Repeated evolution of local adaptation in swimming performance: population‐level trade‐offs between burst and endurance swimming in Brachyrhaphis freshwater fish

Specialization is fundamentally important in biology because specialized traits allow species to expand into new environments, in turn promoting population differentiation and speciation. Specialization often results in trade‐offs between traits that maximize fitness in one environment but not others. Despite the ubiquity of trade‐offs, we know relatively little about how consistently trade‐offs evolve between populations when multiple sets of populations experience similarly divergent selective regimes. In the present study, we report a case study on Brachyrhaphis fishes from different predation environments. We evaluate apparent within/between population trade‐offs in burst‐speed and endurance at two levels of evolutionary diversification: high‐ and low‐predation populations of Brachyrhaphis rhabdophora, and sister species Brachyrhaphis roseni and Brachyrhaphis terrabensis, which occur in high‐ and low‐predation environments, respectively. Populations of Brachyrhaphis experiencing different predation regimes consistently evolved swimming specializations indicative of a trade‐off between two swimming forms that are likely highly adaptive in the environment in which they occur. We show that populations have become similarly locally adapted at both levels of diversification, suggesting that swimming specialization has evolved rather rapidly and persisted post‐speciation. Our findings provide valuable insight into how local adaptation evolves at different stages of evolutionary divergence.     

A Quick Method for Species Identification of Japanese Eel (<Emphasis Type="Italic">Anguilla japonica</Emphasis>) Using Real-Time PCR: An Onboard Application for Use During Sampling Surveys

To compensate for the limited number of morphological characteristics of fish eggs and larvae, we established a convenient and robust method of species identification for eggs of the Japanese eel (Anguilla japonica) using a real-time polymerase chain reaction (PCR) that can be performed onboard research ships at sea. A total of about 1.2 kbp of the mitochondrial 16S ribosomal RNA gene sequences from all species of Anguilla and 3 other anguilliform species were compared to design specific primer pairs and a probe for A. japonica. This real-time PCR amplification was conducted for a total of 44 specimens including A. japonica, A. marmorata, A. bicolor pacifica, and 6 other anguilliform species. Immediate PCR amplification was only observed in A. japonica. We then tested this method under onboard conditions and obtained the same result as had been produced in the laboratory. These results suggest that real-time PCR can be a powerful tool for detecting Japanese eel eggs and newly hatched larvae immediately after onboard sampling during research cruises and will allow targeted sampling efforts to occur rapidly in response to any positive onboard identification of the eggs and larvae of this species.     

Kinematic Comparison of Forward and Backward Swimming and Maneuvering in a Self-Propelled Sub-Carangiform Robotic Fish

We make a thorough kinematic comparison of forward and backward swimming and maneuvering on a self-propelled robot platform that uses sub-carangifbrm swimming as the primary propulsor. An improved Central Pattern Generator （CPG） model allowing free adjustment of phase relationship and directional bias is employed to achieve flexible swimming and smooth transition. Considering the characteristics of forward swimming in carangiform fish and backward swimming in anguilliform fish, various backward swimming patterns for the sub-carangiform robotic fish are suitably created by reversing the direction of propagating propulsive waves. Through a combined use of the CPG control and closed-loop swimming direction control strategy, flexible and precise turning maneuvers in both forward and backward swimming are implemented and compared. By contrast with forward swimming, backward swimming requires a higher frequency or an increased lateral displacement to reach the same relative swimming speed. Noticeably, the phase difference shows a greater impact on forward swimming than on backward swimming. Our observations also indicate that the robotic fish achieves a larger turning rate in forward maneuvering than in backward maneuvering, yet these two maneuvers display comparable turning precision.     

Do blind cavefish have behavioral specializations for active flow-sensing?

Blind cavefish use a form of active sensing in which burst-coast swimming motions generate flow signals detected by the lateral line. To determine if blind cavefish have evolved behavioral specializations for active flow-sensing, including the ability to regulate flow signal production through lateral line feedback, the swimming kinematics of blind and sighted morphs of Astyanax were compared before and after 24?h of familiarization with a novel, dark environment and with and without lateral line functionality. Although both morphs showed little difference in the vast majority of kinematic parameters measured, blind morphs differed significantly from sighted morphs in having a much higher incidence of swim cycle sequences devoid of sharp turns. Both lateral line deprivation and familiarization with the arena led to significant declines in this number for blind, but not sighted morphs. These findings suggest that swimming kinematics are largely conserved, but that blind morphs have nevertheless evolved enhanced abilities to use lateral line feedback when linking swim cycles into continuous, straight trajectories for exploratory purposes. This behavioral specialization can best be understood in terms of the intermittent and short-range limitations of active flow-sensing and the challenges they pose for spatial orientation and navigation.     

