Colonization of freshwater habitats by an introduced crayfish, <Emphasis Type="Italic">Procambarus clarkii</Emphasis>, in Southwest Iberian Peninsula

The introduction of some crustacean species has produced alterations of freshwater environments and declines of native species worldwide. The red swamp crayfish, Procambarus clarkii Girard, was introduced in the Southwest Iberian Peninsula in the 1970’s, producing severe impacts on rice agriculture and on native biota such as macrophytes, gastropods, native crayfish and amphibians. We studied the distribution of P. clarkii in two areas of SW Iberian Peninsula: the Sado River basin (SW Portugal), an area colonized by this species around 1990, and the Parque Natural del Entorno de Doñana (SW Spain), colonized soon after its introduction in the Iberian Peninsula, in the 1970’s. Our main goal was to determine which factors limit crayfish distribution, which could help to identify the most effective management practices to contain its spread. Procambarus clarkii was found in most types of water bodies, including small and shallow ones. Distance to a crayfish source was the single predictor variable explaining crayfish occurrence in most types of habitats and in both areas. The only exception was for the Sado permanent stream points, where crayfish presence was negatively affected by an interaction between elevation and flow velocity. Other habitat characteristics have apparently little or no importance for its successful colonization. Moreover, this study indicated that overland dispersal is apparently a frequent phenomenon in this species. Our findings can be used to determine which habitats are most likely to be colonized by the crayfish and to develop practical measures which may limit its spread and minimize its impacts.     

Morphological asymmetry and behavioral laterality in the crayfish, Procambarus clarkii

Lateral asymmetry is found widely among vertebrates, but is scarcely observed in invertebrates. Here, morphological asymmetry and behavioral laterality of a wild crayfish, Procambarus clarkii, was investigated. The carapace morphology of crayfish showed left–right differences; in some, the right side of the carapace was larger than the left side, while in others, the left side was larger. A bimodal distribution in the direction of escape behaviors induced by a tactile stimulus was also observed. Experimental crayfish were definitively divided into two groups: individuals that frequently jumped leftward (right type) and those that jumped rightward (left type). Moreover, carapace asymmetry and lateralized escape responses were significantly correlated. These results suggest that crayfish exhibit left–right dimorphism in natural populations. The ecological advantages and maintenance mechanisms underlying these behaviors are also discussed.     

Effects of leg movements on the synaptic activity of descending statocyst interneurons in crayfish,Procambarus clarkii

Crustacean postural control is modulated by behavioral condition. In this study, we investigated how the responses of descending statocyst interneurons were affected during leg movements. Intracellular recording was made from an animal whose statoliths had been replaced with ferrite grains so that statocyst receptors could be activated by magnetic field stimulation. We identified 14 morphological types of statocyst-driven descending interneurons. Statocyst-driven descending interneurons always showed an excitatory response to statocyst stimulation on either ipsilateral or contralateral side to the axon. The response of each statocyst-driven descending interneuron to statocyst stimulation was differently modulated by leg movements in different conditions. During active leg movements, six statocyst-driven descending interneurons were activated regardless of whether a substrate was provided or not. In other two statocyst-driven descending interneurons, the excitatory input during leg movements was stronger when a substrate was provided than when it was not. One statocyst-driven descending interneuron received an excitatory input only during leg movements on a substrate, whereas another statocyst-driven descending interneuron did not receive any input during leg movements both on a substrate and in the air. These results suggest that the descending statocyst pathways are organized in parallel, each cell affected differently by behavioral conditions.Abbreviations EMG electromyogram - NGI nonspiking giant interneuron - SDI statocyst-driven descending interneuron     

Histological organization of the intestine in the crayfish Procambarus clarkii

Six longitudinal ridges span the length of the intestine in the crayfish Procambarus clarkii. A simple columnar epithelium with tetralaminar cuticle lines the lumen. Folds of the epithelium overlie a dense irregular connective tissue packed with mixed acinar (alveolar) glands. Mucous secretions are probably involved with formation and lubrication of faecal strings; neither the nature nor the role of the serous secretions is immediately apparent. Aggregations of cells with large cytoplasmic vacuoles, called bladder cells, appear in the subepithelial connective tissue near the tops of the intestinal ridges. The bladder cells are suitably positioned to bolster the integrity of the ridges. Striated muscle of the intestine occurs in inner longitudinal and outer circular layers. The inner longitudinal layer consists of six strips, with one strip associated with the base of each intestinal ridge. The outer circular layer is essentially complete, but there are periodic apertures in this layer on the left and right sides of the intestine, providing nerves and haemolymph vessels with access to the interior of the gut. Based on histological features, and consistent with reports on other crayfish, we conclude that the intestine of P. clarkii has a proctodeal (ectodermal) origin.     

