Secondary Metabolism in the Gill Microbiota of Shipworms (Teredinidae) as Revealed by Comparison of Metagenomes and Nearly Complete Symbiont Genomes

ABSTRACT Shipworms play critical roles in recycling wood in the sea. Symbiotic bacteria supply enzymes that the organisms need for nutrition and wood degradation. Some of these bacteria have been grown in pure culture and have the capacity to make many secondary metabolites. However, little is known about whether such secondary metabolite pathways are represented in the symbiont communities within their hosts. In addition, little has been reported about the patterns of host-symbiont co-occurrence. Here, we collected shipworms from the United States, the Philippines, and Brazil and cultivated symbiotic bacteria from their gills. We analyzed sequences from 22 shipworm gill metagenomes from seven shipworm species and from 23 cultivated symbiont isolates. Using (meta)genome sequencing, we demonstrate that the cultivated isolates represent all the major bacterial symbiont species and strains in shipworm gills. We show that the bacterial symbionts are distributed among shipworm hosts in consistent, predictable patterns. The symbiotic bacteria harbor many gene cluster families (GCFs) for biosynthesis of bioactive secondary metabolites, only <5% of which match previously described biosynthetic pathways. Because we were able to cultivate the symbionts and to sequence their genomes, we can definitively enumerate the biosynthetic pathways in these symbiont communities, showing that ∼150 of ∼200 total biosynthetic gene clusters (BGCs) present in the animal gill metagenomes are represented in our culture collection. Shipworm symbionts occur in suites that differ predictably across a wide taxonomic and geographic range of host species and collectively constitute an immense resource for the discovery of new biosynthetic pathways corresponding to bioactive secondary metabolites. IMPORTANCE We define a system in which the major symbionts that are important to host biology and to the production of secondary metabolites can be cultivated. We show that symbiotic bacteria that are critical to host nutrition and lifestyle also have an immense capacity to produce a multitude of diverse and likely novel bioactive secondary metabolites that could lead to the discovery of drugs and that these pathways are found within shipworm gills. We propose that, by shaping associated microbial communities within the host, the compounds support the ability of shipworms to degrade wood in marine environments. Because these symbionts can be cultivated and genetically manipulated, they provide a powerful model for understanding how secondary metabolism impacts microbial symbiosis.

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Secondary metabolism in the gill microbiota of shipworms (Teredinidae) as revealed by comparison of metagenomes and nearly complete symbiont genomes

10.1101/826933 ◽

2019 ◽

Author(s):

Marvin A. Altamia ◽

Zhenjian Lin ◽

Amaro E. Trindade-Silva ◽

Iris Diana Uy ◽

J. Reuben Shipway ◽

...

Keyword(s):

