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(Pace et al. 1986 ). The molecular tool box is complemented by whole-cell in situ
hybridization with rRNA-targeted fluorescently labeled oligonucleotide probes
(fluorescent in situ hybridization, FISH) for PCR-independent identification and
enumeration (Amann et al. 1995 ), as well as various PCR-based fingerprinting
methods (e.g., DGGE, denaturing gradient gel electrophoresis; T-RFLP, terminal
restriction fragment length polymorphism analysis) (Liu et al. 1997 ; Muyzer et al.
1993 ). Essentially, sequence-based analysis (i.e., cloning/sequencing) allows to
characterize a microbial community at the level of individual phylotypes; however,
this approach has been limited in the past by considering insufficient numbers of
sequences (and replicate samples) causing under-sampling bias (Green and
Bohannan 2006 ; Hughes et al. 2001 ). Fingerprinting methods reflect the diversity
of the whole PCR amplicon pool, and thus, are powerful tools to compare larger
numbers of sample; however, they have limited phylogenetic resolution (Marsh
2005 ). Therefore, this review focuses mainly on studies that characterized the
composition of microbial communities associated with macroalgae using sequence
information (Table 10.1 ).
A number of cultivation studies complemented with molecular cultivation-
independent analysis found only little overlap between the identity of isolates and
predominant sequences retrieved from the algal host (Bengtsson et al. 2010 ; Tujula
et al. 2010 ). For example, the strains of the
-proteobacterial genus Pseudoal-
teromonas have been frequently isolated from Ulvacean algal hosts (Dobretsov
and Qian 2002 ; Egan et al. 2000 , 2001 ; Patel et al. 2003 ), but were not detected by
catalytically amplified reporter deposition (CARD)-FISH on samples of Ulva
australis (Tujula et al. 2010 ).
Sequence-based microbial community composition studies (Table 10.1 )included
a range of macroalgal species representing Rhodophyta ( n
g
¼
5), Phaeophyceae
( n
6), and two functionally defined algae from a coral
reef, crustose calcifying algae, and turf algae. The majority of studies analyzed up
to 300 sequences by conventional Sanger sequencing of clones or DGGE bands, one
study analyzed ~900 clones per individual ( Ulva australis )(Burkeetal. 2011b ), and
one study used pyrosequencing with up to 60,000 sequences from one individual
sample (Barott et al. 2011 ). High-throughput sequencing methods, such as 16S rRNA
gene tag pyrosequencing (Sogin et al. 2006 ), facilitate to retrieve several thousands
(to hundreds of thousands) short sequences (100-400 bp).
Among the microbial phyla encountered sequences representing
¼
4), Chlorophyta ( n
¼
-Proteobacteria
weremost numerous across all studies andmacroalgal species (Table 10.1 ). Frequently
detected were also bacteria associated with the phyla Bacteroidetes, Planctomycetes,
Verrucomicrobia, Cyanobacteria, and
a
-Proteobacteria, and on a few algal species
Actinobacteria, Chloroflexi, and Firmicutes. Up to 22 different phyla were encoun-
tered in a pyrosequencing-based diversity study of four different algae from a coral
reef ecosystem (Barott et al. 2011 ), indicating the high diversity of microbial
communities on macroalgae. Planctomycetal sequences were detected on a relatively
large number of algae (Table. 10.1 ), albeit mismatches of the canonical 27F-primer
used to most planctomycetal 16S rRNA genes typically result in underrep-
resentation of Planctomycetes in clone libraries; when using a specific FISH probe,
d
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