HALO-BLIGHT OF OATS BY CHARLOTTE ELLIOTT Reprinted from JOURNAL OF AGRICULTURAL RESEARCH Vol. XIX, No. 4 : : : : : Washington, D. C, May 15, 1920 mi PUBLISHED BY AUTHORITY OF THB SECRETARY OF AGRICULTURE, WITH THE COOPERATION OF THB ASSOCIATION OF LAND-GRANT COLLEGES WASHINGTON » GOVERNMENT PRINTING OFFICE : 1920 LIBRARY OF CONGRESS APR 6 1922 -o^oft bj u* HALO-BLIGHT OF OATS 1 By Charlotte Elliott Scientific Assistant, Laboratory of Plant Pathology, Bureau of Plant Industry, United States Department of Agriculture INTRODUCTION The present paper is a description and discussion of a bacterial disease of oats which has been the subject of investigation by the writer for the past three years. This "halo-blight" is a disease which occurs to at least some extent each year throughout the oat-growing sections of the central and eastern States and becomes of economic importance during certain seasons when weather conditions are particularly favorable to its development. During the season of 191 8 field observations and specimens of diseased plants from widely separated sections of Wisconsin showed that this disease occurred in practically all the oat fields of the State and was responsible for the abnormal condition prevalent in the early part of that season. DESCRIPTION OF HALO-BLIGHT LESIONS The halo-blight is most conspicuous on the leaves (PI. C; 26), although it may occur on leaf sheaths and glumes (Pi. 29) . Typical well-developed lesions of the disease are oval chlorotic spots ^ to 2 cm. or more in diameter about points of infection which consist of gray-brown col- lapsed tissue measuring from 1 to several millimeters in length. The halolike border is at first only slightly lighter green than the surrounding tissue, but as it becomes older it loses more of its green color and forms oval yellow halos about the central infection areas. Lesions are first visible as light green oval spots 4 to 5 mm. in diameter with central sunken points of infection at first evident only on one side of the leaf. This center of infection increases slowly in size, penetrates 1 The greater part of this work was carried on at the University of Wisconsin during 1917 and 1918 under the direction of Prof. L. R. Jones and was continued in the Pathological Laboratory of the United States Department of Agriculture during 1918 and 1819 under the direction of Dr. Erwin F. Smith. The writer also wishes to acknowledge the courtesy of the Boston Branch of the Association of Collegiate Alumnae whose research fellowship she held during the college year of 1917-18. Journal of Agricultural Research, Vol. XIX, No. 4 Washington, D. C Ma V J 5- ^ 2 ° uc Key No. G-191 (139) 140 Journal of Agricultural Research voi.xrx, No. 4 the leaf tissue, and in a day or two forms a gray or brown dry tissue from 1 to several millimeters in diameter, evident on both sides of the leaf blade. The halolike margin spreads rapidly, becoming uniformly lighter green to yellow or showing concentric markings (PI. 26) of different shades of green and yellow. Occasionally these halolike margins are prolonged at one end into points (PI. C) from 1 to several centimeters long. They may extend as yellow streaks through the center or along the margin to the tip of the leaf, but ordinarily they appear as oval spots, measuring 1 cm. or more in diameter. Marginal infections are common, forming crescent-shaped lesions. These halo- like lesions are conspicuous and characteristic. Except in the central infection area the tissues remain turgid and have a normal appearance except for the paler yellowish color. There is no water-soaked margin about the halo as described by Wolf and Foster (io) 1 for similar lesions of the wildfire disease of tobacco, and the spots do not fall out of the leaves. Exudate does not occur in connection with the lesions. When several lesions occur on the same leaf they often coalesce and produce a general yellowing followed by a breaking across of leaf blades (PI. C) or a shriveling and drying of tips and margins. During periods of warm, dry weather yellow haloed leaf tissue loses its turgidity and color and forms oval, gray-brown dead spots which on some leaves have narrow, brown margins and on others narrow, yellow halolike margins. Very rarely the dead tissue may assume a pinkish or reddish brown color. In separate lesions the oval outline of the dead halo persists, and even when the whole leaf becomes dry and brown, the original halo outlines may still be distinguished. PREVALENCE AND GEOGRAPHICAL DISTRIBUTION Personal observations in Wisconsin and specimens of diseased plants from Ohio, Illinois, Indiana, Minnesota, Tennessee, California, and Virginia have led to the conclusion that halo-blight is present in oat fields every season, scattered lesions occurring on the lower leaves more or less throughout the season and occasionally attacking the panicles. These lesions on the lower leaves are more or less hidden by the fresher upper leaves and so escape observation. Only under particularly favorable weather conditions does the blight develop sufficiently to attract attention or to do serious damage. FIELD WORK IN 1918 During the season of 191 8 weather conditions favorable to halo-blight prevailed in Wisconsin and parts of adjoining States, causing an un- usually severe bacterial blighting. In the experimental plots, halo lesions began to appear on from 1 to 25 per cent of the young plants about the middle of May. By May 25 practically every plant showed 1 Reference is made by number (italic) to "Lirerature cited," p. 172. May 15.19*0 Halo-Blight of Oats 141 some spotting. During the last week in May and the first week in June all untreated plots looked yellowed or slightly browned when viewed from a distance. Practically every first leaf and half of the second leaves were yellowed and dead. Many leaves had yellowed, shriveled tips and margins, and single lesions were abundant on the upper leaves of many varieties. Every field about Madison showed some blighting. Usually the brown, dead leaves were easily seen from the road, and 100 per cent of infection was not at all uncommon. Some fields south of Madison showed distinct yellow spots from a yard to a rod or more in diameter. One field of oats near Monroe, Wis., visited May 29, was so badly blighted as to show from a distance a general yellowing with scattered patches of more marked yellow. Closer examination showed abundant halo lesions, every plant being infected. About 3 per cent of the plants were yellowed throughout, the outer leaves were water-soaked and dead, and some whole plants were stunted to such an extent that their recovery seemed doubtful. On the remaining 97 per cent of the plants the outer two to three leaves were collapsed and dead, and the others showed scattered halo lesions in varying stages of development. Where the blight was farther advanced the leaves were broken over and the tips shriveled and brown. Other leaves showed typical, conspicuous, isolated halo lesions which were central or marginal, covering one-half to the entire width of the leaf blade. The plants in this field showed no marked red- dening. They had been badly beaten by recent driving storms. Two other oat fields in the vicinity showed a normal stand, but the halo-blight was abundant. No plants remained uninfected, but nevertheless none were stunted or entirely yellowed, and chances for recovery were much better than for the field described above. The blight was general through- out the section about Monroe, and the two fields last mentioned probably represented the average. This yellowed condition of oat fields in this section was first evident May 26 and was reported by a number of farmers. From May 3 1 to June 2 oat fields were visited by the writer in five coun- ties of southern Wisconsin. More than 130 fields were inspected, and every one showed halo-blight varying in amount from a fraction of 1 per cent to 100 per cent, the latter being much the more common. The amount varied not only in individual fields but also conspicuously in different counties. In Jefferson County 26 fields were visited. The oats were about half grown. One-fourth of the fields showed only scattered lesions on the lower leaves— an infection of 1 per cent or less. About one-half of the fields showed a general spotting of the lower leaves on from 60 to 100 per cent of the plants. In some cases the infection was in patches from 2 to 6 feet in diameter, where every plant had all but the last one or two leaves badly spotted. A few fields showed general and heavy infection of 100 per cent of the plants. Even the upper leaves were spotted. 142 Journal of Agricultural Research vol. xix, No. 4 The lower leaves were mostly gone, but a general yellowing of the fields was not marked. Only one field was so seriously affected as to show heavy general blighting and large yellow spots 1 to 3 rods across. About 60 per cent infection of the lower leaves was typical for the fields through- out this section. In Dodge County the halo-blight was much more abundant. Of the 37 fields visited all showed at least 20 per cent infection; 5 showed light infection — spotting of the lower leaves of 20 to 50 per cent of the plants. This infection, however, was evident from the road. Over half of these fields showed heavy infection — 60 to 100 per cent — on at least the lower leaves, and yellowed spots in the fields. The plants in these yellowed spots had little normal green leaf area, and as many as 10 per cent of the plants were entirely yellow and stunted. About one-third of the fields showed 100 per cent infection of the lower leaves, the browned tips and margins showing plainly and often giving a brownish tinge to the fields. In two fields the lower two to three leaves were practically dead and the upper leaves so badly spotted as to give a general yellow color to the fields. In all fields visited in Dodge County blight was evident without a close examination and was sufficiently severe to threaten the crop if unfavorable weather conditions continued. New green leaves were just beginning to appear. In Fond du Lac County, farther north, the plants were smaller — 6 to 8 inches high — and the blight was not heavy in most fields. Seven fields showed only traces of blight on lower leaves — 1 to 30 per cent. One showed 100 per cent infection on the lower leaves and another heavy infection — 100 per cent — and a general yellowing of the field. In Columbia and Sauk Counties 10 fields showed a normal blue-green color but had 20 to 100 per cent infection on the lower leaves. Ten other fields showed yellow spots or a general yellowing of the fields. This section was second to Dodge County in the amount of bacterial blight. Reports and specimens of plants from 35 counties in Wisconsin showed that leaf lesions were general throughout the oat-growing sections of the State and that a single disease, the halo-blight, was responsible for the trouble. A similar condition was reported for the oat fields of southern Minnesota, Iowa, northern Illinois, and Indiana. For several years previous to 191 8 this bacterial blight was observed in Wisconsin oat fields, but there was never enough of it to attract par- ticular attention. The cool, cloudy days and frequent rains of the 1918 oat season proved to be just the conditions necessary to favor the develop- ment and spread of the disease. The average rainfall for May, 191 8, was 6.66 inches, or considerably more than the normal for that month and greater than for any year since 1892. At Madison there were only four clear days during the month, and at least four heavy rainstorms were accompanied by strong, driving winds especially favorable to the spread of the disease. During June the weather conditions were much May 15, 1920 Halo-Blight of Oats 143 less favorable for the spread of the bacterial blight. The total precipi- tation in Wisconsin for June was 2.31 inches, or below normal, while the average temperature increased from 58. i° F. in May to 63.9 in June. With this rise in temperature and decrease in rainfall reports came in of improved conditions in the oat fields. The new leaves which came out were unspotted, and by the last of the month all the fields had resumed a normal color and appeared to have almost completely recovered. The badly yellowed field near Monroe was visited again July 2. It had resumed a normal green color throughout with no halo lesions on the upper leaves and only scattered old lesions lower down. The stand was thin and the plants smaller than in adjoining fields. Neighboring fields