Asynchronous swimmeret beating during defense turns in the crayfish, Procambarus clarkii

Swimmeret beating was monitored in freely moving specimens of the crayfish Procambarus clarkii as they exhibited defense turn responses to tactile stimuli. Analysis of videotape records revealed alterations in swimmeret beating during turning responses compared to straight, forward walking. During turns, swimmerets beat with shorter periods and smaller amplitude power strokes than during straight walking. Coordination between swimmerets also changed. Swimmerets on the side toward which the animal turned tended to lag behind their contralateral partners, rather than beat in synchrony as in straight walking, and ipsilateral coordination was loosened relative to straight walking. Asynchronous swimmeret beating accompanied asymmetric motions of the uropods in a manner similar to that observed during statocyst-dependent equilibrium reactions in P. clarkii, but removal of the statoliths did not eliminate turn-associated responses of the swimmerets. The coordinated action of the swimmerets and uropods may contribute to the torque that rotates the animal in the yaw plane. Implications of the observed changes in swimmeret coordination for understanding the underlying neuronal control system are discussed.     

Methyl farnesoate controls adult male morphogenesis in the crayfish, Procambarus clarkii

Adult male crayfish Procambarus clarkii exist in two morphotypes. They continue to molt as adults, switching between Form Is and Form IIs. Form Is are primary reproductive types, with large chelae and spines on the ischiopodites of the third and fourth pair of walking legs. Form IIs are non-reproductive types with smaller chelae and no spines on the ischiopodites. We investigated the hormonal control of these transitions in two ways, by eyestalk ablation and by methyl farnesoate (MF) treatments. Eyestalk ablation accelerates molting and increases MF levels in the blood. MF is a hormone that regulates both reproduction and morphogenesis. MF concentrations were determined in two ways. The hemolymph samples were extracted first, then purified, using normal phase HPLC. The fractions containing MF were collected and analyzed for MF concentration, utilizing both internal and external standards by GC/MS. The other hemolymph samples were analyzed from individual animals by HPLC. The concentrations of ecdysteroids were determined by radioimmunoassay. In the control animals, 4 out of 4 untreated Form I males molted into Form II, while 6 out of 7 Form IIs molted into Form Is. Eight of 8 ablated Form Is molted into Form IIs as expected, while 5 of 5 ablated Form IIs molted into Form IIs, instead of Form Is. MF treatment of intact animals resulted in 6 of 7 Form Is becoming Form IIs and 5 of 6 Form IIs becoming Form IIs. These results were highly significant in comparison of Form I and IIs in each treatment (eyestalk intact, eyestalk ablated and eyestalk intact with MF) by a chi square analysis, P = 0.006, P < 0.0005, and P = 0.013, respectively. MF premolt blood levels suggested that Form IIs were produced in the presence of 1.3 ng/ml MF, while Form Is result from MF levels less than 0.5 ng/ml. Since both eyestalk ablation and MF treatment resulted in the failure of Form IIs becoming Form Is, it was concluded that the control of morphogenesis of primary reproductives (Form Is) depends on a low level of MF prior to the molt, while Form IIs are formed in the presence of increased levels of MF.     

Factors influencing the degradation of photoreceptor membrane in the crayfish,Procambarus clarkii

Summary Light-induced degradation of photoreceptor membrane in the crayfish was studied by quantitative light and electron microscopy. The production of lysosomal organelles within the photoreceptor cells was enhanced by presenting the light stimulus intermittently (i.e., flicker) or by doubling its intensity. The enhancement was seen primarily as an increase in the number and size of multivesicular bodies. As these stimulus conditions are likely to facilitate intracellular Ca⁺⁺ fluxes, the results are compatibl with recent speculations that Ca⁺⁺ ions may regulate membrane degradation. To test the possibility that Ca⁺⁺ acts as a signal coupling receptor stimulation with membrane loss, retinas were incubated in the dark with the ionophore A23187 in the presence or absence of external Ca⁺⁺. The results demonstrate that A23187 produces a Ca⁺⁺-dependent increase in lysosomal organelles, predominantly multivesicular bodies. These data are consistent with a role for intracellular Ca⁺⁺ in the degradative process; however, the exact locus of the effect is unclear.Supported by a grant (BNS 8004587) from the National Science Foundation to G.S.H. The authors gratefully acknowledge the helpful discussions and expert technical assistance of Thomas R. Tokarski     

