Populational genetic structure of free-living maned wolves (Chrysocyon brachyurus) determined by proteic markers.

Electrophoretic analysis of presumptive twenty gene loci products was conducted in hemolisates and plasma samples of twenty-eight maned wolves (Chrysocyon brachyurus) from an area in northeastern São Paulo State, Brazil. The area sampled was divided into three sub-areas, with the Mogi-Guaçu and Pardo rivers regarded as barriers to the gene flow. The polymorphism degree and heterozygosity level (intralocus and average) estimated in this study were similar to those detected by other authors for maned wolves and other species of wild free-living canids. The samples of each sub-area and the total sample exhibited genotype frequencies consistent with the genetic equilibrium model. The values of the F-statistics evidenced absence of inbreeding and population subdivision and, consequently, low genetic distances were found among the samples of each area.

NADP-dependent isocitrate dehydrogenase of Astyanax scabripinnis (Pisces, Characidae) from three altitudes at Grande Stream, Campos do Jordão, SP.

Electrokinetic, thermic, and kinetic properties of products of NADP-dependent isocitrate dehydrogenase (IDHP; EC 1.1.1.42) loci of Astyanax scabripinnis (Pisces, Characidae) collected at three different altitudes (700 m, 1,800 m, and 1,920 m) of Grande Stream at Campos do Jordão, State of São Paulo, Brazil, were analyzed. Two IDHP bidirectionally divergent loci, a single skeletal muscle, the IDHP-A*, and a single liver IDHP-B*, both polymorphic, were detected in the three different altitude populations. The variant allele *128 at the IDHP-A* locus, had its highest frequency detected in the 1,920 m population (0.494). Among the nine variant alleles detected at the IDHP-B* locus (*37, *57, *69, *79, *85, *114, *119, *124, and *140), the *37 and 79 were detected only in 1,800 m population. Chi-square values showed that only the 700 m population is not in Hardy-Weinberg equilibrium for the IDHP-A* locus, while for the IDHP-B* locus, no population is. Homogeneity Chi2 test indicated that the populations are significantly different in their A and B phenotype frequencies. Wright's FST mean value (0.036 and 0.32, IDHP-A* and IDHP-B*, respectively) was 0.178 for the three altitude populations which means that 82% of total genetic diversity was found among individuals of each one of the populations. Stability at environmental temperatures (16 degrees to 21 degrees C), and apparent Km and Vmax values of each A-phenotype skeletal muscle crude extract suggest different roles of A-isoforms during the increased lipogenesis that occurs in fish at low temperatures.

Populational genetic structure of free-living maned wolves (Chrysocyon brachyurus) determined by proteic markers

Os produtos protéicos de 20 locos gênicos foram analisados eletroforeticamente em hemolisados e plasma de 28 lobos-guarás (Chrysocyon brachyurus) de uma área da região Nordeste do Estado de São Paulo, Brasil. A área de estudo foi dividida em 3 subáreas, considerando os rios Mogi-Guaçu e Pardo como barreiras ao fluxo gênico. O grau de polimorfismo e o nível de heterozigosidade (intraloco e média) estimados neste estudo foram semelhantes aos observados por outros autores para lobos-guarás e outras espécies de canídeos de vida livre. As diferentes amostras e a amostra total demonstraram freqüências genotípicas nos locos polimórficos consistentes com o modelo de equilíbrio genético. Os valores da estatística-F evidenciaram ausência de endocruzamento e de estruturação populacional e, como conseqüência, foram encontrados baixos valores de distância genética entre as amostras correspondentes a cada subárea.