Experimental quantification of the swimming performance and behaviour of spawning run river lamprey Lampetra fluviatilis and European eel Anguilla anguilla

Using a large-scale open-channel flume, the swimming ability and behaviour of individual adult European eel Anguilla anguilla and river lamprey Lampetra fluviatilis, species that exhibit anguilliform locomotion, were quantified under complex hydraulic conditions created by a 0·2-0·3 m high under- or overshot weir during four discharge regimes. Fishes were allowed to approach the weirs from both up- and downstream. All fishes passed the undershot weir, independent of discharge and direction of movement, and under high flow (mean ±S.E. 194·63 ± 6·48 l s(-1)) moved upstream against velocities that ranged between 1·75 and 2·12 m s(-1), suggesting greater maximum swimming capability than previously reported. In comparison, passage efficiency during upstream movement was lower for the overshot weir for both L. fluviatilis and A. anguilla. Downstream moving A. anguilla took longer to pass the over- than undershot weir. This study describes a methodology to attain realistic measures of swimming ability and behavioural performance required to develop multispecies fish passage criteria.     

Swimming behavior and ultrastructure of sperm of<Emphasis Type="Italic"> Lygodium japonicum</Emphasis> (Pteridophyta)

Swimming behavior of the sperm of Lygodium japonicum (Pteridophyta) and the associated ultrastructure of the flagellar apparatus were studied by video microscopy, transmission electron microscopy (TEM) and scanning electron microscopy (SEM). The sperm has approximately 70 flagella that emerge from a sinistrally-coiled flagellar apparatus, and swims forward by ciliary beat of these flagella. Backward swimming was not observed even after sperm collided with obstacles. Video microscopy showed that the flagella of the swimming sperm are oriented laterally and oblique-anteriorly. TEM and SEM observations revealed that the basal bodies of these flagella are arranged in at least two rows and oriented in the same directions as observed by video microscopy. These basal bodies (flagella) are categorized into two types according to their orientation: group I (laterally directed) and group II (oblique-anteriorly directed). The directionality of the basal bodies appears to be fixed by electron-dense material around their base. The outer dynein arms of the flagellar axoneme are entirely absent. These morphological characteristics of basal bodies (flagella) may relate to the lack of backward swimming behavior of the sperm. Based on these results, the evolution of swimming behavior in the archegoniates is discussed in connection with lack of backward swimming in a distantly related green alga, Mesostigma viride, and the Streptophyta.     

Disentangling the functional roles of morphology and motion in the swimming of fish

In fishes the shape of the body and the swimming mode generally are correlated. Slender-bodied fishes such as eels, lampreys, and many sharks tend to swim in the anguilliform mode, in which much of the body undulates at high amplitude. Fishes with broad tails and a narrow caudal peduncle, in contrast, tend to swim in the carangiform mode, in which the tail undulates at high amplitude. Such fishes also tend to have different wake structures. Carangiform swimmers generally produce two staggered vortices per tail beat and a strong downstream jet, while anguilliform swimmers produce a more complex wake, containing at least two pairs of vortices per tail beat and relatively little downstream flow. Are these differences a result of the different swimming modes or of the different body shapes, or both? Disentangling the functional roles requires a multipronged approach, using experiments on live fishes as well as computational simulations and physical models. We present experimental results from swimming eels (anguilliform), bluegill sunfish (carangiform), and rainbow trout (subcarangiform) that demonstrate differences in the wakes and in swimming performance. The swimming of mackerel and lamprey was also simulated computationally with realistic body shapes and both swimming modes: the normal carangiform mackerel and anguilliform lamprey, then an anguilliform mackerel and carangiform lamprey. The gross structure of simulated wakes (single versus double vortex row) depended strongly on Strouhal number, while body shape influenced the complexity of the vortex row, and the swimming mode had the weakest effect. Performance was affected even by small differences in the wakes: both experimental and computational results indicate that anguilliform swimmers are more efficient at lower swimming speeds, while carangiform swimmers are more efficient at high speed. At high Reynolds number, the lamprey-shaped swimmer produced a more complex wake than the mackerel-shaped swimmer, similar to the experimental results. Finally, we show results from a simple physical model of a flapping fin, using fins of different flexural stiffness. When actuated in the same way, fins of different stiffnesses propel themselves at different speeds with different kinematics. Future experimental and computational work will need to consider the mechanisms underlying production of the anguilliform and carangiform swimming modes, because anguilliform swimmers tend to be less stiff, in general, than are carangiform swimmers.     

