Comparison of cultivation‐dependent and molecular methods for studying the diversity of anoxygenic purple phototrophs in sediments of an eutrophic brackish lagoon

Abstract

Phototrophic anoxygenic purple bacteria play a key role in many aquatic ecosystems by oxidizing sulfur compounds and low-molecular-weight organic compounds using light as energy source. In this study, molecular methods based upon pufM gene (photosynthetic unit forming gene) were compared with culture-dependent methods to investigate anoxygenic purple phototrophic communities in sediments of an eutrophic brackish lagoon. Thirteen strains, belonging to eight different genera of purple phototrophic bacteria were isolated with a large dominance of the metabolically versatile purple non-sulfur bacteria (eight strains), some purple sulfur bacteria (three strains) and two strains belonging to the Roseobacter clade (aerobic phototrophs). The pufM genes amplified from the isolated strains were not detected by the molecular methods [terminal-restriction fragment length polymorphism (T-RFLP)] applied on in situ communities. An environmental clone library of the pufM gene was thus constructed from sediment samples. The results showed that most of the clones probably corresponded to aerobic phototrophic bacteria. Our results demonstrate that the culture-dependent techniques remain the best experimental approach for determining the diversity of phototrophic purple non-sulfur bacteria whereas the molecular approach clearly illustrated the abundance of organisms related to the Roseobacter clade in these eutrophic sediments.

Anoxygenic phototrophic bacterial diversity within wastewater stabilization plant during ‘red water’ phenomenon

“…The pufM gene was amplified using the primers puf557F and puf750R (Karr et al 2003;Ranchou-Peyruse et al 2006). PCR amplification mixtures contain 5 ll of 109 PCR buffer without MgCl 2 , 3 ll of 25 mM MgCl 2 , 1 ll of each deoxynucleoside triphosphate at 10 mM, 0.5 ll of each forward and reverse primer at 125 lg/ml, 1 ll of DNA template (ca.…”

“…Their higher growth rates and efficiency in organic carbon utilization over strict heterotrophs are likely to make them dynamic and significant contributors to the organic carbon production and cycling within the stabilization ponds (Gasol et al 2008) and it has been shown that these bacteria play a key role in a diverse range of oxic ecosystems (Buchan et al 2005;Oz et al 2005). Among the aerobic anoxygenic phototrophic bacterial group, some representatives may be able to grow under anaerobic conditions (Ranchou-Peyruse et al 2006).…”

“…The cultivationdependent approaches also are critical and may lead to significant gaps in microbial diversity studies (Donachie et al 2007). Different molecular methods based on the pufM gene encoding the M subunit of the photosynthetic reaction center have been used to explore the phototrophic bacterial diversity (Jiao et al 2007) such as gradient gel electrophoresis (Lehours et al 2010), terminal restriction fragment length polymorphism (Ranchou-Peyruse et al 2006), and shotgun sequencing (Yutin et al 2007). Although these DNA-based approaches have given insight into the diversity of anoxygenic phototrophic bacteria (APB), analysis of relevant functional communities is still a challenge.…”

Purple anoxygenic phototrophic bacteria distribution in Tunisian wastewater stabilisation plant exhibiting red water phenomenon

“…L, M and H polypeptides function to bind bacteriochlorophyll and carotenoids such as spirilloxanthin, lycopenal and rhodopinal as well as quinones. The pufM gene has been widely used to investigate Photosynthetic Anoxygenic Purple Bacteria (PAPB) diversity in natural environments (Schwalbach and Fuhrman 2005;Yutin et al, 2005;Okubo et al, 2006, Ranchou-Peyruse et al, 2006. PAPB grow correctly in natural and polluted environments by forming colored microbial mats and by developing massive growth (blooms) (Okubo et al, 2005).…”

Diversity of cultivated and metabolically active aerobic anoxygenic phototrophic bacteria along an oligotrophic gradient in the Mediterranean Sea

“…Molecular analyses based on the pufM gene encoding the M-subunit of the photosynthetic reaction center have explored AAP bacterial diversity using the polymerase chain reaction (PCR) (Béjà et al, 2002;Yutin et al, 2005;, denaturing/temperature gradient gel electrophoresis (DGGE/TGGE) (Yutin et al, 2008;Lehours et al, 2010), terminal restriction fragment length polymorphism (T-RFLP) (Ranchou-Peyruse et al, 2006) and metagenomic analysis (Yutin et al, 2007). Although these DNA-based approaches have given insight into the diversity of AAP bacteria, analysis of relevant functional communities is still a challenge, as DNA could be stable in resting cells and even dead cells and may also be present as extracellular DNA.…”