Avian biodiversity assessment studies in a Neotropical wetland - the combination of sampling methods makes the difference

In studies of avian diversity, many different methods have been applied. Since methodological approaches may affect research results, the choice of a given methodology must be consistent with the scientific objectives. The aim of this study was to investigate how different methodologies with their intrinsic limitations help detect and monitor birds to evaluate how they complement each other in the survey of species. Three different assessment methods, mist nets, point counts, and autonomous acoustic recordings were used to serve this purpose in a study of different Pantanal habitats, such as savannas and forests. The point counts detected more species (126 species) than the two other methods autonomous acoustic recordings (113 species) and mist nets (79 species). We observed significant differences in the number of species detected by mist nets and the other two methods. Each survey method identified exclusive species. When comparing habitats, all three methods showed significant differences in bird species composition. Savannas were richer in bird species than forests, and replacement was the main driver responsible for the differences in beta diversity between the habitats. The three methodologies, when applied together, proved to be complementary in avian species detection.

Association of Kappa casein gene polymorphism with milk production traits in crossbred dairy cows / Associação do polimorfismo do gene Kappa-caseína com características de produção de leite em vacas leiteiras mestiças

Milk's qualitative and technological properties are greatly affected by genetic polymorphisms in the kappa-casein gene, and their polymorphisms may serve as informative markers of yield and composition. Thus, the objective of this study was to detect kappa-casein (kappa-CN) gene polymorphisms and their association with milk production traits in crossbred dairy cows. One hundred healthy crossbred (Friesian x Jenoubi) dairy animals between three and five years old were sampled for blood and milk during their first lactation. The genomic DNA was extracted from whole blood, and restriction fragment length polymorphism (RFLP-PCR) was used to determine the genotype of the kappa-CN gene. As a consequence of the restriction digestion of this fragment with Hind III, it showed three different restriction patterns: BB (453 base pairs uncut), AB (453, 206, and 225 base pairs), and AA (206 and 225 base pairs). Based on genetic diversity, the AB genotype was the most predominant (n = 67), with a frequency of 0.67. A variant genotype of the kappa-CN gene was associated with milk production traits in crossbred dairy cows. Animals with the AA variant produced a higher milk yield and a higher percentage of fat, casein, protein, and solids not fat (SNF) (P≤0.05) (1.397kg, 0.75%, 0.31%, 0.27%, and 0.68%, respectively) than those with the BB variant. A logistic regression analysis confirmed that the kappa-CN genotypes increase milk yield and casein content. Therefore, genetic variants of the kappa-CN gene could be used as genetic markers for improving milk production traits in dairy cattle.(AU)

As propriedades qualitativas e tecnológicas do leite são muito afetadas por polimorfismos genéticos no gene kappa-caseína e esses polimorfismos podem servir como marcadores informativos de rendimento e composição. Assim, o objetivo deste estudo foi detectar polimorfismos do gene kappa-caseína (kappa-CN) e sua associação com características de produção de leite em vacas leiteiras mestiças. Cem animais mestiços (Freisian x Jenoubi) saudáveis, entre três e cinco anos de idade, foram amostrados durante a primeira lactação para sangue e leite. O DNA genômico foi extraído do sangue total e o polimorfismo dos fragmentos de restrição do DNA genômico (RFLP-PCR) foi usado para determinar o genótipo do gene kappa-CN. Em consequência da digestão de restrição deste fragmento com Hind III, ele apresentou três padrões de restrição diferentes: BB (453 pares de bases não cortadas), AB (453,206 e 225 pares de bases) eAA (206 e 225 pares de bases). Com base na diversidade genética, o genótipo AB foi o mais predominante (n = 67), com frequência de 0,67. Genótipo variante do gene kappa-CN foi associado com características de produção de leite em vacas leiteiras mestiças. Animais com a variante AA tiveram maior produção de leite e maior percentual de gordura, caseína, proteína e sólidos não gordurosos (SNG) (P≤0,05) (l,397kg, 0,75%, 0,31%, 0,27% e 0,68%, respectivamente) do que aqueles com variante BB. Uma análise de regressão logística confirmou que os genótipos kappa-CN aumentam a produção de leite e o teor de caseína. Portanto, variantes genéticas do gene kappa-CN podem ser usadas como marcadores genéticos para melhorar as características de produção de leite em bovinos leiteiros.(AU)

Rhizosphere bacterial and fungal communities of healthy and wilted pepper (Capsicum annuum L.) in an organic farming system / Comunidades bacterianas e fúngicas da rizosfera de pimentão sadio e murcho(Capsicum annuum L.) em sistema de cultivo orgânico

ABSTRACT: Rhizosphere microorganisms play an important role in the growth and health of plants. Around the world, diverse soil-borne pathogens attack Capsicum annuum causing significant damage and economic losses. This study determined whether the diversity and composition of microbial communities in the rhizosphere soil of C. annuum plants is significantly changed by wilt disease. We used the 16S rRNA gene for bacteria and the internal transcribed spacer region for fungi to characterize the rhizosphere microbiomes of healthy and wilted plants. The most abundant bacterial phyla were Proteobacteria and Gemmatimonadetes, while the most abundant fungal phyla were Ascomycota and Mucoromycota. The bacterial α-diversity did not show significant differences in richness and diversity, but did show a significant difference in evenness and dominance of species. Rare taxa were present in both healthy and wilted conditions with relative abundances < 1%. In the fungi, all evaluated estimators showed a significant reduction in the wilted condition. The β-diversity showed significant differences in the structure of bacterial and fungal communities, which were segregated according to plant health conditions. The same occurred when comparing the alpha and beta diversity of this study based on organic agriculture with that of other studies based on conventional agriculture. We observed a significant difference with estimators analyzed by segregating rhizosphere communities depending on the farming method used. Finally, the differential abundance analysis did not show significant results in the bacterial communities; however, in the fungal communities, Fusarium, Thanatephorus, Rhizopus, Curvularia, Cladosporium, and Alternaria were more abundant in the rhizosphere of wilted than healthy plants. Species from these genera have been previously reported as phytopathogens of several plants, including C. annuum.