Corazonin promotes tegumentary pigment migration in the crayfish Procambarus clarkii

The undecapeptide corazonin (pGlu-Thr-Phe-Gln-Tyr-Ser-His-Gly-Trp-Thr-AsnNH(2)) elicits a retraction of erythrophore pigment granules and dispersion of leucophore pigment granules in the crayfish Procambarus clarkii. The effects are dose-dependent from 10(-10) to 10(-5)M. Influence on erythrophores is lower than that of Red Pigment Concentrating Hormone (RPCH), which is inactive on leucophores. Corazonin effects are partly blocked by an anti-corazonin antibody, and even less by an anti-RPCH antibody. Corazonin effects are completely suppressed by the calcium chelator BAPTA. Immunoreactive somata and fibers were identified in various regions of the eyestalk (medulla terminalis, medulla interna and medulla externa) with the anti-corazonin antibody. These results suggest the possible existence of a corazonin-like peptide in crustaceans.     

High prevalence of multiple paternity in the invasive crayfish species,Procambarus clarkii

Reproductive strategy is a central feature of the ecology of invasive species as it determines the potential for population increase and range expansion. The red swamp crayfish, Procambarus clarkii, has invaded many countries and caused serious problems in freshwater ecosystems. However, little is known about the effects of environmental conditions on crayfish paternity and offspring traits in the wild. We studied these reproductive characteristics of P. clarkii in wild populations from two different habitats (ponds and ditches) in three locations with different environmental conditions in China. Genotyping of 1,436 offspring and 30 mothers of 30 broods was conducted by using four microsatellites. An analysis of genotyping results revealed that gravid females were the exclusive mother of the progeny they tended. Twenty-nine of 30 mothers had mated with multiple (2-4) males, each of which contributed differently to the number of offspring in a brood. The average number of fathers per brood and the number of offspring per brood were similar (P > 0.05) among six sampling sites, indicating that in P. clarkii multiple paternity and offspring number per brood are independent of environmental conditions studied. Indirect benefits from increasing the genetic diversity of broods, male and sperm competition, and cryptic female choice are a possible explanation for the high level multiple paternity and different contribution of fathers to offspring in this species.     

Historical,human, and environmental drivers of genetic diversity in the red swamp crayfish (Procambarus clarkii) invading the Iberian Peninsula

1. Patterns of genetic diversity in invasive populations can be modulated by a range of factors acting at different stages of the invasion process, including the genetic composition of the source population(s), the introduction history (e.g. propagule pressure), the environmental suitability of recipient areas, and the features of secondary introductions.
2. The North American red swamp crayfish, Procambarus clarkii, is one of the most widely introduced freshwater species worldwide. It was legally introduced into Spain twice, near the city of Badajoz in 1973 and in the Guadalquivir marshes in 1974. Thereafter the species rapidly colonised almost the entire Iberian Peninsula.
3. We used seven nuclear microsatellites to describe the genetic diversity and structure of 28 locations distributed across the Iberian Peninsula and to explain the expansion process of the red swamp crayfish. Additionally, we analysed the relationship between environmental suitability and genetic diversity of the studied locations.
4. The red swamp crayfish had a clear spatial genetic structure in the Iberian Peninsula, probably determined by the two independent introduction events in the 1970s, which produced two main clusters separated spatially, one of which was dominant in Portugal and the other in Spain.
5. The human-mediated dispersal process seemed to have involved invasion hubs, hosting highly genetically diverse areas and acting as sources for subsequent introductions. Genetic diversity also tended to be higher in more suitable environments across the Iberian Peninsula .
6. Our results showed that the complex and human-mediated expansion of the red swamp crayfish in the Iberian Peninsula has involved several long- and short-distance movements and that both ecological and anthropogenic factors have shaped the genetic diversity patterns resulting from this invasion process. Early detection of potential invasion hubs may help to halt multiple short-distance translocations and thus the rapid expansion of highly prolific invasive species over non-native areas.