Secondary Metabolites ◽

Secondary Metabolism ◽

Gene Clusters ◽

The United States ◽

Biosynthetic Gene Cluster ◽

The Philippines ◽

Symbiotic Bacteria ◽

Biosynthetic Pathways ◽

Biosynthetic Gene ◽

Bioactive Secondary Metabolites

AbstractShipworms play critical roles in recycling wood in the sea. Symbiotic bacteria supply enzymes that the organisms need for nutrition and wood degradation. Some of these bacteria have been grown in pure culture and have the capacity to make many secondary metabolites. However, little is known about whether such secondary metabolite pathways are represented in the symbiont communities within their hosts. In addition, little has been reported about the patterns of host-symbiont co-occurrence. Here, we collected shipworms from the United States, the Philippines, and Brazil, and cultivated symbiotic bacteria from their gills. We analyzed sequences from 22 shipworm gill metagenomes from seven shipworm species and from 23 cultivated symbiont isolates. Using (meta)genome sequencing, we demonstrate that the cultivated isolates represent all the major bacterial symbiont species and strains in shipworm gills. We show that the bacterial symbionts are distributed among shipworm hosts in consistent, predictable patterns. The symbiotic bacteria encode many biosynthetic gene cluster families (GCFs) for bioactive secondary metabolites, only <5% of which match previously described biosynthetic pathways. Because we were able to cultivate the symbionts, and sequence their genomes, we can definitively enumerate the biosynthetic pathways in these symbiont communities, showing that ∼150 out of ∼200 total biosynthetic gene clusters (BGCs) present in the animal gill metagenomes are represented in our culture collection. Shipworm symbionts occur in suites that differ predictably across a wide taxonomic and geographic range of host species, and collectively constitute an immense resource for the discovery of new biosynthetic pathways to bioactive secondary metabolites.ImportanceWe define a system in which the major symbionts that are important to host biology and to the production of secondary metabolites can be cultivated. We show that symbiotic bacteria that are critical to host nutrition and lifestyle also have an immense capacity to produce a multitude of diverse and likely novel bioactive secondary metabolites that could lead to the discovery of drugs, and that these pathways are found within shipworm gills. We propose that, by shaping associated microbial communities within the host, the compounds support the ability of shipworms to degrade wood in marine environments. Because these symbionts can be cultivated and genetically manipulated, they provide a powerful model for understanding how secondary metabolism impacts microbial symbiosis.

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Natural Products in Polyclad Flatworms

Marine Drugs ◽

10.3390/md19020047 ◽

2021 ◽

Vol 19 (2) ◽

pp. 47

Author(s):

Justin M. McNab ◽

Jorge Rodríguez ◽

Peter Karuso ◽

Jane E. Williamson

Keyword(s):

Natural Products ◽

Secondary Metabolites ◽

Marine Invertebrates ◽

Symbiotic Bacteria ◽

Chemical Defences ◽

Number Of Species ◽

Bioactive Secondary Metabolites ◽

Potential Sources ◽

Polyclad Flatworms

Marine invertebrates are promising sources of novel bioactive secondary metabolites, and organisms like sponges, ascidians and nudibranchs are characterised by possessing potent defensive chemicals. Animals that possess chemical defences often advertise this fact with aposematic colouration that potential predators learn to avoid. One seemingly defenceless group that can present bright colouration patterns are flatworms of the order Polycladida. Although members of this group have typically been overlooked due to their solitary and benthic nature, recent studies have isolated the neurotoxin tetrodotoxin from these mesopredators. This review considers the potential of polyclads as potential sources of natural products and reviews what is known of the activity of the molecules found in these animals. Considering the ecology and diversity of polyclads, only a small number of species from both suborders of Polycladida, Acotylea and Cotylea have been investigated for natural products. As such, confirming assumptions as to which species are in any sense toxic or if the compounds they use are biosynthesised, accumulated from food or the product of symbiotic bacteria is difficult. However, further research into the group is suggested as these animals often display aposematic colouration and are known to prey on invertebrates rich in bioactive secondary metabolites.

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IMG-ABC: A Knowledge Base To Fuel Discovery of Biosynthetic Gene Clusters and Novel Secondary Metabolites

mBio ◽

10.1128/mbio.00932-15 ◽

2015 ◽

Vol 6 (4) ◽

Cited By ~ 66

Author(s):

Michalis Hadjithomas ◽

I-Min Amy Chen ◽

Ken Chu ◽

Anna Ratner ◽

Krishna Palaniappan ◽

...

Keyword(s):

Secondary Metabolites ◽

Secondary Metabolism ◽

Genomic Data ◽

Gene Clusters ◽

Metagenomic Data ◽

Integrated Analysis ◽

Analysis Tool ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Analysis Tools