RESUMO: Microrganismos na rizosfera desempenham um papel importante no crescimento e saúde das plantas. Em todo o mundo, vários patógenos do solo atacam o Capsicum annuum causando danos significativos e perdas econômicas. Este estudo teve como objetivo determinar se a diversidade e composição das comunidades microbianas no solo da rizosfera de plantas de C. annuum é alterada significativamente pela murcha. Usamos o gene 16S rRNA para bactérias e a região espaçadora transcrita interna para fungos para caracterizar os microbiomas da rizosfera de plantas saudáveis e plantas com murcha. Os filos bacterianos mais abundantes foram Proteobacteria e Gemmatimonadetes, enquanto os filos fúngicos foram Ascomycota e Mucoromycota. A diversidade alfa bacteriana não mostrou diferenças significativas na riqueza e diversidade, mas mostrou uma diferença significativa na uniformidade e dominância das espécies. Táxons raros estavam presentes em condições saudáveis e murchas com abundância relativa < 1%. Em fungos, todos os estimadores avaliados apresentaram redução significativa na condição de murcha. A diversidade beta apresentou diferenças significativas na estrutura das comunidades bacterianas e fúngicas, que foram segregadas de acordo com as condições fitossanitárias. O mesmo aconteceu ao comparar a diversidade alfa e beta deste estudo baseado na agricultura orgânica com a de outros estudos baseados na agricultura convencional. Uma diferença significativa foi observada com os estimadores analisados segregando as comunidades da rizosfera dependendo do método de cultivo utilizado. Por fim, a análise de abundância diferencial não apresentou resultados significativos nas comunidades bacterianas; entretanto, nas comunidades fúngicas, os gêneros Fusarium, Thanatephorus, Rhizopus, Curvularia, Cladosporium e Alternaria foram mais abundantes na rizosfera de plantas murchas do que saudáveis. Várias espécies desses gêneros foram previamente relatadas como fitopatógenos de várias plantas, incluindo C. annuum.

Fecal microbiota profile of ovariohysterectomized cats submitted to estrogen replacement / Perfil da microbiota fecal de gatas ovariohisterectomizadas submetidas à reposição estrogênica

This study aimed to observe the effects of 17 ß-estradiol replacements on the fecal microbiota in spayed cats. Individual samples of fresh feces were collected and stored at -80° C. Sequencing of the V3/V4 regions of the 16S rRNA gene was used, and bioinformatic analysis was performed. Firmicutes/Bacteriodetes ratio was lower in the group receiving estrogen replacement compared to the SHAM group (P = 0,005). Jaccard index (P = 0.123) and Yue & Clayton index (P = 0.094) did not reveal alpha and beta diversity differences. The linear discriminant analysis effect size (LefSe) identified Firmicutes and MegasPhaera as the biomarkers for the SHAM group, and Burkholderiales, Betaproteobacteria, Sutterellaceae, Suterella, Proteobacteria, Proteobacteria unclassified and Collinsella for the group receiving estrogen replacement.(AU)

O objetivo deste estudo foi observar os efeitos da reposição de 17 ß-estradiol na microbiota fecal de gatas castradas. Amostras individuais de fezes frescas foram colhidas e armazenadas a -80°C. Foi realizado o sequenciamento das regiões V3/V4 do gene 16S rRNA e a análise bioinformática. A razão Firmicutes/Bacteriodetes foi menor no grupo que recebeu reposição estrogênica em comparação ao grupo SHAM (P = 0,005). O índice de Jaccard (P = 0,123) e o índice de Yue & Clayton (P = 0,094) não revelaram diferenças na alfa e beta diversidade. A análise discriminatória linear de tamanho do efeito (LefSe) identificou Firmicutes e Megasphaera como biomarcadores para o grupo SHAM, e Burkholderiales, Betaproteobacteria, Sutterellaceae, Suterella, Proteobacteria, Proteobacteria não classificada e Collinsella para o grupo que recebeu reposição estrogênica.(AU)

Fish community turnover in a dammed Andean River over time

We describe the change in the fish community of the Porce River in Magdalena River Basin, Colombia, following the construction of the Porce III hydropower reservoir based on 13 years of monitoring data. The results show a clear reduction of the number of native species, which have been supplanted by colonizing non-native species, especially in the reservoir. Four native species detected prior to dam construction have apparently disappeared, but 12 new species were registered post-construction. We analyzed spatial changes in beta diversity in the aquatic environments surrounding the dam. The new environment generated by the reservoir presents a unique species composition and contributes significantly to the total beta diversity of the system. Altogether three distinct new fish assemblages emerged following reservoir formation and there are now six assemblages where there had previously been three. This dramatic change, already visible within a decade of construction, highlights just how strong of an impact dam construction has on habitats and how rapidly fish communities react in this hotspot for endemic fish diversity. Our findings demonstrate the importance of monitoring fish communities for revealing the impact of damming on river ecosystems and informs potential complementary fish diversity inventories elsewhere in the Magdalena River basin.(AU)

Describimos el cambio en la comunidad de peces del río Porce en la cuenca del río Magdalena en Colombia luego de la construcción del embalse hidroeléctrico Porce III con base en 13 años de datos de monitoreo. Los resultados mostraron una reducción de especies nativas y, el embalse ha sido colonizado por especies no nativas. A lo largo del monitoreo cuatro especies desaparecieron de las capturas, y se registró la aparición de 12 especies nuevas después de la construcción del embalse. Analizamos los cambios espaciales en la diversidad beta en los ambientes acuáticos que rodean la presa. El nuevo entorno generado por el embalse presenta una composición de especies única; en total, surgieron tres nuevos ensamblajes de peces distintos después de la formación del embalse y ahora hay seis ensamblajes. Este cambio, ya visible a una década de la construcción, resalta cuán fuerte es el impacto que tiene la construcción de presas en los hábitats y cuán rápido reaccionan las comunidades de peces. Nuestros hallazgos demuestran la importancia de monitorear las comunidades de peces para revelar el impacto de las represas en los ecosistemas fluviales, además permite complementar los inventarios de diversidad en la cuenca del río Magdalena.(AU)

Effects of small-scale anthropogenic alterations in Euglossini bees at eastern Amazon forest / Efeitos de alterações antrópicas de pequena escala em abelhas Euglossini na floresta da Amazônia oriental

In this study, through a rapid assessment, we evaluated differences in richness, abundance, species composition and beta diversity of Euglossini bees in preserved areas and areas altered by riverine human populations in the Caxiuanã National Forest (northern Amazon). Our hypotheses were: i) the species richness, diversity, abundance and composition of Euglossini bees differ between the preserved and altered areas because many species are intolerant to small anthropic disturbances; and ii) areas altered by riverine populations have higher values ​​of beta diversity (species dissimilarity between sites) than preserved areas, due to local changes in habitat and increased heterogeneity between sites. We sampled two areas (preserved and altered) and, in each area, four transects were sampled with odorous baits. We used a Student's t test to evaluate differences in richness, abundance, and Shannon-Wiener diversity index. We also evaluated differences in composition and differences in beta diversity between areas. We collected 102 specimens, identified in 18 species and three genera, 63 individuals (61.8% of the total) in preserved areas and 39 individuals (38.2%) in altered areas. Euglossa liopoda (Dressler, 1982) was the most abundant in altered areas and Euglossa imperialis (Cockerell, 1922) the most abundant in preserved areas. Species richness was 2.5 times greater in the transects of preserved areas, and species composition also differed between the sampled areas. In contrast, species abundance did not differ between transects of both areas. Despite our small sample size, this study demonstrated that the richness, composition and beta diversity of Euglossini bees can be potentially affected by local changes in habitat caused by small-scale activities provided by traditional Amazonian groups. Our results indicate the urgency for more comprehensive studies aimed to investigate these relationships.

Neste estudo, por meio de uma avaliação rápida, investigamos diferenças na riqueza, abundância, composição de espécies e diversidade beta de abelhas Euglossini em áreas preservadas e áreas alteradas por populações ribeirinhas na Floresta Nacional de Caxiuanã (norte da Amazônia). Nossas hipóteses foram: i) a riqueza de espécies, diversidade, abundância e composição das abelhas Euglossini diferem entre as áreas preservadas e alteradas, uma vez que muitas espécies são intolerantes a pequenos distúrbios antrópicos; e ii) áreas alteradas por populações ribeirinhas apresentam maiores valores de diversidade beta (dissimilaridade de espécies entre locais) do que áreas preservadas, devido às mudanças locais no habitat e aumento da heterogeneidade entre locais. Amostramos duas áreas (preservadas e alteradas) e, em cada área, foram amostrados quatro transectos com iscas odoríferas. Nós utilizamos testes t de Student, para avaliar diferenças de riqueza, abundância e índice de diversidade de Shannon-Wiener. Também avaliamos as diferenças na composição e as diferenças na diversidade beta entre as áreas. Foram coletados 102 espécimes, em 18 espécies e três gêneros, 63 deles (61,8%) em áreas preservadas e 39 (38,2%) em áreas alteradas. Euglossa liopoda (Dressler, 1982) foi a mais abundante nas áreas alteradas e Euglossa imperialis (Cockerell, 1922) nas áreas preservadas. A riqueza foi 2,5 vezes maior nos transectos de áreas preservadas, e a composição de espécies também diferiu entre as áreas amostradas. Em contraste, a abundância de espécies não foi diferente entre os transectos das duas áreas. Apesar de nosso esforço amostral reduzido, nossos resultados demonstram que a riqueza, composição e diversidade beta das abelhas Euglossini podem potencialmente ser afetadas por mudanças locais no habitat causadas por atividades de pequena escala fornecidas por grupos tradicionais da Amazônia. Esses resultados indicam a urgência por estudos mais extensos no sentido de investigar as relações aqui estudadas.

Protected areas and compositional diversity of fish from Serranias Costeiras of the Ribeira de Iguape River basin, Southeast Brazil

The Serranias Costeiras of the Ribeira de Iguape River basin comprise landscapes with steep slopes, where many streams are contained in protected areas. We evaluated the importance of these protected areas for stream fish diversity. We sampled the ichthyofauna during the dry season of 2010, 2018 and 2019 in 36 stream stretches. We used beta diversity measures and estimated species richness and dark diversity in streams from two types of protected areas (full protection, FP and sustainable use, SU) and outside (Out). The altitude-width and velocity gradient of the streams explained the species turnover. The PA type promoted the richness difference, with FP streams having less species richness than SU and outside. Streams from FP presented lower species richness and dark diversity. These results indicate that the few species in FPs are well protected. The FP streams contain a relevant proportion of the regional species pool and, therefore, are essential for conserving fish stream diversity in the study region. On the other hand, streams from SU or Out have higher species richness, but their fish fauna is more vulnerable. Due to longitudinal stream connectivity, we highlight the importance of rethinking the limits of protected areas.(AU)

