Introduction to Freshwater Algae - Freshwater Algae: Identification, Enumeration and Use as Bioindicators - page 12

Table 1.4 Molecular Identification of Algal Species in Aquatic Environmental Samples.

Environmental Samples Techniques References

Picoplankton in Lake Baikal Direct sequencing Semenova and

Kuznedelov (1998)

Flagellate nanoplankton SSU rRNA probes for Paraphysomonas (Chrysophyte) Caron et al . (1999)

Mixed diatom populations
and laboratory cultures

Large subunit rRNA probe for Pseudo-nitzschia (Diatom) Scholin et al . (1997)

Estuarine river samples Use of ITS-specific polymerase chain reaction (PCR)

assays for Pfiesteria (Dinoflagellate)

Litaker et al . (2003)

Laboratory biofilm samples:
blue-greens and unicellular
eukaryotes

Amplification of 16s RNA genes, with denaturing
gradient gel electrophoresis (DGGE)

Droppo et al . (2007)

Diazotrophic blue-green
algae within the Amazon
River freshwater plume

Quantitative PCR (QPCR) analysis of the nif Hgene
(encodes part of the nitrogenise enzyme)

Foster et al . (2007)

Arctic ecosystems:
Freshwater stamukhi lake
and inflow river

Sequencing of 18s rRNA genes to identify major
cryptophyte river populations, plus minor lake
populations of diatoms

Galand et al . (2008)

Lake and river flagellate
samples

Population heterogeneity in Spumella (Chrysophyte) -
SSU rRNA sequences

Pfandl et al . (2009)

Pond samples Population heterogeneity in Desmodesmus

(Chlorophyte) - ITS2 rRNA sequences

Vanormelingen et al .
(2009)

Stream N 2 -ixing
cyanobacteria

Assessment of nitrogenase expression using real-time
reverse transcriptase PCR

Stancheva et al. (2013)

SSU rRNA, small subunit ribosomal RNA; ITS, internal transcribed spacer.

algae. This gene encodes the iron-containing pro-
tein nitrogenase (the key enzyme involved in nitrogen
fixation) and provides a marker for nitrogen-fixing
(diazotrophic) algae in the freshwater environment.
The technique was used to demonstrate that the blue-
green algal symbiont Richelia (associated with the
diatom Hemiaulus hauckii ) was specifically linked to
the Amazon freshwater outflow (river plume) in the
Western Tropical North Atlantic (WTNA) ocean, and
that the H. hauckii-Richelia complex could be used
as a bioindicator for pockets of freshwater within the
WTNA ocean.

As with direct sequencing, this technique has
particular advantages with small unicellular algae,
where there are often relatively few morphological
features available for identification. Caron et al .
(1999) sequenced the small-subunit ribosomal
genes of four species of the colourless chrysophyte
genus Paraphysomonas , leading to the development
of oligonucleotide probes for Paraphysomonas
imperforata and Paraphysomonas bandaiensis .
Molecular probes have major potential for the
detection of nuisance algae, particularly those that
produce toxins. They have been used, for example,
to distinguish toxic from non-toxic diatom species
(Scholin et al ., 1997), where differentiation would
otherwise require the time-consuming application of
SEM and TEM. They have also been used for the
rapid identification of Pfiesteria piscicida , a poten-
tially toxic dinoflagellate that has been the cause
of extensive fish mortalities in coastal rivers of the

Taxon-specific DNA sequences The develop-
ment of taxon-specific oligonucleotide probes from
DNA sequence data, followed by in situ hybridisa-
tion, has considerable potential for the identification
and counting of algae in environmental samples.