Bulletin STATE OF ILLINOIS DEPARTMENT OF REGISTRATION AND EDUCATION DIVISION OF THE NATURAL HISTORY SURVEY THEODORE H. PRISON, ChieJ Vol. XX BULLETIN Article I Initial Studies of American Elm Diseases in Illinois HUBERT A. HARRIS PRINTED BY AUTHORITY OF THE STATE OF ILLINOIS URBANA, ILLINOIS OCTOBER, 1932 STATE OP ILLINOIS DEPARTMENT OP REGISTRATION AND EDUCATION M. P. Walsh, Director BOARD OP NATURAL RESOURCES AND CONSERVATION M. P. Walsh, Chairman WnxiAM Trelease, Biology Henry C. Cowles, Forestry Edson S. Bastin, Geology William A. Noyes, Chemistry John W. Alvord, Engineering Charles M. Thompson, Represent- ing the President of the Univer- sity of Illinois STATE NATURAL HISTORY SURVEY DIVISION Theodore H. Prison, Chief Carroll Chouinard, Editor ScHNEPP & Barnes, Printers Springfield, III. 1932 80883— 1200 CONTENTS PAGE Introduction 1 Acknowledgments 2 Scope of Present Report 3 Previous Outbreaks of Elm Disease in Illinois 3 Character of the Present Elm Malady 6 Importance in Nurseries 9 Importance in Cities i) Methods 12 Isolation of Fungi from Diseased Wood 12 Culturing Fungi from Spores 12 Histological Technique 1 .'J Recording Data 14 General Results of Cultural Tests 15 Specific Diseases 20 Tracheomycosis Associated with Coniothyrium spp 20 Vermicularia Disease 38 Phoma Cankers 41 Sphaeropsis Dieback and Canker 4o Phomopsis Canker 4."J Verticillium Wilt 4.(!—9.5X4.4 — G.9 IX. Cephalosporium In addition to the Fitsaria discussed aljove, isolatiDns ha\e been secured of fungi which are apparently true Cephalosporin. In tlieir early growth upon potato dextrose agar these colonies are hyaline and very slimy on the surface. Later the central portion becomes light yellowish brown and ultimately dark brown or olivaceous. Short, sparse, grayish, aerial hyphae arise in the centi-al portion and to a lim- ited extent over the entire growth. \ slight, irregular zonation also occurs in the central portion of the growth. The conidia are hyaline, single-celled and usually biguttulate, though they often contain three or four small oil globules. They arise either directly from the growing mycelium or upon very short, simple conidiophores. They vary greatly in shape but are usually elongated and slightly cur\ed. b^-om a culture of the fungus grown on potato dextrose agar a range in spore size of 4.3—10.2X1.4—2.9 fx was noted. Chiefly they are about 5..SX2.2 /x. These are not all, however, identical with the Cephalosporium re- cently described by May (1931b) as the cause of a new elm disease in Ohio, Iowa, Missouri, New York and Washington, D. C, since a culture submitted to May for comparison was stated to be dififerent. Alternaria and Stemphylium Altcniaria has been obtained in the isolation trials more times than any other fungus. As its conidia and mycelium occur very comnionl\- on dead material it is believed to be saprophytic or, at most, only a weak parasite. Its mycelium does not penetrate into the woody or deeper tissues of the stem but is limited to the outer, corky tissue of the cortex. Consequently no extensive injury could result from its presence. Very young twigs which have just died and which have a very shriveled ap- pearance have been observed to be almost completely covered by the black, dusty spore masses of Alternaria. Since the mycelium of Alter- naria penerates the bark tissues to some extent, it is very difiicult to destroy by any method of surface sterilization before platings are made of samples of disease, and this readily accounts for the numerous occurrences of the fungus in platings of wood slices u])on which the bark was left intact. The fungus has never grown out of slices from which the bark had been removed previous to plating. 52 ILLINOIS NATURAL HISTORY SURVEY BULLETIN The importance of Stemphylium in its relationship to ehii diseases is also undetermined. It has not been isolated as frequently as Alter- nar'ia, though it usually occurs as a growth decidedly secondary to that causing the disease. Our observations indicate that StcinphyUiim, like Alternaria, is also essentially bark inhabiting, since its mycelium grows in the outer tissues of the bark and not in the wood. Anthostomella Anthostoinella is the only ascomycetous fungus which has been ob- tained directly in culture. It has been secured twice and in both in- stances apparently grew out of the bark. In one case it was isolated from the dead wood of a Aloline elm along with Nigrospora and Fu- sariiiiii. Living, diseased wood from the same tree yielded Vermicu- laria. In the other instance it was isolated along with Stemphylium and Alternaria from an American elm. The perithecia of Anthostoinella, shown in figure 29, are produced very abundantly in culture upon corn meal agar. They form upon the plated wood slices and the glass sides of the plate as well as upon the agar itself. They are black, flask shaped, and provided w'ith