ABSTRACTIn the discovery of secondary metabolites, analysis of sequence data is a promising exploration path that remains largely underutilized due to the lack of computational platforms that enable such a systematic approach on a large scale. In this work, we present IMG-ABC (https://img.jgi.doe.gov/abc), an atlas of biosynthetic gene clusters within the Integrated Microbial Genomes (IMG) system, which is aimed at harnessing the power of “big” genomic data for discovering small molecules. IMG-ABC relies on IMG's comprehensive integrated structural and functional genomic data for the analysis of biosynthetic gene clusters (BCs) and associated secondary metabolites (SMs). SMs and BCs serve as the two main classes of objects in IMG-ABC, each with a rich collection of attributes. A unique feature of IMG-ABC is the incorporation of both experimentally validated and computationally predicted BCs in genomes as well as metagenomes, thus identifying BCs in uncultured populations and rare taxa. We demonstrate the strength of IMG-ABC's focused integrated analysis tools in enabling the exploration of microbial secondary metabolism on a global scale, through the discovery of phenazine-producing clusters for the first time inAlphaproteobacteria. IMG-ABC strives to fill the long-existent void of resources for computational exploration of the secondary metabolism universe; its underlying scalable framework enables traversal of uncovered phylogenetic and chemical structure space, serving as a doorway to a new era in the discovery of novel molecules.IMPORTANCEIMG-ABC is the largest publicly available database of predicted and experimental biosynthetic gene clusters and the secondary metabolites they produce. The system also includes powerful search and analysis tools that are integrated with IMG's extensive genomic/metagenomic data and analysis tool kits. As new research on biosynthetic gene clusters and secondary metabolites is published and more genomes are sequenced, IMG-ABC will continue to expand, with the goal of becoming an essential component of any bioinformatic exploration of the secondary metabolism world.

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Characterizing the Pathogenic, Genomic, and Chemical Traits ofAspergillus fischeri, a Close Relative of the Major Human Fungal PathogenAspergillus fumigatus

mSphere ◽

10.1128/msphere.00018-19 ◽

2019 ◽

Vol 4 (1) ◽

Cited By ~ 11

Author(s):

Matthew E. Mead ◽

Sonja L. Knowles ◽

Huzefa A. Raja ◽

Sarah R. Beattie ◽

Caitlin H. Kowalski ◽

...

Keyword(s):

Secondary Metabolites ◽

Aspergillus Fumigatus ◽

Secondary Metabolism ◽

Human Disease ◽

Chemical Characterization ◽

Gene Clusters ◽

Invasive Disease ◽

Close Relative ◽

Low Oxygen ◽

Content Type

ABSTRACTAspergillus fischeriis closely related toAspergillus fumigatus, the major cause of invasive mold infections. Even thoughA. fischeriis commonly found in diverse environments, including hospitals, it rarely causes invasive disease. WhyA. fischericauses less human disease thanA. fumigatusis unclear. A comparison ofA. fischeriandA. fumigatusfor pathogenic, genomic, and secondary metabolic traits revealed multiple differences in pathogenesis-related phenotypes. We observed thatA. fischeriNRRL 181 is less virulent thanA. fumigatusstrain CEA10 in multiple animal models of disease, grows slower in low-oxygen environments, and is more sensitive to oxidative stress. Strikingly, the observed differences for some traits are of the same order of magnitude as those previously reported betweenA. fumigatusstrains. In contrast, similar to what has previously been reported, the two species exhibit high genomic similarity; ∼90% of theA. fumigatusproteome is conserved inA. fischeri, including 48/49 genes known to be involved inA. fumigatusvirulence. However, only 10/33A. fumigatusbiosynthetic gene clusters (BGCs) likely involved in secondary metabolite production are conserved inA. fischeriand only 13/48A. fischeriBGCs are conserved inA. fumigatus. Detailed chemical characterization ofA. fischericultures grown on multiple substrates identified multiple secondary metabolites, including two new compounds and one never before isolated as a natural product. Additionally, anA. fischerideletion mutant oflaeA, a master regulator of secondary metabolism, produced fewer secondary metabolites and in lower quantities, suggesting that regulation of secondary metabolism is at least partially conserved. These results suggest that the nonpathogenicA. fischeripossesses many of the genes important forA. fumigatuspathogenicity but is divergent with respect to its ability to thrive under host-relevant conditions and its secondary metabolism.IMPORTANCEAspergillus fumigatusis the primary cause of aspergillosis, a devastating ensemble of diseases associated with severe morbidity and mortality worldwide.A. fischeriis a close relative ofA. fumigatusbut is not generally observed to cause human disease. To gain insights into the underlying causes of this remarkable difference in pathogenicity, we compared two representative strains (one from each species) for a range of pathogenesis-relevant biological and chemical characteristics. We found that disease progression in multipleA. fischerimouse models was slower and caused less mortality thanA. fumigatus. Remarkably, the observed differences betweenA. fischeriandA. fumigatusstrains examined here closely resembled those previously described for two commonly studiedA. fumigatusstrains, AF293 and CEA10.A. fischeriandA. fumigatusexhibited different growth profiles when placed in a range of stress-inducing conditions encountered during infection, such as low levels of oxygen and the presence of chemicals that induce the production of reactive oxygen species. We also found that the vast majority ofA. fumigatusgenes known to be involved in virulence are conserved inA. fischeri, whereas the two species differ significantly in their secondary metabolic pathways. These similarities and differences that we report here are the first step toward understanding the evolutionary origin of a major fungal pathogen.