Os riachos das Serranias Costeiras da bacia do rio Ribeira de Iguape estão na região do estado de São Paulo com maior quantidade de áreas preservadas e com ictiofauna muito particular. Avaliamos a importância das áreas preservadas na diversidade da ictiofauna. Durante o período de seca de 2010, 2018 e 2019 coletamos a ictiofauna em 36 trechos de riachos em dois tipos de áreas protegidas (Proteção Integral, FP e Uso Sustentável, SU) e fora (Out). Utilizamos medidas de diversidade beta e estimamos a riqueza de espécies e a diversidade escura. A ictiofauna regional apresentou alta diversidade beta. O gradiente de largura e velocidade dos riachos e altitudinal explicou a substituição de espécies. A diferença de riqueza foi promovida pelo tipo de UC sendo que os riachos FP possuem menor riqueza de espécies que os SU e fora das UCs. Os riachos inseridos em FP contêm uma proporção relevante do pool regional de espécies e, portanto, são importantes para a conservação da diversidade de riachos na região de estudo. Por outro lado, os riachos que estão em SU ou Out possuem maior riqueza de espécies e sua fauna de peixes está mais vulnerável. Devido à conectividade longitudinal dos riachos ressaltamos a importância de repensar os limites das unidades de conservação.(AU)

Plant diversity in Sabkha ecosystems of arid region: spatial and environmental drivers / Diversidade de plantas em ecossistemas Sabkha da região árida: drivers espaciais e ambientais

This study investigated the effects of spatial and environmental factors and their interactions on plant species composition in salt marsh (Sabkha) ecosystem located in arid region (Saudi Arabia). The plant species and environmental variables were investigated in 38 sites located in three regions. A total of 15 environmental variables were measured in each site and the geographical coordinates were used to extract spatial variables (using PCNM). A total of 81plant species were reported from 38 sites. The three regions showed patterns of homogeneity of multivariate dispersions (i.e. beta diversity). The PCNM analysis extracted 18 PCNM vectors and only 3 vectors were retained after forward selection. The spatial variables (selected PCNM vectors) explained only 3.21% of the variation in species composition of plants (using variation partitioning technique). However, eight environmental variables were selected after forward selection (Lead, Copper, total organic matter, Potassium, Magnesium, pH, Zinc and Iron, F= 4.72, P<0.05) and explained 19.61% of the total variation in the species composition. In conclusion, the plant communities in Sabkhas were not spatially structured due to the low percentage of variation explained by the spatial variables (PCNM vectors). The environmental variables were corresponded to the high fraction of variation explained. On the other hand, Sabkhas in Saudi Arabia are considered a hot spot for diversity not only for plants but for other animals (birds, vertebrates and invertebrates). Therefore, immediate conservation plans should be implemented to reduce the adverse effect of urbanization, industrialization as well as other anthropogenic activities.

Este estudo investigou os efeitos de fatores espaciais e ambientais e suas interações na composição de espécies vegetais no ecossistema de sapal (Sabkha) localizado na região árida (Arábia Saudita). As espécies vegetais e variáveis ​​ambientais foram investigadas em 38 locais localizados em três regiões. Um total de 15 variáveis ​​ambientais foi medido em cada local, e as coordenadas geográficas foram usadas para extrair as variáveis ​​espaciais (usando PCNM). Um total de 81 espécies de plantas foi relatado em 38 locais. As três regiões mostraram padrões de homogeneidade de dispersões multivariadas (ou seja, diversidade beta). A análise PCNM extraiu 18 vetores PCNM e apenas 3 vetores foram retidos após a seleção direta. As variáveis ​​espaciais (vetores PCNM selecionados) explicaram apenas 3,21% da variação na composição de espécies das plantas (utilizando a técnica de partição de variação). No entanto, 8 variáveis ​​ambientais foram selecionadas após seleção direta (chumbo, cobre, matéria orgânica total, potássio, magnésio, pH, zinco e ferro, F = 4,72, P < 0,05) e explicaram 19,61% da variação total na composição de espécies. Em conclusão, as comunidades vegetais em Sabkhas não foram espacialmente estruturadas devido à baixa porcentagem de variação explicada pelas variáveis ​​espaciais (vetores PCNM). As variáveis ​​ambientais corresponderam à alta fração de variação explicada. Por outro lado, os Sabkhas na Arábia Saudita são considerados um hot spot de diversidade não apenas para plantas, mas para outros animais (aves, vertebrados e invertebrados). Portanto, planos de conservação imediatos devem ser implementados para reduzir os efeitos adversos da urbanização, industrialização e outras atividades antrópicas.

PCR-RFLP Based genetic diversity of Plasmodium vivax genotypes in district Mardan, Pakistan

Abstract Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3 and Pvmsp-3genes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3 and Pvmsp3 genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp-3 genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3genes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3 and Pvmsp-3 genes of P. vivax isolates by using PCR/RFLP from District Mardan and showed a remarkable level of genetic diversity in the studied genes of circulating parasites in the study area. The results of this study will contribute in future studies about the genetic structure of parasite and vaccine development against the malaria.

