Multilocus isozyme systems in African lungfish,Protopterus annectens: distribution,differential expression and variation in dipnoans

Distribution of ADH, ALP, FBALD, GAPDH, G3PDH, G6PDH, GPI, LDH, MDH, PGM, and SOD was identified in retina, heart, muscle, liver, kidney, gills, brain, gut, lung and ovary of the African lungfish. Data are compared with patterns previously described in dipnoans and other vertebrates. The number of loci expressed for all enzymes was found to be similar to those of diploid Actinopterygii. Differences in the number of loci expressed in Amphibia were found for ALP, sG3PDH, GPI, LDH, MDH and SOD. Differences in tissue distribution were noted in ALP due to the absence of an intestinal-specific form typical of teleostean fish, amphibians, reptiles and birds, and in GPI and MDH, due to the tissue expression, as in primitive fish. There were also differences in LDH, where a third locus (LDH-C*) was expressed in the gills of Protopterus annectens and not in the retina or liver tissues, as in teleosts. LDH-A₄ was most common in all the tissues. Major differences were noted in the tissue patterns of protein expression in the three dipnoans compared. As expected, the least divergence was found between the two species belonging to the same family (Lepidosirenidae). The highest index of divergence was observed between Neoceratodus forsteri and Lepidosiren paradoxa, belonging to the families Ceratontidae and Lepidosirenidae, respectively. The divergence is revealed by changes at the enzyme and morphological levels. These results suggest that P. annectens occupies an interesting systematic position, its biochemical characteristics distinguishing it from N. forsteri, L. paradoxa, the advanced fish and amphibians.     

Genetic variability in Hoplias malabaricus (Osteichthyes: Erythrinidae) in fluvial and lacustrine environments in the Upper Paraná river floodplain (Paraná State,Brazil)

Genetic variability in Hoplias malabaricus, from two localities in the upper Paraná River floodplain, was investigated by starch and polyacrylamide gel electrophoresis. A total of 52 specimens were analyzed for 14 enzymatic systems. Twenty-three gene loci of 13 enzymatic systems (AAT, ACP, ADH, GDH, G6PDH, GPI, IDH, LDH, MDH, MEP, PGM, PER, and SOD) were analyzed by starch gel electrophoresis (Penetrose-30). The EST system was analyzed by polyacrylamide gel electrophoresis, and one polymorphic locus was found (EST-1). Twenty-four loci were detected. The proportion of polymorphic loci was 37.5% in the lagoon and 33.3% in the river. Significant differences in allele frequencies of five loci were found between specimens from the two environments. Expected mean heterozygosity (H _e = 0.14) is the same in the river and lagoon, however, Nei's genetic distance (D) between the population of the two locations was 0.049.     

青草鲢鳙四种鱼同工酶的比较研究

采用聚丙烯酰氨垂直板状连续电泳方法,对青草鲢鳙６种组织、１０种同工酶和蛋白质进行了电泳研究,并结合作者以前做的工作,对所研究的共２１种同工酶和蛋白质在４种鱼中的组织分布,位点表达及活性作了分析和总结。结果表明：４种鱼同工酶的种内组织分布差异大于种间差异;ＡＤＨ、ＡＭＹ、ＣＡＴ、ＥＳＴ、ＭＥ、ＰＯＸ、ＳＤＨ存在不同程度的种间差异,可作为种类的遗传标记;４种鱼ＭＤＨ、ＧＯＴ、ＡＬＰ、ＡＯ、ＳＯＤ同工酶谱非常相似,只在活性上略有差异;ＬＤＨ、Ｇ６ＰＤＨ和ＣＯＸ的酶谱特征可作为青草和鲢鳙两个亚科的鉴别标记。     

Biochemical comparison of two Hypostomus populations (Siluriformes, Loricariidae) from the Atlântico Stream of the upper Paraná River basin, Brazil