were just heading out, but this field would be 10 days to 2 weeks late. Other fields showing yellow spots were reported to have resumed a normal color, but plants in spots previously yellowed were at least a week behind the others in development. This change of weather conditions in June came at an opportune time. Continued cloudy, rainy weather would undoubtedly have destroyed many plants and reduced the yield. As it was, reports for the two seasons of 1917 and 191 8 show an increase per acre for the whole State of 2.2 bushels in 1918, but this increase would undoubtedly have been more than doubled but for the presence of halo- blight. Following the unusually severe bacterial blight of the early part of the season, blasting of panicles was also unusually abundant and gen- eral throughout Wisconsin oat fields during 1918. In extreme cases as many as 25 to 50 per cent of the spikelets in a head were undeveloped. Counts of 30 panicles in a severely blighted spot gave an average of 29 spikelets per panicle and 31 per cent blasting. Counts of 30 panicles from a part of this same plot not severely halo-blighted gave an average of 34 spikelets per panicle and 20 per cent blasting. On six panicles sent in from Lincoln County the numbers of normal and blasted spikelets were as follows : Panicle No. Number of normal spikelets. Number of blasted spikelets. I 2 3 4 5 6 36 24 38 IO 34 16 2S °34 28 8 20 19 1 Top blasted. The blasted spikelets are mostly in the lower half of the panicle, but occasionally the upper half is blasted as in No. 2. All the experimental plots showed considerable blasting and numerous empty spikelets. Counts of 36 panicles of Wisconsin No. 14 oats from treated seed showed an average of 1 1 per cent of the spikelets blasted, 144 Journal of Agricultural Research vol. xix.no. 4 varying from o to 30 per cent. Counts of 40 similar panicles showed 21 per cent of the spikelets blasted. Experiments carried on during the summer of 1918 indicate that this blasting is probably not due to the bacterial disease but to the unusual meteorological conditions which favored the development and spread of the bacterial blight. BACTERIAL ISOLATION EXPERIMENTS Oat plants showing typical lesions of halo-blight were collected from fields around Madison, Wis., and from other points in the State, from Tennessee, Urbana, 111., Lafayette, Ind., Wooster, Ohio, Davis, Calif., and Arlington Farm, Va. Twenty-eight isolations were made from these lesions, and 36 isolations from halo lesions produced by inoculations in the field and greenhouse. Most of these isolations were from leaf lesions, but a few were made from lesions on glumes (PI. 29). The first isolations were made by washing the leaf tissue through 10 sterile water blanks, crushing on a sterile slide, transferring to broth, and plating from this broth suspension. Later isolations were made by dipping the tissue for a second in 95 per cent alcohol, then into 1 to 1,000 mercuric chlorid (HgCl 2 ) for one minute, washing through three sterile water blanks, and proceeding as in the earlier method. This later method proved to be more satisfactory, but a comparison of the results from both methods proved interesting. From all these isolations, with the exception of two from glumes, typi- cal white colonies of the halo organism were obtained. These appeared on potato agar in from 1 to 3 days. When the first method of isolation was used, without sterilizing the surfaces of the tissues, yellow colonies appeared on the plates with the white colonies in 25 per cent of the isolations from natural infections and in 22 per cent of the isolations from inoculation experiments. When the surfaces of the lesions were sterilized in mercuric chlorid for one minute no yellow colonies were obtained. Twelve isolations were made from natural infections, using mercuric chlorid ; and a still larger number were made from lesions due to inoculation experiments. One set of isolations was made by placing the leaf tissue in the mercuric chlorid for only 30 seconds. This leaf tissue had been sprayed with a mixed culture of yellow and white organisms. Yellow colonies appeared on the plates with the white colonies, but the yellow colonies were not nearly so numerous as on plates poured from, tissue which had not been sterilized If the tissue had remained in the mercuric chlorid for 60 seconds instead of 30 seconds no yellow colonies would have appeared. These yellow organisms appear to be surface saprophytes and do not occur within the tissues. The yellow colonies were mostly of one kind, judged by their appear- ance on agar plates — round, smooth, shining, lemon-yellow with entire margins — and they appeared on the potato agar in from one to two days. This type of colony was chosen for inoculation experiments. May is, iswo Halo-Blight of Oats 145 The white colonies of halo-producing organisms from natural infections were all alike on beef-peptone agar, but on potato agar two only of the man}/ isolations gave colonies of a slightly different character, like that designated in this paper as "stock. " On potato agar most of the isolations gave raised, umbonate colonies of a butyrous consistency with thin margins, entire or slightly undu- late. This was the usual type of colony isolated. The two varying isolations were from a leaf lesion from Lafayette, Ind., and a glume lesion on Wisconsin No. 14 oats in an experimental plot. (See PI. 31, C; 32, A.) The colonies were thicker and of an equal thickness out to the margin; the margin was slightly undulate, and the consistency of the colony was like that of boiled starch or gelatin. They gave a more rapid and abundant growth on potato agar than the common type. This second type of colony is the same as an isolation made in 191 6 by Mr. Reddy from a halo lesion on oats and kept as a stock culture at Madison, Wis. The pathogenicity of each of these 28 isolations from natural infec- tions was tested and proved by one or more inoculation experiments. Mr. Reddy's oat stock culture and isolation No. 36 (the common form) from a leaf lesion from Wooster, Ohio, were used as representatives of the two types of white colonies in the inoculation and cultural work and are designated respectively as "stock" and "36." INOCULATION EXPERIMENTS 1. Inoculation experiments were carried on at Madison, Wis., in experimental plots out of doors and in the greenhouses. The plants in the field were in various stages of development, from half gro^ra to fully headed; and those in the greenhouse were from 4 to 8 inches high. The uninjured plants were sprayed with water suspensions of organisms from agar slants 2 days to 1 week old. The greenhouse plants were then placed in damp chambers for 48 hours. Plants sprayed in the field were covered with water-proofed translucent (glassine) bags for the same length of time. Control plants were sprayed with sterile water and treated in the same manner. Oat plants of Wisconsin No. 1, Wis- consin No. 5, and Wisconsin No. 14 were used for greenhouse inocula- tions. Wisconsin No. 14 was used more often than the others because it proved to be more susceptible than any other variety. Occasionally halo lesions appeared at the end of the first 48 hours, when the plants were removed from the damp chamber; but usually none appeared until 3 to 4 days after inoculation. On young plants the lesions were often so numerous that centers of infection appeared in rows where the organ- isms had entered the stomata. The halolike discolorations around these points of sunken tissue were at first only slightly lighter green than the normal tissue but quickly became more marked until about a week after inoculation, when the tissue was a distinct yellowish green 146 Journal of Agricultural Research vol. xix.No. 4 to yellow. Numerous confluent lesions quickly killed the leaf tips and margins, which shriveled, turned brown, and died. Isolated lesions de- veloped in the same way into distinct oval spots of yellow tissue 1 cm. or more in diameter with small dead centers. Infection was always abundant on inoculated oat plants. (See PI. 27.) Cultures proved by inoculation experiments to be pathogenic were kept as stock cultures. In this way 21 such cultures were obtained. 2. Since both yellow and white colonies were isolated from leaf sec- tions showing halo lesions, inoculations were made with pure cultures of each and also with mixed cultures of yellow and white colonies for comparison with inoculation work done by Thomas F. Manns (3, p. 107, PL I). In 25 inoculation experiments pure cultures of the white halo organisms produced abundant and typical infections. In 13 tests, pure cultures of the yellow organisms produced no lesions whatsoever. Twelve sets of inoculations were made with mixed cultures by com- bining the 2 white halo organisms, No. 36 and stock, with 4 different isolations of yellow organisms. Isolation 39a from a leaf lesion from Urbana, 111., was the yellow organism most often used. Separate pure cultures of yellow and white organisms were used for control inocula- tions. The cultures were mixed just before the inoculations were made for the reason that long-continued attempts to grow mixed cultures in broth or on various agars were not successful. 1 In the 12 inoculation tests with the yellow and white mixed cultures typical halo infections were produced, but the lesions were only one-half to three-fourths as abundant as on plants inoculated with pure cultures of the white organisms. The development of lesions from mixed cultures was also somewhat retarded, the infections being evident from one to two days later than tnose obtained from the pure white cultures. These inoculation experiments showed plainly that the white organism alone is responsible for the production of the halo lesions while the yellow organisms used are neither parasites nor favorable to parasitism. 3. In June, 191 8, field inoculations were made on the following 13 Wisconsin varieties: Wisconsin No. 1, 3, 4, 5, 7, 13, 14, 15, 22, 25, 49, 52, and 62. The plants were just beginning to head out, and the experi- ment was carried on to test the pathogenicity of the white organisms on mature leaves and on panicles, and the effects of possible lesions upon the development of the panicles, spikelets, and kernels. Water suspen- sions of the halo organisms were sprayed into unopened sheaths upon uninjured bundles of plants, the tops of which were drawn together and tied close so as to be covered with bags, and upon bundles of plants 1 For two months mixed cultures of white and yellow organisms were grown on potato agar and in +10 beef-peptone broth. Plates poured from these cultures when they were 5 days old showed a few white and many yellow organisms in the broth cultures and about equal numbers of yellow and white on agar. Plates poured from these cultures ?% weeks later showed no growth of either white or yellow colonies from the agar and showed pure cultures of the yellow organisms from the broth. On the contrary, separate pure cultures of the same organisms held for the same time in these media and under the same conditions gave abundant and characteristic colonies on the plates poured. May is, 19=0 Halo-Blight of Oats 147 injured with a scalpel or drawn between the fingers to rub off the bloom. Bundles of control plants were treated in the same manner and sprayed with sterile water. All inoculated and control plants were covered with glassine bags for 48 hours, as stated above. Characteristic halo lesions appeared on all the varieties inoculated except Wisconsin No. 4. Only uninjured plants of this variety were inoculated. Five other varieties (No. 22, 25, 49, 52, and 62) showed no lesions on uninjured plants, but all varieties showed fairly abundant spotting of leaves and sheaths of plants which had had the bloom removed or had been cut with a scalpel. Some of these leaves were almost entirely yellowed with lesions. On 6 varieties lesions appeared on uninjured plants, but the lesions were not nearly so abundant as on injured leaves and panicles. Wisconsin No. 7 was the only variety in which the panicles were entirely out of the sheaths. In this variety every spikelet of the injured panicles showed halo lesions which stood out as oval yellow spots on the glumes. About half of the spikelets in these panicles were not filled out. Spikelets of untreated panicles of the same variety were also poorly filled out. Under favorable conditions the panicles appear to be just as susceptible to halo- blight as the leaves. Wisconsin No. 14 also showed heavy spotting of injured panicles. Uninjured spikelets of two varieties were halo-spotted when the suspension was sprayed into the unopened sheath. Though none of the controls showed any halo lesions, both water- sprayed controls and inoculated plants showed considerable sterility, amounting to from one-fifth to one-half of the spikelets in a panicle. Untreated heads of the same varieties and in the same plots showed either no sterility at all or only traces at the base of the panicle. This sterility was particularly abundant when either the water suspension or sterile water was sprayed into unopened sheaths or sheaths just opening at the top. The fact that both controls and inoculated plants showed the same amounts of sterility would indicate that the sterility was not due to the effects of the organism. Excessive moisture around the developing spikelets while these were still inclosed within the sheath offers the most plausible explanation for this sterility. In the same way heavy rains at the time oat fields are heading out probably account for the sterility commonly observed in oat fields. This set of field inoculations has led to the following conclusions : 1. Leaves and panicles of oat plants approaching maturity are sus- ceptible to halo infection under favorable conditions. 