Resumo O Plasmodium vivax é o parasita da malária humana mais comum nos países asiáticos, incluindo o Paquistão. O presente estudo foi desenhado para explorar a diversidade genética de genótipos de Plasmodium vivax baseados nos genes Pvmsp-3 e Pvmsp-3, usando marcadores de ensaios alélicos nested PCR e RFLP de isolados de campo no distrito de Mardan, Paquistão. Amostras de sangue de 200 pacientes com malária por P. vivax foram coletadas após assinatura do termo de consentimento livre e esclarecido. A diversidade genética em produtos de PCR nested foi determinada por polimorfismo de fragmento de restrição (RFLP) utilizando as enzimas de restrição Alu1 e PstI para a digestão dos produtos dos genes alfa e beta, respectivamente. Para análise da diversidade genética das variantes subalélicas dos genes Pvmsp3 e Pvmsp3, o teste Qui-quadrado foi realizado utilizando o software de programação Minitab 18. O valor P = 0,05 foi considerado estatisticamente significativo. Para os genes Pvmsp-3, após eletroforese em gel de produtos digeridos, quatro genótipos distintos foram obtidos de um total de 50 amostras; tipo A: 35 (70%) (1,5-2,0 kb), 12 do tipo B (24%) (1,5-1,7 kb), 2 do tipo C (4%) (0,5-1,5) e um para o tipo D (2%) (0,5-0,65 kb), que podem ser caracterizados em nove padrões alélicos (A1-A4, B1-B3, C1, D), em que A3 permaneceu como o mais predominante. Para Pvmsp-3genes, três genótipos distintos foram obtidos a partir de 50 amostras; 40 (80%) do tipo A (1,5-2,5 kb), 9 (18%) do tipo B (1,0-1,5 kb) e 1 (2%) do tipo C (0,65 kb), que podem ser caracterizados em seis padrões alélicos (A1-A3, B1-B2 e C1). Os mais dominantes no tipo A foram o alelo A1, observados em 46%, enquanto, no tipo B, os mais dominantes foram B1 (10%). Este estudo é o primeiro relato de epidemiologia molecular e variação genética em Pvmsp-3. Os genes Pvmsp-3 de isolados de P. vivax utilizando PCR/RFLP do Distrito Mardan mostraram um nível notável de diversidade genética nos genes estudados de parasitas circulantes na área de estudo. Os resultados desse estudo contribuirão em estudos futuros sobre a estrutura genética do parasita e o desenvolvimento de vacinas contra a malária.

PCR-RFLP Based genetic diversity of Plasmodium vivax genotypes in district Mardan, Pakistan / Diversidade genética baseada em PCR-RFLP de genótipos de Plasmodium vivax no distrito de Mardan, Paquistão

Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3α and Pvmsp-3ßgenes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3α and Pvmsp3ß genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp3α genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3ßgenes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3α and Pvmsp-3ß genes of P. vivax isolates by using PCR/RFLP from District Mardan and showed a remarkable level of genetic diversity in the studied genes of circulating parasites in the study area. The results of this study will contribute in future studies about the genetic structure of parasite and vaccine development against the malaria.

O Plasmodium vivax é o parasita da malária humana mais comum nos países asiáticos, incluindo o Paquistão. O presente estudo foi desenhado para explorar a diversidade genética de genótipos de Plasmodium vivax baseados nos genes Pvmsp-3α e Pvmsp-3ß, usando marcadores de ensaios alélicos nested PCR e RFLP de isolados de campo no distrito de Mardan, Paquistão. Amostras de sangue de 200 pacientes com malária por P. vivax foram coletadas após assinatura do termo de consentimento livre e esclarecido. A diversidade genética em produtos de PCR nested foi determinada por polimorfismo de fragmento de restrição (RFLP) utilizando as enzimas de restrição Alu1 e PstI para a digestão dos produtos dos genes alfa e beta, respectivamente. Para análise da diversidade genética das variantes subalélicas dos genes Pvmsp3α e Pvmsp3ß, o teste Qui-quadrado foi realizado utilizando o software de programação Minitab 18. O valor P = 0,05 foi considerado estatisticamente significativo. Para os genes Pvmsp3α, após eletroforese em gel de produtos digeridos, quatro genótipos distintos foram obtidos de um total de 50 amostras; tipo A: 35 (70%) (1,5-2,0 kb), 12 do tipo B (24%) (1,5-1,7 kb), 2 do tipo C (4%) (0,5-1,5) e um para o tipo D (2%) (0,5-0,65 kb), que podem ser caracterizados em nove padrões alélicos (A1-A4, B1-B3, C1, D), em que A3 permaneceu como o mais predominante. Para Pvmsp-3ßgenes, três genótipos distintos foram obtidos a partir de 50 amostras; 40 (80%) do tipo A (1,5-2,5 kb), 9 (18%) do tipo B (1,0-1,5 kb) e 1 (2%) do tipo C (0,65 kb), que podem ser caracterizados em seis padrões alélicos (A1-A3, B1-B2 e C1). Os mais dominantes no tipo A foram o alelo A1, observados em 46%, enquanto, no tipo B, os mais dominantes foram B1 (10%). Este estudo é o primeiro relato de epidemiologia molecular e variação genética em Pvmsp-3α. Os genes Pvmsp-3ß de isolados de P. vivax utilizando PCR/RFLP do Distrito Mardan mostraram um nível notável de diversidade genética nos genes estudados de parasitas circulantes na área de estudo. Os resultados desse estudo contribuirão em estudos futuros sobre a estrutura genética do parasita e o desenvolvimento de vacinas contra a malária.

Effect of Dietary Tetramethylpyrazine on Egg Production, Nutrient Retention and Cecal Bacterial Diversity in Aged Laying Hens

This study aimed to investigate the effect of tetramethylpyrazine (TMP) supplementation on egg production, nutrient retention and cecal microbiota diversity using 288 commercial Hy-Line brown hens as of wk 75 to 86. Four treatments consisted of TMP addition at 0 (control, basal diet), 100, 150 and 200 mg/kg of diet. The results showed that diets supplemented with TMP addition improved egg-laying rate as of wk 77 compared to the control, which led to an increase (p<0.001) of egg mass by 97-225 g/hen throughout the whole trial, and a linear increase (p=0.003) of egg mass to the incremental TMP doses was found. At wk 86, the apparent digestibilities of dry matter and crude protein were enhanced (p<0.05), exhibiting consistent linear increases (p≤0.033) with the TMP doses. However, TMP did not cause alpha and beta diversity of cecal microbiota. The results suggest that TMP can be an additive to improve egg production and nutrient digestibility of aged laying hens.(AU)

PCR-RFLP Based genetic diversity of Plasmodium vivax genotypes in district Mardan, Pakistan / Diversidade genética baseada em PCR RFLP de genótipos de Plasmodium vivax no distrito de Mardan, Paquistão

Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3α and Pvmsp-3βgenes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3α and Pvmsp3β genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp 3α genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3βgenes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3α and Pvmsp-3β genes of P. vivax isolates by using PCR/RFLP from District Mardan and [...].