     

Serotonergic regulation of crustacean hyperglycemic hormone secretion in the crayfish, Procambarus clarkii

These studies investigate if crustacean hyperglycemic hormone (CHH) is involved in 5-hydroxytryptamine (5-HT)-induced hyperglycemia. Eyestalk ganglia with intact X-organ-sinus gland complex were dissected from the crayfish Procambarus clarkii and incubated under various experimental conditions. Incubation media were then analyzed for the presence of released hyperglycemic factor using an in vivo bioassay. The results show that 5-HT enhanced release of hyperglycemic factor in a dose-dependent manner. This stimulatory effect of 5-HT was significantly decreased by adding ketanserin or methysergide (both 5-HT receptor antagonists) into incubation of eyestalk ganglia. Further, activity of the 5-HT-released hyperglycemic factor could be eliminated by adsorption of incubation media with anti-CHH serum but not by preimmune or anti-5-HT serum. These results confirm the hypothesis that 5-HT enhances release of CHH, which in turn elicits hyperglycemic responses. It is probable that 5-HT activates an excitation-secretion coupling mechanism by interacting with receptors located on the X-organ neurosecretory cells.     

Morphology of descending statocyst interneurons in the crayfish Procambarus clarkii Girard

Summary The morphological features of descending interneurons that responded to the artificial bending of statolith hairs were assessed with intracellular recording and staining techniques. Seven statocyst interneurons were identified on the basis of their structure and response characteristics and designated as interneurons S1 to S7. All seven identified interneurons project to the optic lobe, where the optic nerve also projects, and to the dorsal part of the tritocerebrum, where the eyestalk motoneurons originate. All except interneuron S6 also extend their major branches to other neuropilar regions. S2 projects to the dorsal part of the deutocerebrum, where the statocyst nerve terminates, and S3 to the dorsal part of deutocerebrum and the antennal lobe. Four other interneurons (S1, S4, S5, S7) also extend their branches to the parolfactory lobe to which the statocyst nerve projects as well as to the deutocerebrum and antennal lobe. The extensive dendritic projections of S1–S7 suggest that they are complex multimodal interneurons rather than simple relay interneurons, receiving at least visual and statocyst sensory information. The function of the antennal lobe branches, however, has yet to be determined since the functional role of antennal input in equilibrium control is unknown.     

养殖条件下克氏原螯虾亲虾的形态性状研究

为了解人工养殖的克氏原螯虾(Procambarus clarkii)亲虾的生长和形态性状,为人工放养亲本的选择提供参考依据,测定了342尾体重大于20 g的克氏原螯虾的体重(W)、全长(TL)、头胸甲长(CL)、头胸甲宽(CW)、螯足长(ChL)和螯足宽(ChW),比较了雌性和雄性生长和形态参数的差异性,模拟了各参数间的回归方程,并探讨了各参数间的相关性。结果显示,雌性个体体重、螯足长和螯足宽的均值显著低于雄性,而头胸甲长和头胸甲宽的均值则显著高于雄性,全长和丰满度在雌雄之间差异不显著。雌性和雄性个体的优势全长范围均为90～119. 99 mm,优势体重范围分别为20. 0～39. 9 g和30. 0～44. 9 g。克氏原螯虾全长-体重的关系为W_♀=0. 000 01×TL~(3. 381 3)、W_♂=0. 000 01×TL~(3. 230 9),雌性和雄性均为正异速生长;头胸甲长-体重的关系为W_♀=0. 000 1×CL~(3. 215 5)、W_♂=0. 000 2×CL~(3. 179 8);头胸甲长-全长的关系为TL_♀=2. 928 6+2. 027×CL、TL_♂=-5. 036 9+2. 370 1×CL;头胸甲宽-头胸甲长的关系为CL_♀=5. 479 2+1. 942 3×CW、CL_♂=9. 646 8+1. 765 6×CW;螯足宽-螯足长的关系为ChL_♀=-13. 721+6. 679×ChW、ChL_♂=-4. 828+6. 148 7×CW。相关性分析表明,总体上6个参数之间均有显著的正相关关系,其中与体重相关性较高的为全长和头胸甲长与宽。     

A novel calcium-binding peptide from the cuticle of the crayfish, Procambarus clarkii

A novel peptide named calcification-associated peptide (CAP)-2 was isolated from the exoskeleton of the crayfish, Procambarus clarkii. CAP-2 consists of 65 amino acid residues and has a 44% sequence identity with CAP-1 characterized previously. It has a chitin-binding domain observed in many arthropod cuticle proteins. CAP-2 showed inhibitory activity on calcium carbonate precipitation and chitin-binding ability. A CAP-2 cDNA was cloned using RT-PCR and RACE and the open reading frame encoded a precursor peptide consisting of a signal peptide and CAP-2. RT-PCR revealed that CAP-2 mRNA was exclusively expressed in the epidermal tissue during the postmolt stage, the site and stage being associated with calcification. Calcium-binding assay using recombinant CAP-2 revealed that this peptide had affinity for calcium ions with a Kd value of about 1 mM. All these results suggest that CAP-2 serves as a nucleator or a regulator in the calcification of the exoskeleton.     