2. Infection takes place more readily on injured than on uninjured parts of the plants. 3. Some varieties are more susceptible to infection than others. Green- house inoculations on young plants also led to this conclusion. 4. Although both natural and artificial halo infection may occur on heads, these infections are not responsible for the blasting of oat heads. Sterility is due probably to physiological rather than pathological con- ditions. 148 Journal of Agricultural Research voi.xix.No. 4 CULTURAL CHARACTERS I. — STOCK HALO ORGANISM Morphology. — The organism is a motile rod with rounded ends (PI. 34, B, E), sometimes occurring singly or paired, but usually in short to long chains (PI. 34, C). Organisms grown on beef-peptone agar and potato agar and stained with Ribbert's capsule stain, gentian violet, and carbol fuchsin measure from 1 to 4 fx in length and from 0.4 to 0.8 n in width, with an average of 0.65 by 2.3 /x. Stained by the Van Ermengen method from 24-hour cultures on beef-peptone agar, the organism shows from one to several polar flagella about the same length as the organism or only a little longer (PI. 34, E). No spores have been observed, although special staining methods with hot carbol fuchsin and methylene blue were used. Capsules are formed on both potato and beef-peptone agar and were stained with Ribbert's capsule stain (PI. 34, D). Compact pseudozoogloeae are not formed — that is, there is little or no viscidity. No branched forms have been observed. Nutrient broth. — Beef-peptone bouillon ( + 10) shows light clouding in 24 hours at 25 C. In 5 days there is moderate uniform clouding, and a flocculent white film or pellicle forms on the surface and falls to the bottom of the tube in small white flakes. On further shaking the flakes disappear. In older cultures there may be no pellicle but merely a slight ring around the surface. The clouding is never very heavy, and the thin surface film soon disappears. The medium is gradually changed in color until at the end of 60 days it is a deep amber brown. 1 The odor of decay is distinct with more or less of the penetrating smell of ammonia. The sediment in cultures from recent isolations is loosely flocculent. There was a somewhat viscid swirl in some of the old broths containing sodium chlorid. Rectangular crystals form at the surface. Broth plus absolute alcohol. — To 10 cc. of +15 beef- peptone bouillon absolute alcohol was added to make 4, 5, 6, and 7 per cent. There was heavy clouding in 4 and 5 per cent, moderate clouding in 6 per cent, and slight clouding in two out of three tubes of 7 per cent. Agar stroke. — On + 10 beef-peptone agar slants growth in 2 days is moderate, flat, undulate, white, shining, translucent, slightly contoured, butyrous. The medium is slightly browned. There is a slight odor of decay. On potato-dextrose agar slants the growth in 2 days is abundant, slightly undulate, raised, glistening, smooth, opaque, white, of gelatinous consistency (PI. 30, B, b). The medium is unchanged and there is no odor. Agar colonies. — (1) On poured plates of +10 2 beef-peptone agar from + 10 broth cultures, colonies appear after 30 hours at 25 C. as tiny translucent dots. When 2 days old the colonies are 1 to 2 mm. in diame- 1 Ridgway, Robert, color standards and color nomenclature. 43 p., 53 col. pi. Washington, D.C. 1912. 8 Fuller's scale. May 15, i 9 :o Halo-Blight of Oats 149 ter, white, smooth, shining, round, with a denser center. When 11 days old they are more or less irregularly circular, 5 to 10 mm. in diameter in thin-sown plates, and flat with slightly raised margins. Microscopically, -.nth low powers, the internal structure is filamentous, the margin consist- ing of folded parallel strands or chains. The margin is undulate. Deep colonies are lens-shaped and opaque. The medium may be slightly browned. The markings in Plate 31, A, B, E, F, are characteristic of these colonies under a hand lens, and they do not disappear as the colonies grow older. Very similar markings appear in young colonies of some of the softrot organisms. A half dozen of these softrot organisms tried on oats have not produced an}' halo lesions. (2) On plates of potato-dextrose agar the colonies grow more rapidly. They are raised, white, shining, opaque, with only slightly undulate or entire margins, and of the same gelatinous consistency described above (Pl. 3 i,C; 32, A.) Gelatin colonies. — Growth slow, circular, crateriform, margins en- tire to undulate with folded strands (PI. 31, G), liquefaction saucer-shaped, rather slow. Gelatin stab. — Growth on + 10 peptone gelatin at 22 C. is slow, best at the top, with only a slight filiform growth along line of stab, liquefaction crateriform. At the end of 2 days a small pit is formed at the surface 3 mm. deep. At the end of 6 days liquefaction extends two-thirds of the way across a 20-mm. tube, and the pit is 10 mm. deep. At the end of 18 days liquefaction covers the surface to a depth of 14 mm. At the end of 60 days (at 20 ) the tube is only half liquefied. Potato cylinders. — At 25 C. there is moderate growth in 24 hours and slight darkening of the medium. At the end of 4 days growth is abundant, flat, smooth, glistening, butyrous to slimy, of a cream color, and the medium is a uniform dark gray color. At the end of 20 days there is a decided odor of decay. There is feeble diastasic action on starch. Smith's potato starch jelly (5). — Growth moderate, diastasic action feeble, medium stained a light bluish green. Traces of dextrin. Starch agar. — To melted tubes of +10 beef-peptone agar sterile potato starch was added and plates poured. Tests with iodin showed no diastasic action. Litmus sugar agars. — One per cent lactose, maltose, dextrose, sac- charose, and galactose were used in beef-peptone litmus agar. Change of medium to a bright red showed considerable acid production with dex- trose and galactose; with saccharose there was less acid produced; and with lactose and maltose there was no evidence of acid production. Re- duction of litmus took place to a slight extent under the streaks with dextrose and maltose. Milk. — In fresh isolations a soft curd forms in from 5 to 7 days, fol- lowed by slow peptonization of the curd, which is completed in from 5 150 Journal of Agricultural Research vol. xix,No. 4 to 6 weeks. In old isolations curd usually is absent. The medium does not become viscid or slimy. The liquid at the top of the tube is some- times a yellowish green but more often brown. This brown color may be confined to a surface layer a few millimeters deep or may extend through- out the liquid medium. In old tubes the fluid is coffee-colored. It is unlike any color in Ridgway, but is somewhat like his moss brown. Litmus milk. — At room temperature the medium begins to turn slightly blue in 2 days, beginning at the top; and in 5 or 6 days it is fre- quently stratiform, being deepest blue at the top. Reduction begins at the end of a week, the tubes becoming cream-colored throughout, and clearing at the top or showing reduction only at the bottom of the tube for a depth of 1 cm. or more. There is no curdling, and clearing is com- plete in 2 weeks. At the end of 2 months the tubes are a deep blue-black and sometimes of a gelatinous consistency. At no time is there any red- dening. Methylene blue in milk. — In fresh isolations reduction begins in 3 days and is completed in 7 days, except for a rim of blue at the top 1 mm. deep. Curdling takes place in 1 week; peptonization begins soon after and is completed in 5 weeks, the clear liquid being yellowish to neuvider green, especially toward the top. Cohn's solution. — Growth is very slight, appearing in 24 hours and increasing slightly the second day. In a week clearing begins, and at the end of 3 or 4 weeks there is no clouding and only a little precipitate. Nonfluorescent. No crystals. Uschinsky's solution. — The medium shows light clouding in 24 hours. In 48 hours a thin flocculent white film has formed over the surface and shakes down in fine particles. In 4 days there is moderate clouding, a slight surface film, and the medium is a pale turtle green. In 2 weeks a heavy white rim has formed around the surface of the liquid. When the cultures are 6 weeks old there is considerable white precipitate — fluid, not viscid — and slides stained in carbol fuchsin show a network of long chains (PI. 34, C). Fluorescence persists in old cultures. Fermi's solution. — Light clouding occurs in 24 hours. In 4 days the're is moderate clouding and a delicate surface film which shakes down in fine flocculent particles. In 4 days as much growth as in Uschinsky. In 2 weeks the clouding is heavy, the medium has a greenish tinge, and there is a heavy white surface pellicle 2 to 3 mm. deep, which shakes down in strings of fine white particles. There is considerable white pre- cipitate — a heavy growth. In 3 weeks the white surface pellicle and the precipitate become cream-colored. No chains are formed. Greening first visible after about 2 weeks. At end of a month surface pellicle and precipitate tan color. Clouding and pellicle twice as abundant as in Uschinsky. LoEFFLER's blood serum. — Growth moderate, filiform to slightly undulate, flat, glistening, smooth, medium slightly browned beneath the streak. No liquefaction, not even after 2 months. May is, 1920 Halo-Blight of Oats 151 Soyka's rice medium. — The growth and medium in all cases except one were cream colored. A culture marked "stock b" turned the medium a buff-pink. Nutrient broth plus carbon compounds. — To tubes of +10 beef- peptone bouillon 1 per cent asparagin was added and to other tubes 1 per cent asparagin plus 1 per cent dextrose were added. The growth was equally good in both kinds of media. The organisms seem to obtain their carbon as readily from asparagin as from dextrose. Indol production. — Feeble or absent in beef-peptone bouillon or 1 per cent peptone water containing 0.5 disodium phosphate and 0.1 mag- nesium sulphate. Hydrogen sulphid. — Hydrogen sulphid is not produced. Lead ace- tate paper suspended over broth cultures is not blackened, and the medium is unchanged when streaks are made on lead carbonate agar plates. Ammonia production. — Moderate. Made tests with Nessler's reagent. Nitrate in nitrate broth. — No gas is produced in fermentation tubes. Nitrates are not reduced. Tests were made at the end of 9 days and at the end of 2 months. Temperature relations. — The maximum temperature for growth, tested on beef broth and on agar and potato, is 31 ° C. The minimum temperature for growth is below o°. Tubes surrounded with ice showed clouding. The optimum temperature for growth is 24 to 25 . The thermal death point is between 47 and 48 . Moisture relations. — The organisms are very readily killed by drying. Smears were made from 5-day-old broth cultures to sterile cover glasses and placed in sterile Petri dishes. Pieces of these cover glasses transferred to sterile bouillon after 3 hours showed growth in all. All were dead at end of 24 hours. In a repetition, transfers after 6 hours gave no growth. Fermentation tests: (i) Potato juice. — Undiluted potato juice was expressed afte; passing the pared tubers through a meat grinder. Moderate clouding in open arm of fermentation tubes. No growth in closed