O Plasmodium vivax é o parasita da malária humana mais comum nos países asiáticos, incluindo o Paquistão. O presente estudo foi desenhado para explorar a diversidade genética de genótipos de Plasmodium vivax baseados nos genes Pvmsp-3α e Pvmsp-3β, usando marcadores de ensaios alélicos nested PCR e RFLP de isolados de campo no distrito de Mardan, Paquistão. Amostras de sangue de 200 pacientes com malária por P. vivax foram coletadas após assinatura do termo de consentimento livre e esclarecido. A diversidade genética em produtos de PCR nested foi determinada por polimorfismo de fragmento de restrição (RFLP) utilizando as enzimas de restrição Alu1 e PstI para a digestão dos produtos dos genes alfa e beta, respectivamente. Para análise da diversidade genética das variantes subalélicas dos genes Pvmsp3α e Pvmsp3β, o teste Qui-quadrado foi realizado utilizando o software de programação Minitab 18. O valor P = 0,05 foi considerado estatisticamente significativo. Para os genes Pvmsp 3α, após eletroforese em gel de produtos digeridos, quatro genótipos distintos foram obtidos de um total de 50 amostras; tipo A: 35 (70%) (1,5-2,0 kb), 12 do tipo B (24%) (1,5-1,7 kb), 2 do tipo C (4%) (0,5-1,5) e um para o tipo D (2%) (0,5-0,65 kb), que podem ser caracterizados em nove padrões alélicos (A1-A4, B1-B3, C1, D), em que A3 permaneceu como o mais predominante. Para Pvmsp-3βgenes, três genótipos distintos foram obtidos a partir de 50 amostras; 40 (80%) do tipo A (1,5-2,5 kb), 9 (18%) do tipo B (1,0-1,5 kb) e 1 (2%) do tipo C (0,65 kb), que podem ser caracterizados em seis padrões alélicos (A1-A3, B1-B2 e C1). Os mais dominantes no tipo A foram o alelo A1, observados em 46%, enquanto, no tipo B, os mais dominantes foram B1 (10%). Este estudo é o primeiro relato de epidemiologia molecular e variação genética em Pvmsp-3α. Os genes Pvmsp-3β de isolados de P. vivax utilizando PCR/RFLP do Distrito Mardan mostraram um nível notável de diversidade genética nos genes estudados [...].

PCR-RFLP Based genetic diversity of Plasmodium vivax genotypes in district Mardan, Pakistan / Diversidade genética baseada em PCR RFLP de genótipos de Plasmodium vivax no distrito de Mardan, Paquistão

Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3α and Pvmsp-3βgenes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3α and Pvmsp3β genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp 3α genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3βgenes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3α and Pvmsp-3β genes of P. vivax isolates by using PCR/RFLP from District Mardan and [...].(AU)

O Plasmodium vivax é o parasita da malária humana mais comum nos países asiáticos, incluindo o Paquistão. O presente estudo foi desenhado para explorar a diversidade genética de genótipos de Plasmodium vivax baseados nos genes Pvmsp-3α e Pvmsp-3β, usando marcadores de ensaios alélicos nested PCR e RFLP de isolados de campo no distrito de Mardan, Paquistão. Amostras de sangue de 200 pacientes com malária por P. vivax foram coletadas após assinatura do termo de consentimento livre e esclarecido. A diversidade genética em produtos de PCR nested foi determinada por polimorfismo de fragmento de restrição (RFLP) utilizando as enzimas de restrição Alu1 e PstI para a digestão dos produtos dos genes alfa e beta, respectivamente. Para análise da diversidade genética das variantes subalélicas dos genes Pvmsp3α e Pvmsp3β, o teste Qui-quadrado foi realizado utilizando o software de programação Minitab 18. O valor P = 0,05 foi considerado estatisticamente significativo. Para os genes Pvmsp 3α, após eletroforese em gel de produtos digeridos, quatro genótipos distintos foram obtidos de um total de 50 amostras; tipo A: 35 (70%) (1,5-2,0 kb), 12 do tipo B (24%) (1,5-1,7 kb), 2 do tipo C (4%) (0,5-1,5) e um para o tipo D (2%) (0,5-0,65 kb), que podem ser caracterizados em nove padrões alélicos (A1-A4, B1-B3, C1, D), em que A3 permaneceu como o mais predominante. Para Pvmsp-3βgenes, três genótipos distintos foram obtidos a partir de 50 amostras; 40 (80%) do tipo A (1,5-2,5 kb), 9 (18%) do tipo B (1,0-1,5 kb) e 1 (2%) do tipo C (0,65 kb), que podem ser caracterizados em seis padrões alélicos (A1-A3, B1-B2 e C1). Os mais dominantes no tipo A foram o alelo A1, observados em 46%, enquanto, no tipo B, os mais dominantes foram B1 (10%). Este estudo é o primeiro relato de epidemiologia molecular e variação genética em Pvmsp-3α. Os genes Pvmsp-3β de isolados de P. vivax utilizando PCR/RFLP do Distrito Mardan mostraram um nível notável de diversidade genética nos genes estudados [...].(AU)

Mesoscale bird distribution pattern in montane phytophysiognomies along an ecotone between two hotspots

Brazil has a high diversity of birds and presents the largest number of threatened bird species in the neotropical region. Even so, there are gaps in the bird knowledge, especially in ecotonal montane regions. Given this panorama, this study aimed to analyse the bird community distribution (richness, composition, and beta diversity between phytophysiognomies) of an ecotonal montane landscape of southeastern Brazil, with the purpose of detecting substitution patterns of bird species on a meso-scale. Using bird data performed during the years 1998 to 2015 in 46 sampling points, we found high bird richness in montane phytophysiognomies along an ecotone between Cerrado and Atlantic Forest hotspots. The composition present species of both domains, with high turnover component. We highlight the field environments and candeais are considered homogeneous and threathened, which would directly affect birds. The present study contributes to future conservation strategies, as it demonstrates ecotonal regions as transition zones and reinforces the need to consider as particular ecological units. These ecotonal regions are key locations for understanding ecological patterns in response to environmental changes or phytophysiognomies. Knowing how partitioning of the composition occurs within an environmental mosaic is essential to understand the limits and distributions of the species and conserve them.