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Unlocking Fungal Cryptic Natural Products

Natural Product Communications ◽

10.1177/1934578x0900401113 ◽

2009 ◽

Vol 4 (11) ◽

pp. 1934578X0900401 ◽

Cited By ~ 8

Author(s):

Yi-Ming Chiang ◽

Kuan-Han Lee ◽

James F. Sanchez ◽

Nancy P. Keller ◽

Clay C. C. Wang

Keyword(s):

Natural Products ◽

Secondary Metabolites ◽

Secondary Metabolism ◽

Analytical Methods ◽

State Of The Art ◽

Molecular Genetic ◽

Biosynthetic Pathways ◽

Fungal Metabolites ◽

Potential Sources ◽

Fungal Genomes

Recent published sequencing of fungal genomes has revealed that these microorganisms have a surprisingly large number of secondary metabolite pathways that can serve as potential sources for new and useful natural products. Most of the secondary metabolites and their biosynthesis pathways are currently unknown, possibly because they are produced in very small amounts and are thus difficult to detect or are produced only under specific conditions. Elucidating these fungal metabolites will require new molecular genetic tools, better understanding of the regulation of secondary metabolism, and state of the art analytical methods. This review describes recent strategies to mine the cryptic natural products and their biosynthetic pathways in fungi.

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High Potential for Secondary Metabolite Production of Paracoccus marcusii CP157, Isolated From the Crustacean Cancer pagurus

Frontiers in Microbiology ◽

10.3389/fmicb.2021.688754 ◽

2021 ◽

Vol 12 ◽

Author(s):

Janina Leinberger ◽

Jonas Holste ◽

Boyke Bunk ◽

Heike M. Freese ◽

Cathrin Spröer ◽

...

Keyword(s):