Burrowing behavior and ecology of the crab-eating Indian snake eel Pisoodonophis boro

Synopsis The snake eel Pisoodonophis boro burrows, causing leaks in the embankments and damaging the paddy fields and salt pans near estuaries. Field observations and laboratory experiments were made to study this behavior. P. boro was burrowing to eat the fiddler crab Uca annulipes in the mud flats. The eel showed a patchy distribution within the Uca zone. Salinity and the physical nature of the deposits controlled the distribution of the eel. Eel population density was low when the estuary was completely filled with neritic waters during the summer and fresh water during the monsoon period. The region of greatest abundance contained a good mixture of sand, silt and clay. Eels were not found where medium and fine sand formed the bulk of the substratum. The laboratory experiments showed that P. boro preferred loam soil although it could invariably burrow into hard substratum like sand for protection. The eel adapted itself to the experimental substrates ranging from sand to fine clay. However, their natural distribution was determined by Uca distribution. As U. annulipes is not found either in salt pans or in paddy fields P. boro rarely occurs in these habitats.     

Proliferation of elongate fishes in the deep sea

It was hypothesized that energetically efficient anguilliform swimming and axial elongation in fishes is favoured in the deep sea and predicted that the degree of elongation of the body form of fishes would increase with depth. An index of fish shape was derived from the relationship between length and mass. This was combined with data on abundance of c. 266 fish species from 389 research trawl tows made at depths of between 300 and 2030 m in the north‐east Atlantic Ocean. The degree of elongation of the fish increased with depth to c. 1250 m before levelling off. The strength of this phenomenon varied between higher level taxa, being most apparent in the Gadiformes and Osmeriformes, and weak or absent in the Perciformes and Selachimorpha. The advantage of efficient elongate body forms may explain why certain taxa such as the grenadiers (Macrouridae) have dominated the deep sea, some have restricted depth ranges, e.g. the sharks, skates and rays, and others are almost entirely absent, e.g. the flatfishes (Pleuronectiformes).     

Swimming ability and burrowing time of two cirolanid isopods from different levels of exposed sandy beaches

Most of the macroinfauna from sandy beaches is highly mobile, emerging out of the sediment when the tide rises, and using the swash to migrate up and down the beach face or feed (searching for prey or carrion). After swash excursions, they usually burrow back into the sediment, maintaining zonation at low tide. Therefore, the different species abilities to emerge, move around and burrow under different swash climates and sediment conditions are expected to influence observed distribution patterns. Nonetheless, few attempts have been made to understand behavioral mechanisms of these organisms in moving fluids.In this study, we used a flume tunnel to investigate the orientation, swimming ability and burrowing time of two similar species of cirolanid isopods, Excirolana armata Dana and Excirolana braziliensis Richardson, under current velocities ranging from 5 to 30 cm·s⁻¹. E. armata inhabits middle levels of dissipative to intermediate beaches, while E. braziliensis is found towards the upper level of a wider range of beach states. Both species oriented countercurrent above a threshold velocity, which turned out to be significantly lower for E. armata than for E. braziliensis. E. armata proved to be a stronger swimmer as shown by the higher velocities surmounted, and the less drags experienced at the highest current velocity. E. armata also burrowed faster than E. braziliensis. Burrowing time was affected by sediment grain size and water content, but not by water flow. Once organisms managed to begin burrowing under different flow conditions, they were not affected by current velocity. Nonsaturated sand precluded burial, while coarse sand retarded it. Differences in the observed patterns of across-beach distribution may thus be the result of species-specific behavioral responses to swash climate, manifested in swimming ability, burying and orientation in directional flows.     