Mesoscale bird distribution pattern in montane phytophysiognomies along an ecotone between two hotspots

Brazil has a high diversity of birds and presents the largest number of threatened bird species in the neotropical region. Even so, there are gaps in the bird knowledge, especially in ecotonal montane regions. Given this panorama, this study aimed to analyse the bird community distribution (richness, composition, and beta diversity between phytophysiognomies) of an ecotonal montane landscape of southeastern Brazil, with the purpose of detecting substitution patterns of bird species on a meso-scale. Using bird data performed during the years 1998 to 2015 in 46 sampling points, we found high bird richness in montane phytophysiognomies along an ecotone between Cerrado and Atlantic Forest hotspots. The composition present species of both domains, with high turnover component. We highlight the field environments and candeais are considered homogeneous and threathened, which would directly affect birds. The present study contributes to future conservation strategies, as it demonstrates ecotonal regions as transition zones and reinforces the need to consider as particular ecological units. These ecotonal regions are key locations for understanding ecological patterns in response to environmental changes or phytophysiognomies. Knowing how partitioning of the composition occurs within an environmental mosaic is essential to understand the limits and distributions of the species and conserve them.(AU)

Biotic differentiation in headwater creeks after the massive introduction of non-native freshwater aquarium fish in the Paraíba do Sul River basin, Brazil

This study evaluated fish beta diversity in six headwater creeks located in the area affected by the largest ornamental aquaculture center implemented in the Minas Gerais State, southeastern Brazil. We sampled fish assemblages in 2017 and 2018 to investigate changes in assemblage structure (species richness and beta diversity), comparing these data with the historic species pool. We recorded 60 fish species, of which 16 were native and 44 non-native with 19 translocated, and 25 exotic. The exotics Poecilia reticulata, Xiphophorus maculatus, X. variatus, Danio rerio, and Misgurnus anguillicaudatus were the most widely distributed in the headwater creeks. The Contamination Index showed that most creeks had high proportional contamination by exotic species (above 60%). Beta diversity increased from historical to contemporary periods in all creeks due to the introduction and differential colonization pressure of several non-native translocated and exotic species, indicating biotic differentiation. Temperature and number of ponds were the main preditors of change in beta diversity in the headwater creeks during the contemporary period. In summary, we observed that invaders have induced substantial changes to fish communities under influence of environmental filters. Our results support the hipothesis that aquaculture is a main driver of fish non-native fish introduction and native biodiversity loss in the Neotropics.(AU)

Este estudo avaliou a diversidade beta de peixes em seis riachos de cabeceira localizados em uma área afetada pelo maior centro de aqüicultura ornamental do Brasil, localizado em Minas Gerais. Amostramos assembleias de peixes em 2017 e 2018 para investigar mudanças na estrutura (riqueza de espécies e diversidade beta), comparando esses dados com tendências históricas de composição de comunidades. Registramos 60 espécies de peixes, sendo 16 nativos e 44 não nativos: 19 translocados e 25 exóticos. Os exóticos Poecilia reticulata, Xiphophorus maculatus, X. variatus, Danio rerio e Misgurnus anguillicaudatus foram os mais distribuídos nos riachos. O Índice de Contaminação mostrou que a maioria dos riachos apresentou alta contaminação proporcional por espécies exóticas (acima de 60%). A diversidade beta aumentou do período histórico para o contemporâneo em todos os riachos devido à introdução e pressão de colonização de várias espécies não-nativas translocadas e exóticas, indicando diferenciação biótica. Temperatura da água e número de tanques de piscicultura foram os principais fatores de mudança na diversidade beta dos riachos no período contemporâneo. Os não-nativos induziram mudanças em nível de comunidades e sob influência de variáveis ambientais. Os resultados mostram que a aquicultura é um dos principais vetores da introdução de peixes não-nativos e perda de biodiversidade nativa nos Neotrópicos.(AU)

Biotic differentiation in headwater creeks after the massive introduction of non-native freshwater aquarium fish in the Paraíba do Sul River basin, Brazil

This study evaluated fish beta diversity in six headwater creeks located in the area affected by the largest ornamental aquaculture center implemented in the Minas Gerais State, southeastern Brazil. We sampled fish assemblages in 2017 and 2018 to investigate changes in assemblage structure (species richness and beta diversity), comparing these data with the historic species pool. We recorded 60 fish species, of which 16 were native and 44 non-native with 19 translocated, and 25 exotic. The exotics Poecilia reticulata, Xiphophorus maculatus, X. variatus, Danio rerio, and Misgurnus anguillicaudatus were the most widely distributed in the headwater creeks. The Contamination Index showed that most creeks had high proportional contamination by exotic species (above 60%). Beta diversity increased from historical to contemporary periods in all creeks due to the introduction and differential colonization pressure of several non-native translocated and exotic species, indicating biotic differentiation. Temperature and number of ponds were the main preditors of change in beta diversity in the headwater creeks during the contemporary period. In summary, we observed that invaders have induced substantial changes to fish communities under influence of environmental filters. Our results support the hipothesis that aquaculture is a main driver of fish non-native fish introduction and native biodiversity loss in the Neotropics.(AU)

