Reproductive Isolation and Distinct Population Structures in Two Types of the Freshwater Shrimp Palaemon Paucidens
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Evolution,42(4), 1988, pp. 804-813 REPRODUCTIVE ISOLATION AND DISTINCT POPULATION STRUCTURES IN TWO TYPES OF THE FRESHWATER SHRIMP PALAEMON PA UCIDENS SEINENCHOW,' YOSHIHISA Fujio, AND TADASHINOMURA Departmentof Fishery Science, Tohoku University, Amamiyamachi, Sendai 980, JAPAN Abstract.-Twenty local populations of the Japanese freshwater shrimp Palaemon paucidens were electrophoreticallyand morphologicallysurveyed. Based on the diagnosticdistributions of some alleles at Gpi, Mpi, Mdh-J,and Mdh-2, these populationswere largelyclassified into two types(A and B). The A type occurredin lakes, ponds, and rivers,while the B type was observed only in rivers. Average Nei's genetic distance (D) between the two types fell into the subspecies range (D = 0.1 186). The coefficientof gene differentiation,GsT, varied considerablybetween the two types.In 12 populations of the A type,with a GST value of 0.281, nine pond and lake populations showed a higherGsT (0.246) than the threeriver populations (0.151). On the otherhand, GsT was 0.036 forthe eightlocal populations of the B type.The lower rostrumtooth numberhad a mode of two in typeA and threein typeB. Type-A populationslargely varied in the upperrostrum tooth numberand egg size but typeB did not. Under laboratoryconditions, mating frequently occurred withineach type,but not between types.Furthermore, no embryonicdevelopment was observed in the fewcases of intertypemating. These resultsindicate thatthe A and B typeshad experienced cladogenic separationwith pre- or postmatingisolation, whereafterthe A type,under geographic isolation,underwent genetic and phenotypicdifferentiation, while the B type,under extensive gene flow,did not undergodifferentiation. Received February6, 1987. Accepted January13, 1988 It has been long recognizedthat intraspe- caughtin the same area, we concluded that cific genetic differentiationof sexually re- the typesare reproductivelyisolated in the producingspecies is promotedby geograph- singleriver basin. Differencesin eggsize and ic and reproductiveisolation. Cladogenic rostrumtooth number between the types are separation accompanying intrinsicrepro- apparentlymaintained by the reproductive ductive isolation is now generallyaccepted isolation. Additional analysis of the allelic as speciation. Even so, genetic differentia- distributionssuggested that the geneticdis- tion can occur in geographicallyisolated tance between the types fell into the range populations without such intrinsicrepro- of subspecies, according to the rough cri- ductive isolation. Thus, the origin,nature, teriongiven by Nei (1975) (Chow and Fujio, and functionof isolatingmechanisms play 1985c). Nishino's (1980) discoveryof great importantroles in evolutionarychanges. variation in egg size among local popula- A previous paper provided biochemical tions of this species led us to presume a evidence of two typesof common Japanese highlysubdivided and geneticallydifferen- freshwatershrimp Palaemon paucidens in- tiated population structureand convinced habitingthe same watersystem (Chow and us thatan extensivesurvey of the local pop- Fujio, 1985a). The two types,named A and ulationswould be worthwhile.In thispaper, B, were clearly distinguishedby the pres- we present evidence for reproductiveiso- ence or absence of diagnosticalleles at the lation and distinctpopulation structures be- glucosephosphate isomerase (Gpi), man- tweenthe two typesof Palaemon paucidens nosephosphateisomerase (Mpi), and malate and discuss the processes involved in di- dehydrogenase(Mdh-1 and Mdh-2) loci. vergence between and within these two Since both typeshave almost the same re- types. productiveseason and since mature males and femalesof each typeare simultaneously MATERIALS AND METHODS All samples of 20 local populations of P. paucidens were collectedwith dip nets dur- I Present address: Marine Resources Research In- ing the months of March-September in stitute,Charleston, SC 29412. 1983-1986. Sitesand theirnames are shown 804 REPRODUCTIVE ISOLATION 805 *~HNP 30 km IHPO KMP 2 km FIG. 1. Map showingcollection localities ofPalaemon paucidens. Miyagi Prefecture:IHP = Ishihoga Pond, HNP = Hinata Pond, KMP = Katamase Pond, HKP = Hanukizawa Pond, TTP = Tomita Pond, ATP = Aburata Pond, KKR = Kitakami River, NRR = Naruse River, HRR = Hirose River, NTR = Natori River, ABR = Abukuma River, SIR = ShiroishiRiver; Ibaragi Prefecture:NKR = Naka River; Tokyo: SPP = Sanpoji Pond, TMR = Tama River; Chiba Prefecture:KYR = Kiyosumi River, OBR = Ohbiso River; Ishikawa Prefecture: LK = Lake Kahokugata; Shiga Prefecture:LB = Lake Biwa; Hiroshima Prefecture:ASR = Ashida River. Symbols show the A (0) and B ( H)types. in Figure 1. Live samples