arm and no gas. (2) Milk. — At the end of a week the milk at the open end had cleared without evident curdling. Two days later the milk in the closed end had curdled. This curd was gradually peptonized, about a third of it remain- ing at the end of 2 months. The cleared liquid in the open arm was browned — a chestnut to auburn brown at the surface and gradually changing to a lighter shade through the open arm and a third of the way up the closed arm. No gas was formed. (3) Carbon compounds. — Tests were made in the fermentation tubes with 2 per cent solutions of dextrose, saccharose, maltose, lactose, man- nit, glycerin, and levulose in 2 per cent water solutions of Difco's and Witte's peptones. Bacillus coli Escherich was used as a control and produced gas in the closed arm. The oat organism produced no gas and 152 Journal of Agricultural Research vol. xix, No. 4 did not grow in the closed arms of tubes containing maltose or lactose. In tubes containing saccharose, glycerin, and mannit there was growth at first only in the open arm, with a sharp line of demarcation between open and closed arms. At the end of a week clouding began to appear in the closed arms of tubes containing these three substances. In 3 weeks there was light clouding throughout the closed arms in saccharose, moderate clouding throughout the closed arms in mannit, and in glycerin heavy clouding to within an inch of the top of the closed arm with light clouding on up to the top. In a later test there was again light clouding throughout the closed arms of tubes containing saccharose. In two later tests a moderate clouding appeared in the closed arms of tubes of saccharose and dextrose in from 4 to 7 days. In a later test of mannit and glycerin there was no ciouding in the closed arm. Tests for ammonia with Nessler's reagent gave a positive reaction in solutions of maltose, saccharose, mannit, glycerin, and lactose, but only traces of ammonia or negative reactions in cultures containing dextrose. Titrations with phenolphthalein as an indicator show a higher total titrable acidity in the cultures than in the controls in saccharose and dextrose. These solutions were also acid to litmus as compared with controls in two later experiments. The hydrogen-ion concentrations, determined after about 6 weeks by the colorimetric method, were as follows: Control, P H = 4.8; dextrose, P H = 4.8; maltose, P H =7; saccharose, P H = 6.4; and lactose, P H =4-8. The organisms grew best in saccharose, levulose, and dextrose, showing heavy growth in the open arm and slight to moderate growth in the closed arm. This organism is evidently a facultative anaerobe when certain sugars are available. Toleration of acids. — Transfers were made to tubes of +10 beef- peptone broth containing 0.1 per cent and 0.2 per cent of citric, tartaric, and malic acids. There was good growth in 0.1 per cent of each acid but only slight growth or none at all in 0.2 per cent. Toleration of sodium chlorid. — Neutral beef-peptone bouillon con- taining, respectively, 2, 3, 4, 5, 6, and 7 per cent of sodium chlorid was inoculated from potato agar slants. There was slight clouding of 2 per cent after 3 days. None of the stronger solutions clouded, but slides made from a stringy white precipitate and stained with carbol fuchsin showed that long chains of cells had been formed in all strengths of sodium chlorid. A second test was made, using neutral broth with 0.5, 1, 1.5, 2, 3, and 4 per cent solutions of sodium chlorid and inoculating from broth cultures. There was slight clouding in 1 per cent at the end of 2 days, slight clouding in 1.5 per cent at the end of 3 days, moderate clouding in 1.5 per cent at the end of 5 days. At the end of 7 days there was slight clouding in 2 per cent and moderate clouding and a stringy swirl of pre- cipitate in 2 per cent at the end of 19 days. Stained slides of precipitate from 1.5 and 2 per cent solutions showed a network of long chains. In the second test there was no growth in solutions of more than 2 per cent. May is, 1920 Halo-Blight of Oats 1 53 Optimum reaction and Toleration limits. — Beef-peptone bouillon was adjusted to each of the following reactions with sodium hydroxid and hydrochloric acid +20, +15, +10, +5,0, —5, —6, —13, —16, and — 22. These were uniformly inoculated from broth cultures and kept at 24 C. At the end of 24 hours there was light clouding in —5,0, +5, and +10. Subsequent clouding occurred in —6 and +15. A stringy precipitate formed in — 13, and on — 15 a thin surface film developed and the medium was slightly darkened. At the end of 48 hours the clouding in + 5 was slightly heavier than in +10, and the flocculent surface film slightly heavier. Clearing began in 3 weeks. At that time +10 was browned, +15 slightly deeper brown, and +5 and —5 showed a greenish tinge. The optimum reaction for growth is, therefore, + 5 Fuller's scale, although +10 and +15 are also favorable reactions. In later tests the limits of growth on agar were +27 and — 17, and in bouillon +27 and — 18, when the agar was reinoculated from an alkaline culture. Volatile acids. — Tests for volatile acids were negative. Cultures were grown in tap water containing 1 per cent Witte's peptone and 1 per cent dextrose. The steam from these cultures gave an alkaline reaction to litmus although the liquid was acid to litmus. Freezing. — Six plates were poured in +15 agar from 24-hour +15 broth cultures. This 24-hour culture was exposed for 1 hour in salt and crushed ice and then six more plates were poured. Eighty-seven per cent of the organisms were killed by this treatment. Effect OF sunlight. — The organism is sensitive to sunlight; 80 per cent were killed by 15 minutes' exposure on ice in thinly sown beef- peptone agar plates. Vitality on culture media. — Typical colonies of this organism have been obtained from + 10 beef-peptone agar slants which have stood for 11 months and from broth cultures 10 months old. These were tested by inoculation on young oat plants and gave abundant and typical halo lesions. Loss of virulence. — Loss of virulence on culture media has not been observed in cultures carried for more than 3 years. Group number. 1 — 221.2323023. The name Bacterium coronafaciens , n. sp., is suggested for this organism. TECHNICAL DESCRIPTION Bacterium coronafaciens, n. sp. A motile rod with rounded ends and polar flagella; single, in pairs or long chains, average measurement 2.3 by 0.65 n; no spores, zoogloea, or involution forms; capsules are formed; slightly facultative anaerobic. On nutrient agar colonies are white, round becoming irregularly circular, fiat with slightly raised margins, surface smooth or slightly contoured; deep colonies are lens-shaped and opaque. Its proteolytic 1 Society of American Bacteriologists, descriptive chart. Indorsed by the society for general use at the annual meeting Dec. 31, 1914. Prepared by the committee on revision of chart identification of bacterial species. 154 Journal of Agricultural Research vol. xix, No. 4 power is moderate; gelatin is liquefied slowly, beginning in 2 days and not complete in 60 days; reduction of litmus occurs in milk, and the casein is digested without curdling; milk curdles in 5 days, and peptonization is completed in 5 weeks. No acid is produced in milk. Oxidations of proteins are incomplete; ammonia is produced ; hydrogen sulphid, gas, and indol are not produced. Nitrates are not reduced. There is slight diastasic action on potato cylinders. Good growth in Uschinsky's solution and in Fermi's solution. Growth in Cohn's solution is scanty. Maximum tempera- ture for growth is 31 C, minimum below o°, optimum 24 to 25 , thermal death point between 47 ° and 48 . Tolerates sodium hydroxid to —18 Fuller's scale and hydro- chloric acid to +27. The optimum reaction for growth is +5 Fuller's scale. Gram- negative, not acid-fast, stains readily and uniformly with gentian violet and methy- lene blue. Stains more or less irregularly with carbol fuchsin (often polar staining). Sensitive to drying; 87 per cent killed by freezing, 80 per cent killed by sunlight. Vitality on culture media long. Pathogenic on varieties of cultivated oats and to a slight degree on wheat, rye, and barley, producing oval halolike lesions of chlorotic tissue surrounding dead brown centers of infection. Beef-peptone agar and beef bouillon are favorable media for prolonged growth. Growth on potato agar brings out more distinguishing characteristics. II. — ISOLATION NO. 36 This isolation was made from a halo lesion on oats obtained from Wooster, Ohio, in June, 191 7. It has the same group number as the stock halo organism just described but differs from it in the characters mentioned below. The differences, though not very marked, seem to be fairly constant, while the lesions from which the cultures were isolated and which they produce in inoculation work can not be distinguished. The stock organism seems to be slightly more virulent. Morphology. — The organism occurs singly or in twos but seldom in long chains (Pi. 34, A). Stained by Ribbert's capsule stain it measures from 1.1 to 3 ix in length and from 0.5 to 0.8 ix, in width, not including the capsule, with an average measurement of 0.66 by 2.1 /*. BEEF agar plates. — On +10 beef-peptone agar, the surface colonies remain round, and the margin tends to remain entire (Pi. 31, D). Potato-dextrose agar stroke. — Two-day-old slants from broth show moderate flatter growth, which is filiform and dull, with more or less wrinkling on the surface. The growth is somewhat translucent and of a butyrous to slightly membranous consistency (Pi. 30, B, a). Gelatin stab. — Liquefaction is more rapid, being complete in 40 days. Toleration of sodium chlorid. — Same as stock, but slides from a 2 per cent solution stained with carbol fuchsin show only a few scattered short chains. Litmus milk. — Litmus is not reduced. Methylene blue. — Digestion of casein a little slower than with stock. Uschinsky's solution. — No chains on slide stained with carbol fuchsin. Cohn's solution. — Clouding heavier than with stock. Crystals are formed on the sides of the tubes. Starch agar. — The organism showed a feeble diastasic action on Starch. May is, 1920 Halo-Blight of Oats 155 Temperature; relations. — Thermal death point is between 47 and 48°C. Strain 36 usually gives a greenish tinge to bouillon cultures, which in old cultures contrasts strongly with the brown of old "stock" cultures. On ordinary beef-peptone agar the two strains can not be distinguished but on potato-dextrose agar there is considerable difference in amount of growth, and they are noticeably different in consistency. The most important differences perhaps are in size and in nonformation of chains. The rods of No. 36 are shorter and plumper. They seem to be two strains of the same organism. III. — YELLOW ORGANISM Morphology. — The organism is a motile rod with rounded ends and one to several polar flagella. It occurs singly or in short chains. When grown for 24 hours on beef-peptone agar and stained by the Duckwall modification of Pitfield method, it has an average measurement of 3.5 by 1.4 ix, varying in length from 2.3 to 3.7 /*, and in width from 0.98 to 2.1 n. No spores have been found. Beef-peptone agar plates. — Colonies appear after 24 hours on + 10 beef -peptone agar; in 2 days they measure 2 mm. in diameter and are a translucent light yellow. When a week old, surface colonies are circular, 4 to 5 mm. in diameter, raised, smooth, lemon-yellow, with entire translucent margins. Microscopically the internal structure is finely granular. Deep colonies are lens-shaped and opaque. Beef-peptone agar stroke. — Growth in two days is moderate, fili- form, flat, glistening, slightly contoured, translucent, light orange- yellow, with a faint odor. Consistency is butyrous, and medium is unchanged (Pi. 30, B, c). The organism lives at least three or four months on beef -peptone agar. Potato agar stroke. — In two days the growth is abundant, filiform, fiat, spreading, glistening, smooth, opaque, light orange-yellow. The medium is unchanged, and the consistency butyrous. Gelatin stabs. — At 22 ° C. growth in + 10 nutrient peptone gelatin is moderate. The liquefaction at first is saccate along the stab and later stratiform. Liquefaction is completed in 40 days. The surface growth has a pinkish tinge, but the precipitate is yellow. Beee-peptone broth. — There is moderate clouding in +10 beef- peptone broth in 24 hours at 25 C, very heavy clouding in 48 hours, and a slight flocculent surface growth. In 3 days there is a