raised ostioles, and their walls are some- what roughened and carbonous. A range of 204—394 jj. has been noted in their size in culture. In figure 30 a single ascus with its eight ascospores is show'n. The asci are c y 1 i n d r i c. hyaline, eight-spored, aparaphysate, and open by an apical pore. They measure 158—180X 12 — Id ft. The ascospores are brown, one-celled and elliptic, wath a size range of 16.6—26.6X8.3 — 13.3 ,x but chiefly 19.9X10 fi. No imperfect spore form has occurred in our cultures. Cytospora Canker Dead stems and branches of elms are commonly covered by the pustules of Cytospora, but it has as yet been obtained only once as an isolation from plated wood slices. Discolored wood, which often occurs below dead areas occupied by this Fig. 30.—Photomicrograph, X 400, of an ascus and spores of Anthosto- mella, showing the regular end-to- end arrangement of the eight spores. The apical pore of the ascus is not clearly visible. HARRIS: INITIAL STUDIES OF ELM DISEASES 53 fungus, generally either gives no fungous growth in culture or yields other fungi. Cytospora apparently does not fruit readily in culture. Numerous platings have been made from single-spore isolations and an abundant vegetative growth was secured but thus far only a single pycnidium has been formed on agar. It does, however, produce its pycnidia in great abundance when grown upon sterilized elm twigs. The stromata are depressed-conical, gregarious, subepidermal, eventually erumpent, and dark gray with the disc paler, and the conidia are allantoid, 3.3—7.3X.8—2.1 ix but chiefly 6.GX1.1 ^, hyaline. The fungus is apparently C. aiiihicns Sacc. Diploma Canker Cankers bearing the pycnidia of a Diplodia have been found sev- eral times on diseased elms. The fungus has been grown in artificial culture from single spores segregated by methods previously described, but it has never been isolated directly from slices of diseased wood. In attempting to determine the proper species name for the Diplodia on elms in Illinois, it was found that, though D. Uliui had l)een described by Dearness (1916), the spores as measured by him were much smaller than the spores produced by the Illinois material. -\ portion of Dearness' type specimen was obtained for examination, and it was found that the spore measurements given in Dearness' description were in error, at least so far as the part of his material examined by us was concerned. A comparison is given in table IV of the measurements given in the description of Diplodia Ulini and of the measurements obtained by us from 100 spores of the type material and of the Illinois elm fungus. Table IV. — Spore Measurements of Diplodia Ulmi Dearn. 54 ILLINOIS NATURAL HISTORY SURVEY BULLETIN Even the minimum spore length measurements taken from the type samples exceeds the maximum spore length given in the descrip- tion. The measurements of spore width, however, nearly approximated those given in the description. From the measurements given in the tahle there is no reason to doubt that the Illinois fungus is properly designated as Diplodia Ulnti Dearness, though the description of the species requires emending as follows. Pycnidia scattered but abundant, 250—400 fi in diameter, occur- ring singly or in small groups, subglobose, immersed in the cortex, raising the epidermis in small, scarcely visible pustules which are ruptured by the minute ostiole. Conidia brown, elliptic, constricted at the septa, 19.3—29.2X8.3—11.6 //.. Mean spore size, 23.6X10.3 fi. On corn meal agar, Diplodia Ulini produces a whitish mycelial growth provided, in its early stages of development, with sparse, gray- ish, aerial hyphae. Later the mycelium darkens, becoming green and finally black. Eventually, as the agar is utilized, the entire growth assumes a charred appearance. During the early growth of a culture numerous round, definitely carbonized, cellular structures are formed which have the shape of rudimentary pycnidia, but they never de- velop an ostiole and always remain sterile, when crushed yielding only masses of oil globules. When cultures are kept until the agar sub- stratum becomes dry and hard, true pycnidia which do yield the char- acteristic brown, two-celled spores occur in aggregations at various places in the thallus. The full process of pycnidial formation has not been observed, but it is probable that the cellular structures mentioned above furnish a pseudostroma upon which the pycnidial aggregations are formed later. NiGROSPORA This fungus has been isolated thrice from diseased elms. Each time it grew from wood slices which were plated with the bark still upon them. Other fungi which we consider of more importance in causing elm diseases were obtained from the same material which yielded Nigrospora. The identification of this fungus was established for us by E. W. Mason, of the Imperial Mycological Institute, as Nigrospora