Two syntopic morphotypes of the genus Hypostomus - H. nigromaculatus and H. cf. nigromaculatus (Atlantico Stream, Paraná State) - were compared through the allozyme electrophoresis technique. Twelve enzymatic systems (AAT, ADH, EST, GCDH, G3PDH, GPI, IDH, LDH, MDH, ME, PGM and SOD) were analyzed, attributing the score of 20 loci, with a total of 30 alleles. Six loci were diagnostic (Aat-2, Gcdh-1, Gpi-A, Idh-1, Ldh-A and Mdh-A), indicating the presence of interjacent reproductive isolation. The occurrence of few polymorphic loci acknowledge two morphotypes, with heterozygosity values He = 0.0291 for H. nigromaculatus and He = 0.0346 for H. cf. nigromaculatus. F(IS) statistics demonstrated fixation of the alleles in the two morphotypes. Genetic identity (I) and distance (D) of Nei (1978) values were I = 0.6515 and D = 0.4285. The data indicate that these two morphotypes from the Atlantico Stream belong to different species.     

Allozyme Variation in a Threatened Freshwater Fish, Spotted Murrel (Channa punctatus) in a South Indian River System

Samples of the spotted murrel (Channa punctatus) were collected from three rivers of Tamil Nadu and Kerala. The allozyme variation of C. punctatus was investigated by polyacrylamide gel electrophoresis. Eighteen enzymes were detected, but only 10 (EST, PGM, G3PDH, G6PDH, SOD, GPI, ODH, GDH, XDH, and CK) showed consistent phenotypic variations. Allele frequencies were estimated at the 18 polymorphic loci representing 10 enzymes. Two rare alleles, EST-4*C and G6PDH-2*C, were noted in the Tamirabarani and Kallada populations but were absent in the Siruvani population. The allele frequencies of the Tamirabarani and Kallada populations were similar, except for a few loci. Among the three populations, the maximum genetic distance (0.026) and FST (0.203) were found between the geographically distant Siruvani and Kallada populations. Overall the study showed that among the three populations, the Tamirabarani and Kallada have similar genetic structures.     

山西省六个负蝗种群的遗传多样性

用等位酶电泳分析方法对短额负蝗(A tractom orpha sinensis)和奇异负蝗(A tractom orpha p ereg rina)各3个自然种群10种酶(AAT,CK,G 3PD,HEX,IDH,LDH,M DH,M E,PG I,PGM)进行检测。结果显示:两种负蝗在某些基因座上共享常见的等位基因,如A a t-1-b、A a t-2-b、G 3p d-a、Ck-1-b和Ldh-b;除个别基因座在部分种群符合H ardy-W e inberg平衡外,两种负蝗的大多数基因座的基因型频率显著偏离H ardy-W e inberg平衡。此外,奇异负蝗M e-c(0.318~0.740)、短额负蝗H ex-1-a(0.800~1.000)及Ldh-b(0.487~0.750)等位基因频率呈现出由北向南递增的趋势,表明M e和H ex、Ldh基因座上的等位基因频率与地理分布存在一定的相关关系。短额负蝗平均每个基因座的等位基因数(A)为1.9~2.3、多态基因座百分率(P)为56.3%~68.8%、平均观察杂合度(Ho)为0.072~0.096;而奇异负蝗的相应值依次为A=1.7~2.2,P=43.8~56.3%,Ho=0.070~0.107。从A、P和Ho3个参数可知,短额负蝗遗传多样性明显高于奇异负蝗。6个负蝗种群的平均观察杂合度均明显低于H ardy-W e inberg平衡预期值,表明6个负蝗种群均出现了杂合体缺乏现象。短额负蝗3种群I值为0.971~0.996,奇异负蝗3种群I值为0.982~0.995,短额负蝗与奇异负蝗I值为0.379~0.451,表明种内遗传相似度明显高于种间,从种间I值可知奇异负蝗和短额负蝗属于近缘种。根据R oger's遗传距离进行的聚类分析表明,两种负蝗可分为两支,且两种负蝗的遗传距离与地理距离均存在明显的相关趋势。两种负蝗的平均FST值都不显著偏离0值(奇异负蝗FST=0.087,p>0.05,短额负蝗FST=0.045,p>0.05),表明该两种负蝗种群间的分化不明显。     

Selection of genotypes of Schistosoma mansoni and their maintenance by sporocyst transplantation