heavy mem- branous pellicle which breaks up when shaken and sinks to the bottoii of the tube. The precipitate is abundant and finely granular. Clearing begins in about 2 weeks. Toleration of sodium chlorid. — Tables of neutral beef-peptone bouil- lon containing respectively 2, 3, 4, 5, 6, and 7 per cent of sodium chlorid were inoculated from potato agar slants. In 24 hours there was 156 Journal of Agricultural Research voi.xix,No. 4 clouding in 2,3,4, and 5 per cent solutions. In 3 days there was a very slight clouding in 6 and 7 per cent solutions. A stringy yellow precipi- tate formed in the 4, 5, 6, and 7 per cent. Slides made from 2 and 3 per cent solutions and stained with carbol fuchsin showed long chains of cells. There were no long chains in the 4, 5, 6, and 7 per cent. In a second test a delicate pink surface film, not previously observed, formed in 0.5, 1, 1.5, 2, 3, and 4 per cent solutions; and a pink stringy precipi- tate formed in 2 and 3 per cent, becoming a brick red in 4 per cent. Potato cylinders. — At 25 ° C. there was slight growth in 24 hours and a slight graying of the medium. In 4 days there was abundant yellow growth, and the medium had become slightly browned. Growth was filiform, flat, raised, glistening, somewhat contoured, orange-yellow to red on top. There was no odor, and the consistency was butyrous. There was no action on the starch. Milk. — Milk titrating +18 on Fuller's scale was inoculated from 9- day-old potato agar slants. A slight yellow surface film was formed in 2 days. At the end of 1 week yellow precipitate was evident. Curdling began in 3 weeks. There was a slight separation of curd and whey at the end of 2 months. The solid curd gradually dried down without any evidence of peptonization. Litmus milk. — Complete reduction occurs in 24 hours, leaving the medium cream -colored. Shaking tended to restore the color. After about a week some of the reduced tubes were steamed, whereupon the original lavender color returned. Curdling occurred in 3 weeks. There was no evidence of digestion at the end of 2 months. Methylene blue in milk. — Reduction takes place in 24 hours. In 3 weeks there is curdling and the blue color begins to return at the tops of the tubes. No petonization. Uschinsky's solution. — There is moderate clouding in 24 hours at 25 C. and a membranous surface film. At the end of 2 days there is a fairly heavy light yellow surface film. In 4 days there is heavy clouding and a heavy surface film and yellow precipitate. Slides stained with carbol fuchsin show many short chains. Fermi's solution. — There is moderate clouding in 24 hours at 25 C. In 2 days there is a fairly heavy light yellow surface film. In 4 days the clouding is heavy and there is a heavy orange-colored surface film 2 mm. thick. At the end of a week this pellicle is 4 mm. thick. Clearing begins in 2 weeks, and yellow strands extend from the heavy pellicle to the bottom of the tube. At the end of 3 weeks the pellicle is 1 cm. thick. In 4 weeks the medium has a greenish tinge. No chains were observed on slides stained with carbol fuchsin. Cohn's solution. — There is a slight clouding at the end of 2 days at 25 C At the end of 4 days the clouding is still very light, and there is just a trace of surface growth. Rhomboid crystals are formed on the tube above the liquid. Growth is very slight in comparison with that in Fermi's solution. May is, 1920 Halo-Blight of Oats 157 Blood serum . — Growth was moderate, filiform, slightly raised, orange- yellow, smooth, shining. In 2 weeks the center of the growth became red, but the author was unable to verify this change in 191 9. The medium was unchanged. Litmus sugar agars. — In 24 hours there is a slight reddening of litmus dextrose agar and in 3 days reduction has begun in the lower end of the tubes, the upper two-thirds being rose red. Litmus-lactose and litmus-maltose agar show reduction in the lower ends of the tubes in 3 days. These tubes are red through the center and blue at the top. At the end of a week all agars are colorless at the bottom of the tubes, red in the center, and blue toward the top. Growth is abundant. At the end of 2 weeks the colony begins to turn red. Starch agar. — There is no diastasic action on starch. Indol. — Indol production is feeble. Nitrate bouillon. — No gas is produced in fermentation tubes. Nitrates are not reduced. Ammonia. — Ammonia production is moderate. Hydrogen sulphid. — No hydrogen sulphid is produced. Tests were made with lead-acetate paper over broth and with lead-carbonate agar. Optimum reaction and toleration limits. — By the use of sodium hydroxid and hydrochloric acid, using phenolphthalein as indicator, beef-peptone bouillon was adjusted to each of the following reactions: + 25, +20, +15, +5, o, —5, —6, —13, —15, and —22. These were uni-' formly inoculated from broth cultures and kept at 24 C. In 24 hours there was clouding in all except +20 and +25. At the end of 3 days there was clouding in all except + 25. The clouding in + 20 wa"s slight. At the end of 1 week there was no growth in +25, light clouding in + 20, — 15, and — 22, and heavy clouding in all the other reactions, with precipitation and surface growth. In 3 weeks there was clearing in — 15 and —22, but a viscid yellow precipitate. There was never any growth in +25. The optimum reaction for growth is +5 Fuller's scale. Gas formation and aerobism. — Tests were made in fermentation tubes in the presence of the following carbon compounds: dextrose, sac- charose, lactose, maltose, mannit, and glycerin. A 2 per cent solution of each was made in a 2 per cent water solution of Dif co peptone. Bacil- lus coli Escherich was used as a control and produced gas in each solution. No gas was produced by the yellow organism. There was clouding in the open arm of all tubes in 2 days, the heaviest growth being in saccha- rose and maltose. In 3 days clouding began in the closed arm of tubes containing saccharose and mannit. At the end of a week there was clouding in the closed arm of all tubes — heavy in glycerin and mannit, light in dextrose, and moderate in the others. Tests for ammonia with Nessler's reagent gave a positive reaction in all sugars — slight in glycerin, and moderate in the others. Titrations with phenolphthalein as indica- tor showed no acid production. The hydrogen-ion concentrations were 158 Journal of Agricultural Research vol. xix.no. 4 determined by the colorimetric method at the end of 6 weeks. The P H for dextrose was for maltose 5 to 5.2, for saccharose 4.6, for lactrose 7, for glycerin 4.8, and for mannit 6. Controls and Bacillus coli Escherich showed a P H of 4.8 throughout. Temperature relations. — The maximum temperature for growth is above 38 C. The minimum temperature for growth is 3 . The optimum temperature for growth is 24 to 25 . The thermal death point is 48 to 50 . Tests were made by the same methods as those used for the halo organisms. Vitality on culture media. — The organism lives for 2 months on beef-peptone agar at room temperatures. It is nonpathogenic. Group number. — The group number is 221.3333533, according to the descriptive chart of the Society of American Bacteriologists. TECHINAL DESCRIPTION A motile rod, with rounded ends, one polar flagellum or several, single or occasion- ally in short chains; average measurement 3.5 by 1.4 a*; no spores, pseudozoogloeae, or involution forms; facultative anaerobic. On beef- peptone agar the colonies are round, raised, smooth, lemon-yellow with entire translucent margins; deep colonies, lens-shaped and opaque. Liquefaction of gelatin begins in 2 days and is complete in 40 days. There is reduction in litmus milk in 24 hours and delayed curdling with- out subsequent peptonization ; milk is curdled in 3 weeks without subsequent peptoni- zation; ammonia production moderate; indol production feeble; does not produce hydrogen sulphid or other gas; no diastasic action on starch; grows moderately in Uschinsky's solution, and very copiously in Fermi's solution. Growth slight in Cohn's solution. Maximum temperature for growth is above 38 C, minimum 3 , optimum 24 to 25 , thermal death point 48 to 50 . Tolerates sodium hydroxid to below —22 Fuller's scale, and hydrochloric acid to +20 Fuller's scale. The opti- mum reaction for growth is +5 Fuller's scale. Gram-negative; not acid-fast; stains readily with carbol fuchsin, gentian violet, and methylene blue. Nonpathogenic to oats. OVERWINTERING AND DISSEMINATION There is evidence from three sources that the organism causing halo- blight winters over on the seed: (1) the presence of typical halo lesions on the glumes and lemmas of maturing spikelets (Pi. 29) ; (2) the early appearance of the disease on seedlings grown on soil not previously sown to oats (PI. 28) ; and (3) the great difference in amount of blight in oat plots from treated and untreated seed. (1) natural and artificial infections of spikelets In 1 91 8 at the time the oat plants were heading out it became evident from observations of the plot of Wisconsin No. 14 and from artificial inoculation of Wisconsin No. 7 that the spikelets were also susceptible to infection with the halo organism. After the Wisconsin No. 7 plants had headed out a number of uninjured heads were sprayed with a water suspension of the organism. Another bundle of heads, bruised by draw- ing between the fingers, was similarly sprayed; and both were covered with glassine bags for two days. When the bags were removed infec- tions were already appearing on the bruised spikelets as light green May is, 1930 Halo-Blight of Oats 159 discolorations on the glumes. A week after inoculation every spikelet of these panicles showed distinct typical halo lesions. Many halo lesions also appeared on the uninjured spikelets. Injured and uninjured con- trols sprayed with sterile water and treated in a similar way showed no halo lesions. Early in July natural infections on the spikelets of Wisconsin No. 14 oats were observed. Flag leaves were found which showed either scattered halos or yellow halo tissue the length of the blade and sheath. Where sheaths surrounding the heads were badly haloed, every spikelet in the panicle showed infection. If there is one single lesion on a glume it appears as a typical light green to yellow halo about the point of infec- tion. When the whole glume is infected the tissue becomes yellow and translucent between the veins. Only a few such complete infections of panicles were found. Further observations showed that infections on a small percentage of the spikelets in a panicle were not uncommon even when there were no lesions on the sheaths below. Wind and rain might easily spread the infection directly from lower leaves to panicles. Isolations from the glumes showing these lesions and from the parts inside the glumes gave typical halo organisms. Table I, which records the counts on 42 panicles in one corner of a Wisconsin No. 14 plot, will give some idea of the percentage of blighted spikelets. Table I, — Number of blighted and blasted spikelets on oats naturally infected with halo-blight Panicle No. I 2 3 4 5 6 7 8 9 10 11 12 13 14 *5 16 !7 18 19 20 21 32 23 Number of spikelets per panicle. Number of blighted spikelets per panicle. Number of blasted spikelets per panicle. 58 O 6 54 O ir 77 I 5 66 O 14 55 O 22 55 12 55 17 60 11 42 14 64 I 13 90 2 16 5i I 9 61 O 22 79 36 25 18 l8 23 87 O 4 5° O 13 39 O 21 54 4 34 5° 5 35 55 2 14 62 23 69 20 Panicle No. 24. 25- 26. 27. 28. 29. 3°- 3* • 32. 33- 34- 35- 36. 37- 38. 39- 40. 4i- 42. Total. Average . . . Per cent. .. Number of spikelets per panicle. 67 33 80 65 45 72 85 46 SO 55 45 7i 65 109 52 69 66 60 2,387 59+ Number of blighted spikelets per panicle O 25 o o 29 o 5 o 165 4 6+ Number of blasted spikelets per panicle. 