Secondary Metabolites ◽

Elevated Temperatures ◽

Proteolytic Enzymes ◽

Natural Habitat ◽

Antibacterial Properties ◽

Gene Clusters ◽

Natural Environments ◽

Phylogenetic Distance ◽

Cancer Pagurus ◽

Bioactive Secondary Metabolites

Secondary metabolites are key components in microbial ecology by mediating interactions between bacteria and their environment, neighboring species or host organisms. Bioactivities can be beneficial for both interaction partners or provide a competitive advantage only for the producer. Colonizers of confined habitats such as biofilms are known as prolific producers of a great number of bioactive secondary metabolites and are a potential source for novel compounds. We investigated the strain Paracoccus marcusii CP157, which originates from the biofilm on the carapace of a shell disease-affected Cancer pagurus specimen, for its potential to produce bioactive secondary metabolites. Its closed genome contains 22 extrachromosomal elements and several gene clusters potentially involved in biosynthesis of bioactive polyketides, bacteriocins, and non-ribosomal peptides. Culture extracts of CP157 showed antagonistic activities against bacteria from different phyla, but also against microalgae and crustacean larvae. Different HPLC-fractions of CP157 culture extracts had antibacterial properties, indicating that several bioactive compounds are produced by CP157. The bioactive extract contains several small, antibacterial compounds that partially withstand elevated temperatures, extreme pH values and exposure to proteolytic enzymes, providing high stability toward environmental conditions in the natural habitat of CP157. Further, screening of 17 Paracoccus spp. revealed that antimicrobial activity, hemolysis and production of N-acyl homoserine lactones are common features within the genus. Taking into account the large habitat diversity and phylogenetic distance of the tested strains, we hypothesize that bioactive secondary metabolites play a central role in the ecology of Paracoccus spp. in their natural environments.

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Complete Genome Sequence of the Lignocellulose-Degrading Actinomycete Streptomyces albus CAS922

Microbiology Resource Announcements ◽

10.1128/mra.00227-20 ◽

2020 ◽

Vol 9 (21) ◽

Author(s):

Anna Tippelt ◽

Markus Nett ◽

M. Soledad Vela Gurovic

Keyword(s):

Secondary Metabolites ◽

Complete Genome Sequence ◽

Sunflower Seed ◽

Gene Clusters ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Carbohydrate Active Enzymes ◽

Content Type ◽

Bioactive Secondary Metabolites ◽

Streptomyces Albus

ABSTRACT Streptomyces albus CAS922 was isolated from sunflower seed hulls. Its fully sequenced genome harbors a multitude of genes for carbohydrate-active enzymes, which likely facilitate growth on lignocellulosic biomass. Furthermore, the presence of 27 predicted biosynthetic gene clusters indicates a significant potential for the production of bioactive secondary metabolites.

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IMG-ABC v.5.0: an update to the IMG/Atlas of Biosynthetic Gene Clusters Knowledgebase

Nucleic Acids Research ◽

10.1093/nar/gkz932 ◽

2019 ◽

Cited By ~ 5

Author(s):

Krishnaveni Palaniappan ◽

I-Min A Chen ◽

Ken Chu ◽

Anna Ratner ◽

Rekha Seshadri ◽

...

Keyword(s):

Secondary Metabolites ◽

Secondary Metabolism ◽

Gene Clusters ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Microbial Genomes ◽

Initial Release ◽

Systematic Identification ◽

Biomedical Potential ◽

Biosynthetic Machinery

Abstract Microbial secondary metabolism is a reservoir of bioactive compounds of immense biotechnological and biomedical potential. The biosynthetic machinery responsible for the production of these secondary metabolites (SMs) (also called natural products) is often encoded by collocated groups of genes called biosynthetic gene clusters (BGCs). High-throughput genome sequencing of both isolates and metagenomic samples combined with the development of specialized computational workflows is enabling systematic identification of BGCs and the discovery of novel SMs. In order to advance exploration of microbial secondary metabolism and its diversity, we developed the largest publicly available database of predicted BGCs combined with experimentally verified BGCs, the Integrated Microbial Genomes Atlas of Biosynthetic gene Clusters (IMG-ABC) (https://img.jgi.doe.gov/abc-public). Here we describe the first major content update of the IMG-ABC knowledgebase, since its initial release in 2015, refreshing the BGC prediction pipeline with the latest version of antiSMASH (v5) as well as presenting the data in the context of underlying environmental metadata sourced from GOLD (https://gold.jgi.doe.gov/). This update has greatly improved the quality and expanded the types of predicted BGCs compared to the previous version.

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Variation Among Biosynthetic Gene Clusters, Secondary Metabolite Profiles, and Cards of Virulence Across Aspergillus Species

Genetics ◽

10.1534/genetics.120.303549 ◽

2020 ◽

Vol 216 (2) ◽

pp. 481-497 ◽

Cited By ~ 1

Author(s):

Jacob L. Steenwyk ◽

Matthew E. Mead ◽

Sonja L. Knowles ◽

Huzefa A. Raja ◽

Christopher D. Roberts ◽

...

