Phylogenetic and functional gene analysis of the bacterial and archaeal communities associated with the surface microlayer of an estuary

Abstract

The surface microlayer (SML) is the thin biogenic film found at the surface of a water body. The SML is poorly understood but has been shown to be important in biogeochemical cycling and sea-air gas exchange. We sampled the SML of the Blyth estuary at two sites (salinities 21 and 31 psu) using 47 mm polycarbonate membranes. DNA was extracted from the SML and corresponding subsurface water (0.4 m depth) and microbial (bacteria and archaea) community analysis was performed using denaturing gradient gel electrophoresis of 16S rRNA gene PCR amplicons. The diversity of bacterial functional genes that encode enzyme subunits for methane monooxygenase (pmoA and mmoX) and carbon monoxide dehydrogenase (coxL) was assessed using PCR, clone library construction and restriction fragment length polymorphism (RFLP) analysis. Methanotroph genes were present only in low copy numbers and pmoA was detected only in subsurface samples. Diversity of mmoX genes was low and most of the clone sequences detected were similar to those of mmoX from Methylomonas spp. Interestingly, some sequences detected in the SML were different from those detected in the subsurface. RFLP analysis of coxL clone libraries indicated a high diversity of carbon monoxide (CO)-utilizing bacteria in the estuary. The habitats of the closely related coxL sequences suggest that CO-utilizing bacteria in the estuary are recruited from both marine and freshwater/terrestrial inputs. In contrast, methanotroph recruitment appears to occur solely from freshwater input into the estuary.

The sea-surface microlayer is a gelatinous biofilm

“…More recent evidence published in The ISME Journal supports the observations made in the North Sea by Franklin et al (2005) and shows distinct bacterioneuston communities are also present in estuarine surface microlayers (Cunliffe et al, 2008). Denaturing gradient gel electrophoresis comparison of bacterial and archaeal 16S rRNA genes from surface microlayer and subsurface water samples at two sites along the estuary showed that bacteria form distinct surface microlayer-specific community structures.…”

Correlating carbon monoxide oxidation with cox genes in the abundant Marine Roseobacter Clade

“…King (2003) made a wider PCR-based survey of coxL genes across strains from two phyla (Proteobacteria and Actinobacteria) and also showed monophyletic groupings similar to 16S rRNA gene phylogeny. The concordance of coxL gene and 16S rRNA gene phylogeny has significant practical implications if coxL genes are used as molecular markers instead of 16S rRNA genes to study CO oxidiser assemblage structure in situ King, 2004, 2005;Cunliffe et al, 2008).…”

Eukarya 18S rRNA gene diversity in the sea surface microlayer: implications for the structure of the neustonic microbial loop

“…The bacterial community present in the surface microlayer is known as the bacterioneuston and has a different community structure compared with that of subsurface water below (Cunliffe et al, 2008, 2009a, Franklin et al, 2005. Monitoring of the bacterioneuston community structure during a fjord mesocosm experiment, using Bacteria 16S rRNA gene terminal restriction fragment length polymorphism and denaturing gradient gel electrophoresis (DGGE), showed distinct and consistent differences between the bacterioneuston and the bacterioplankton communities (Cunliffe et al, 2009c).…”

“…Community samples were collected from the surface microlayer using a metal mesh screen (Garrett Screen; 16 mesh stainless steel screen, size 275 Â 275 mm, sampling depth 0 to 400 mm) and subsurface water (sampling depth 0.75 m). For each sample, 250 ml of water was filtered using a Sterivex-GS filter unit (pore size 0.2 mm; Millipore, Watford, UK) and DNA was extracted from the filter in a sucrose buffer using lysozyme, proteinase K, sodium dodecyl sulphate and phenol-chloroform (Cunliffe et al, 2008). The re-suspended DNA was diluted in molecular grade water to a concentration of 30 ng ml À1 and stored at À20 1C.…”