NUCLEAR BEHAVIOR IN THE BASIDIUM OF FOMES ANNOSUS

Meiotic division in Fomes annosus is similar to that reported for other higher fungi. Nuclei in dikaryotic cells prior to fusion in the basidia are long, thin, and double-stranded with paired heterochromatic areas. Various stages of prophase are similar to those in higher plants. At metaphase I and II seven pairs of chromosomes are aligned in a circle and the chromatids migrate to opposite poles established by two centrioles. The centrioles function in the movement of the nuclei in the basidium, nuclear alignment prior to fusion, establishment of poles for division, and the migration of the nuclei into the basidiospore. After nuclei migrate into the basidiospore, they soon divide, producing a binucleate spore. FOMES ANNOSUS (Fr.) Karst. is being investigated extensively because of the losses it causes to pine. There has been some work on the asexual nuclear behavior of this fungus (Roll-Hansen, 1940; Brushaber, Wilson, and Aist, 1967; Griffin and Wilson [in press]), but no information is available concerning nuclear activity in the basidium. In F. annosus asexual nuclear division occurs in a manner first described for Ceratocystis fagacearum (Bretz) Hunt (Griffin and Wilson, in press) and subsequently found in other fungi (Aist and Wilson, 1965, 1967; Wilson, Brushaber, and Aist, 1966; Brushaber et al., 1967). Chromosomes are associated in a bar of chromatin along one wall of the hypha at metaphase and the chromosomes migrate independently to the opposite wall. Division is always perpendicular to the long axis of the hypha. MATERIALS AND METHODS-Basidiocarps of F. annosus were collected in the duff attached to the base of shortleaf pines (Pinus echinata Mill.) near Fayetteville, Arkansas. This material was imbedded in paraffin, sectioned, and stained using rather extreme procedures for imbedding because of the toughness of basidiocarps of F. annosus. The following procedure was used: Small blocks (5 X 10 X 20 mm) of the basidiocarp were fixed in Carnoy's fluid for 6 hr, washed in water 12 hr, dehydrated through an ethyl alcohol series (10, 25, 50, 95, 100%), placed in an ethyl alcohol-xylene mixture (50-50) and transferred into pure xylene. A block of paraffin was suspended in the xylene for 24 hr, after which half of the xylene-paraffin mixture was poured off and more paraffin added. The blocks of the basidiocarp were heated to 59 C for 24 hr, and placed in pure paraffin maintained at 59 C for 1 week during which time it was transferred twice I Received for publication 6 February 1967. Published with the approval of the Director of the Arkansas Agricultural Experiment Station. into pure paraffin. The infiltrated blocks wet e then imbedded in paraffin using standard procedures. Sections were cut at 2A4M and attached to No. 1 coverslips with Carnoy's fluid. The paraffin was removed from the sections with xylene (4-10 min) and then stained, using the HCl-Giemsa technique described previously (Aist and Wilson, 1965). RESULTS-Vegetative nuclei of F. annosus are usually long, thin, and double-stranded. The two strands have paired heterochromatic regions and are usually connected at one end. Before fusion the centrioles precede two paired nuclei as they move distally in the basidium (Fig. 1). Each of these nuclei is double-stranded and pairing is apparent between heterochromatic regions of each nucleus. The heterochromatic regions of the four strands become arranged into continuous bands perpendicular to the long axis of the basidium (Fig. 2). The strands lose their distinctiveness and a diffuse interphase nucleus is evident (Fig. 3). Each nucleus in the dikaryotic cell has a nucleolus prior to fusion. Following nuclear fusion, one large nucleolus is evident at the distal end of the basidium and it persists through anaphase I. After this it becomes faint and disappears. We observed the five classical stages of the meiotic nucleus in F. annosus with only slight variations (Fig. 3-7). Leptotene appears as thin, barely visible chromonemata, most of which are oriented parallel to the long axis of the basidium (Fig. 3). In a zygotene-like phase, the chromosomes and heterochromatic regions become more distinct and homologous chromosomes are not visible (Fig. 4). At the pachytene-like stage, the chromosomes are visibly double and the heterochromatin of each strand is paired (Fig. 5). Distinct chromosomes are not visible. As the diplotene-like stage