RESUMO
Abstract Polymerase chain reaction (PCR) assays targeting 16S rRNA genes followed by DNA sequencing are still important tools to characterize microbial communities present in environmental samples. However, despite the crescent number of deposited archaeal DNA sequences in databases, until now we do not have a clear picture of the effectiveness and specificity of the universal primers widely used to describe archaeal communities from different natural habitats. Therefore, in this study, we compared the phylogenetic profile obtained when Cerrado lake sediment DNA samples were submitted to 16S rDNA PCR employing three Archaea-specific primer sets commonly used. Our findings reveal that specificity of primers differed depending on the source of the analyzed DNA. Furthermore, archaeal communities revealed by each primer pair varied greatly, indicating that 16S rRNA gene primer choice affects the community profile obtained, with differences in both taxon detection and operational taxonomic unit (OTU) estimates.
Resumo A amplificação de genes que codificam o rRNA 16S por reação em cadeia da polimerase (PCR) e o seu subsequente sequenciamento consistem em uma ferramenta importante na caracterização de comunidades microbianas presentes em amostras ambientais. No entanto, apesar do crescente número de sequências de DNA de Archaea depositadas em bancos de dados, a especificidade e efetividade dos iniciadores de PCR descritos como universais e amplamente utilizados na descrição desse grupo ainda não está clara. Neste estudo foram comparados os perfis filogenéticos de comunidades de arqueias obtidos a partir amostras de DNA de sedimentos lacustres do Cerrado submetidas a ensaios de PCR empregando três pares de iniciadores específicos para Archaea, comumente utilizados neste tipo de estudo. Nossos resultados indicam que as comunidades de arqueias detectadas com cada par de iniciadores apresentaram grande variação filogenética, sugerindo que a escolha de iniciadores dirigidos ao gene de rRNA 16S tem efeito significativo no perfil da comunidade descrita, com diferenças tanto em relação aos táxons detectados, como nas estimativas de unidades taxonômicas operacionais (OTU).
RESUMO
The phylum Firmicutes comprises seven classes where most species are either aerobic or anaerobic endospore former. Inside Firmicutes, species allocated in the genus Bacillus and related genera are collectively named aerobic endospore-forming bacteria (AEFB), and the soil is their major reservoir. AEFB have great importance in health, agriculture, and biotechnology although the more studied species are Bacillus subtilis and the human pathogens Bacillus cereus and Bacillus anthracis. AEFB have great importance in health, agriculture, and biotechnology; although the knowledge about these organisms is based on few species, notably, Bacillus subtilis, Bacillus cereus, and Bacillus anthracis. In this work, we generated partial 16S rRNA gene sequences of both strands of 192 AEFB strains isolated from soils of Distrito Federal, Brazil (SDF strains). The resulting consensus sequences were used to obtain taxonomic assignment and establish the phylogenetic relationships among these strains. Through this approach, we could observe that classified SDF strains were distributed among genera Bacillus (169 strains; 88.02%), Paenibacillus (11; 5.73%), Lysinibacillus (6; 3.13%), Brevibacillus (4; 2.08%), Terribacillus (1; 0.52%), and Rummeliibacillus (1; 0.52%). Phylogenetic trees revealed these 192 SDF strains can be segregated into eight groups spanning families Bacillaceae and Paenibacillaceae belonging to the order Bacillales. To expand the knowledge about the diversity of these SDF strains, further studies regarding characterization with different methodologies are underway
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