Reynolds number limits for jet propulsion: a numerical study of simplified jellyfish

The Scallop theorem states that reciprocal methods of locomotion, such as jet propulsion or paddling, will not work in Stokes flow (Reynolds number=0). In nature the effective limit of jet propulsion is still in the range where inertial forces are significant. It appears that almost all animals that use jet propulsion swim at Reynolds numbers (Re) of about 5 or more. Juvenile squid and octopods hatch from the egg already swimming in this inertial regime. Juvenile jellyfish, or ephyrae, break off from polyps swimming at Re greater than 5. Many other organisms, such as scallops, rarely swim at Re less than 100. The limitations of jet propulsion at intermediate Re is explored here using the immersed boundary method to solve the 2D Navier-Stokes equations coupled to the motion of a simplified jellyfish. The contraction and expansion kinematics are prescribed, but the forward and backward swimming motions of the idealized jellyfish are emergent properties determined by the resulting fluid dynamics. Simulations are performed for both an oblate bell shape using a paddling mode of swimming and a prolate bell shape using jet propulsion. Average forward velocities and work put into the system are calculated for Re between 1 and 320. The results show that forward velocities rapidly decay with decreasing Re for all bell shapes when Re<10. Similarly, the work required to generate the pulsing motion increases significantly for Re<10. When compared to actual organisms, the swimming velocities and vortex separation patterns for the model prolate agree with those observed in Nemopsis bachei. The forward swimming velocities of the model oblate jellyfish after two pulse cycles are comparable to those reported for Aurelia aurita, but discrepancies are observed in the vortex dynamics between when the 2D model oblate jellyfish and the organism. This discrepancy is likely due to a combination of the differences between the 3D reality of the jellyfish and the 2D simplification, as well as the rigidity of the time varying geometry imposed by the idealized model.     

Evidences of habitat displacement between two common soft-bottom SW Atlantic intertidal crabs

The intertidal burrowing crab Chasmagnathus granulatus Dana is the dominant species in soft sediments and vegetated intertidal areas along the SW Atlantic estuaries (southern Brazil 28°S to the northern Argentinean Patagonia 41°S) where it produces dense and extensive burrowing beds. The mud crab Cyrtograpsus angulatus Dana coexists with Ch. granulatus in this area, but it also inhabits areas to the south (northern and central Argentinean Patagonia). A survey covering both areas showed that C. angulatus rarely live in burrows when coexisting with Ch. granulatus, but form large burrowing beds when not coexisting with Ch. granulatus. When both species coexisted, burrowing beds of C. angulatus are restricted to sandy-muddy areas. Only rarely are burrows of C. angulatus found within Ch. granulatus beds. However, when Ch. granulatus were experimentally excluded within their burrowing beds, new settlers of C. angulatus made burrows and maintained them until they reached large size. Paired (inside and outside Ch. granulatus burrowing bed) sampling during high tide using beach nets showed that C. angulatus rarely venture inside the Ch. granulatus crab beds. Other field experiments showed that adults Ch. granulatus always displace C. angulatus from burrows. Furthermore, in several sites located south of the limit of distribution of Ch. granulatus at the Patagonian coast, soft bare intertidals are dominated by burrowing beds of C. angulatus mixed with the congener C. altimanus Dana. Together, these evidences suggest that the mud crab C. angulatus is displaced from soft bottom areas by the burrowing crab Ch. granulatus. It is an example of competitive exclusion through aggressive interference in soft-bottom habitats when the shared resource is the access to sediment surface, a two-dimensional well-defined resource.     

Diel activity patterns in Metapenaeus and Penaeus juveniles

Small (5–10.9 mm carapace length), medium (11–15.9 mm), and large (16–20.9 mm) juveniles of Metapenaeus anchistus, Metapenaeus sp., Penaeus monodon and P. merguiensis were stocked individually in glass tanks provided with sand substrate, sea water, artificial bamboo shelter, aeration and food. The seven activity types (recorded for each shrimp hourly for 24 h) were classified as below (burrowing) or above substrate (swimming, walking, stationary, in shelter, feeding and cleaning). Shrimp juveniles exhibited a strong diel periodicity — emergence and activity at night and burrowing in the day. The chi-square test showed that type of activity (above/below substrate) was associated with period (light/dark). Diurnal burrowing was greater among Metapenaeus than Penaeus; inversely, above substrate activities were more frequent for Penaeus species compared to Metapenaeus. Feeding was the major above substrate and nocturnal activity for M. anchistus, Metapenaeus sp. and P. monodon. Only P. Monodon used the shelter consistently. Frequency of the 7 activity types was dependent on juvenile size for Penaeus, e.g., the preference for shelters shifted to burrowing with increase in size in P. monodon. Results are discussed in relation to the importance of mangrove habitats in providing shelter to penaeids, in particular the mangrove-associated P. monodon and P. merguiensis.     