Este estudo avaliou a diversidade beta de peixes em seis riachos de cabeceira localizados em uma área afetada pelo maior centro de aqüicultura ornamental do Brasil, localizado em Minas Gerais. Amostramos assembleias de peixes em 2017 e 2018 para investigar mudanças na estrutura (riqueza de espécies e diversidade beta), comparando esses dados com tendências históricas de composição de comunidades. Registramos 60 espécies de peixes, sendo 16 nativos e 44 não nativos: 19 translocados e 25 exóticos. Os exóticos Poecilia reticulata, Xiphophorus maculatus, X. variatus, Danio rerio e Misgurnus anguillicaudatus foram os mais distribuídos nos riachos. O Índice de Contaminação mostrou que a maioria dos riachos apresentou alta contaminação proporcional por espécies exóticas (acima de 60%). A diversidade beta aumentou do período histórico para o contemporâneo em todos os riachos devido à introdução e pressão de colonização de várias espécies não-nativas translocadas e exóticas, indicando diferenciação biótica. Temperatura da água e número de tanques de piscicultura foram os principais fatores de mudança na diversidade beta dos riachos no período contemporâneo. Os não-nativos induziram mudanças em nível de comunidades e sob influência de variáveis ambientais. Os resultados mostram que a aquicultura é um dos principais vetores da introdução de peixes não-nativos e perda de biodiversidade nativa nos Neotrópicos.(AU)

Nestedness of stream insects in Subtropical region: importance of inter-annual temporal scale

ABSTRACT Stream insects use in different ways the stream habitats which affect patterns of species occurrence on stream. The nestedness is one of these patterns being an ecological mechanism that generates beta diversity. In this study, we verify the effects of temporal environmental variations on nestedness patterns of Chironomidae assemblages on subtropical streams. We collected chironomid larvae during winter and summer during three years (2010, 2011 and 2012) in Brazilian Subtropical low-order streams (Erechim, state of Rio Grande do Sul, Brazil). We measured environmental variables in each stream. To assess if assemblages will show nested patterns intra- and interannual we used a NODF metric. We observed nestedness only in one year, this pattern was due to variation of limnological factors, which are directly associated with climatic conditions (precipitation and water temperature) and agricultural management (e.g. dissolved oxygen and nutrients). Climatological events affect Chironomidae community through the time. The variations on precipitation values could be one of the mainly factors that influence nestedness for this community. Finally, studies on nesting on time scales should consider inter-annual comparisons, as climatic variations are more evident.

Quantity - but not diversity - of secreted peptides and proteins increases with age in the tree frog Pithecopus nordestinus

Amphibians inhabit the terrestrial environment, a conquest achieved after several evolutionary steps, which were still insufficient to make them completely independent of the aquatic environment. These processes gave rise to many morphological and physiological changes, making their skin (and cutaneous secretion) rich in bioactive molecules. Among the tree frogs, the secretion is composed mainly of peptides; but alkaloids, proteins and steroids can also be found depending on the species. The most known class of biologically active molecules is the antimicrobial peptides (AMPs) that act against bacteria, fungi and protozoans. Although these molecules are well-studied among the hylids, AMPs ontogeny remains unknown. Therefore, we performed peptidomic and proteomic analyses of Pithecopus nordestinus (formerly Phyllomedusa nordestina) in order to evaluate the peptide content in post-metamorphosed juveniles and adult individuals. Methods: Cutaneous secretion of both life stages of individuals was obtained and analyzed by LC-MS/MS after reduction and alkylation of disulfide bonds or reduction, alkylation and hydrolysis by trypsin. Results: Differences in the TIC profile of juveniles and adults in both treatments were observed. Moreover, the proteomic data revealed known proteins and peptides, with slight differences in the composition, according to the life stage and the treatment. AMPs were identified, and bradykinin-potentiating peptides were observed in trypsin-treated samples, which suggests a protein source of such peptide (cryptide). Conclusion: In general, skin secretion contents were similar between juveniles and adults, varying in quantity, indicating that the different stages of life are reflected in the number of molecules and not on their diversity.(AU)

Quantity - but not diversity - of secreted peptides and proteins increases with age in the tree frog Pithecopus nordestinus

Amphibians inhabit the terrestrial environment, a conquest achieved after several evolutionary steps, which were still insufficient to make them completely independent of the aquatic environment. These processes gave rise to many morphological and physiological changes, making their skin (and cutaneous secretion) rich in bioactive molecules. Among the tree frogs, the secretion is composed mainly of peptides; but alkaloids, proteins and steroids can also be found depending on the species. The most known class of biologically active molecules is the antimicrobial peptides (AMPs) that act against bacteria, fungi and protozoans. Although these molecules are well-studied among the hylids, AMPs ontogeny remains unknown. Therefore, we performed peptidomic and proteomic analyses of Pithecopus nordestinus (formerly Phyllomedusa nordestina) in order to evaluate the peptide content in post-metamorphosed juveniles and adult individuals. Methods: Cutaneous secretion of both life stages of individuals was obtained and analyzed by LC-MS/MS after reduction and alkylation of disulfide bonds or reduction, alkylation and hydrolysis by trypsin. Results: Differences in the TIC profile of juveniles and adults in both treatments were observed. Moreover, the proteomic data revealed known proteins and peptides, with slight differences in the composition, according to the life stage and the treatment. AMPs were identified, and bradykinin-potentiating peptides were observed in trypsin-treated samples, which suggests a protein source of such peptide (cryptide). Conclusion: In general, skin secretion contents were similar between juveniles and adults, varying in quantity, indicating that the different stages of life are reflected in the number of molecules and not on their diversity.(AU)