The Translocation of Antibiotics in Higher Plants: II. THE MOVEMENT OF GRISEOFULVIN IN BROAD BEAN AND TOMATO

The uptake and translocation of griseofulvin from water cultureby broad bean and tomato has been studied; observations werealso made on its decay in broad bean. In most cases griseofulvinwas determined by bioassay. Where possible the bioassay waschecked by chemical estimations and was found to be adequate. The amount of griseofulvin taken up by the broad bean was proportionalto the volume of water transpired for any single concentrationof the treating solution and the decay was exponentially relatedto the time of exposure in the tissues. The accumulation ofgriseofulvin in the tissue had, therefore, an exponential componentbut within the limits of error there was constant relationshipbetween accumulation and transpiration over the time periodsused in the trials. The rate of accumulation in tomato was alsoconstant. The amount of griseofulvin accumulated by both beanand tomato after a definite time was a linear function of theconcentration of the treating solution. There were two processes involved in the uptake of griseofulvinby the broad bean: (a) an initial rapid entry into the rootswhich was inhibited by sodium azide and dinitrophenol at concentrationswhich did not reduce transpiration; and (b) a prolonged uptakelinearly related to transpiration which was not affected bythese concentrations of the inhibitors.     

STUDIES ON SYSTEMIC FUNGICIDES. I. FUNGICIDAL PROPERTIES OF THE ARYLOXYALKYLCARBOXYLIC ACIDS

A method is described for assessing the systemic activity of compounds in checking the infection of broad bean ( Vicia faba ) by the fungi Botrytis cinerea or B. fabae. Treatment consisted in allowing the roots of young seedlings to stand in a solution containing 10 p.p.m. of the chemical for 2–3 weeks. The plants, together with controls, were then inoculated and when symptoms had had time to develop, the degree of chocolate spot infection was assessed. Several methods of disease assessment were examined and are critically discussed.
Certain phenoxyalkylcarboxylic acids tested by this method consistently gave a reduction in the mean size of fungus lesions on the bean leaves, clearly indicating systemic fungicidal action. The most promising substances were 2:4:6-trichlorophenoxyacetic, pentachlorophenoxyacetic and pentachlorophenoxy iso butyric acids. Further experiments with these compounds involving soil treatment, stem injection and spray application are described, and in most cases systemic fungicidal activity was clearly demonstrated. Certain compounds caused visible damage to the plants or resulted in a reduction in growth.
The results presented indicate that phenoxy acids can be fairly readily translocated in bean plants and that they tend to accumulate in actively growing tissues. It is considered unlikely, however, that they persist for long periods in plant tissue.
In the soil, the compounds appeared to remain effective for a considerable time, particularly the less soluble pentachloro acids, suggesting that soil application might provide a safe and useful method of treatment.     

OBSERVATIONS ON THE EFFECT OF GROWTH-STIMULATING COMPOUNDS ON THE HEALING OF WOUNDS ON APPLE TREES

Wounds were made at various times of the year on the trunks and large branches of apple trees with a 1 in. bit and treated with a number of organic compounds in lanoline paste. The healing of these wounds was followed through two growing seasons by tracing and measuring the area of exposed wood at different times after wounding.
Callus growth was practically confined to the summer months. Development of callus was improved by applications of lanoline, and the lanoline effect could be further enhanced in the early part of the first growing season following treatment by the addition of certain growth-stimulating compounds including 4-chloro-3:5-dimethylphenoxyacetic acid and 2:4-dichlorophenoxyacetic acid. Indolyl-3-butyric acid probably also caused some stimulation. After the period of initial stimulation the rate of callusing was approximately the same on treated as on untreated wounds.     

OBSERVATIONS ON APPLE CANKER

Field observations on leaf-scar canker of apple confirm its importance in the southwest of England: on a heavily infected tree more than 5% of the current year's growth may be damaged. Leaf scars may become infected by spores of Nectria galligena which are sucked into the tracheids of a freshly exposed scar where they develop in a relatively protected environment. These freshly exposed scars become much less susceptible to infection after approximately one hour.
Infections established in leaf scars may develop immediately into cankers or they may be confined by a suberized or gum barrier. Mycelium from some of these confined lesions may spread into the surrounding healthy tissue through growth cracks when the buds develop. The lesions extend throughout the winter and spring; usually the scars earliest inoculated are the first to show cankers. It seems possible that infections of this type might be controlled by eradicant sprays applied just before leaf-fall.     

The Translocation of Antibiotics in Higher Plants: I. ISOLATION OF GRISEOFULVIN AND CHLORAMPHENICOL FROM PLANT TISSUE

Griseofulvin and chloramphenicol have been recovered from thetissues of broad-bean plants grown in solutions containing theseantibiotics. Partial separation of the compounds from other constituentsof solvent extracts of plant tissue was effected by counter-currentdistribution in a carbon tetra chloride methanol : water system.Pure griseofulvin and chloramphenicol were then obtained byrecovery of the solute in appropriate tubes followed by chromatographyon an activated alumina column; they were identified by mixedmelting point determinations and comparison of their infra-redspectra. A method for estimating griseofulvin in plant tissue extractsbased on counter current distribution and examination of thefractions spectrophotometrically has been developed, and usedto strike a balance between the gnseofulvin entering the plantand that present in the tissues after treatment. Estimates of chloramphenicol in the tissues by bioassay agreewell with the chemical determinations.     

Water Pathways in Higher Plants: I. FREE SPACE IN WHEAT LEAVES

A technique has been developed for the study of pathways ofwater movement in the xylem and free space of wheat leaves.Plants were treated with Lead-EDTA chelate through either theroots or the leaves; after treatment the lead was precipitatedin situ as lead sulphide with hydrogen sulphide gas and itslocation determined by light and electron microscopy. Bulk water movement was in the lumen of the xylem, where therewas always a heavy deposit of lead sulphide after root treatments.Outside the xylem the deposits were confined to the cell wallsand were most dense in the middle lamella. Deposits were notfound in the cells themselves. The main zones of water loss,marked by heavy deposits of lead sulphide, were associated withthe stomata, the junctions of the periclinal walls of the epidermalcells, and the cuticle, leaf hairs, and specialized epidermalcells with pitted walls associated with the vascular bundles. Entry of lead chelate into the leaves was adequately describedby a diffusion model. The free space seemed to be located mainlyin the water of hydration of the pectin middle lamella and wasesmated to occupy 3 to 5 per cent of the volume of the tissue.     

Water Pathways in Higher Plants: II. WATER PATHWAYS IN BOOTS

The technique for studying the pathways of water movement described,in the first paper of this series has been applied to the transpirationstream in the roots of higher plants. Free space is confinedto the cell walls except in flooded tissue where intercellularspaces are also included. Errors involved in free-space estimatesare discussed and free-space volumes of 5.4 per cent have beenobtained for wheat root cortex and 4.1 per cent for carrot xylemparenchyma. The main water-absorbing regions of roots beginimmediately behind the elongating zone, where the first xylemelements are fully differentiated, and end when the endodermisundergoes secondary wall development. In the cortex the transpirationstream is located mainly in the cell wall. Calculations indicatethat the symplastic pathway is of only minor importance in transpiringplants. At the endodermis the free-space pathway is blockedby the Casparian strip and all water and solute entering thestele must pass through the lumen of the endodermal cells. Thepermeability of individual endodermal cells varies considerablyboth between cells of the same species and between those ofdifferent species. Once inside the endodermis, the transpirationstream returns to the cell-wall pathway until it reaches thexylem vessels where it enters the lumen of the mature xylemelements.     

Observations on apple canker; the anatomy of the stem canker

The development of cankers caused by Nectria galligena on apple is described; the pathogen exploits all the tissues outside the xylem and will also penetrate the xylem to an appreciable depth, invading the xylem parenchyma, vessels and fibres.
Spread in the peripheral tissues is checked to some extent by successive barriers laid down by wound phellogens, but the barriers are eventually passed by the mycelium. There appear to be two distinct ways in which these barriers are passed: there may be direct penetration which appears to be associated with an aggregation of the mycelium into blocks, and symptoms suggestive of mechanical rupture; or there is an alternative route in which the pathogen grows in the lumen of the fibres and emerges behind the barrier.
In the xylem the spread of the pathogen is checked by tyloses and gumming in the vessels, and by gumming in the parenchymatous tissues, but in the fibres there appears to be no defence mechanism, and the spread beyond the gum barriers was usually recorded in the xylem fibres. It is suggested that the presence of the pathogen in the xylem fibres might provide the explanation of the formation of cankers on partially healed pruning cuts, and this type of canker is described in some detail.
The development of the canker depends on the balance between the development of the pathogen and the resistance of the host, and there is some discussion on the relation of this balance to the control of the disease.     

Water Pathways in Higher Plants: III. THE TRANSPIRATION STREAM WITHIN LEAVES

Water in the transpiration stream is distributed throughoutthe leaves in the vascular bundles. In wheat, water appearsto be confined to the main veins by the mestome sheath and toenter the mesophyll through the walls of the smaller veins.Within the mesophyll the water in the transpiration stream movesin the free space of the cell walls to the evaporating surfacesof the leaf. The lead chelate, which is used to trace the transpirationstream, accumulates at the final points of evaporation at themargin of the leaf. Lead chelate accumulates beneath and onthe surface of the cuticle, being partly associated with theanticlinal walls of the epidermal cells, the walls of the stomatalguard cells and specialized epidermal cells. Chelate does notaccumulate at the base of substomatal cavities, indicating thatthe cuticle of the epidermis is the main evaporating surfaceof the leaf. The behaviour in broad bean, laurel, and plantainis essentially the same. The rate of peristomatal and cuticulartranspiration is closely related to the size of the stomatalaperture. Conditions which control stomatal aperture also causechanges in the dimensions of the epidermal cells.