NADP-dependent isocitrate dehydrogenase of Astyanax scabripinnis (Pisces, Characidae) from three altitudes at Grande Stream, Campos do Jordão, SP

Foram analisadas propriedades eletrocinéticas, térmicas e cinéticas dos produtos dos locos que codificam o isocitrato desidrogenase NADP-dependente (IDHP; E.C. 1.1.1.42) como Astyanax scabripinnis (Pisces, Characidae) em três diferentes alturas do Ribeirão Grande, Estado de São Paulo, Brasil. Nas populaçäes das três altitudes foram detectados dois locos da IDHP, bidirecionalmente divergentes, um único em músculo esquelético, o IDHP-A*, e um único em fígado, o IDHP-B*, ambos polimórficos. O único alelo variante no loco IDHP-A*, A*128, mostrou maior freqüência na população de 1.920 m (0,494). Dentre os nove alelos variantes detectados no loco IDHP-B* (*37, *57, *69, *79, *85, *114, *119, *124 e *140), os alelos *37 e *79 apareceram somente na população de 1.800 m. Valores de Qui-quadrado revelaram que para o loco IDHP-A* somente a população de 700 m não está em equilíbrio de Hardy-Weinberg, enquanto para o loco IDHP-B* nenhuma população se encontra em equilíbrio. Qui2 da homogeneidade revelou que as populaçäes são significativamente diferentes em suas freqüências fenotípicas para ambos os locos. Valor médio de FST de Wright (0,036 e 0,320, IDHP-A* e IDHP-B*, respectivamente) foi de 0,178 para populaçäes das três altitudes, o que significa que 82% da diversidade gênica total foi detectada entre indivíduos da mesma população. Estabilidades em temperatura ambiente (16§ a 21§C) e valores de Km e Vmax aparentes de extratos de músculo esquelético de cada fenótipo-A sugerem diferentes papéis das isoformas-A na lipogênese aumentada que ocorre nos peixes em baixas temperaturas.

Isoform expression in the multiple soluble malate dehydrogenase of Hoplias malabaricus (Erythrinidae, Characiformes).

Kinetic properties and thermal stabilities of Hoplias malabaricus liver and skeletal muscle unfractionated malate dehydrogenase (MDH, EC 1.1.1.37) and its isolated isoforms were analyzed to further study the possible sMDH-A* locus duplication evolved from a recent tandem duplication. Both A (A1 and A2) and B isoforms had similar optima pH (7.5-8.0). While Hoplias A isoform could not be characterized as thermostable, B could as thermolabile. A isoforms differed from B isoform in having higher Km values for oxaloacetate. The possibly duplicated A2 isoform showed higher substrate affinity than the A1. Hoplias duplicated A isoforms may influence the direction of carbon flow between glycolisis and gluconeogenesis.

Isoform expression in the multiple soluble malate dehydrogenase of Hoplias malabaricus (Erythrinidae, Characiformes)

Kinetic properties and thermal stabilities of Hoplias malabaricus liver and skeletal muscle unfractionated malate dehydrogenase (MDH, EC 1.1.1.37) and its isolated isoforms were analyzed to further study the possible sMDH-A* locus duplication evolved from a recent tandem duplication. Both A (A1 and A2) and B isoforms had similar optima pH (7.5-8.0). While Hoplias A isoform could not be characterized as thermostable, B could as thermolabile. A isoforms differed from B isoform in having higher Km values for oxaloacetate. The possibly duplicated A2 isoform showed higher substrate affinity than the A1. Hoplias duplicated A isoforms may influence the direction of carbon flow between glycolisis and gluconeogenesis

Astyanax scabripinnis (Pisces: Characidae) hemoglobins: structure and function

Electrophoretic patterns of hemoglobins, Root effect, Bohr effect in blood and stripped hemoglobin, Hb-O2 affinity GTP modulation of Astyanax scabripinnis (lambari), caught at three different altitudes in Ribeiräo Grande, near Campos do Jordäo (Säo Paulo), are described. All populations showed the same electrophoretic patterns: two cathodal components in starch gel. Normal Bohr effect values were found in these three populations both in blood (phi = -0,11) and stripped hemoglobin (phi = -0,12). Different blood O2 affinities collected in fish of these 3 populations were detected. GTP has a large influence on Hb-O2 binding properties in A. scabripinnis. Stripped hemoglobin shows small Root effect. The addition of triphosphated nucleotides increases this effect. GTP is more effective than ATP on enhancing Root effect. Oxygen availability in water can be the factor responsible for differences found in blood O2 affinity