sphacrica (Sacc.) Mason. Mason (1937) believes that the proper generic designation of this kind of fungus, one species of wdiich causes a severe cob rot of corn and is known in the United States as Basisporitiin gallarum, is Nigrospora. In its early development from platings upon corn meal agar the fungus appears as a hyaline growth with very little aerial mycelium. Later, as the thallus grows older, it assumes a brownish tint and be- comes covered by a rather compact cottony growth of white to slightly grayish, aerial mycelium. In old cultures the mycelium from which HARRIS: INITIAL STUDIES OF ELM DISEASES 55 conidiophores arise is very dark brown and has thicker walls than the ordinary vegetative mycelium, which is either light brown or hyaline. As ma}' be seen in figure 'SlA the spores of Nigrospora are formed in great abundance in culture. The conidiophores, figure 3 IB, may be either short and simple or rather long and several-septate. They are always swollen or enlarged at the apex, which is the point of spore attachment. The conidia are globose or nearly so,, smooth, brown when young and opaque black when mature. They usually arise singly on conidiophores though occasionally a conidiophore may branch at its apex so that two or three conidia occur there. From a culture of the fungus grown upon corn meal agar a range in spore size of 14.0—30.7 fj. was noted, though the spores were chiefly 16.6 fx. A B Fig. 31. — Xigrosijora siJhaerica. one of the unusual saprophytic fungi iso- lated from diseased elms. A.—Mycelium and spores X 60, as produced in corn meal agar culture. B.—A septate sporophore with a spore at its apex. X 550. Xylaria A Xylaria isolated from wood slices taken from a diseased elm in Illinois furnished one of our most interesting finds. Various species of the genus Xylaria are associated with and believed to be the cause of serious root rots of apple trees, maple, basswood, Cottonwood, black walnut and white and red oaks. Outward symptoms shown by infected trees are dwarfing and discoloration of the crown ; identification of the disease generally depends upon the presence of fruiting bodies of the fungus upon infected roots and in the debris at the base of the tree. It is, however, very unusual for the fungus to penetrate into the upper branches of attacked but still living trees, and so far as we are aware it has never before been isolated directly from small twigs situated toward the ends of living branches. 56 ILLINOIS NATURAL HISTORY SURVEY BULLETIN Fig. 32.—Petri-dish culture of Xylaria, showing its white, cottony my- celium and a number of black stromata in various stages of early develop- ment, after 30 days on acidified potato dextrose agar. An early stage in the development in culture of the elm Xylaria is shown in figure 32. In its early growth the mycelium is dense, cottony and pure white. Later, after having spread over the entire culture plate, the mycelium darkens, ultimately forming very black, irregular blotches either at the edge of the plate, or around or upon the wood slices from which it grew. At the same time that the black patches are formed, or even while the mycelium is still white, stalk- like structures such as are shown in figure 32 begin to grow up from the mycelium. The fungus is shown in figure 33 as it appeared six weeks after the stage pictured in figure 32. The tips of the stalks have remained white, although as they grew the lower portions gradually turned black. Also, a considerable area in the mycelium around the base of the stalks eventually turned black. The stromata, that is, the stalks, of this fungus exhibit the same very strong positive phototropism that is characteristic of most Xylariae. HARRIS: INITIAL STUDIES OF ELM DISEASES 57 Fig. 33.—Petri-dish culture of Xylaria. showing the mycelium ami stro- matal branches after two and one-half months on acidified potato dextrose agar. The positive phototropic reaction of the stromata during growth is evident. Abrupt bending- of tbe stalks with their tendency to elongate hori- zontally, shown in figure 33, is evidence of this reaction. This culture was ke])t at the far side of a room rather dimly lighted by windows on one side, first in one position and then in the opposite with refer- ence to the source of light, and the striking changes in direction of growth shown in the picture resulted from shifting the position of the culture. Small ovoid, hyaline, single-celled conidia are borne along the white upper portion of the stromata about ten days after they aj^pear. In our cultures the stromata were generally unbranched. but a few became forked near the tips. Perithecia have not developed, though swellings in the stromata have begtm to form. Other Fungi Other fungi which have been isolated from diseased elms are HcliiiiiitJiosporiuui, Rhahdnspora and Epicoccum. The first has been 58 ILLINOIS NATURAL HISTORY