Schistosoma mansoni was isolated by hatching eggs obtained from a naturally infected Rat in Grand Etang, Guadeloupe; fifty Biomphalaria glabrata were exposed to five miracidia each. The resulting cercariae were used to infect laboratory mice which were later sacrificed to provide worms for enzyme analyses and eggs for further infections. Seven enzymes in extracts of individual worms were examined by isoelectric focusing in polyacrylamide gels: AcP, G6PDH, PGM, GPI and HK showed no variation, whereas MDH and LDH proved to be polymorphic. Two MDH loci were recognised, MDH-2 was invariant whereas two alleles were assumed at the MDH-1 locus. It was not possible to make a genetic interpretation of the complex banding pattern of LDH, although 4 types (LDH-A, -B, -C, -D) were observed. Of the snail infections, one batch of snails was exposed to 5 miracidia per snail in the normal way whereas other snails were each exposed to a single miracidium. The latter were sacrificed to provide sporocysts to transplant into further groups of recipient snails. Cercariae from the recipient snails were used to infect mice and the adult worms were analysed and compared with the normally passaged material. In this way, three lines, defined by the possession of particular MDH and LDH types, were selected from the originally polymorphic population; two were identical. The combination of single miracidium infections and enzyme typing has illustrated the possibility of selecting parasite lines of known genotype; transplantation of sporocysts from snail to snail has demonstrated that such lines can be maintained exclusively in the intermediate host.(ABSTRACT TRUNCATED AT 250 WORDS)     

Enzymatic urea adaptation: lactate and malate dehydrogenase in elasmobranchs

Lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) electrophoretic tissue patterns of two different orders of Elasmobranchii: Carchariniformes (Galeus melanostomus and Prionace glauca) and Squaliformes (Etmopterus spinax and Scymnorinus licha) were studied. The number of loci expressed for these enzymes was the same of other elasmobranch species. Differences in tissue distribution were noted in LDH from G. melanostomus due to the presence of an additional heterotetramer in the eye tissue. There were also differences in MDH. In fact, all the tissues of E. spinax and G. melanostomus showed two mitochondrial bands. Major differences were noted in the number of isozymes detected in the four compared elasmobranchs. The highest polymorphism was observed in E. spinax and G. melanostomus, two species that live in changeable environmental conditions. The resistance of isozymes after urea treatment was examined; the resulting patterns showed a quite good resistance of the enzymes, higher for LDH than MDH, also at urea concentration much greater than physiological one. These results indicated that the total isozyme resistance can be considered higher in urea accumulators (such as elasmobranchs) than in the non-accumulators (such as teleosts).     

Phenotypic polymorphism and allele differentiation of isozymes in fodder beet,multigerm sugar beet and monogerm sugar beet

Summary Thirteen enzymes (MDH, SDH, LAP, PGM, PX, IDH, GPI, 6PGD, APH, GOT, GDH, ME and SOD) of 3 cultivated beet (B. vulgaris L.) gene pools, comprising 12 accessions of fodder beet, 11 of old multigerm sugar beet and 10 of modern monogerm sugar beet, were investigated using horizontal starch gel electrophoresis. Eleven accessions of primitive or wild B. vulgaris were also included for the comparison of isozymes. Variation in isozyme phenotypes was investigated to detect diversity in the three cultivated forms of beet. Phenotypic variation was observed in all except ME and SOD, which were monomorphic. A high degree of phenotypic polymorphism (Pj) was found in GDH, PGM, IDH, APH and MDH. Differences in phenotypic polymorphism in MDH, GPI and PX were recognized between fodder beet and both sugar beet groups. Average polymorphism for 13 enzymes in both sugar beets was significantly higher than that in fodder beet. For 13 enzymes, the existence of high isozyme diversity in both sugar beet gene pools was revealed. Allele frequencies in 13 alleles of five enzyme-coding loci, Lap, Px-1, Aph-1, Got-2 and Gdh-2, were investigated. New alleles, Px-1 ¹ and Got-2 ¹, were found in fodder beet accessions. No significant differences of average allele frequencies of five loci between fodder beet and both sugar beets were recognized. Several unique alleles and different isozyme phenotypes were observed in the accessions of B. vulgaris ssp. macrocarpa and ssp. adanensis. Future utilization of cultivated beet gene pools for sugar beet breeding is discussed from the viewpoint of genetic resources.     

Some aspects of isozymes of lactate dehydrogenase, malate dehydrogenase and glucosephosphate isomerase in fish

1. The present paper reports some aspects of the isozymes of LDH, MDH and GPI in fish. 2. In Petromyzontiformes LDH is encoded by a single Ldh-A gene locus. In Myxiniformes and in most vertebrates LDH is encoded by two gene loci, A and B. A third Ldh-C locus is characteristic of the bony fishes Actinopterygii. 3. In fish the MDH isozymes are generally encoded by three gene loci Mdh-M, Mdh-A and Mdh-B. 4. In most diploid bony fish the GPI is controlled by two independent gene loci Gpi-A and Gpi-B. 5. The relationships of isozymes with evolution of vertebrates, tissual specificity, ontogenetic changes, with physiological and metabolic roles are discussed.     

Differences in isoenzyme patterns of axenically and monoxenically grown Acanthamoeba and Hartmannella

Axenically and monoxenically grown Acanthamoeba castellanii, Acanthamoeba polyphaga and different isolates of Hartmannella vermiformis strains were examined by polyacrylamide isoelectric focusing in the pH range 3–10. Isoenzyme patterns of acid phosphatase (AP), propionyl esterase (PE), malate dehydrogenase (MDH), alcohol dehydrogenase (ADH), glucose phosphate isomerase (GPI) and phosphoglucomutase (PGM) were compared. Zymograms were used to reveal differences in typical isoenzyme patterns between axenically and monoxenically grown amoebae and to compare axenically grown A. castellanii, A. polyphaga and H. vermiformis. Comparison of zymograms for AP, PE and MDH between axenically grown Acanthamoeba and Hartmannella strains revealed different isoenzyme patterns. Acanthamoeba showed strong bands for ADH and extremely weak bands for GPI and PGM, while Hartmannella lacked ADH but possessed bands for GPI and PGM.\par Comparison of zymograms from axenically and monoxenically grown amoebae revealed a lower intensity and even lack of typical isoenzyme bands in lysates from monoxenic cultures. The observed changes in typical isoenzyme patterns induced by the bacterial substrate can influence the correct isoenzymatic typing of different strains in clinical and phylogenetic studies.     

鲇鱼不同组织同工酶的组织特异性的初步研究

采用聚丙烯酰胺垂直凝胶电泳法(PAGE)对鲇鱼的9种组织的5种同工酶(LDH、MDH、ADH、SUDH和EST)进行了分析,结果表明:5种同工酶在各种组织均有不同的表达程度:在心、肝、卵巢和肌肉中的表达频率最高,在脑和眼中最低。LDH和MDH在各个组织中均为强表达,与以往研究结果比较,认为鲇鱼LDH同工酶不存在C位点。鲇鱼同工酶组织特异性较明显。     

Metabolic background for establishment of the subpopulation structure in early ontogenesis in the Atlantic salmon (the case of energy of carbohydrate metabolism)

The activity of some enzymes involved in energy and carbohydrate metabolism was studied in Atlantic salmon embryos at the eyed egg stage and in salmon fingerlings (0+) from two trophic–ecological groups: the Varzuga River bed and two tributaries, the Pyatka and Sobachii rivers (Kola Peninsula). It has been demonstrated that heterogeneity of embryos was most evident in the case of cytochrome c oxidase (CO), malate dehydrogenase (MDH), glycerol-1-phosphate dehydrogenase (G1PDH), and glucose-6-phosphate dehydrogenase (G6PDH), while the lowest level of heterogeneity was observed for lactate dehydrogenase (LDH) and aldolase. A positive correlation was revealed between the activities of CO, LDH, MDH, and G1PDH. It was noted that G6PDH showed a negative correlation with almost all enzymes under study. It was found that salmon juveniles inhabiting the tributaries were characterized by high LDH, aldolase, and G1PDH activity and lower activity of G6PDH compared to the juveniles inhabiting the main river bed. Notably, the differences in the activity of the enzymes involved in aerobic metabolism between the two groups of fingerlings under analysis were observed only in the autumn.     

Enzymatic patterns in the embryonic development of the cricket,Acheta domesticus L.