17 8 14 7 10 1 o 47 8 7 4 14 6 o 8 6 13 587 14+ 24+ 160 Journal of Agricultural Research vol. xix.no. 4 In this case 6 per cent of the spikelets are blighted. This accords with the percentage of primary lesions usually observed on seedlings in the field. The sheaths below the panicles numbered 15, 32, and 35 were badly yellowed with halo lesions. (2) PRIMARY LESIONS ON THE FIRST LEAVES OF SEEDLINGS These primary lesions have been observed by the writer on more than 30 varieties of oats in Wisconsin in two different years. They may appear as halos on any part of the leaf blade, but they more often occur on the tips or margins of the leaves as shown by Plate 28. (3) EXPERIMENTS WITH TREATED AND UNTREATED SEED During the season of 191 7 two plots of oats were planted on soil which had not previously been planted to oats. Untreated seed of each of 33 Wisconsin varieties was planted in April, and in May seed of the same 33 varieties was planted after having been soaked for 2% hours in 1 to 320 formalin (1 pint to 40 gallons). Every one of the 33 varieties from untreated seed showed halo-blight to at least some extent, the amount decreasing as the hot weather came on. Wisconsin No. 14 showed the heaviest blighting, and Wisconsin No. 25 was also heavily spotted. Throughout the season not a single lesion was found on the 33 varieties from treated seed. In April, 1 91 8, three parallel plots of oats were planted on soil not previously planted to oats. Thirty-three Wisconsin varieties of untreated 191 6 seed were planted in the first plot, 44 Wisconsin varieties of untreated 191 7 seed were planted in the second plot, and 44 Wisconsin varieties of treated 191 7 seed were planted in the third plot. Also treated seed of Wisconsin No. 14 was planted as a fourth plot on the experimental ground where oats were grown in 191 7. This seed was treated by soaking for 3 hours previous to planting in 1 to 320 formalin. Counts of infections appearing in these plots were begun just as the second leaf was coming out. On May 16, 17, and 18 primary lesions were appearing on the first leaves of plots from untreated 1916 and 1917 seed, the number of primary infections varying from less than 1 per cent to 8 per cent in each plot. These primary lesions on the 191 6 plot would indi- cate that the organism may live for two years on the seed. No lesions were found at this time on the plot from 191 7 treated seed. Counts were made again in the untreated 191 7 plots on May 25, four or five days after heavy driving rains, the normal incubation period for halo lesions. Practically all the first leaves were found to be spotted, and lesions were also appearing on the upper leaves. The condition in the 191 6 plot at this time was about the same and continued to parallel that of the 191 7 plot. At this same time — 9 days after the first appearance of the disease on the untreated plots — scattered halo spots and yellowed leaf tips were beginning to appear on the treated plot, evidently by infection from the May is, 1920 Halo- Blight of Oats 161 neighboring untreated plots, one of which was only 3 feet away. On May 24 and 25 there were more driving rains, and on the twenty-eighth the effects of these storms were evident. Secondary lesions in the untreated plots were so abundant that no attempt was made to count them. Many of the first leaves were completely yellowed and dead, and lesions on sec- ond and third leaves were so numerous that tips, margins, and even whole leaves were becoming yellowed. On varieties where infections were not so abundant the second leaves showed only scattered lesions. On the treated plot the primary lesions were still few, and there was here very striking evidence of the way in which the organism spreads about a center of infection. More or less circular spots of infected plants could be dis- tinguished with the more heavily spotted plants in the center. The amount of infection in this treated plot gradually increased until most of the first and second leaves showed some spotting, but in none of the vari- eties was there more than half as much blighting as in the untreated plots. In the third treated plot, Wisconsin No. 8 showed only scattered lesions on the lower leaves and none on the upper. In the untreated plot of this variety the lower leaves were practically destroyed and the upper so badly spotted that they showed a yellow-brown color at a distance. There were similar but less marked differences in other varieties. Through June there was very little rain. The amount of blight gradually de- creased until at heading time, about the first of July, very few halo lesions could be found, and the upper leaves were practically unspotted. No halo lesions were observed on the fourth plot from Wisconsin No. 14 treated seed until about the twenty-fifth of the month, when two or three centers of infection began to appear as small yellow spots. These spread rapidly after each rain until one of them stood out as a distinct yellow spot irregularly 5 by 1 o feet in diameter. The plants in this spot at heading time were 4 or 5 inches shorter than the more normal plants about them and headed out about a week later. Subsequently scattered lesions occurred on lower leaves throughout the plot and undoubtedly came either from the first infections observed or from the neighboring plots. If these primary infections had been produced by soil organisms they would probably have been much more general. Either sterilization of seed was not complete or else the infection came from the neighbor- ing plots. An experiment with hot-air treatment of seed gave additional proof that the organism is seed-borne. A plot from Graber oats heated to ioo° C. for 30 hours showed no lesions throughout the season. There was not a single spot. The plot from untreated Graber oats showed an abundance of halo lesions through May and June. On every plant there was some spotting and many lower leaves were yellowed and dead. This early appearance of lesions on seedlings grown on new soil, the appearance of typical halo lesions on the glumes and lemmas of the l62 Journal of Agricultural Research Vol. XIX, No. 4 developing spikelets from which the halo organism was isolated, and finally, the absence of the disease on plants from sufficiently treated seed all lead to the conclusion that this is a seed-borne disease. HOSTS OTHER THAN OATS Field observations and artificial inoculation experiments indicate that the halo-blight organism of oats does not readily infect other hosts. No halo lesions similar to those appearing on oats have been observed in the field on wheat, barley, corn, or timothy. In Jefferson and Dodge Counties, Wis., fields of oats and barley were planted so close together that the plants were intermingled at the margins. In both places the oat plants were heavily spotted with halo lesions, but even where these spotted oat leaves came in contact with the barley leaves not a halo could be found on barley. At Arlington Farm, Va., one halo lesion was found on a rye plant growing among infected oat plants, but no plates were made. The field was half oats and half rye, and although practically all the oat plants were spotted no other lesions could be found on rye. Six different sets of inoculation experiments were carried on in the greenhouse during the winter of 1917-18 to test the pathogenicity of the halo-blight organism on wheat, rye, barley, and corn. The methods of inoculation were the same as those described above. The organisms used were stock and No. 36. The results are given in Table II. Table II. — Inoculations on other plants with halo-blight from oats Wheat Rye Barley Spelt Corn Oats, Wisconsin 14. Controls Experi- ment I, stock. + + + Experi- ment II, stock. Experi- ment III, stock. + + + + + + + + + + + + Experi- ment IV, stock. Experi- ment V, stock. + + + + + + + Experi- ment VI, No. 36. + + + + + + + + + Slight infection. + + Moderate infection. + + + Heavy infection. — No infection. Halo lesions were obtained on wheat in two different experiments, in the second of which the halo lesions were not so large but almost as numerous as on oats. In three out of six experiments halo lesions were produced on rye. In the first, infection was so heavy that there was a general wilting of the leaves. Typical white organisms were isolated from these leaves which on reinoculation produced halo lesions on oats but not on rye. Halos on barley were obtained in three out of six inoculation experi- ments. There were eight halos in the first experiment and two in the second. In the third experiment, six leaves had one or more halos. May is. 1920 Halo-Blight of Oats 163 Reisolation from the first halos gave typical white colonies which on sub- culture and reinoculation produced halos on barley and oats. No halos were obtained on corn in four experiments, and no halos were obtained on broom corn in later experiments. Oat plants inoculated at the same time always showed abundant infection. It is evident that the halo-blight organism may attack wheat, rye, and barley to a slight extent; but in Wisconsin, at least, halo lesions in the field rarely, if ever, appear on anything but oats. VARIETAL SUSCEPTIBILITY All observed varieties of cultivated oats are attacked by the halo-blight to some extent. Wisconsin No. 14, both in the field and in the greenhouse, is more susceptible than any other variety and shows more lesions in later stages of development, especially on the flag leaf, rachis, and spike- lets. Two varieties, Wisconsin No. 13 and Wisconsin No. 15, grown in the fields on either side of Wisconsin No. 14 during 1917, showed considerable resistance. Although leaves of Wisconsin No. 14 were badly spotted, the leaves of Wisconsin No. 13 and 15, which came in contact with them, showed little spotting. In the first plot (from 191 6 untreated seed), described above, Wisconsin No. 128 showed only six primary infections, while Wisconsin No. 124 showed 169. In the second plot (from 1917 untreated seed) some varieties showed only slight secondary infections, others moderate, and some heavy infection. Inoculation experiments in the greenhouse also brought out differences in varietal resistance. Wisconsin No. 1,5, and 14 were used for several experiments, Wisconsin No. 14 always showing so much heavier infection than either of the other two that Wisconsin No. 1 and 5 were no longer used. Wisconsin No. 1 showed more resistance to infection than Wis- consin No. 5. While certain varieties are more susceptible than others under ordinary conditions and show fewer primary lesions at the beginning of the season, as above indicated, the differences are not marked in severe blight years as the season advances. RELATION OF ORGANISM TO HALOED TISSUE The oval outline of the halo, its rapid spread from the point of infec- tion, and the fact that the haloed tissue remains normal, apparently, except for loss of color, have led to the conclusion that the discoloration is probably due to some diffusible substance produced by the bacteria rather than to their immediate presence. To determine whether or not the bacteria were equally distributed throughout the lesions, isolations were made from pieces of tissue cut from the centers of lesions and from points at varying distances from the center as shown in the following diagrams. Isolations were made after treatment with mercuric chlorid as described above. The distribution of bacteria throughout the halo lesions is shown in Plate 33 and Table III. 164 Journal of Agricultural Research Vol. XIX. No. a V, ^ § ^ S ft! •4 pg < I I + + I I + °. T ° +00 1 1 1 l ++ ++ GO May is, 1920 Halo-Blight of Oats 165 The first two lesions used were produced by artificial inoculation. The last three were natural infections from the experimental plots. In all of the five isolations only the plates from the centers of the lesions showed any growth at all, and these plates were heavily seeded with typical white colonies. The only exception is the one colony on a plate from isolation IV, section 2. In isolation I the broth cultures from sec- tions outside the center did not even cloud. In other isolations where broth cultures from sections outside the center clouded, subsequent plates showed that the clouding was sometimes due to the growth of the halo organism and sometimes to contamination. The bacteria are evidently abundant only in the centers of the lesions, and if any do occur outside in the halo they are very few in number. This indicates that the discoloration of the halo-tissue is due only indi- rectly to the presence of bacteria, and that some enzym or toxic by- product destroys the chlorophyl. A suggestion of what this by-product might be was obtained from some plates of potato-dextrose agar on which colonies of the blight organism were growing. When colonies of the stock organism were 3 days old distinct halos appeared in the agar about the colonies as illustrated in Plate 32, B. The agar around these colonies was less translucent than that outside the halos and was dis- tinct in outline. These halos in the agar increased in diameter from day to day, showing the concentric circles of growth illustrated in the plate and characteristic of the lesions on oat leaves. Acetic acid dropped on the agar-plate halos cleared them in a minute or two. Drops of am- monium carbonate [(NH^COJ and ammonium chlorid [NH 4 C1] on