SURVEY BULLETIN | isolated twice and the other two once each. The conidia and sporo- I dochia of Epicocciim have been observed to occur frequently upon the ,! bark of elms, and it is not surprising that it should have been obtained ' in culture. The significance of Helminthosporiuui and Rhahdospora \ has not been determined. Bacteria ] A little over 8 per cent of the isolation tests made of diseased ; elms furnished bacterial growths. During the course of the investiga- \ tions of the dying of elms in Europe, bacteria were encountered by ' various workers. Brussofif (1925), in particular, isolated a bacterium, i which he called Micrococcus Uliiii, from trees showing symptoms of i wilt and dieback. A considerable controversy resulted, following pub- lication of his report in 1925, as the fungus Graphimn Uhiii had been ; described in 1922 by Schwarz as the cause of the same (or a very simi- ' lar) disease. During the controversy and later, as a result of further I research, Graphium Ulmi was determined to be the cause of the Dutch j elm disease and Brussoff's work received more discredit than was due ' it. Brussoff's evidence points definitely to the existence of bacterial i elm pathogens, and this is further borne out by Buisman's (Wester- i dijk en Buisman, 1929) description of Pseiidomonas lignicola as the , cause of one elm disease in Holland. ' In view of the present incomplete knowledge of elm bacteria, the isolations obtained from diseased trees appear to have a potential im- : portance that exceeds our present results with them. : Bacterial colonies have been obtained in isolation tests for the > most part when the summer temperatures were too high for fungus ] growth—at least, at such times the growth of most of our fungi was ' very slow, and some ceased growing. | Tests of the pathogenicity of several of the bacterial isolations ; were made upon elm seedlings grown in the laboratory. Inoculations were made in early July and the inoculated seedlings were kept under | observation until late October. Negative results were obtained with i all, but the conditions under which the trials were made were by no j means ideal and it is not safe to conclude that these organisms are i nonpathogenic. • INOCULATION TRIALS ; With so large a number of fungi isolated from elms, it is readily ' seen that much study and experimentation with each organism will be I necessary before the exact relationship existing between it and the I so-called "elm wilt" can be ascertained and its importance definitely j established. | As stated before, certain of the fungi that have been obtained are ' HARRIS: INITIAL STUDIES OF ELM DISEASES 59 ])rol)ably entirely saprophytic, others may be weak parasites capable ot injurious attack only after the tree is in a weakened condition, and others must certainly be highly aggressive parasites. The facts can be determined only after thorough inoculation tests of the individual organisms have been made. During the past summer inoculations were attempted under as near an approach to field conditions as possible. Thirteen of the dif- ferent fungi which had been secured from diseased elms were tested. For this purpose 15U0 young American elms grown from seedling stock were used. The trees, grown out of doors, occupied approximate- !)• a quarter of an acre. At the time the first trials were made they were between two and three feet high and quite vigorous. They had been planted in April, in rows about the same distance apart as elms set in the nursery. X'arious methods of inoculation were tried. Some of the trees were sprayed thoroughly with suspensions of spores. Other inocula- tions were made by placing fragments of plate cultures in the axils of unfolding leaves and shoots. Direct inoculation of the stems was also attempted by inserting cultures in incisions made in the bark and in holes bored in the stem. Adequate care was always taken to cover the inoculum against chance contamination and to maintain an abundance of moisture for growth of the fungus. Up to this writing these ex- periments have given no results. Leaf fall occurred before any external symptoms indicating positive reactions became manifest. It is alto- gether probable that a rather long period may be required before evi- dence of the success of the inoculations appears. In some cases another, or perhaps even two more, seasons' growth may be needed before any .symptoms of disease become evident, while in other cases the success of inoculation trials may be dependent upon the season of the year in which the trials are made. PRELIMINARY CONTROL EXPERIMENTS Tin-: problem of finding a means for controlling these elm diseases is a decidedly complex question, since there are so many separate diseases to be combated. If control measures are developed whicli are successful against one disease they may not be equally useful against the others. However, some encouragement is to be had from the fact that, at least in nursery stock, the diseases caused by Coiiiotliyriitiii and Plwiiia are of outstanding importance. They are very widely dis- tributed and abundant, and a means for controlling them will greatly reduce the losses of elm stock in nurseries. During the summer of 1931 two experiments u])on the control of elm diseases were undertaken in nurseries. Both experiments were 60 ILLINOIS NATURAL HISTORY SURVEY BULLETIN Plot I (Check) O He&lihy Trees Plot 2 Plot 3 Plot 4 (Sprayed) (Sprayed3.n(/Pra//ec/J (Pruned) I Orig/na I Tree Inpeciion E Or/ginAl Sucker Inrecft ^l^itr Tree Infer Hon ^L^fer Suc>ter Infe c lion 'ion Fig. 34.