Summary The activity of the enzymes G6PDH, 6PGDH, GAPDH, LDH, CS, MDH, NADP-linked ICDH, GOT, GPT, and GLDH was tested in different stages of developing eggs and in just hatched larvae of crickets. A malic synthase could not be found. The enzymatic pattern changes considerably during this period. Considering this pattern, and especially the quotient (GAPDH)/CS, it seems justified to state that, up to the formation of the germ band, the energy is derived mainly from oxidation of glucose in the citric acid cycle. Later on, lipids substitute partly for glucose. Anaerobic energy production seems to be limited to the stage of the dorsal closure. By inhibiting the formation of the germ band, the activity of GAPDH, and CS is not changed significantly. The rise in activity of G6PDH is correlated with the increased rate of r-RNA synthesis. The increase of activity of MDH and NADP-linked ICDH corresponds to the differentiation of the germ band. These enzymes, together with GOT, GPT, and GLDH should be involved in the synthesis of amino acids from carbohydrates during the later part of development. In the hatched larvae the activity of all enzymes has risen considerably. This stage is characterized by the fact that G6PDH, 6PGDH, GAPDH, LDH, CS, and NADP-linked ICDH display the same activity.This work was supported by the Stiftung für Krebs- und Scharlachforschung (Strebelstiftung).     

Enzymes in Schistosoma intercalatum and the relative status of the Lower Guinea and Zaire strains of the parasite

Seven enzyme systems [phosphoglucomutases (PGM), glucose phosphate isomerases (GPI), glucose-6-phosphate dehydrogenases (G6PD), malate dehydrogenases (MDH), laetate dehydrogenases (LDH), acid phosphatases (AcP) and hexokinases] in extracts of adult worms from two isolates of Schistosoma intercalatum, one from Zaire and one from Cameroun, were compared by isoelectric focusing. Two systems (GPI and PGM) were also compared in extracts of cercariae. Distinctive differences between the strains were found in the LDH system and even more marked differences in the G6PD and PGM systems (the latter were apparent in both adult worm and cercarial extracts). These observations are discussed in conjunction with existing evidence on the results of intermediate host infection experiments and of experimental hybridization between the two strains of S. intercalatum. In turn these aspects are discussed in the light of what is known about other species of African Schistosoma. It is concluded that any definite decision on the relative status of the two strains of S. intercalatum is still premature.     

Expression of the enzyme-encoding genes under the influence of colchicine and triton X-100 in nongerminating seeds of sugarbeet (<Emphasis Type="Italic">Beta vulgaris</Emphasis> L.)

The expression of the enzyme-coding genes, controlling glucose-phosphate isomerase (GPI), malate dehydrogenase (MDH), and alcohol dehydrogenase (ADH), was examined in nongerminating seeds of sugarbeet after Triton X-100 (TX-100) and colchicine treatment. Two types of changes revealed included modification of the enzymatic loci expression (change of the isozyme electrophoretic mobility) and inactivation of standard profiles. In the MDH and GPI systems, these processes were found to be associated. Complete isozyme modification was accompanied with the disappearance of standard profiles. In the ADH system, the treatment with TX-100 and colchicine gave rise to two independent processes, including silencing of the Adh1 locus and the appearance of the ADH isozymes with abnormal electrophoretic mobility, which were probably the products of the Adh2 locus. It was suggested that the effect of TX-100 and colchicine on the expression of the enzyme-encoding genes examined depended on the intracellular localization of the encoded enzymes.     

Genetic structure of two Turkish brown trout populations

The genetic structure of two brown trout Salmo trutta populations living in Lake Abant in Bolu and Üzüm River in Antalya was determined by examining 15 enzyme coding loci ( AAT, ADH, LDH, MDH, MEP, GPI, PGM and SOD ) using starch gel electrophoresis. Population specific mobilities were observed for the fixed alleles of LDH-B2, mMEP-2 and SOD-1 loci. Polymorphisms in sAA T-4, GPI-B2 loci were observed within the populations. Average heterozygosity of Abant and Antalya populations was 0.0358 and 0.0224 respectively. For LDH-C which is the post glaciation marker locus, the ancestral allele * 105 was found to be fixed in both of the populations. Nei's genetic distance between the two populations was 0.2507 which is the level of genetic distance often found between different species. This difference seems to be due to the presence of unique alleles in the LDH-B2, mMEP-2 and SOD-1 loci of the Abant population, indicating that the conservation of the Abant population and its heterozygosity is of prime importance.     