were transferred exceed 0.95. Observed heterozygosity(h.) to the laboratory,except forthose of Lake was obtained fromdirect counts of hetero- Kahokugata which were frozen on dry ice zygotes,while expected heterozygosity(he) fortransport. was calculated as he = 1 - : xi2,where xi Rostrum tooth number and egg volume is the frequencyof the ith allele at each lo- were adopted as the morphological char- cus. Mean heterozygosities and He) for acters. Ten eggs at the pre-eye stage of (H. each local population were obtained from development were measured from each mother shrimp. The egg volume (V) was the formulas,H. = : ho/rand He = : he! calculated from the formula V==rh2/6, r, where r is the number of loci. Allele-fre- where I and h are lengthand width of an quency data were used to calculate genetic egg in mm, respectively.(Data forrostrum distance(D) (Nei, 1972) and a coefficientof toothnumber and eggvolume in the Natori gene differentiation(GST) (Nei, 1973). and Hirose riverswere derived fromChow Mating experiments were designed to and Fujio [1985a].) Afterthese measure- compare the matingpropensities of mature ments,some individuals were rearedin 40- males and femaleswhen they are pairedwith liter aquaria at 250C for mating experi- potential mates fromthe same versus dif- ments,and otherswere wrapped and stored ferentlocal populations and of the same at -800C forelectrophoresis. versus differenttypes. In decapod crustacea Horizontal starch-gelelectrophoresis and (especially Caridea), mature females molt nomenclatureof alleles followedChow and priorto matingand spawning.In captivity, Fujio(1985a, 1985b).Allele-frequencydata the mature females of P. paucidens were for the Natori, Kitakami, Naruse, Hirose, distinguishedby fully developed ovaries, Abukuma, Naka, Ohbiso, and Kiyosumi whichcould be observed throughthe trans- rivers are from Chow and Fujio (1985a, lucent exoskeleton. Mature females were 1985b). A locus was assumed to be poly- takenout ofcommunal holdingaquaria and morphicin each local population if the fre- maintained individually; these usually quency of the most common allele did not underwent a prespawning molt within a 00 O TABLE 1. Summarized allele distributionsat fivediagnostic loci of 20 local populations.Localities and abbreviationsare as given in Figure 1. Symbols: #= most common allele (P _ 0.50); * = otheralleles (0.05 _ P < 0.50). Rare alleles (P < 0.05) are not shown here.Detailed allele-frequencydata is available upon request fromthe authors. A-typelocalities B-typelocalities Locus Allele IHP HNP KMP HKP TTP ATP NTR SPP TMR LK LB ASR KKR NRR HRR ABR SIR NKR OBR KYR Gpi A # # # # # # # * # # # # B * * * * * * * # * * # # # # # # # # Q MPI A' * * A # * * # # # # # # # # # C B * # # * * * * * C * * * * * * * * * # * # # # # # # D * * # * * * * * * > Mdh-J B # # # # # # # # # # # # # # # # # # # C' * C * * * * * * * * * Mdh-2 A # # # * # * * B * * * # * # # # # # # # # # # # # # # # 6Pgd A * * * * * * * # # * * B # # # # # # # # # # # # # # # # # # REPRODUCTIVE ISOLATION 807 week. Each molted female was then trans- rangedfrom 0.0602 to 0.1945 witha mean ferredto an aquarium containing 10-20 of 0.1186. The dendrogramdrawn by the males, obtained fromeither the same or a UPGMA methods(Sneath and Sokal, 1973) differentlocality. The aquarium was then and based on the genetic-distancevalues is observedfor five minutes for signs of sexual shown in Figure 2. There are two major behavior (chasing, mounting,and mating clustersat a discriminantlevel of0.1, clearly by males). Females were transferredfrom separatingA and B types.The diversitylevel one aquarium to another until mating oc- among the A-type populations was large, curred.Females thatdid notmate with males and one subcluster(LK and LB) was quite fromany otherlocalities were finallypaired differentfrom the other localities. The D with males fromthe same localityin order values betweenthe subclusterof LK and LB to evaluate whetherthey were still sexually and otherA-type populations ranged from attractiveor not. If a femalewas not mated, 0.0548 to 0.1057, with a mean of 0.0770; the matingexperiment was abandoned for D values betweenthe subclusterand B-type thatfemale. After mating, females were iso- populations range from 0.1645 to 0.1879 lated and maintained individuallyto con- with a mean of 0.1770. Formation of this firmfertilization. distinctsubcluster is due to the diagnostic allele distributionat the 6Pgd locus. The RESULTS diversitylevel among the B-type popula- Genetic Variation and Differentiation.- tions was extremelysmall. The difference Out of the total of 15 loci encoding 1 1 en- in diversitylevels betweeneach typeis fur- zymes, eight(Aat-2, aGpd-1, aGpd-2, Est, thershown by the coefficientof gene differ- Idh, Ldh, Me-i, and Me-2) were mono- entiation, GST (see Table 2). For the 12 morphicin all local populations,and