—Diagram of the nursery plots used in Experiment No. 1. Trees are represented by squares, open squares sliowing liealthy trees, black squares originally-infected trees, and crossed squares trees infected after treatments were begun. Diseased suckers growing from stumps are shown by half black squares, as indicated in the legend. HARRIS: INITIAL STUDIES OF ELM DISEASES 61 performed chiefly as preliminary tests which would serve as guides for more intensive ones to follow, though it was, of course, desired to ohtain as much information as possible relative to means of com- bating the various diseases. The two experiments were conducted with the cooperation and aid of the Swain-Nelson nursery and the Freeman nursery, both of which firms willingly and courteously contributed the use of their elm blocks to the experiments, as well as much valuable assistance in other phases of the work. Experiment No. 1 A diagram of the plot of trees used in the experiment conducted at the Swain-Nelson nursery is shown in figure 34. The trees were divided into four plots, each of which included three rows. As each row contained 100 trees, there were 300 in each plot and a total of I'^OO trees in the experiment. Plot 1 served as a check. It received no treatment ])ut indicated the amount of increase in infection taking place during the course of the experiment. The trees in Plot 2 were sprayed with Devoe's stand- ard preparation of commercial Bordeaux mixture, which contains, ac- cording to the manufacturers, a metallic copper equivalent of 13 per cent or a copper oxide equivalent of 16.35 per cent. This spray was used in a 3-3-50 concentration. Plot 3 also received a thorough spraying with the same material, and in addition all badly diseased trees, all diseased parts of trees and all dead limbs were pruned out as carefully and completely as possible. In Plot 4 only pruning was practiced, no spray being applied. This experiment was begun July "29, 1931. and the final observa- tions were made September 21, 1931. During this interval Plots 2 and 3 received two sprays, the first at the beginning of the experiment or August 1, and the other, August 24. The results obtained from this experiment are inconclusive, due in l)art to the fact that only a very small number of new infections ap- peared in the check plot during the course of the experiment and also to the fact that sometime previous to the beginning of the experiment the nursery owners had removed all trees that appeared to them to be diseased. In the rows used for the experiment there was, of course, some difference in the number of diseased trees, but this number was uniformly small, ranging from G to 9 per cent, and the possibility of infection coming in from outside sources was not restricted. For a re- liable interpretation of results from an experiment such as this, it is desirable to have a fairly large number of new infections occur in the check plot. Most of the infections in these plots previous to treat- ment were in the form of diseased shoots or suckers arising from the bases of diseased trees which previously had been cut down. 62 ILLINOIS NATURAL HISTORY SURVEY BULLETIN Results of the experiment are presented in table V, which shows \ the per cent of increase of disease above that originally present in each j plot. Tahle V. — Increase in Infection After Treatment in Experiment No. 1. i Plot HARRIS: INITIAL STUDIES OF ELM DISEASES 63 EXPKKIMKNT No. 'i The second experiment in controlling "elm wilt" was conducted at the Freeman nursery. A diagram of the trees used in this experi- ment is shown in figure 35. The trees were divided, as in the first experiment, into four plots of three rows each. There were ."JO trees in each row, making a total of 130 trees in each plot and GOO trees in the entire experiment. The treatments employed were the same as in the first experiment. and the arrangement of the treatments hy plots was as follows: I'lcjt 1 was subjected to pruning, Plot 3 was pruned and sprayed. Plot 3 was sprayed, and Plot 4 served as a check. Pruning and spra}'ing were done in the same way as the first experiment, the spray used being also a 3-3-50 concentration of Devoe's commercial Bordeaux mixture. The experiment was begun August 3, 1931, and final obser- vations were made September 23, 1931. Plots 3 and 3 were s])ra}ed August 5 and again August 35. The results obtained are given in table VI. Table VI. — Increase in Infection After Treatment in BxPEHiarENT No. 2. Plot 64 ILLINOIS NATURAL HISTORY SURVEY BULLETIN Plot I Plot 2 Plot 3 Plot -f (Pruned) (Pruned^ndSpr^^/ed) (Sprayed) (Check) a Healthy Trees Orlg/nal Infection ^ UtcrInfection Fig. 35.—Diagram of the nursery plots used in Experiment No. 2Trees are represented by squares, open squares showing healthy trees'black squares trees diseased before, and crossed squares trees diseasedsubsequent to the beginning of treatment HARRIS: INITIAL STUDIES OF ELM DISEASES 65 The fact that under conditions of heavy infection the lowest per- centages of new cases of disease occurred in the plots treated by spraying, or by s])raying and i)runing, alTords hope that the most satisfactory control may be developed along this line. A more thor- ough experimentation with spraying appears especially justifiable, since increase in number of diseased trees was 50 per cent less in the plot treated by spraying alone than it was in the check, while in the pruned and sprayed plot the increase was only about 40 per cent of that in the check. Relation to the Problem of Control An important fact to be considered in connection with the results of these two experiments is the concentration of Bordeaux mixture used. The 3-3-50 concentration was employed, not because it was thought to be strong enough to afiford maximum, or even satisfactory, protection but because no one knew definitely what concentration elm foliage could endure without injury. The trees employed in the experi- ments were of considerable commercial value to their owners. In the experiments, however, no elm suffered any injury from the spray, and we are therefore assured that a more strongly fungicidal concen- tration could be used without endangering the trees. These experiments have been concerned with preventing occur- rence of new cases of disease rather than with curing trees after they have become diseased. The facts concerning effect. ])robable time and place of infection, and spread of infection presented, especially in con- nection with the important Coniothyrhivn disease of nursery elm stock, seem to show that attempts to cure sick trees are not likely to meet with satisfactory success, especially since trimming out, or amputating, diseased parts is the only curative measure that can be applied. It is, of course, obvious that if diseased parts of trees could be cut out so as certainly to remove all internal infection and at the same time leave the tree in salable condition from the nurserymen's point of view, the problem of control would be solved, exce])t for the great cost of doing the work. Rut there is no indication at the present time that trimming performed by ordinary workmen can be depended upon to accomplish that result. There is. also, the obvious possibility of removing diseased trees from nursery blocks. This, of course, will destroy infection present at the time the removal is performed, but the healthy trees left in the blocks still remain exposed to infection from outside sources, and year b}- year a varying proportion of these trees will become diseased and have to be removed. The cumulative effect of the |)rocess will be a gradual but certain demolition of the elm block and a conse- quent loss to the nurseryman, which will depend ui)on the age at which he is able to sell his trees. Trimming, and its possible effectiveness, needs further detailed 66 ILLINOIS NATURAL HISTORY SURVEY BULLETIN study before it can be recommended. Tbe nurseryman will recognize that its use, together with the immediate removal and destruction of badly diseased trees, is a protective measure which, if taken, will eliminate many sources of infection in the elm block itself. But, in addition, protection must be secured against the introduction of infec- tion from outside sources, and it is this at which a spraying program is aimed. Once the disease establishes itself within a tree, no amount of spraying or other kind of treatment now known will afford a cure. A claim has been made by a European pharmacist (Anonymous, 1931) that a remedy has been discovered which will cure elm trees even after they have become badly diseased. The injection of this remedy into the roots, its subsequent absorption by the root system and conduction into aflfected parts of the tree supposedly effects the cure. It is very doubtful whether practical results can be had from the use of this treatment. May and Liming (1931) state that in no instance have carefully controlled experiments shown any of these injected substances to be of value. Beneficial effects may be secured by pruning, provided all infec- tion is removed. In the experimental work this was indicated by the fact that a further progress of disease was not observed, up to the time when our final check was made, in any trees treated by pruning. Proof of this mvist depend, however, upon what happens during another sea- son's growth, as it has been found in other trees treated by pruning that unless complete removal of the infection is accomplished the disease will continue through the tree and eventually kill it. 