The palmitoleate: a natural selective denaturant of enzymes

A study has been carried out in order to explain the enzyme-palmitoleate interaction. The highly purified and crystalline enzymes representative of fundamental metabolic pathways were: alcohol dehydrogenase (ADH), lactate dehydrogenase (LDH), malate dehydrogenase (MDH), isocitrate dehydrogenase (ICDH), glucose-6-phosphate dehydrogenase (G6P-DH), alkaline phosphatase. The enzyme-palmitoleate interaction was studied as a phenomenon time-independent (inhibition) and time-dependent (inactivation). Palmitoleate inhibited remarkably LDH, MDH, ICDH and G6P-DH. A kinetic analysis of the inhibitory action of palmitoleate on LDH and MDH was also carried out. Inactivation studies have shown that ADH and alkaline phosphatase are not sensitive to palmitoleate action, unlike the other enzymes. A comparison was made between the action of palmitoleate and that of a synthetic anionic detergent, sodium dodecyl sulfate (SDS).     

Genetic Variation of Mantled Howler Monkeys (Alouatta palliata) from Costa Rica1

We examined genetic diversity of howler monkeys (Alouatta palliata) from Costa Rica. Blood samples of howler monkeys were collected at various locations in Costa Rica, and electrophoresis of total plasma proteins yielded no variation. We also conducted starch gel electrophoresis of red cell isozymes and did not find variation for any of the 14 loci analyzed (i.e., ACP, ADA, CA2, EST, GPI, IDH, LDH‐1, LDH‐2, MDH, PGD, PGM‐1, PGM‐2, SOD, and TPI). These findings were compared with the levels of genetic variation for A. seniculus and A. belzebul from one Brazilian population. Four of the 14 isozymes (ADA, GPI, PGD, and SOD) showed more than one allele for these species. Both A. seniculus and A. belzebul from Brazil showed similar levels of genetic variation. The potential causes of the low genetic variation in A. palliata from Costa Rica are discussed.     

An isoenzyme study in the genus Lotus (Fabaceae). Experimental protocols and genetic basis of electrophoretic phenotype

Summary An isoenzyme survey of some taxa in the genus Lotus (Fabaceae) was undertaken to increase the number of genetic markers available to breeders and to students of Lotus phylogeny. Twenty-one enzymes were examined using starch gel electrophoresis and nine buffer systems. Clear, consistent banding patterns were obtained for PGI, TPI, MDH, IDH (NADP), PGM, 6-PGDH, and ME. Clear but inconsistent banding patterns were obtained for FDP, G₃PDH (NADP), -EST, LAP, MDH, DIA, and NADHDH. Phenotypes of the seven consistently resolved enzyme systems were obtained for different tissues for each of several genotypes at different stages of development. Variation in enzyme phenotypes of the same individuals under different growth conditions indicated the presence of different isozymic forms of these enzymes. Shoot tissue of plants over 6 weeks of age was found to be suitable material for further genetic studies, since phenotype for this tissue was constant despite changes in growing conditions. A formal genetic analysis of segregation and/or recombination of allozymes for the enzymes PGM, TPI, MDH, IDH, and 6-PGDH was undertaken. Isoenzyme phenotypes were examined for the diploids L. alpinus Schleich., L. burttii Sz. Borsos, L. conimbricensis Brot., L. ornithopodioides L., L. tennis Waldst. et Kit., and L. uliginosus Schkuhr; and for the diploid interspecific hybrids L. alpinus x L. conimbricensis, L. burttii x L. ornithopodioides, and L. japonicus x L. alpinus. Several new loci were identified for Lotus, namely, Idh1, Idh2, Mdh3, Pgi1, Pgi2, Tpi1, Tpi2, and 6-Pdgh1. Duplications of loci of IDH, MDH, PGI, and 6-PGDH were detected in the diploid (2n=12) interspecific hybrid L. japonicus x L. alpinus.