Cyanobacterial Diversity in Natural and Artificial Microbial Mats of Lake Fryxell (McMurdo Dry Valleys, Antarctica): a Morphological and Molecular Approach

Abstract

Currently, there is no consensus concerning the geographic distribution and extent of endemism in Antarctic cyanobacteria. In this paper we describe the phenotypic and genotypic diversity of cyanobacteria in a field microbial mat sample from Lake Fryxell and in an artificial cold-adapted sample cultured in a benthic gradient chamber (BGC) by using an inoculum from the same mat. Light microscopy and molecular tools, including 16S rRNA gene clone libraries, denaturing gradient gel electrophoresis, and sequencing, were used. For the first time in the study of cyanobacterial diversity of environmental samples, internal transcribed spacer (ITS) sequences were retrieved and analyzed to complement the information obtained from the 16S rRNA gene. Microscopy allowed eight morphotypes to be identified, only one of which is likely to be an Antarctic endemic morphotype. Molecular analysis, however, revealed an entirely different pattern. A much higher number of phylotypes (15 phylotypes) was found, but no sequences from Nodularia and Hydrocoryne, as observed by microscopy, were retrieved. The 16S rRNA gene sequences determined in this study were distributed in 11 phylogenetic lineages, 3 of which were exclusively Antarctic and 2 of which were novel. Collectively, these Antarctic sequences together with all the other polar sequences were distributed in 22 lineages, 9 of which were exclusively Antarctic, including the 2 novel lineages observed in this study. The cultured BGC mat had lower diversity than the field mat. However, the two samples shared three morphotypes and three phylotypes. Moreover, the BGC mat allowed enrichment of one additional phylotype. ITS sequence analysis revealed a complex signal that was difficult to interpret. Finally, this study provided evidence of molecular diversity of cyanobacteria in Antarctica that is much greater than the diversity currently known based on traditional microscopic analysis. Furthermore, Antarctic endemic species were more abundant than was estimated on the basis of morphological features. Decisive arguments concerning the global geographic distribution of cyanobacteria should therefore incorporate data obtained with the molecular tools described here.

Hidden levels of phylodiversity in Antarctic green algae: further evidence for the existence of glacial refugia

“…Micro-organisms form an interesting test case in the light of the refugium hypothesis in that they are expected, within the limits of their environmental tolerance, to show ubiquitous distribution patterns as a result of their virtually unlimited dispersal capacity (but see ongoing discussion on microbial biogeography in, for example, Martiny et al 2006). Molecular studies of cyanobacteria, using 16S rRNA gene sequencing, suggest that most isolates have a long association with the Antarctic environment (Taton et al 2003(Taton et al , 2006. Likewise, morphological studies on diatoms suggest that in some areas at least 40 per cent of the species are Antarctic endemics (Schmidt et al 1990;Sabbe et al 2004;Spaulding et al in press).…”

Late Holocene changes in cyanobacterial community structure in maritime Antarctic lakes

“…The cyanobacterial sequences were grouped into operational taxonomic units (OTUs) with the software DOTUR, using the average-neighbor method (Schloss and Handelsman 2005). We define the OTUs as groups of sequences that exhibit more than 97.5 % similarity with each other, using E. coli positions 405-780, except in some cases where the sequences were slightly shorter (Taton et al 2003).…”

“…The PCR program was similar to that reported in Fernandez-Carazo et al (2011) with the following modifications: 1.2 units of Super Taq Plus (HT Biotechnology, Cambridge, UK) and 3-4 lL of DNA were used per reaction. All the primers are listed in Taton et al (2003) except 16S369F (GGCAG-CAGTGGGGAATTTTC), which was designed and tested to improve the specificity of the cyanobacterial amplification. The T and C at positions 366 and 369 (Escherichia coli positions), respectively, are specific for cyanobacteria and different in many other bacteria.…”

“…DGGE and phylogenetic analysis DGGE was run in 6.5 % acrylamide gels with a (40-65 %) denaturing gradient using a Dcode gene system (Bio-Rad), as described in Taton et al (2003). DGGE gels were performed two times for most of the samples to assess reproducibility and detect potential contamination.…”

“…1). DGGE allows relatively rapid and inexpensive inventory of the most abundant taxa in microbial communities and was successfully used to study present-day cyanobacterial communities in Antarctic lakes (Taton et al 2003;Verleyen et al 2010). It separates PCR products of equal length, but different sequence (and thus melting temperature), on an acrylamide gel containing a gradient of chemical denaturants (Muyzer 1999).…”