Part C | 17.3
466 Part C Marine Genomics
consisting of several (although nonculturable) bacteria. A recent 16S rRNA gene sequence-based study in
the southeastern Atlantic region generated 133 bacterial
sequences from the sponges Hymeniacidon heliophila
and Polymastia janeirensis [17.62]. These two sponge
species appear to share several bacterial taxa affiliated
with Cyanobacteria and Proteobacteria. An analysis
of 254 archaeal 16S rRNA gene partial sequences
from the sponges Hymeniacidon heliophila, Polymastia
janeirensis, Paraleucilla magna, and Petromica citrina
suggested that Crenarchaeota is well represented in sequence databases [17.63]. Silva et al. provided the first
large-scale analysis of the taxonomic and metabolic diversity of the microbiome of the sponge A. brasiliensis.
These results demonstrated that a complex microbiome
exists within this sponge, which presents a particular metabolic profile [17.64]. It also indicates the A.
brasiliensis microbiome is unique, differing from the
microbiomes present in the water column surrounding
these sponges and those associated with other marine
organisms. Within the taxonomic signature of the A.
brasiliensis microbiome, an enrichment of Betaproteobacteria (Burkholderia) and Gammaproteobacteria
(Pseudomonas and Alteromonas) were detected, which
indicates the specificity of the species.
17.3.2 Functional Genes Related
to Natural Products
of Bacterial Symbionts in Sponges
Natural products isolated from sponges are a key source
of new biologically active compounds. However, the
development of these compounds into drugs has been
held back by the difficulties to achieve a sustainable
supply of these, often complex, molecules for preclinical and clinical development. Increasing evidence
implicates microbial symbionts as the source of many
of these biologically active compounds, but the vast
majority of the sponge microbial community remains
uncultured [17.65]. Metagenomics offers a biotechnological solution to this problem. Metagenomes of
sponge microbial communities have been shown to
contain genes and gene clusters typical for the biosynthesis of biologically active natural products. Heterologous expression has also led to the isolation
of secondary metabolism gene clusters from uncultured microbial symbionts. Combining a metagenomic approach with heterologous expression holds
much promise for the sustainable exploitation of the
chemical diversity present in the sponge microbial
community.
Polyketides are considered as an important class
of bacterial bioactive secondary metabolites. Efforts
have been focused on isolating PKS gene clusters from sponge-associated bacterial metagenomic libraries [17.66]. Piel et al. [17.67] successfully employed this approach and identified the putative onnamide PKS gene cluster from a marine sponge T. swinhoei metagenomic library. Onnamide A (Fig. 17.4(1)),
found in marine sponges of the genus Theonella, is
very similar in structure to pederin (Fig. 17.4(2)) from
Paederus beetles. A major difference is the addition
of a terminal arginine, potentially added by NRPS
domains, in the sponge compounds. The very clear
homology between pederin and onnamide genes was
used to identify the candidate onnamide gene cluster.
The trans-AT feature of this gene cluster was used in
the cloning strategy. Interestingly, PCR primers were
designed for the PKS genes involved in pederin biosynthesis to identify the closely related onnamide gene
cluster.
The dysidenin (Fig. 17.4(3)) group of nonribosomal peptide natural products was isolated from marine
sponges of the genera Dysidea and Lamellodysidea,
which harbor photosynthetic symbiotic cyanobacteria
of the genus Oscillatoria. Synthesis of these nonribosomal peptides was not successfully determined [17.68].
The identification of the barbamide (Fig. 17.4(4))
biosynthetic gene cluster from L. majuscula facilitated
identification of candidate dysidenin biosynthetic genes
within marine sponges [17.69]. This family of compounds contains a very rare chemical motif, a multiply
chlorinated leucine residue. The PCR primers were designed for this chemical motif, targeting the halogenase
gene. This feature was used to isolate candidate halogenases from marine sponges containing the dysidenin
family of compounds.
Fisch et al. [17.70] found that general KS primers
amplify too many diverse PKS gene sequences, making it very difficult to identify the correct gene cluster.
Hence, they applied a nested PCR strategy to amplify the specific gene cluster. First, the degenerate
KS primers were used to amplify the KS region followed by the application of a trans-AT specific PCR
primer pair. The primers were designed such that one
primer was specific to 84% of all trans-AT KS, while
the second primer was specific for certain chemical motifs in the resulting polyketide product. Using
this strategy, probable gene clusters were identified
for the pederin relatives mycalamide (Fig. 17.4(5))
and psymberin (Fig. 17.4(6)) from marine sponge
metagenomes.