'Sieve removal of all external manifestations of disease is not sufficient. In the case of the Conioth.yrhim disease, actual infection in the interior of the wood is known to extend an indeterminate distance below the lower limits of the externally visible cankers, and even beyond visible ivood discoloration. Pruning is to be recommended, therefore, only when the number of diseased trees is so small that it can be done effectively at low cost. Cleanliness in the nursery cannot be disregarded as a practical step in lessening the number of sources of infection and the amount of infectious material. The crowded condition which conmionly prevails in nurser}' elm blocks probably influences the prevalence and severity of disease. The moist environment created by closely planted trees favors fungous growth and probably spore production, but since it is usually the top parts of the trees which first become affected, it is evident that ex- posure to sunlight and free circulation of air are necessary for infec- tion. HARRIS: INITIAL STUDIES OF ELM DISEASES 67 Some varieties of the American elm are evidently more resistant to disease than others. No experimental work has heen done in this connection hut our ohservations in nurseries where the Moline and Vase elms are cultivated show that these varieties are always quite free from disease. That these varieties may he attacked hy fungous parasites is evident, however, from the fact that we have isolated a J^crmicularia from a Moline elm. SUMMARY Results of the present investigations concerning the nature of the elm malady in Illinois show, primarily, that the so-called "elm wilt" is not a single disease hut is in reality several different diseases. These diseases manifest themselves in a numher of forms, such as twig hlights. cankers, diehacks and true wilts. They are statewide in dis- trihution and occur in nursery, parkway and lawn trees. During the past few years thousands of elms have been lost in nurseries and many valuable trees standing in cities have been killed. Most important of these diseases in Illinois is one that appears to be caused by Coiiiolhyrium. It manifests itself outwardly as a twig blight and canker and internally as a mycosis of the water conducting tissues. Other important diseases are caused by Plionia, S/^Iuicro/^sis, Vennicularia and J\vficil!iitiii. The Dutch elm disease has never been found in Illinois, though the other diseases have been popularly mistaken for it. Nearly "300 diseased trees have received laboratory examinations during the course of the investigation and over 500 isolations of fungi have been secured, among which are 20 genera and about 30 species. The pathogenicity of these various fungi is as yet not completely de- termined, though infection experiments are progressing at the present time. The problem of controlling these diseases will prove complex, since several diseases, rathel- than one. have to be considered. In jire- liminary experiments conducted in lun'series, Bordeaux mixture has given encouraging results, and i)runing when carefully practiced has proved valuable also. LITERATURE CITED AxDEKsox, Patl J., and H. W. Anderson 191?. The chestnut blight fungus and a related saprophyte. Pennsylvania Chestnut Tree Blight Commission, Bulletin No. 4. 26 pp., 6 figs. 68 ILLINOIS NATURAL HISTORY SURVEY BULLETIN Literature Cited—cont'd Anonymous 1931 La luttre contre la maladie des ormes. La Tribune Horticole, Vol. XVI, No. 755, pp. 322-323. Brooks. F. T., and G. O. Searle 1922 An investigation of some tomato diseases, pp. 192-194. The British Mycologlcal Society Transactions, Vol. VII, pp. 173-179. Bbussoff, a. 1925 Die hollandlsche Ulmenkrankheit—eine Bakterlosis. Centralblatt fiir Bacterlologle, Parasltenkunde und Infek- tlonskrankheiten, Zweite Abtellung, Vol. 63, pp. 256-257. 1 pi. BuisMAx, Christine 1932 Ceratostomella Ulnii, de geslachtelijke vorm van Gh-apMum Ulmi Schwarz. Tidschrift over Plantenzlekten 38, pp. 1-8. 3 pis. Chordat, F. 1926 Recherches experimentales sur la mutation chez les champignons, pp. 66-136. Bulletin de la Societe Botanique de Geneve, Second series, Vol. XVIII, pp. 41-144. Illus. Davis, W. K. 1930 Single spore isolation. Proceedings of the Iowa Academy of Science for 1930, Vol. XXXVII, pp. 151-159. Dearness, Johx 1916 New or noteworthy species of fungi. Mycologia, Vol. VIII, pp. 98-107. 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