Functional properties of the three hemoglobin phenotypes of Nelore cattle

Blood and stripped hemoglobin from Nelore cattle individuals were submitted to oxygen equilibrium experiments in both gasometric and spectrophotometric methods. No difference was detected in oxygen affinity and Bohr effect among Hb A, Hb B and Hb AB in experiments with both blood and stripped hemoglobin, in the presence and absence of the chloride ion. However, n values (Hill plots) were higher in Hb B and Hb AB (>2) than in Hb A (<2)

Astyanax scabripinnis (Pisces: Characidae) hemoglobins: structure and function.

Electrophoretic patterns of hemoglobins, Root effect, Bohr effect in blood and stripped hemoglobin, Hb-O2 affinity GTP modulation of Astyanax scabripinnis (lambari), caught at three different altitudes in Ribeirão Grande, near Campos do Jordão (São Paulo), are described. All populations showed the same electrophoretic patterns: two cathodal components in starch gel. Normal Bohr effect values were found in these three populations both in blood (phi = -0.11) and stripped hemoglobin (phi = -0.12). Different blood O2 affinities collected in fish of these 3 populations were detected. GTP has a large influence on Hb-O2 binding properties in A. scabripinnis. Stripped hemoglobin shows small Root effect. The addition of triphosphated nucleotides increases this effect. GTP is more effective than ATP on enhancing Root effect. Oxygen availability in water can be the factor responsible for differences found in blood O2 affinity.

Adaptative features of enzymes from family sciaenidae (Perciformes)--I. Studies on soluble malate dehydrogenase (s-MDH) and creatinine kinase (CK) of fishes from the south coast of Uruguay.

1. Electrophoretic analysis of the soluble malate dehydrogenase (s-MDH) and creatine kinase (CK) isozyme patterns of three species of temperate fishes (Scianidae, Perciformes) indicates at least two loci for s-MDH, Mdh-A and Mdh-B, and four CK, Ck-A, Ck-B, Ck-C and Ck-D. 2. The subunits encoded by these loci occurred at different levels in different tissues and organs analyzed. 3. Through electrophoretic analysis the products of these loci showed different behaviour to changes in temperature. 4. Relative activities of s-MDH and CK isozymes were compared by Klebe's (1975) method to determine pattern of divergence of duplicated gene expression in the three studied species.

Adaptative features of enzymes from family sciaenidae--II. Studies on phosphoglucose isomerase (PGI) of fishes from the south coast of Uruguay.

1. The electrokinetic and thermostability properties of phosphoglucose isomerase (PGI) in three species of temperate fishes (Perciformes, Acanthopterygii)--Cynoscion striatus, Macrodon ancylodon and Micropogonias furnieri--have been analyzed in order to study the adaptative 2. Unlike most diploid fishes the PGI of these species seemed to be encoded by three PGI loci. 3. The subunits encoded by these loci occurred at different levels in the different tissues and organs analyzed. 4. Genetic variants at two loci (Pgi-A and Pgi-B2) were detected in Cynoscion striatus, and at one (Pgi-A) in Micropogonias furnieri. 5. The product of these loci could be separated in three PGI regions based on their electrophoretic distribution and thermostability properties. 6. The more anodal region (including isozymes and allozymes) was more thermolabile than the less anodal one, which is predominant in a single tissue skeletal muscle. 7. Relative activities of PGI isozymes were compared by Klebe's method to determine pattern of divergence of duplicated gene expression in the three species studied.

Studies on the influence of nitrite on methemoglobin formation in Amazonian fishes.

Twenty one species of fishes, collected from the Rio Solimões and a tributary lake in the Amazon Basin near Manaus, showed a wide range of methemoglobin formation 1 hr after a dose of 30 mg/kg of sodium nitrite i.p. Methemoglobin formation in two experimental fishes, Brycon cf. melanopterum and Semaprochilodus insignis, maintained in tanks in our INPA laboratory, was studied in detail. Both fishes survived a dose of 10 mg/kg of nitrite i.p. but usually died within 3 hr of a dose of 30 mg/kg with levels of blood methemoglobin in excess of 80%. Methemoglobin produced in vitro by addition of nitrite to fresh blood was slowly reduced back to hemoglobin over a period of several hours at room temperature. Hemoglobin in hemolysates was auto-oxidized to methemoglobin at pH 6.1 and below but not at 6.9 and above.

Adaptative features of enzymes from family Sciaenidae--III. Studies on lactate dehydrogenase (LDH) of fishes from the south coast of Uruguay.

1. Electrophoretic analyses of lactate dehydrogenase isozyme patterns of three species of temperate fish belonging to family Sciaenidae (order Perciformes) indicates that at least three LDH loci--Ldh-A, Ldh-B and Ldh-C are active. 2. The subunits encoded by these loci occurred at different levels in the different tissues and organs analyzed. 3. The products of these loci show different response to changes in temperature. The results were obtained by electrophoretic analyses. 4. Thermostability of skeletal muscle, heart and eye lactate dehydrogenase incubated at 60 degrees C for different periods of time were examined. 5. Relative activities of LDH isozymes were compared by Klebe's method to determine pattern of divergence of duplicated gene expression in the three species studied.

Biological aspects of Amazonian fishes--VI. Hemoglobins and whole blood properties of Semaprochilodus species (Prochilodontidae) at two phases of migration.

Electrophoretic patterns of hemoglobins, red blood cell phosphates, Bohr effect of blood and stripped hemoglobin, ATP and GTP modulation of Hb-O2 affinity of two species belonging to the genus Semaprochilodus (S. insignis and S. taeniurus--jaraquis), captured at two different locations in Rio Negro, near Manaus (Amazonas), are described. High Bohr effect values were found in these two species and in jaraqui-açu (a possible hybrid between them). In S. insignis a higher Bohr effect for migrating individuals was observed, while S. taeniurus showed the reverse. The main erythrocytic phosphates found in these two species are ATP and GTP. They have a large influence on the O2-Hb binding properties in both species. The GTP/ATP ratio showed large differences between catching places. The two species have different electrophoretic patterns. Semaprochilodus insignis showed four anodal components which showed quantitative variation between catching places; S. taeniurus had three anodal components and a similar quantitative variation.

Electrophoretic patterns of hemoglobin and oxygen binding properties of blood of anostomidae fishes from Parana-Pardo-Grande hydrographic basin (São Paulo State, Brazil).

The electrophoretic patterns of hemoglobin (Hb) and the functional properties of blood of seven migratory fish species, in two different kinds of habitat during both the rainy and dry seasons, were studied. All species showed multiple hemoglobins and very similar electrophoretic patterns except for Schizodon nasutus, which showed two cathodal components. The functional properties of blood reflect those electrophoretic patterns, since all species showed very little or no Bohr effect during each situation studied. Data on Leporinus obtusidens blood (during the rainy season), from both lotic and lentic environments, showed a normal Bohr effect (phi = -0.073). Similarly, Schizodon nasutus and Leporinus friderici from a lotic environment during the dry season were compared and showed a reverse Bohr effect (phi = 0.042). In the other 14 situations, only Leporinus elongatus from a lentic environment during the rainy season showed a statistically significant difference in log P50 values from the other fishes.

Hemoglobin, hematology, intraerythrocytic phosphates and whole blood Bohr effect from lotic and lentic Hypostomus regani populations (São Paulo-Brasil).