466 Part C Marine Genomics
consisting of several (although nonculturable) bacteria. A recent 16S rRNA gene sequence-based study in
the southeastern Atlantic region generated 133 bacterial
sequences from the sponges Hymeniacidon heliophila
and Polymastia janeirensis [17.62]. These two sponge
species appear to share several bacterial taxa affiliated
with Cyanobacteria and Proteobacteria. An analysis
of 254 archaeal 16S rRNA gene partial sequences
from the sponges Hymeniacidon heliophila, Polymastia
janeirensis, Paraleucilla magna, and Petromica citrina
suggested that Crenarchaeota is well represented in sequence databases [17.63]. Silva et al. provided the first
large-scale analysis of the taxonomic and metabolic diversity of the microbiome of the sponge A. brasiliensis.
These results demonstrated that a complex microbiome
exists within this sponge, which presents a particular metabolic profile [17.64]. It also indicates the A.
brasiliensis microbiome is unique, differing from the
microbiomes present in the water column surrounding
these sponges and those associated with other marine
organisms. Within the taxonomic signature of the A.
brasiliensis microbiome, an enrichment of Betaproteobacteria (Burkholderia) and Gammaproteobacteria
(Pseudomonas and Alteromonas) were detected, which
indicates the specificity of the species.
17.3.2 Functional Genes Related
to Natural Products
of Bacterial Symbionts in Sponges
Natural products isolated from sponges are a key source
of new biologically active compounds. However, the
development of these compounds into drugs has been
held back by the difficulties to achieve a sustainable
supply of these, often complex, molecules for preclinical and clinical development. Increasing evidence
implicates microbial symbionts as the source of many
of these biologically active compounds, but the vast
majority of the sponge microbial community remains
uncultured [17.65]. Metagenomics offers a biotechnological solution to this problem. Metagenomes of
sponge microbial communities have been shown to
contain genes and gene clusters typical for the biosynthesis of biologically active natural products. Heterologous expression has also led to the isolation
of secondary metabolism gene clusters from uncultured microbial symbionts. Combining a metagenomic approach with heterologous expression holds
much promise for the sustainable exploitation of the
chemical diversity present in the sponge microbial
community.
Polyketides are considered as an important class
of bacterial bioactive secondary metabolites. Efforts
have been focused on isolating PKS gene clusters from sponge-associated bacterial metagenomic libraries [17.66]. Piel et al. [17.67] successfully employed this approach and identified the putative onnamide PKS gene cluster from a marine sponge T. swinhoei metagenomic library. Onnamide A (Fig. 17.4(1)),
found in marine sponges of the genus Theonella, is
very similar in structure to pederin (Fig. 17.4(2)) from
Paederus beetles. A major difference is the addition
of a terminal arginine, potentially added by NRPS
domains, in the sponge compounds. The very clear
homology between pederin and onnamide genes was
used to identify the candidate onnamide gene cluster.
The trans-AT feature of this gene cluster was used in
the cloning strategy. Interestingly, PCR primers were
designed for the PKS genes involved in pederin biosynthesis to identify the closely related onnamide gene
cluster.
The dysidenin (Fig. 17.4(3)) group of nonribosomal peptide natural products was isolated from marine
sponges of the genera Dysidea and Lamellodysidea,
which harbor photosynthetic symbiotic cyanobacteria
of the genus Oscillatoria. Synthesis of these nonribosomal peptides was not successfully determined [17.68].
The identification of the barbamide (Fig. 17.4(4))
biosynthetic gene cluster from L. majuscula facilitated
identification of candidate dysidenin biosynthetic genes
within marine sponges [17.69]. This family of compounds contains a very rare chemical motif, a multiply
chlorinated leucine residue. The PCR primers were designed for this chemical motif, targeting the halogenase
gene. This feature was used to isolate candidate halogenases from marine sponges containing the dysidenin
family of compounds.
Fisch et al. [17.70] found that general KS primers
amplify too many diverse PKS gene sequences, making it very difficult to identify the correct gene cluster.
Hence, they applied a nested PCR strategy to amplify the specific gene cluster. First, the degenerate
KS primers were used to amplify the KS region followed by the application of a trans-AT specific PCR
primer pair. The primers were designed such that one
primer was specific to 84% of all trans-AT KS, while
the second primer was specific for certain chemical motifs in the resulting polyketide product. Using
this strategy, probable gene clusters were identified
for the pederin relatives mycalamide (Fig. 17.4(5))
and psymberin (Fig. 17.4(6)) from marine sponge
metagenomes.