sterile plates of the same potato agar produced in a few minutes halos similar in size and appearance to those produced by the colonies of bacteria. Acetic acid also cleared these halos. Litmus was added to melted potato agar at the rate of one drop of a saturated solution to 10 cc. of the agar, and plates were poured. Streaks of stock were made across the agar as soon as it had hardened. Similar streaks on plain potato agar produced distinct halos about them in two days. In the same time the litmus potato agar had turned a distinct blue for 1 cm. or more on all sides of the growth. It seems probable, therefore, that the ammonia produced by the blight organism is responsible for the destruction of the chlorophyl and for the halolike lesions produced in the oat plants. Stained sections of haloed leaf tissue also show bacteria only in the center of the lesion. The bacteria at first are intercellular, but later they destroy the cell walls and cause the collapse of the tissue. The collapsed tissue is evident as the dead brown centers of lesions. (See H. 35, C.) 1 66 Journal of Agricultural Research vol. xix,n .4 COMPARISON WITH OTHER SIMILAR BACTERIAL DISEASES Seasons of excessive rainfall and of abnormal conditions in the oat fields similar to those of 191 8 have been recorded for 1890 and 1907-8. For the earlier record we are indebted to Galloway and Southworth, of the United States Department of Agriculture (1), and for the later work to Thomas F. Manns, of the Ohio Experiment Station (j). WORK OF GALLOWAY AND SOUTHWORTH In 1890 Galloway and Southworth (/) published a preliminary note on what they termed "a new and destructive oat disease." This disease appeared in May and June of that season and was so widespread and severe as to threaten to destroy the entire oat crop of the eastern and central States. The signs described were a browning of the tips of the lower leaves, which spread until in a short time all the leaves were dead and brown. Bacteria were found in these lesions. The account of the disease by these authors, however, is too meager to afford any basis for judgment as to whether or not it was the disease here described. During the seasons of 1906- 1909 blade-blight of oats was recorded again over a fairly wide area, and in 1907 it was so severe in some fields as to occasion a loss of from one-half to two-thirds of the crop. In 1908 the blight was threatening at one time but eventually caused little loss. The accounts of the disease from southern Canada and central and eastern States are of the same general kind. They mention a general yellowing of the lower leaves of young plants, the yellow color changing to a brown or red under weather conditions unfavorable to the organism, such as a sudden change from cool, cloudy weather to bright sunshine and higher temperature. The fields are often described as having a rusted appear- ance because of this reddening of the blades. The trouble was attributed to various causes — to insects, to bacteria, to fungi, and to unfavorable weather conditions. In 1908 Dr. Erwin F. Smith discovered this disease at Arlington Farm, Va., photographed it, cut sections, and made cultures of the organ- ism on various media, but did not publish upon it nor make any inocu- lations, although it is quite certain from the type of the disease and the nature of the cultures that he had the same organism here described. This was perhaps its first isolation in pure culture. No other serious research work was undertaken until Thomas F. Manns carried on his investigations during the seasons of 1908-9 at the Ohio Experiment Station. WORK OP THOMAS F. MANNS, I906-I909 Manns (3) states that — the disease manifests its presence by changes in color varying from a light yellowing, which apparently checks but little the growth of the oats, to a pronounced redden- ing, which in severe cases kills the blades, leaving only the younger leaves and the central axes alive. May is. 19=0 Halo-Blight of Oats 1 67 The primary yellowing sooner or later changes to a mottled red or brown. In another place he says: The preliminary effects of this disease is a yellowing, beginning either as small, round lesions on the blade, or as long, streak lesions extending throughout the blade or even the whole length of the culm and blade. Occasionally it begins at the tips and works back into the culm; again the upper leaves often break down through a weak- ened condition of the plant from defoliation below. When lesions work back from the leaves to the culm a general yel- lowing and collapse of all the foliage may result. In 1909 — the disease in the majority of infected leaves began as small yellow spots on different parts of the blades. When these points of infection were numerous, the infected areas quickly became confluent, and the collapsed leaf showed a brownish mottled appear- ance. These brief statements are the only references in the bulletin (exclu- sive of PI. XIII) to anything at all corresponding to the lesions char- acteristic of the blight here described, and there is much that is contra- dictory. His colored figures as well as most of his text indicate an entirely different disease, but his Plate XIII shows that this halo-disease formed at least a part of the phenomenon under consideration. The distinct reddening which he describes and which he illustrates in Plates X and XI was not observed anywhere in Wisconsin even in the worst blight year, 191 8. A distinct reddening of oat leaves was observed in our plots but was not due to the halo-blight. Two unsuccessful attempts were made by the writer to isolate bacteria from these reddened leaves. Manns attributes the severity of the outbreak in 1907 to the abnormally low temperatures of April, May, June, and July and to the unusual amount of rainfall during those months and gives convincing climato- logical data in support of his conclusion. He states that the results of artificial inoculation in the greenhouse also support this theory that cool, humid weather conditions favor the disease. Through isolation and inoculation experiments Manns came to the conclusion that the blade-blight of oats was due to two species of bacteria living in symbiotic relations within the host tissue (Pseudomonas avenae Manns and Bacillus avenae Manns). His isolations were made by ster- ilizing the blades in 2 to 1,000 mercuric chlorid solutions for 1 to 1% minutes and following this by four washings in sterile water. He states that in practically all isolations from diseased oats these two bacteria were found to be more or less abundant, and when occurring together they could be plainly seen on the agar poured plates in from 2 to 3 days. The yellow organism {Bacillus avenae Manns) always appeared first. As a rule, the white organism predominated. Inoculations were made by Manns in several ways: (1) Directly from crushed leaves; (2) by hypodermic injection, using separate pure cultures of the white and the yellow organism; (3) by hypodermic in- jection, using the two cultures mixed (3, PI. X); (4) by spraying mixed 1 68 Journal of Agricultural Research voi.xix,No. 4 cultures on injured and uninjured leaves; (5) by root inoculations with- out wounds, using mixtures of the two organisms; and (6) by means of grain aphids. He reports that inoculations in the field and in the greenhouse showed that the yellow organism when used alone produced no lesions and that the white organism when used alone produced only "limited and non- typical lesions," which formed slowly, extended from ^ to 1 inch from the point of infection, and then remained checked. When a mixture of the two organisms was used the lesions appeared in from 10 to 12 days and spread rapidly. From these results he concludes that the disease is a symbiosis, the white organism requiring the presence of the yellow organism to be actively pathogenic. He also states that the virulence and viability of the white organism on artificial culture media depend greatly upon association with t«he yellow organism and that the pathogenic action of the white organism was more marked when carried over winter in mixed culture with the yel- low organism than when carried over separately. After nine months in pure culture t'he white organism failed in several instances to grow. Manns sta'tes that endospores occur. These were stained with hot carbol fuchsin from 2-months-old cultures. The figure of these spores in his Plate IX is too indistinct to be of any value in verifying his state- ment. His white organism is described as a short motile rod with polar flagella. These are three to five times the length of the rods in his Plate IX, fig. 4, and one to six times those in his text figure No. 1. The rods measure in the majority of cases 0.75 by 1.5 ju. They are rarely in chains of three to four. The thermal death point is 6o° C. The optimum temperature is 20 to 30 . He states that his organism is pathogenic on oats, corn, timothy, barley, wheat, and bluegrass. The group number for his white organism is given as 11 1.2223032. Manns' yellow organism is a bacillus with the group number 222.2223532. Manns suggests the probability of the organism's wintering over in the soil and so being distributed to the leaves by spattering rains. He states that there is no doubt that on seedlings lesions sometimes start on the roots or on that part of the stem in contact with the soil. He does not describe these lesions. The possibility that the disease is seed- borne is not mentioned. Manns' descriptions of individual lesions are so meager and his descrip- tions of general signs so inclusive as to lead to grave doubt about his hav- ing worked with a single bacterial disease. There is no doubt, however, that he sometimes had typical halo-blight lesions, because of his Plate XIII, but with this exception there is no conclusive evidence from either his text or figures that he had this disease under observation; and the May is, 1920 Halo-Blight of Oats 169 result of his inoculations as indicated on his colored plates is quite con- tradictory. The chief differences between the two white organisms Pseudomonas avenae Manns and Bacterium corona faciens, n. sp., are summarized below : 6. PSEUDOMONAS AVENAE MANNS. Produces typical blight lesions only when used with Bacillus avenae Manns (a yellow organism). Spreads throughout the lesion when used alone. Virulence and viability on artificial media dependent upon association with Bacillus avenae Manns. Viability and virulence greatly re- duced by a number of transfers. Growth feeble on artificial media. (See 3, PL VIII, fig. j.) Liquefaction of gelatin stabs begins in 7 to 12 days. 7. Pitting of gelatin colonies begins in 7 days. 8. Visible growth in broth in 3 days. 9. Manns does not record browning of broth or other media. 10. Milk not coagulated in 30 day 3. 11. Acid to litmus milk. 12 . No reduction of litmus milk recorded. 13. Strictly aerobic. 14. No ammonia produced. 15. Nitrates reduced. 16. Limits of growth, —5 to +15. 17. Thermal death point 6o° C. 18. Internal structure of agar colonies amorphous. In hanging drop there are few motile organisms. Growth viscid on agar. Produces Clostridium forms in one week on nutrient glucose agar. 22. Produces endospores. 23. Does not form long chains. Shorter and thicker than Bacterium coronafaciens. Average size 0.75 by i-5 M- Lives over in the soil. 26. Pathogenic on oats, corn, timothy, barley, wheat, and bluegrass. Group number 11 1.2223032. IQ 20. 21. 24. BACTERIUM CORONAFACIENS, N. SP. i. Produces typical halo-blight lesions when used in pure culture. 2. Found only about the point of infec- tion and not throughout the halo. 3. Virulence and viability not dependent on another organism. 4. Viability and virulence not reduced by transfer. 5. Growth abundant on artificial media. (See PI. 30, A, B, a, b.) 6. Liquefaction begins in 3 days. 7. Pitting begins in 3 days. 8. Visible growth in 1 day. 9. Broth and other media turned brown. 10. Milk usually coagulated in 5 to 7 days. 11. Alkaline to litmus milk. 12. Litmus milk reduced. 13. Facultative anaerobic. 14. Ammonia produced. 15. Nitrates not reduced. 16. Limits of growth, —18 to +2 7. 17. Thermal death point 47 to 48 C. 18. Internal structure of agar colonies not amorphous. (See PI. 31.) Active motile organisms in hanging drop. Growth butyrous. No Clostridium forms observed in any medium. Does not produce endospores. Forms chains and long filaments. Average size 0.65 by 2.3 /*. 19 23- 24. 