Hemoglobin electrophoretic pattern, whole blood Bohr effect, intraerythrocytic phosphates and some hematological parameters were determined and compared between lotic and lentic Hypostomus regani specimens. The electrophoretic patterns of specimens from both kinds of environments showed four anodic fractions. The relative concentration of the fraction I (the most anodal) is significantly different between specimens from lotic and lentic ambients. Both lentic and lotic specimens presented a low Bohr effect: phi = -0.012 to lentic habitat and phi = -0.187 to lotic one. ATP/GTP ratio are higher than 1.0 for lotic environment specimens and lower than 1.0 for lentic ones.

Biological aspects of amazonian fishes I. Red blood cell phosphates of schooling fishes (genus Semaprochilodus: Prochilodontidae).

Red blood cell soluble phosphates from Semaprochilodus taeniurus, S. insignis and a possible hybrid jaraqui-açu, from Amazon basin, were studied by column chromatography. The Semaprochilodus school and the jaraqui-açu shows an intermediate percentage of total phosphorus content as GTP but not as ATP. GTP/ATP ratio for jaraqui-açu and S. taeniurus were 1.5 and for S. insignis, 3.0.

Adaptative features of ectothermic enzymes--IV. Studies on malate dehydrogenase of Astyanax fasciatus (Characidae) from Lobo Reservoir (Såo Carlos, Såo Paulo, Brasil).

1. Skeletal muscle and heart supernatant malate dehydrogenase (s-MDH) from a subtropical fish, Astyanax fasciatus consists of three electrophoretically anodal bands. Each band is a dimer (AA, AB and BB) and two loci are active. 2. In A. fasciatus tissue extracts, A and B subunits are present at differing quantitative levels and their activities are almost season-independent. However, the relative activity of each homodimer in relation to total s-MDH estimated by densitometry of gels or of each homodimer purified by chromatography varies with temperature. The more anodic homodimer is thermolabile and the less anodic one is thermostable. 3. The pH optimum of s-MDH is 7.5, of AA is 6.5 and of BB is 7.8. 4. The BB isozyme is more sensitive to high concentrations of substrate and has a Km temperature-independent. The AA isozyme is not inhibited by high concentrations of oxaloacetate and shows a Km temperature-dependent with a fourteenfold increase between 20 degrees and 40 degrees C.

Adaptative features of ectothermic enzymes--I. Temperature effects on the malate dehydrogenase from a temperate fish Leiostomus xanthurus.

1. Following electrophoresis the s-MDH activity of Leiostomus xanthurus and many other species of fish and amphibian appears in three sharp, equally-spaced, anodal bands. Each band is a dimer (AA, AB and BB) and two loci are active. 2. In Leiostomus tissue extracts A and B subunits are present are differing quantitative levels and their activities can be modified by changes in environment temperature. 3. Thermostability and thermal dependency tests show that, similar to what occurs during acclimatization, the AA isozyme is more stable to heat than is the BB isozyme. The BB isozyme is activated by low temperatures and is rapidly inactivated by high temperatures. 4. Extracts from a variety of fishes, amphibians, reptiles and birds suggest that when only one or two s-MDH bands are present, they behave as dose the AA homodimer in Leiostomus Xanthurus, i.e., are stable at elevated temperatures.

Adaptative features of ectothermic enzymes--II. The effects of acclimation temperature on the malate dehydrogenase of the spot. Leiostomus xanthurus.

1. Isozyme patterns and thermostability of the skeletal muscle and heart malate dehydrogenase (s-MDH) from the spot acclimated to different temperatures were examined. 2. No changes in isozyme patterns were seen for MDH in any of the tissues examined in response to 15 degrees and 20 degrees C acclimation. 3. The A-homodimer, which was more thermostable showed an increase in its relative activity, whereas the B-homodimer, which was more heat sensitive, showed a decrease during warm acclimation. However, after different periods of low temperature incubation, these samples showed a decrease in their subunit ratios. 4. Concerning the effect to thermal acclimation on the thermostability of MDH, it was found that skeletal muscle samples of the 30 degrees C-acclimated spot were more stable to heat than the 20 degrees and 15 degrees C-acclimated fishes.