Myoseptal architecture of sarcopterygian fishes and salamanders with special reference to Ambystoma mexicanum

During axial undulatory swimming in fishes and salamanders muscular forces are transmitted to the vertebral axis and to the tail. One of the major components of force transmission is the myoseptal system. The structure of this system is well known in actinopterygian fishes, but has never been addressed in sarcopterygian fishes or salamanders. In this study we describe the spatial arrangement and collagen fiber architecture of myosepta in Latimeria, two dipnoans, and three salamanders in order to gain insight into function and evolution of the myoseptal system in these groups. Salamander myosepta lack prominent cones, and consist of homogenously distributed collagen fibers of various orientations that never form distinct tendons. Fiber orientations are difficult to homologize with those of fish myosepta. The myosepta of Latimeria and dipnoans (Protopterus and Neoceratodus) illustrate that major changes in architecture occurred in the sarcopterygian clade (loss of horizontal septum), in the rhipidistian (dipnoans + tetrapods) clade (loss of epineural and epipleural tendon), and in tetrapods (loss of lateral tendons and myoseptal folding). When compared to fishes, the myosepta of wholly aquatic salamanders (Ambystoma mexicanum, Amphiuma tridactylum, Necturus maculosus) do not have the lateral tendons we suppose serve to transfer muscular forces posteriorly. We propose that alternative structures (most conspicuously present in Ambystoma) perform this function: posteriorly the relative amount of connective tissue increases considerably, and myosepta are disintegrated to horizontal lamellae of connective tissue. The structures thought to be involved in modulation of body stiffness in fishes during swimming are also absent in salamanders. Our data also have implications for the hypothesis that salamander hypaxial myosepta are designed to increase shortening amplification of the hypaxial muscle fibers. The posterior hypaxial myosepta of all three salamander species possess only mediolaterally directed collagen fibers, which would indeed amplify the shortening of the associated muscle.     

Can bivalve veligers escape feeding currents of adult bivalves?

While the stock of introduced Pacific oysters (Crassostrea gigas) increased in the Oosterschelde estuary (SW Netherlands), so did the filtration pressure of all bivalve species together. In the same period, stocks of native bivalves declined slightly. The expansion of Pacific oysters in Dutch estuaries might be partially due to better abilities of their larvae to avoid or escape filtration, compared to larvae of native bivalves. In this context, escape and swimming abilities of Pacific oyster larvae and the larvae of the native blue mussel (Mytilus edulis) were compared.Swimming behaviour of C. gigas larvae and larvae of M. edulis was recorded in still water and in a suction current mimicking a bivalve feeding current, in a horizontal and in a vertical plane. Larval swimming behaviour in a suction flow field was reconstructed by subtracting local water movement vectors from the total movement of larvae, yielding movement paths due to larval swimming alone.Swimming speeds and the rate of displacement in vertical direction of C. gigas and M. edulis larvae were related to larval shell length, and to the pitch of up- or downward swimming.Larvae of both species did not show escape reactions in a suction flow field. With increasing shell length, larval swimming speeds of both species increased significantly. Swimming speeds of C. gigas larvae were significantly higher than swimming speeds of M. edulis larvae, resulting in a faster vertical displacement. The ability to migrate to more favourable water layers faster may offer C. gigas an advantage over native bivalves with slower swimming larvae.     