Keyword(s):

Secondary Metabolites ◽

Secondary Metabolism ◽

Secondary Metabolite ◽

Immune Cell ◽

Sequence Divergence ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Host Immune System ◽

Biosynthetic Gene ◽

Genetic Determinants

Aspergillus fumigatus is a major human pathogen. In contrast, Aspergillus fischeri and the recently described Aspergillus oerlinghausenensis, the two species most closely related to A. fumigatus, are not known to be pathogenic. Some of the genetic determinants of virulence (or “cards of virulence”) that A. fumigatus possesses are secondary metabolites that impair the host immune system, protect from host immune cell attacks, or acquire key nutrients. To examine whether secondary metabolism-associated cards of virulence vary between these species, we conducted extensive genomic and secondary metabolite profiling analyses of multiple A. fumigatus, one A. oerlinghausenensis, and multiple A. fischeri strains. We identified two cards of virulence (gliotoxin and fumitremorgin) shared by all three species and three cards of virulence (trypacidin, pseurotin, and fumagillin) that are variable. For example, we found that all species and strains examined biosynthesized gliotoxin, which is known to contribute to virulence, consistent with the conservation of the gliotoxin biosynthetic gene cluster (BGC) across genomes. For other secondary metabolites, such as fumitremorgin, a modulator of host biology, we found that all species produced the metabolite but that there was strain heterogeneity in its production within species. Finally, species differed in their biosynthesis of fumagillin and pseurotin, both contributors to host tissue damage during invasive aspergillosis. A. fumigatus biosynthesized fumagillin and pseurotin, while A. oerlinghausenensis biosynthesized fumagillin and A. fischeri biosynthesized neither. These biochemical differences were reflected in sequence divergence of the intertwined fumagillin/pseurotin BGCs across genomes. These results delineate the similarities and differences in secondary metabolism-associated cards of virulence between a major fungal pathogen and its nonpathogenic closest relatives, shedding light onto the genetic and phenotypic changes associated with the evolution of fungal pathogenicity.

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Genome, Transcriptome, and Functional Analyses of Penicillium expansum Provide New Insights Into Secondary Metabolism and Pathogenicity

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-09-14-0261-fi ◽

2015 ◽

Vol 28 (3) ◽

pp. 232-248 ◽

Cited By ~ 109

Author(s):

Ana-Rosa Ballester ◽

Marina Marcet-Houben ◽

Elena Levin ◽

Noa Sela ◽

Cristina Selma-Lázaro ◽

...

Keyword(s):

Secondary Metabolites ◽

Secondary Metabolism ◽

Plant Pathogens ◽

Citrus Fruit ◽

Genomic Analysis ◽

Pathogenic Fungi ◽

Gene Clusters ◽

Penicillium Expansum ◽

Pome Fruit ◽

Penicillium Species

The relationship between secondary metabolism and infection in pathogenic fungi has remained largely elusive. The genus Penicillium comprises a group of plant pathogens with varying host specificities and with the ability to produce a wide array of secondary metabolites. The genomes of three Penicillium expansum strains, the main postharvest pathogen of pome fruit, and one Pencillium italicum strain, a postharvest pathogen of citrus fruit, were sequenced and compared with 24 other fungal species. A genomic analysis of gene clusters responsible for the production of secondary metabolites was performed. Putative virulence factors in P. expansum were identified by means of a transcriptomic analysis of apple fruits during the course of infection. Despite a major genome contraction, P. expansum is the Penicillium species with the largest potential for the production of secondary metabolites. Results using knockout mutants clearly demonstrated that neither patulin nor citrinin are required by P. expansum to successfully infect apples. Li et al. ( MPMI-12-14-0398-FI ) reported similar results and conclusions in MPMI's June 2015 issue.

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