Macrobrachium rosenbergii (de Man 1879): the antennal gland and the role of pheromones in mating behaviour

The freshwater prawn, Macrobrachium rosenbergii (de Man, 1879) is an important aquaculture species but one that has the disadvantage of heterogeneous individual growth (HIG) according to different morphotypes. Chemical cues, especially, pheromones, are one of the most important communication types between individual prawns, along with visual and tactile methods. Testing pheromones, whilst restricting other cues, may therefore lead to a better understanding of the influence of these communicatory compounds on the prawn reproductive process. The three principle objectives of this study were therefore: 1) To examine the effect of moult stage and morphotype on pheromone-induced sexual behaviour 2) To examine the role of pheromone / urine concentrations on sexual attraction behaviour 3) To describe the functional morphology of the antennal gland and examine its possible role in pheromone production and release. Identical bioassay tanks were designed and constructed to study the reproductive behaviour of prawns. Experiments were set up to examine responses to pheromone release by live prawns over 30 minutes and behavioural response observations were made with the aid of a Closed-Circuit Videotape System (CCVS). Results were statistically analysed using a repeated measures general linear model (GLM). Three trials were designed to test the effect of moult stage of both males and females and male morphotypes on sexual attraction behavioural responses. Twelve prawns were used for each trial and each prawn was used five times (1 no-pheromone control and 4 for experimental tests). The first trial studied the effect of female moult stages (pre-, inter and newly-moulted) on sexual attraction behaviour of blue claw (BC) male. Results of this trial showed that newly-moulted females spent significantly (p<0.05) less time approaching the BC male than the pre- and inter-moult females. The second trial studied the effect of male moult stage (pre-, inter and newly-moulted) on sexual attraction to receptive females. Results showed that the time taken by the inter-moult males was (p<0.05) less than the pre- and newly-moulted males in approaching the newly-moulted female. The third trial tested the effect of male morphotypes (small male, SM, orange claw, OC and dominant blue claw, BC) on sexual attraction behaviour towards newly-moulted females. Results showed that the BC male was significantly more attractive (p<0.05) than other morphotypes to newly-moulted females and that the OC male was the least attractive. The role of moulting stage for both male and female prawns on reproductive response behaviour was investigated. Because BC males responded significantly faster towards newly-moulted female more than to either pre-or inter-moult females, results of the first trial suggest that BC males are able to use different chemical cues to gather information about a conspecific’s gender and can differentiate female’s moult stages. Since BC males responded significantly faster towards newly-moulted females more than to either pre-or inter-moult females, this suggests that females at this particular stage released a distinct sexual pheromone or concentration of pheromone that differed from those pheromones released by both pre- and inter-moult females. In contrast, newly-moulted females prefer the inter-moult BC males which indicate that females have an ability to distinguish the moult status of BC males. Furthermore, it indicates that pheromone characteristics change with the moult status of BC males. Also, newly-moulted females are most likely to be avoiding the potential costs of mate guarding with soft shell BC males. Results obtained from the third trial suggested that a newly-moulted female can discriminate male morphotypes (SM, OC and BC) from their pheromone cues. This indicates that male morphotypes release pheromones which differ from each other in some way. Newly-moulted females responded positively to both SM and BC males with different levels of attraction with the greatest attraction to BC males to BC males suggesting that pheromone released from the BC male may carry information relating to dominance status. Urine is believed to be one of the main carriers of pheromone and is usually released from the antennal gland. Different urine concentrations (0.1, 1.0, 2.0, 3.0, 5.0 and 10µl l-1) of collected urine from BC males were used to test the sexual attraction behaviour of receptive newly-moulted females. Also, the attractant capability of fresh urine following exposure to different temperature regimes (cooled at 4ºC, frozen at -70ºC and heated at 70ºC) was tested. Since newly-moulted female M. rosenbergii were attracted to BC male urine, this indicates the existence of sex pheromone in the fresh urine. Also, it was found that the sexual response of females to fresh urine of BC males was directly proportional to urine concentration with faster responses observed with increasing urine concentrations. At the three fresh urine concentrations 0.1 µl l-1, 1.0 µl l-1 and 2.0 µl l-1, statistical analysis indicated no significant difference (p>0.05) between these three concentrations while a significant (P<0.05) response was to concentrations more than 3.0 µl l-1. This may indicate that these three concentrations were not sufficient to elicit attraction behaviour in newly-moulted females. A concentration of 3.0 µl l-1 of fresh urine is suggested to be a sufficient concentration to elicit a significant sexual attraction under laboratory conditions. Response of newly-moulted female prawns to the various temperature treatments tested declined in response to nominally increasingly degradative treatments. Also, statistical analysis showed that temperature treatment and concentration added both had a significant effect on the response of females. The greatest degradation of urine attractiveness was found with the 70ºC heat treatment. It can be concluded that the pheromone components of prawn urine are friable when exposed to high temperatures. Using light and transmission electron microscopes, ultrastructural observation of the antennal gland (AG) of M. rosenbergii suggests that it has four distinct regions, the coelomosac, the nephridial tubules, the labyrinth and the bladder. Morphological and functional descriptions of each of these regions were compared with those of other aquatic Crustacea.