25 25. Lives over winter on the seed. 26. Pathogenic on oats, barley, wheat, and rye. Group number 221.2323023 A bacterial disease producing lesions similar to those of the halo-blight of oats has been described from tobacco (10). The lesions are similar to 170 Journal of Agricultural Research Vol. XIX. No. 4 the halos of oats in that they form "circular chlorotic areas" 2 to 3 cm. in diameter with minute brown centers. The oat lesions, however, have no water-soaked borders, and the affected tissues do not fall out as in tobacco wildfire. A white organism has been isolated from these lesions which differs from the halo-blight organism in the points mentioned below : HALO-BUGHT ORGANISM. One to several polar flagella. Single to long chains. 2.3 by 0.65 fx. Capsules. Odor in agar stroke. Casein not precipitated in litmus milk. Ammonia produced. Thermal death point 47 to 48° C. TOBACCO ORGANISM. One polar flagellum. Single to chains of five elements. 3.3 by 1.2 p. No capsules. No odor in agar stroke. Casein precipitated in litmus milk. Ammonia not produced. Thermal death point 65 ° C. The halo lesion so characteristic of this oat disease does not occur in the blackchaff disease of wheat (6-9) or the bacterial blight of barley (2), while the oat disease lacks the translucent water-soaked stripes of these diseases as well as the exudate so abundant in both. R. H. Rosen has recently published a preliminary note on a bacterial dis- ease of foxtail (4), which he thinks may be similar to the halo-blight of oats. His description of lesions as dark brown spots or streaks, however, makes it probable that if it is similar to either bacterial disease of oats it would resemble stripe-blight rather than halo-blight. The writer has not observed halo lesions on foxtail and in two sets of field inoculations has obtained no infections on foxtail with the halo organism. CONTROL MEASURES The evidence that the halo-blight of oats is seed-borne seems conclu- sive. However, no practical method of seed treatment has, as yet, been found which will entirely control the disease. Treatment with formalin for smut controls halo-blight to a marked extent but not entirely. In 1917, treated seed of 33 Wisconsin varieties did not show a halo lesion throughout the season, while the same untreated varieties all showed some halo-blight. In 1918, 44 treated varieties of Wisconsin oats devel- oped primary lesions which, however, were later and fewer than on the same untreated varieties. Even when the blight was most severe it was only about half as heavy in the treated plots as in the untreated. The plot from Wisconsin No. 14 treated seed showed very few primary lesions and little secondary spotting except in patches about these pri- mary lesions. This would indicate that soaking for three hours in 1 to 320 formalin kills many but not all of the organisms on the seed. In Jefferson County, Wis., where most of the seed was treated for smut, the blight during the 191 8 season was much less abundant than in Dodge County, where seed treatment was not general. May 15, 1920 Halo-Blight of Oats 171 Another method of seed treatment is being developed at Wisconsin which in 191 8 entirely controlled halo-blight. The treated seed was heated in a gas oven at ioo° C. for 30 hours. The plot from this treated seed did not show a single halo lesion even during the time when other oats were most severely attacked. The plot from untreated seed of the same variety showed primary infections on 10 per cent of the plants and 100 per cent secondary infections on the lower leaves during May and the first two weeks in June. While oats in good condition success- fully withstand this treatment of 30 hours at ioo° C, a similar treatment for a shorter period would perhaps be just as effective. The commercial application of this treatment has not as yet been worked out. SUMMARY A bacterial disease known as halo-blight was unusually severe in its attack on oats throughout Wisconsin during the 191 8 season, and reports of a similar disease were received from southern Minnesota, Iowa, north- ern Illinois, and Indiana. Such epidemics occur under particularly favorable weather conditions, disappearing with the advent of weather conditions more favorable to the development of the host plant. Typical lesions of halo-blight are characterized by halolike margins of chlorotic tissue about a center of dead tissue. Isolations from these lesions have constantly given a typical white organism. Yellow organisms also appear from isolations when the surf- ace of the tissue has not been sterilized. Inoculation experiments have shown conclusively that the white organ- ism alone is responsible for the production of typical lesions. The yellow organism is evidently a surface saprophyte. Since few if any organisms are found outside the central infection area, the halo is thought to be due to a diffusible substance, probably am- monia. The organisms live over winter on the seed, producing primary lesions on the first leaves of seedlings. From these lesions the organisms are carried to other leaves by wind and rain. It seems probable that the percentage of blasting on oat panicles varies with the severity of the halo-blight from season to season. This blasting seems to be due to the same unfavorable weather conditions which favor the development of the bacterial blight rather than to the disease itself. Halo-blight lesions from natural infections have never been observed on any hosts except oats and rye. Artificial inoculations show that the halo organism may be slightly pathogenic on wheat, rye, and barley. When the halo-blight is not too severe, different varieties of oats show differences in susceptibility to the disease. 172 Journal of Agricultural Research vol. xix, No. 4 The organism isolated and described by the writer has the group number 221.2323023. No other white organism used by the writer has produced anything similar to the halo lesions. Other white organ- isms have in fact produced no lesions on oats. Three strains of softrot organisms with internal markings very much like those of oat colonies have been used, and also the white organism, Bacterium atrofaciens McC, which produces lesions on wheat. The name of Bacterium coronafaciens n. sp. is suggested for this white halo-producing organism. Treatment with 1 to 320 formalin, as for smut, checks but does not entirely control the disease. A hot-air treatment for 30 hours at ioo° C. does control the blight. LITERATURE CITED 1) Galloway, B. T., and Southworth, E. A. 1890. PRELIMINARY NOTES ON A NEW AND DESTRUCTIVE OAT DISEASE. In Jour. Mycol., v. 6, no. 2, p. 72-73. 2) Jones, L. R., Johnson, A. G., Reddy, C. S. 1917. bacterial blight OP barley. In Jour. Agr. Research, v. 11, no. 12, p. 625-644, illus., 4 pi. (1 col.). Literature cited, p. 643. 3) Manns, Thomas F. 1909. THE BLADE BLIGHT OF OATS, A BACTERIAL DISEASE. Ohio Agr. Exp. Sta. Bui. 210, 167 p., illus., 15 pi. Literature cited, p. 166. 4) Rosen, H. R. 1919. A preliminary note on A bacterial disease OP foxtail. In Science, n. s. v. 49, no. 1264, p. 291. 5) Smith, Erwin F. 1905. bacteria in relation to plant diseases, v. 1. Washington, D. C. (Carnegie Inst. Washington Pub. 27.) 6) 7) 1917. black chaff of wheat. In U. S. Dept. Agr. Bur. Plant Indus. Plant Disease Survey Bui. 2, p. 40. 1917. A NEW disease of wheat. In Jour. Agr. Research, v. 10, no. r, p. 51-54. pi. 4-8. 8) 1918. black chaff OF wheat. In U. S. Dept. Agr. Bur. Plant Indus. Plant Disease Survey Bui., v. 2, no. 6, p. 98-99. 9) Jones, L. R., and Reddy, C. S. 1919. THE black chaff of wheat. In Science, n. s. v. 50, no. 1280, p. 48. 10) Wolf, F. A., and Foster, A. C. 1918. tobacco wildfire. In Jour. Agr. Research, v. 12, no. 7, p. 449-458, illus., pi. 15-16. Halo-Blight of Oats D LATE C Vol. XIX, No. 4 PLATE C Halo lesions on flag leaves of Wisconsin No. 14 oats. Natural infections from Hill Farm, Madison, Wis. Photographed June, 1917. PLATE 26 Typical isolated halo lesions. Natural infection on Graber oats. Photographed June 24, 1918. Natural size4 Halo-Blight of Oats Plate 26 Journal of Agricultural Research Vol. XIX, No. 4 Halo-Blight of Oats Plate 27 Journal of Agricultural Research Vol. XIX, No. 4 PLATE 27 Halo lesions on Wisconsin No. 14 oats produced by spraying with a water suspen- sion of the stock organism May 26, 1917. Photographed May 31, 1917. PLATE 28 Infe&tion from untreated 1916 seed of Wisconsin No. 124 oats. Planted April 24, 1918. Photographed May 17, 1918. Halo-Blight of Oats Plate 28 Journal of Agricultural Research Vol. XIX, No. 4 Halo-Blight of Oats Plate 29 Journal of Agricultural Research Vol. XIX, No. 4 PLATE 29 Spikelets of Wisconsin No. 14 oats: A. — Left and center spikelets show natural infection with halo-blight. Tips yellowed and translucent. Spikelet at right normal, unspotted. Photographed July 17, 1918. B. — Upper spikelet shows typical isolated halo lesion near base. Lower spikelet normal, unspotted. PLATE 30 A. — Two per cent +5 glucose Difco peptone beef bouillon agar slant of No. 36. Three-day colony. Photographed August 29, 1919. Natural size. B. — Two per cent potato-dextrose agar slants, a, No. 36, white culture, consistency butyrous; b, stock, white culture, consistency of boiled starch; c, No. 39a, yellow culture. Halo-Blight of Oats Plate 30 Journal of Agricultural Research Vol. XIX, No. 4 Halo-Blight of Oats Plate 31 Journal of Agricultural Research Vol. XIX, No. 4 PLATE 31 A. — Three-day colony of stock on 2 per cent dextrose-potato agar. Photographed February 17, 1919, by oblique transmitted light. X 10. B. — Two-day colonies of stock on + 10 beef-peptone agar. Photographed March 26, 1919, by oblique transmitted light. X 10. C. — Five-day colony of stock on potato-dextrose agar. Colony of boiled starch consistency. Photographed January, 1918, by reflected light. X 7. D. — Five-day colony of No. 36 on + 10 beef-peptone agar. Photographed October 1, 1918, by oblique transmitted light. X 10. E. — Three-day colony of stock on 2 per cent glucose Difco peptone beef bouillon agar. Photographed October 7, 1919, by oblique transmitted light. X 10. F. — Seven-day colony of stock on -f 1 5 beef -peptone agar. Photographed March 3 1 , 1919, by oblique transmitted light. X 10. G. — Margin of 3-day colony of stock on + 15 gelatin. Photographed September 30, 1919- X 75- PLATE 32 A. — Five-day colonies of stock on potato-dextrose agar. Colonies of boiled starch consistency. (For single colony see PI. 31, C.) Photographed by reflected light, Natural size. B. — Three-day colony of stock on potato-dextrose agar. Halo about colony. Photographed February 17, 1919, by oblique transmitted light. X 5. Halo-Blight of Oats Plate 32 Journal of Agricultural Research Vol. XIX, No. 4 Halo-Blight of Oats Plate 33 Journal of Agricultural Research Vol. XIX, No. 4 PLATE 33 Isolations from sections of halo lesion. The lesion was from an artificial inocula- ion with oat stock organism made February 25, 1918. Isolates were made March k, igi8 on potato agar. The sections were dipped in alcohol and then submerged for one minuTe in 1 to 1,000 mercuric chlorid. The plate from secUon c xs the only one showing colonies of bacteria. A.— Poured plate of isolation from section a of lesion. B — Poured plate of isolation from section b of lesion. C— Poured plate of isolation from section c of lesion. D— Poured plate of isolation from section d of lesion. E— Poured plate of isolation from section e of lesion. Photographed March 15, 1918. PLATE 34 A. — No. 36 from 24-hour potato-dextrose agar slant; carbol fuchsin stain. X 620. B.— Stock from 24-hour potato-dextrose agar; Ribbert's capsule stain. X 620. C. — Stock from 4-day potato-dextrose agar; carbol fuchsin stain, showing long chains. X 620. D. — Stock from 3-day potato-dextrose agar; Ribbert's capsule stain. X 1.550. E. — Stock from i-day + 15 beef-peptone agar; Van Ermengem stain. X 1.550. F. — No. 36 from i-day + 5 beef-peptone agar; Caesar-Gil stain. X 1,550. Halo-Blight of Oats Plate 34 Journal of Agricultural Research Vol. XIX, No. 4 Halo-Blight of Oats Plate 35 Journal of Agricultural Research Vol. XIX, No. 4 PLATE 35 Sections of oat leaves through halo lesions, showing bacteria in the tissues. Fixed in Gilson's fixative and stained with carbol fuchsin. A. — Bacteria in substomatal cavity, showing method of entrance of bacteria into the leaf tiasue. Cut 15 n thick. X 700. B. — Bacteria in substomatal cavity. Cut 15 n thick. X 1, 650. C. — Section of older lesion, showing bacteria between the cells. In the upper part of this section the tissue is disintegrating at about the point of infection. Photographed August 26, 1919. X i>55Q- WUSk