Dataset Information

A Metagenomic and Amplicon Sequencing Combined Approach Reveals the Best Primers to Study Marine Aerobic Anoxygenic Phototrophs.

ABSTRACT: Studies based on protein-coding genes are essential to describe the diversity within bacterial functional groups. In the case of aerobic anoxygenic phototrophic (AAP) bacteria, the pufM gene has been established as the genetic marker for this particular functional group, although available primers are known to have amplification biases. We review here the existing primers for pufM gene amplification, design new ones, and evaluate their phylogenetic coverage. We then use samples from contrasting marine environments to evaluate their performance. By comparing the taxonomic composition of communities retrieved with metagenomics and with different amplicon approaches, we show that the commonly used PCR primers are biased towards the Gammaproteobacteria phylum and some Alphaproteobacteria clades. The metagenomic approach, as well as the use of other combinations of the existing and newly designed primers, show that these groups are in fact less abundant than previously observed, and that a great proportion of pufM sequences are affiliated to uncultured representatives, particularly in the open ocean. Altogether, the framework developed here becomes a better alternative for future studies based on the pufM gene and, additionally, serves as a reference for primer evaluation of other functional genes.

SUBMITTER: Gazulla CR

PROVIDER: S-EPMC10497671 | biostudies-literature | 2023 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

A Metagenomic and Amplicon Sequencing Combined Approach Reveals the Best Primers to Study Marine Aerobic Anoxygenic Phototrophs.

Gazulla Carlota R CR Cabello Ana María AM Sánchez Pablo P Gasol Josep M JM Sánchez Olga O Ferrera Isabel I

Microbial ecology 20230506 3

Studies based on protein-coding genes are essential to describe the diversity within bacterial functional groups. In the case of aerobic anoxygenic phototrophic (AAP) bacteria, the pufM gene has been established as the genetic marker for this particular functional group, although available primers are known to have amplification biases. We review here the existing primers for pufM gene amplification, design new ones, and evaluate their phylogenetic coverage. We then use samples from contrasting ...[more]

PMID: 37148309

Similar Datasets

Project description:BackgroundAerobic anoxygenic phototrophic (AAP) bacteria are heterotrophic bacteria that supply their metabolism with light energy harvested by bacteriochlorophyll-a-containing reaction centers. Despite their substantial contribution to bacterial biomass, microbial food webs, and carbon cycle, their phenology in freshwater lakes remains unknown. Hence, we investigated seasonal variations of AAP abundance and community composition biweekly across 3 years in a temperate, meso-oligotrophic freshwater lake.ResultsAAP bacteria displayed a clear seasonal trend with a spring maximum following the bloom of phytoplankton and a secondary maximum in autumn. As the AAP bacteria represent a highly diverse assemblage of species, we followed their seasonal succession using the amplicon sequencing of the pufM marker gene. To enhance the accuracy of the taxonomic assignment, we developed new pufM primers that generate longer amplicons and compiled the currently largest database of pufM genes, comprising 3633 reference sequences spanning all phyla known to contain AAP species. With this novel resource, we demonstrated that the majority of the species appeared during specific phases of the seasonal cycle, with less than 2% of AAP species detected during the whole year. AAP community presented an indigenous freshwater nature characterized by high resilience and heterogenic adaptations to varying conditions of the freshwater environment.ConclusionsOur findings highlight the substantial contribution of AAP bacteria to the carbon flow and ecological dynamics of lakes and unveil a recurrent and dynamic seasonal succession of the AAP community. By integrating this information with the indicator of primary production (Chlorophyll-a) and existing ecological models, we show that AAP bacteria play a pivotal role in the recycling of dissolved organic matter released during spring phytoplankton bloom. We suggest a potential role of AAP bacteria within the context of the PEG model and their consideration in further ecological models.

Project description:Anoxygenic phototrophic bacteria (APBs) occur in a wide range of aquatic habitats, from hot springs to freshwater lakes and intertidal microbial mats. Here, we report the discovery of a novel niche for APBs: endoliths within marine littoral carbonates. In a study of 40 locations around Isla de Mona, Puerto Rico, and Menorca, Spain, 16S rRNA high-throughput sequencing of endolithic community DNA revealed the presence of abundant phylotypes potentially belonging to well-known APB clades. An ad hoc phylogenetic classification of these sequences enabled us to refine the assignments more stringently. Even then, all locations contained such putative APBs, often reaching a significant proportion of all phototrophic sequences. In fact, in some 20% of samples, their contribution exceeded that of oxygenic phototrophs, previously regarded as the major type of endolithic microbe in carbonates. The communities contained representatives of APBs in the Chloroflexales, various proteobacterial groups, and Chlorobi The most abundant phylotypes varied with geography: on Isla de Mona, Roseiflexus and Chlorothrix-related phylotypes dominated, whereas those related to Erythrobacter were the most common in Menorca. The presence of active populations of APBs was corroborated through an analysis of photopigments: bacteriochlorophylls were detected in all samples, bacteriochlorophyll c and a being most abundant. We discuss the potential metabolism and geomicrobial roles of endolithic APBs. Phylogenetic inference suggests that APBs may be playing a role as photoheterotrophs, adding biogeochemical complexity to our understanding of such communities. Given the global extent of coastal carbonate platforms, they likely represent a very large and unexplored habitat for APBs.IMPORTANCE Endolithic microbial communities from carbonates, which have been explored for over 2 centuries in predominantly naturalistic studies, were thought to be primarily composed of eukaryotic algae and cyanobacteria. Our report represents a paradigm shift in this regard, at least for the marine environment, demonstrating the presence of ubiquitous and abundant populations of APBs in this habitat. It raises questions about the role of these organisms in the geological dynamics of coastal carbonates, including coral reefs.

Dataset Information

A Metagenomic and Amplicon Sequencing Combined Approach Reveals the Best Primers to Study Marine Aerobic Anoxygenic Phototrophs.

Publications

A Metagenomic and Amplicon Sequencing Combined Approach Reveals the Best Primers to Study Marine Aerobic Anoxygenic Phototrophs.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets