A CYTOLOGICAL AND NUTRITIONAL STUDY OF POLYCHYTRIUM AGGREGATUM. I. CYTOLOGY

Abstract

THE WORK on the morphology and classification of Polychrytrium aggregatum previously described (Ajello, 1942) has been extended by cytological studies. These clarify various points bearing on relationships to similar forms and on classification. Maintaining P. aggregatum in culture was formerly a critical obstacle. However, adequate material at various stages of development became available when it was found to grow on chitinous substrata as developed by Karling (1945). This paper describes cytological studies with special reference to phylogeny and taxonomy. MATERIALS AND METHODS.-The strain of P. aggregatum used throughout this study was originally isolated from a Brazilian mud sample collected by Karling in 1943. Unifungal cultures were established by introducing several heavily infected pieces of chitin into a culture dish containing autoclaved soil and charcoal water. Fragments of purified chitin were placed at the bottom of the dish from time to time to maintain growth and to obtain material for cytological study at various stages of development. Generally, after 18 hours, germination stages of the zoospores were available. Within 24-48 hours, sporangial development had progressed far enough to provide abundant material for the study of mitosis, and by 72 hours, the bait was overgrown with mature thalli which could either be fixed or used for obtaining zoospores. The material was fixed in a variety of fluids: Navashin's, Allen and Wilson's modification of Bouin's fluid, chrom-acetic acid, and Flemming's medium and weak solutions. The best results were obtained with Navashin's and Allen and Wilson's fluids after 24hour fixation. Zirkle's, Benda's and Champy-KuIl's fixatives were used for study of mitochondria and these preparations were stained either with Haidenhain's iron-haematoxylin or by Benda's and Champy-Kull's methods. Janus Green B was prepared and used as a vital, mitochondrial dye according to the following method: A drop of 0.1 per cent Janus green B dissolved in 95 per cent ethyl alcohol was placed on a clean cover slip rimmed with vaseline and allowed to dry. Test material suspended in water was then placed upon the cover slip, inverted, and sealed onto a clean glass slide. The preparation was then ready for microscopic examination. For nuclear details, Newton's gentian violet stain proved