THE LOCOMOTOR BEHAVIOR OF LYGODIUM AND MARSILEA SPERM

Flagella of living sperm of the ferns, Lygodium japonicum (Thunb.) Sw. and Marsilea vestita Hook. and Grev., beat three dimensionally with a continuous traveling helical wave. The wave is propagated from base to tip of the flagellum. Flagella of Lygodium and Marsilea complete 65 and 30 beat cycles per sec, respectively. Each flagellum circumscribes an open conicoid oriented in a latero-posterior direction. Only dead sperm have anteriorly directed flagella as illustrated in plant morphology textbooks. FIXED AND STAINED fern sperm have been observed and described adequately by light and electron microscopy (Lagerberg, 1906; Manton, 1959; Rice and Laetsch, 1967; Duckett and Bell, 1971). However, little is known about the locomotor behavior of living fern sperm. High speed cinemicrography and frame by frame analysis have made it possible to observe the locomotor behavior of rapidly swimming sperm of Lygodiun and Marsilea. MATERIALS AND METHODS-Large numbers of antheridia were obtained by growing the gametophytes of Lygodium japonicum (Thunb.) Sw. for four weeks on a 1.5 % agar medium containing the minerals of Bold (1967b) and 10-5 M gibberellic acid (GA3). Sperm of the water fern, Marsilea vestita, Hook. and Grev. were obtained from sporocarps by methods described by Rice and Laetsch (1967). Lygodium sperm were released from antheridia into distilled water and Marsilea sperm into Sparkletts brand spring water. Vaseline-supported coverslips were made in order to insure free sperm mobility and to prevent evaporation. Cinemicrography was done with a Red Lake Lo Cam high speed camera mounted on a Zeiss phase contrast microscope. Eastman 4 x Negative film exposed at 400 frames per sec was analyzed with an L and W photo,analyzer by tracing successive individual frames projected upward by means of a mirror onto a glass-topped table. RESULTS-Sperm of both Lygodium and Marsilea have an anterior nuclear coil bearing flagella and a posterior cytoplasmic vesicle (Fig. 1, 2). The anterior coil of Lygodium consists of 2-3 'Received for publication 1 October 1973. Supported by Grant GB-24840 from the National Science Foundation to T. L. Jahn. gyres, while that of Marsilea has 9-11 gyres. Although it is not possible to determine the number of flagella by cinemicrography, Manton (1959) reported that another fern sperm (Pteridium), which is similar in structure to Lygodium, does have approximately 40 flagella, and Mizukumi and Gall (1966) reported that Marsilea sperm possess about 100 flagella. The flagella of viable sperm are always directed in a latero-posterior direction. Anterior orientation of the flagella indicates a lack of locomotor control and approaching death. When released from the antheridium, Lygodium sperm swim at rates of 150t per sec in large helices and often abruptly change direction. Marsilea sperm swim more slowly at rates of 120 f per sec, traversing a constant unidirectional path. High speed cinemicrography shows that the flagella of both Lygodium and Marsilea beat three dimensionally with a continuously traveling helical wave (Fig. 1-3). An individual flagellum circumscribes an open conicoid oriented in a lateroposterior direction. Only one undulatory wave is propagated from base to tip along the flagellum during each beat cycle. Flagella of Lygodium and Marsilea complete 65 and 30 beat cycles per sec, respectively. Since the continuous wave form cannot be resolved into components, no power or recovery stroke can be observed. The flagella function in a coordinated manner. All flagella do not beat at the same time. A metachronal or synchronous wave is initiated at the apex of the anterior coil and proceeds down the spiral. With the completion of four metachronal waves or after each individual flagellum has completed four beat cycles, the body of Lygodium sperm makes a complete rotation. Marsilea sperm rotate more slowly, completing a rotation after eight metachronal waves or flagellar beat cycles.