The granular glands of anuran skin secrete an array of bioactive molecules that protect a frog against pathogens and predators. The skin also harbors a microbial community. Although there is evidence to suggest that the microbiota complement the innate immune defense systems against pathogen infection, the effect of the frog bioactive molecules on its resident microbiota has not yet been fully investigated. In the present study, the skin microbiota of Phyllomedusa distincta obtained from two different geographical areas was evaluated with molecular and culture-based approaches. The antagonistic effects exhibited by the host's microbiota and by a novel dermaseptin peptide isolated from P. distincta skin were investigated. Four isolated bacterial colonies displayed antimicrobial activity against known frog pathogens. Our results were consistent with the hypothesis that microbiota from P. distincta may interact with pathogenic microorganisms to protect a frog's health. On the other hand, the novel dermaseptin peptide exhibited an antimicrobial effect on pathogens as well as on some of the bacteria obtained from the skin microbiota. The richness of bacteria on P. distincta skin was further investigated by 16S rRNA gene clone libraries, which revealed that the family Enterobacteriaceae was prevalent, but a high variability at the species level was observed among individual frogs. Differences observed on the microbiota of frogs from contrasting habitats indicated an influence of the environment on the structure of the skin microbiota of P. distincta.

The cutaneous microbiota of amphibians can be defined as a biological component of protection, since it can be composed of bacteria that produce antimicrobial compounds. Several factors influence skin microbial structure and it is possible that environmental variations are among one of these factors, perhaps through physical-chemical variations in the skin. This community, therefore, is likely modified in habitats in which some ecophysiological parameters are altered, as in fragmented forests. Our research goal was to compare the skin bacterial community of four anuran species of the Atlantic Forest of Brazil in landscapes from two different environments: continuous forest and fragmented forest. The guiding hypotheses were: 1) microbial communities of anuran skin vary among sympatric frog species of the Atlantic forest; 2) the degree to which forested areas are intact affects the cutaneous bacterial community of amphibians. If the external environment influences the skin microbiota, and if such influences affect microorganisms capable of inhibiting the colonization of pathogens, we expect consequences for the protection of host individuals. We compared bacterial communities based on richness and density of colony forming units; investigated the antimicrobial potential of isolated strains; and did the taxonomic identification of isolated morphotypes. We collected 188 individual frogs belonging to the species Proceratophrys boiei, Dendropsophus minutus, Aplastodiscus leucopygius and Phyllomedusa distincta, and isolated 221 bacterial morphotypes. Our results demonstrate variation in the skin microbiota of sympatric amphibians, but only one frog species exhibited differences in the bacterial communities between populations from fragmented and continuous forest. Therefore, the variation we observed is probably derived from both intrinsic aspects of the host amphibian species and extrinsic aspects of the environment occupied by the host. Finally, we detected antimicrobial activity in 27 morphotypes of bacteria isolated from all four amphibian species.

The composition of the skin microbiota of amphibians is related to the biology of host species and environmental microbial communities. In this system, the environment serves as a microbial source and can modulate the hosted community. When habitats are fragmented and the environment disturbed, changes in the structure of this microbial community are expected. One important potential consequence of fragmentation is a compromised protective function of the microbiota against pathogenic microorganisms. In this study, the skin microbiota of the amphibian Proceratophrys boiei was characterized, evaluated for relationships with environmental variables and environmental sources of microbial communities, and its diversity evaluated for frog populations from fragmented and continuous forests. In addition, the antimicrobial activity of this skin community was studied in frogs from both forest types. Culture methods and 16S rRNA high-throughput gene sequencing were used to characterize the microbial community and demonstrated that the skin microbiota of P. boiei is more closely related to the soil microbial communities than those inhabiting water bodies or fragment matrix, the unforested area around the forested fragment. The microbial diversity and abundance of P. boiei skin microbiota are different between continuous forests and fragments. This community is correlated with environmental variables, especially with temperature of microhabitat and distance to human dwelling. All individuals of P. boiei harbored bacteria capable of inhibiting the growth of pathogenic bacteria and different strains of the pathogenic fungus Batrachochytrium dendrobatidis, and a total of 27 bacterial genera were detected. The results of this study indicate that the persistence of populations of this species will need balanced and sustained interactions among host, microorganisms, and environment. K E Y W O R D Samphibian, antimicrobials, environmental microbes, fragmented forests, metabolites, skin microbiota 902 | de ASSIS et Al.

Revisão ib.usp.br/revistaRevista da Biologia (2012) 8: 45-48 A perda da biodiversidade é uma consequência dos impactos ambientais provocados pela atividade humana nas áreas naturais (Keisecker e Blaustein, 2001; Pinto e Assis, 2005; Myers e col. 2000). Individualmente, os fatores apontados são bem conhecidos, destacando-se as mudanças climáticas, o aumento da incidência de radiação ultravioleta (UV-B), a introdução de espécies exóticas, a contaminação por agentes tóxicos e o desmatamento (Blaustein e Kiesecker, 2002 Resumo. O significado das mudanças climáticas para os anfíbios, em sinergismo com outros impactos antropogênicos, está atrelado às possíveis mudanças dos microclimas nos habitats ocupados. Nesse contexto, os eventos de epidemias merecem destaque, uma vez que, é provável que as modificações no meio ambiente contribuam para a crescente disseminação de doenças. A primeira proteção dos anfíbios contra muitas doenças é a pele, junto com as suas secreções e microbiota, e são primordiais porque muitos dos agentes infecciosos envolvidos atuam nesse tecido. As principais barreiras são a microbiota residente e as secreções cutâneas, que possuem efeito antibiótico contra organismos patogênicos conhecidos. O ambiente modula as caracterís-ticas inerentes a esses componentes, e por meio desses, mudanças nos padrões microclimáticos poderiam determinar os níveis de vulnerabilidade das populações de anfíbios aos patógenos. Palavras-chave. Anfíbios, microclima, microbiota, secreções cutâneas.Abstract. The meaning of climate change for amphibians, in synergy with other anthropogenic impacts, is linked to possible changes in the microclimate in the habitats occupied. In this context, the events of epidemics are highlighted, since it is likely that changes in the environment contribute to the increasing spread of diseases. The first protection of amphibians, against several diseases, is the skin with is associate secretions and microbiota, all of them essential because many infectious agents operate through the skin. The main barriers are the resident microflora and skin secretions, which have an antibiotic effect against pathogenic organisms known. The environment modulates the inherent characteristics of these components and at the same time, changes in climatic patterns may dictate significant changes in these components and, consequently, make amphibians more susceptible to pathogens. Keywords. Amphibians, microclimate, cutaneous microflora, skin secretions. Microbiota, secreções cutâneas e microclima: consequências para os anfíbiosMicroflora, skin secretions and microclimate: implications for amphibians Ananda Brito de Assis Departamento de Fisiologia, Instituto de Biociências, Universidade de São PauloAnfíbios são vulneráveis às mudanças no ambiente e, para estes, a ocorrência de epidemias tem sido mencionada como causa de declínio e extinção de populações (Carey e col., 1999;Schumacher, 2006). Uma maior vulnerabilidade dos anfíbios às modificações ambientais provavelmente está relacionada às características de...

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