Freshwater habitat use by a moray eel species,Gymnothorax polyuranodon,in Fiji shown by otolith microchemistry

Freshwater eels of the Anguillidae are diadromous because they migrate between ocean and freshwater environments, but other anguilliform fishes are generally considered to be strictly marine species. A few marine eels of the Muraenidae and Ophichthidae have occasionally been found in freshwater or estuaries, indicating that anguillids are not the only anguilliform eels that can use freshwater in some parts of the world. The moray eel Gymnothorax polyuranodon is one species that is known to be present in freshwater in the Indo-Pacific, but its life history is unknown. One way to evaluate what types of habitats are used by fishes is to determine the ratio of strontium (Sr) to calcium (Ca) in their otoliths, because this can show if they have used freshwater or saltwater environments. To evaluate the patterns of freshwater use by this unusual species of marine eel, the otolith Sr/Ca ratios of four G. polyuranodon (275–344 mm) caught in a freshwater stream of Fiji were analyzed. The consistently low Sr/Ca values (0–4) indicated upstream movement after settlement and freshwater or estuarine residence of all four individuals. These eels did not appear to have entered freshwater just for a short time period, which is consistent with other reports that this species is present in estuarine and freshwater habitats. This suggests that G. polyuranodon may be a catadromous species of marine eel. The similarities and differences between the life histories of anguillid eels and the few marine eels that have evolved the ability to invade freshwater habitats is discussed in relation to the evolutionary origin of diadromy in anguilliform fishes that originated in the marine environment.     

The Effect of Drag and Attachment Site of External Tags on Swimming Eels: Experimental Quantification and Evaluation Tool

Telemetry studies on aquatic animals often use external tags to monitor migration patterns and help to inform conservation effort. However, external tags are known to impair swimming energetics dramatically in a variety of species, including the endangered European eel. Due to their high swimming efficiency, anguilliform swimmers are very susceptibility for added drag. Using an integration of swimming physiology, behaviour and kinematics, we investigated the effect of additional drag and site of externally attached tags on swimming mode and costs. The results show a significant effect of a) attachment site and b) drag on multiple energetic parameters, such as Cost Of Transport (COT), critical swimming speed (U_crit) and optimal swimming speed (U_opt), possibly due to changes in swimming kinematics. Attachment at 0.125 bl from the tip of the snout is a better choice than at the Centre Of Mass (0.35 bl), as it is the case in current telemetry studies. Quantification of added drag effect on COT and U_crit show a (limited) correlation, suggesting that the U_crit test can be used for evaluating external tags for telemetry studies until a certain threshold value. U_opt is not affected by added drag, validating previous findings of telemetry studies. The integrative methodology and the evaluation tool presented here can be used for the design of new studies using external telemetry tags, and the (re-) evaluation of relevant studies on anguilliform swimmers.     

Morphological specializations in heterocongrinae (Anguilliformes: Congridae) related to burrowing and feeding

The remarkable lifestyle of heterocongrines has drawn the attention of many authors in the past, though no or little attention has been paid to the morphology of the tail and the head of these species. In order to examine the true nature of possible morphological specializations of the head and tail and their relation to their tail-first burrowing habit and/or feeding mode, a detailed myological and osteological study of Heteroconger hassi and Heteroconger longissimus was performed. The osteological similarities of the cranial skeleton between H. hassi and H. longissimus are striking. Most of the cranial muscles show no variation in presence, insertion or origin between these two species except for the adductor mandibulae complex, the adductor hyomandibulae and the intermandibularis. The adductor mandibulae complex is small, compared to that of other anguilliform species, and is probably related to their suction-dominated feeding mode and a diet, comprising mainly small, soft prey items. Heterocongrinae have undergone several morphological specializations in the tail for their tail-first burrowing lifestyle. The skeleton and musculature of the tail of H. hassi and H. longissimus are similar. In both species the caudal skeleton is highly reduced and fortified, forming a firm, pointed burrowing tool. Intrinsic caudal musculature is reduced and some muscles (interradials, supracarinalis) are even absent.     

A mitochondrial DNA based phylogeny of weakfish species of the Cynoscion group (Pisces: Sciaenidae)

We infer the phylogeny of fishes in the New World Cynoscion group (Cynoscion, Isopisthus, Macrodon, Atractoscion, Plagioscion) using 1603 bp of DNA sequence data from three mitochondrial genes. With the exception of Plagioscion, whose position was ambiguous, the Cynoscion group is monophyletic. However, several genera examined are not monophyletic. Atlantic and Pacific species of Cynoscion are interspersed in the tree and geminate species pairs are identified. Intergeneric relationships in the group are clarified. Our analysis is the first comprehensive phylogeny for the Cynoscion group based on molecular data and provides a baseline for future comparative studies of this important group.