Distribution of synapses on two types of ascending interneurons in the crayfish,Procambarus clarkii

Summary About 60 pairs of ascending interneurons are present in the terminal ganglion of the crayfish Procambarus clarkii (Girard). Some of these interneurons have been impaled intracellularly, characterized physiologically, and then labeled with horseradish peroxidase (HRP) to examine the distribution and ultrastructure of synapses. A close relationship between ultrastructure and physiological properties has been found between two types of interneurons, which either have a pre-motor effect upon motor neurons or have no such effect. In one interneuron with a pre-motor effect (6D2), input and output synapses are intermingled on thicker branches, whereas only input synapses are found on small diameter branches. Only input synapses have been observed on the branches in another interneuron with-out a pre-motor effect (6B1). No differences in branch morphology are found in these two interneurons. Interneuron 6D2 contains large numbers of small round agranular vesicles, but the same type of synaptic vesicles is rarely seen in interneuron 6B1, which has no output synapses. Our results indicate a good correlation between the synaptic distribution and pre-motor effects of interneurons in the terminal ganglion.Abbreviations A6, 7 Sixth and seventh abdominal segment of the terminal ganglion - AVC anterior ventral commissure - DC I dorsal commissure I - DIT dorsal intermediate tract - DMT dorsal medial tract - eLG extra lateral giant interneuron - LVT lateral ventral tract - LG lateral giant interneuron - LVT lateral ventral tract - MDT median dorsal tract - MG medial giant interneuron - MoG motor giant neuron - MVT median ventral tract - PVC posterior ventral commissure - R1s sensory fiber tract of nerve root 1 - R3m motor fiber tract of nerve root 3 - R4–7 nerve roots 4–7 - SC I,II sensory commissure I,II - VC I,III ventral commissure I, III - VIT ventral intermediate tract - VLT ventral lateral tract - VMT ventral medial tract     

Morphogenesis and pattern formation in the retina of the crayfish Procambarus clarkii

Pattern formation and ommatidial differentiation in the crayfish retina were analyzed using confocal, light and electron microscopy. Optic primordia first appear in the embryo as round elevations covered by a surface epithelial layer. Retinal differentiation begins with a wave of mitotic activity that moves across this epithelium from lateral to medial. Ommatidial cell clusters are visible at the surface along a transition zone, which lies at the interface of the medial undifferentiated retina and the lateral patterned retina. This zone is 8–10 cells wide and composed of small uniform cell profiles. Lateral to the transition zone the initial ommatidial cell clusters form staggered rows across the surface. Each first row cluster contains eight retinula cells surrounded by four cone, two corneagenous and two distal pigment cells. Ommatidial clusters in the first nine rows show significant changes in their organization, which are visible at the surface of the retina. In row 10 the retinula cells recede from the surface and the cone cells close in above them creating a constant cell pattern at the surface. Rhabdome development begins distally and extends downward as the retinula cluster recedes from the surface. Movement of the retinula cells inward and enlargement of the cone and corneagenous cells at the surface creates a two-tiered organization characteristic of each ommatidium. Comparison of retinal pattern formation and differentiation in the crayfish with retinal morphogenesis in Drosophila and other insects show several similarities between the two arthropod groups.     

Micro-scale fluid and odorant transport to antennules of the crayfish, Procambarus clarkii

A numerical model was developed to determine advective-diffusive transport of odorant molecules to olfactory appendages of the crayfish, Procambarus clarkii. We tested the extent of molecule transport to the surfaces of aesthetasc sensilla during an antennule flick and the degree of odorant exchange during subsequent flicks. During the rapid downstroke of a flick, odorant molecules are advected between adjacent aesthetascs, while during the slower return stroke, these odorants are trapped between the sensilla and molecular diffusion occurs over a sufficient time period to transport odorants to aesthetasc surfaces. During subsequent flicks, up to 97.6?% of these odorants are replaced with new odorant molecules. The concentration of molecules captured along aesthetasc surfaces was found to increase with increased gap spacing between aesthetascs, flick speed, and distance from the proximal end of the aesthetasc, but these changes in morphology and flicking kinematics reduce the animal's ability to take discrete samples of the odorant-laden fluid environment with each flick. Results suggest that antennule flicking allows discrete sampling of the time- and space-varying odorant signal, and high concentration odorant filaments can be distinguished from more diffuse, low concentration filaments through changes in both the timing and the encounter rate of odorant molecules to aesthetasc surfaces.