin Park Rootstock Experi- ment. There appear to be no genetic incompatibilities between the Valencia orange scion and the wide range of root- stock varieties in the 1948 plantings at Baldwin Park. The growth of the non- inoculated trees was very satisfactory. The present indications as to the tolerance of the various combinations to quick de- cline are in many instances tentative. In many cases several of the inoculated trees in certain top-root combinations are now showing symptoms of quick decline, while other similar trees are still healthy. Al- though a few of the noninoculated trees have more recently developed symptoms as a result of natural infection, not enough are affected to permit conclusions to be drawn from these trees alone. The first effects of the inoculations made in 1948 were evident during the following winter. Forty-three of the in- oculated trees suffered frost injury to the tops and trunks, as compared with 14 of the check trees. Combinations on lemon, lime, and shaddock stocks were the more tender. In July, 1949, the first symptoms of quick decline appeared on a few combi- nations, but they were not well defined. Additional combinations continued to express symptoms through the summer, and the severity of symptoms increased. In regard to the time of symptom appear- ance and the rate of symptom progres- sion, it was evident that the inoculated trees on some rootstocks are much more affected by artificial inoculations with quick decline than are others. On November 11-12, 1949, and also at subsequent dates, all of the trees were graded. The standards used are as fol- lows: grade = healthy; grade 0.5 = sug- gestive of the first stages of quick decline — reduced growth, small leaves, slightly off-color, off-bloom; grade 1 = definite quick decline — marked stunting, open growth, off-color, small leaves, slight vein clearing, off-bloom; grade 2 = marked stunting — yellowing of leaves, vein clear- ing, slight defoliation, off-bloom, fruit set, wilting; grade 3 = marked yellowing of the leaves, vein clearing, medium de- foliation, excessive bloom or off-bloom, fruit set, wilting; and grade 4 = severe defoliation — excessive bloom or off- bloom, fruit set, no growth, tree nearly dead or dead. Not all of these symptoms are required in each case for grading. Individual trees showing advanced de- cline symptoms were re-examined to de- termine if the observed symptoms were due to gummosis, dryness of soil, gopher injury, mechanical girdling, a variant rootstock, or other causes than quick decline. When such evidence was found, the records of those trees were not used. Trees showing symptoms of grade 1 are probably definitely not tolerant to quick decline, but combinations are listed as susceptible only when at least three trees showed grades of 0.5 or higher and the growth of the inoculated trees was con- siderably reduced. Classification is des- ignated as uncertain only if one or two of the inoculated trees of a combination were showing symptoms of grade 0.5, or if symptoms and tree growth measure- ments did not substantiate one another. Combinations are designated therefore as tolerant, susceptible, or uncertain. All frost-damaged trees were excluded. [14] The grading of May 5-6, 1950, was made during the flowering period. The presence of prolific flowering on young trees serves as an excellent symptom of the disease. The last grading reported herein was made on August 1-2, 1950. Results of this grading are presented in Table 2. This table also presents a sum- mary of the rate of growth of the inocu- lated trees, relative to the growth of non- inoculated trees, between August, 1948, and August, 1950. Measurements of the diameter of the scion were taken 4 inches above the bud union. In November, 1949, 12.45 per cent of the inoculated trees showed symptoms of quick decline, but only 0.19 per cent of the noninoculated trees. The small num- ber of noninoculated ttees that developed symptoms during the first year may be partly attributed to low insect popula- tions in the trial plots. By May, 1950, 27.37 per cent of the inoculated trees was affected, and 5.88 per cent of the check trees. The August, 1950, inspection re- vealed 41.60 per cent of the inoculated trees to be affected, and 10.12 per cent of the trees that were not artificially in- oculated. Trees on some roots, such as sour or- ange or shaddock, appeared to be much more affected by natural infection than others, such as trees on roots of the man- darin hybrids or trifoliate orange hy- brids. Differences in tolerance to natural and to artificial infection have also been observed by Costa, Grant, and Moreira (13). The data in Table 2 also show that the majority of the artificially inoculated trees has made less growth than those that were not inoculated by budding. In the case of some stocks, this has been the only observed effect of inoculation. This was noticeable with trees on sweet orange roots, on some of the mandarin roots, and on the roots of some of the trifoliate hybrids. In only a very few cases did the inoculated trees make greater growth than those which were not inoculated, and in these cases the increase is con- sidered so small as to be within the realm of experimental error. The significance of the effects of inoculation on the growth of trees that developed no other symp- toms of quick decline has not been en- tirely established; it is not believed to be due to mechanical effects. The possibility exists that some other virus, such as psorosis, which is known to reduce tree growth, was also introduced with the buds that were used for inoculation. The re- duction in growth has been more severe during the second year than it was the first year following inoculations. Sour Oranges. Nineteen varieties of sour orange were used as rootstocks in these trials. Some trees of all of these combinations showed symptoms by No- vember, 1949. By August, 1950, a greater number of trees were affected, and the degree of symptom expression was more advanced. In these combinations it was interesting to follow the rate of symptom development in the inoculated trees and in the check trees infected by the vector. In November, 1949, 36.8 per cent of the inoculated trees showed symptoms, and only 1.1 per cent of the noninoculated trees. By May, 1950, the percentage of visibly affected trees was 60.3 for the inoculated and 10.9 for the noninocu- lated. In August, 1950, the number of trees showing symptoms had increased to 70.7 per cent for the inoculated trees and 20.7 per cent for the noninoculated. Trees on certain varieties of roots ap- peared to show symptoms sooner than did those on some other varieties (see table 2) . The average growth of the inoculated trees of each combination was less than that of the noninoculated. Workers in Brazil (25, 27) also found sweet oranges to be susceptible on all the varieties of sour orange that they have tested as root- stocks. The possibility that a variety of sour orange may be found which is toler- ant to quick decline seems highly im- probable. 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