COPULATORY BEHAVIOUR OF MALE PANAGRELLUS REDIVIVUS

BY

C. L. DUGGAL Department of Zoology and Applied Entomology, Imperial College Field Station, Ashurst Lodge, Silwood Park, Ascot, Berkshire, England

The copulatory behaviour of Panagrellus redivivus is described and that of the male related to its age and reproductive state. Some observations are made on the structure of the male reproductive system. Male P. redivivus show both pre- and post-insemination coiling around the female and use their spicules to probe and open the female gonopore, and as a channel for the sperm. The frequency of copulation of male nematodes and its effect on the number of sperm produced and on the nematodes' life-span was examined and related to development and longevity of ageing virgin males. The number of sperm shed into the uterus of the female increased with increasing intervals between copulations.

Anya (1976) and Lee & Atkinson (1976) reviewed the reproductive physi- ology of nematodes. It is apparent that little is known of copulatory behaviour of nematodes and the process of copulation has been described in few species. In particular, few attempts have been made to quantify the copulatory behaviour of males during their life span. Phillipson (1970) suggested that male Nippostrongylus bra.rilien.ri.r could inseminate one female every 2 hr during the first 20 hr of adult life, while Fisher (1972) observed that male ApbelencbAJ avenae copulated 18-26 times in 5 weeks and noted that the interval between successive copulations appeared to increase with age. In male Rhabditis pellio the mean number of matings per day decreased with age (Somers, Shorey & Gaston, 1977 ) . In the present work the copulatory behaviour of male Panagrellus redivivu.r Goodey of different ages is described.

MATERIALSAND METHODS

P. redivivus was cultured monoxenically (Duggal, 1978). The behaviour of nematodes was observed in 50 mm diameter plastic Petri dishes containing 1.5 % (w/v) Davis agar inoculated with Escherichic¡ coli. All experiments were at 25°. To study the copulatory behaviour of adult male P. redivivu.r, a moulting fourth-stage male and four newly moulted virgin females were observed on an agar plate with E. coli. The behaviour of the males during and after its final moult was recorded. Four males were studied. To estimate the rate of copulation, the number of sperm transferred and the life span of a normally copulating male, a fourth-stage moulting male was placed with eight newly moulted, virgin females on an agar plate containing E. coli. At 258

12 hr intervals the number of females with which it had copulated and the number of sperm and fertilized eggs present in each was recorded. After each 12 hr period, the male was transferred to another plate with a fresh batch of newly moulted, virgin females and E. coli. During the day the behaviour of the male was also observed. Each male was observed until it died. Five males were studied. The development and life span of virgin males was examined by placing a fourth stage male on an agar plate with E. coli and observing its development at 12 hr intervals for 4 days after the final moult; the male being transferred to a fresh agar plate every 24 hr. Five males were studied. To determine the mortality rate of virgin males, about 70 fourth-stage males were separated from the main culture and allowed to moult. After 12 hr a batch of 50 males was separated; the nematodes were transferred to fresh agar plates every 24 hr and during each transfer the number of dead worms was recorded. This experiment was repeated three times. To determine the effects of short periods of isolation, fourth-stage males were kept individually on agar plates and the exact time of the final moult recorded. Six, 12, 18, 24 or 30 hr after moulting, an isolated male was placed with five newly moulted females and its copulatory activity observed. The duration of pre-insemination coiling, the number of sperm transferred, and the duration of post-insemination coiling and attachment were recorded. Four males or more were studied per isolation period. To determine the effects of long periods of isolation, two hundred fourth- stage males were allowed to moult on agar plates with E. coli. After 12 hr the moulted males were transferred to new plates. Thereafter, at 24 hr intervals ten males were transferred individually to fresh agar plates with five newly moulted, virgin females for 1 hr. The number of females with which they had copulated was recorded and the copulated females separated for 24 hr so that fertilization of their oocytes could occur; the number of eggs fertilized was then recorded. This experiment was repeated three times. RESULTS

T'he reproductive system of P. i,ediviz,u.v P. redivizwJ males are monorchic (Hcchler, 1971, and Fig. la ) . The reflexed anterior of the testis, the germinal zone, is followed by a wider growth zone and the seminal vesicle. The latter region is very distinctive in P. rediviv¡/J when many sperm are present, and has a granular appearance. Around the seminal vesicle is a band of seven to eight clear cells (Fig. 1a) previously undescribed of un- known function. All mature sperm were stored in the seminal vesicle before copulation. Between the seminal vesicle and the vas deferens previously un- described aggregates of small globules were often found also of unknown func- tion. These aggregates increased in number as the interval between copulations increased.