The Marine Microbiome: An Untapped Source of Biodiversity and Biotechnological Potential

501 Pages·2016·13.37 MB·English

by  Lucas J. Stal

Lucas J. Stal · Mariana Silvia Cretoiu
Editors
The Marine
Microbiome
An Untapped Source of Biodiversity and
Biotechnological Potential
The Marine Microbiome
Lucas J. Stal Mariana Silvia Cretoiu
(cid:129)
Editors
The Marine Microbiome
An Untapped Source of Biodiversity
and Biotechnological Potential
123
Editors
Lucas J.Stal Mariana Silvia Cretoiu
RoyalNetherlandsInstituteforSeaResearch RoyalNetherlandsInstituteforSeaResearch
andUtrechtUniversity andUtrechtUniversity
University of Amsterdam Yerseke
Yerseke TheNetherlands
TheNetherlands
ISBN978-3-319-32998-7 ISBN978-3-319-33000-6 (eBook)
DOI 10.1007/978-3-319-33000-6
LibraryofCongressControlNumber:2016938664
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Foreword
Microbial Cultures are Still Essential in the Era
of High-Throughput Sequencing
We live in an era when it is possible to sequence any sample, taken from any
environment, and to know which organisms are present. It is an extraordinary
achievement. In little over 40 years, from the very innovative scientific advances
initiated by Frederick Sanger and others, and with the equally important develop-
mentoftechnology,wehaveapresent-daycapabilitytogeneratemetagenomesand
metatranscriptome libraries, and to provide that information to every microbiolo-
gist. Why then, should microbiologists still be interested in isolating microorgan-
isms from the natural environment, manipulating samples to form axenic cultures,
and then working with those cultures in the very artificial environment of the
laboratory? Surely, all the information that anyone could ever require can be
gleaned from sequencing DNA and RNA. Are microbiologists too wedded to a
culturing approach that, although very successful in the past, may now have out-
lived its usefulness? My view is that this is not the case and, more than ever, we
needtobringimportantmicrobesintoculture.Butthereisacaveat—wehavetobe
selective and not attempt to culture everything.
In a sense, high-throughput sequencing has introduced new challenges for
microbiologists. When marine scientists first turned their attention to the microbi-
ologyoftheoceans,itseemedarelativelytractableproblem.Microbiologists,such
as Claude ZoBell, using approaches based on classical microbiology methods,
estimatedbacterialnumberfromthecoloniesthatdevelopedonanutrientagarplate.
Acoreassumptionofmicrobiologistsisthatasinglebacterialcellwillgrowtoform
asinglecolony;earlymarinemicrobiologistshadeveryexpectationthatthiswould
beaveryquantitativeapproachtoestimatehowmanybacteriawereinthesea.They
found that there were only a few 100 bacteria per mL of seawater—so suggesting
that bacteria were not at all a significant component of the pelagic ecosystem and
v
vi Foreword
wereapparentlymuchlessabundantthanphytoplankton.Butinthe1970swiththe
introduction of epifluorescence microscopy, microbiologists discovered that cul-
turingtechniqueswereunderestimatingthenumberofbacteriaintheeuphoticzone
by4or5ordersofmagnitude;i.e.,therewerenotjustafewhundredbacteriapresent,
but millions of bacteria per mL. Most marine bacteria were not growing in the
culture mediaand the numbers were grosslyunderestimated.
High-throughput sequencing techniques now allow us to describe that massive
diversity. We can do many things with these data: infer the biogeochemical func-
tion of the bacterial assemblage, track changes in species richness with time, and
describenovelsequencesthatarenotpresentlyindatabases.Therearethelimitsto
what sequence data can reveal; but many of these challenges can easily be
addressedifthereisaccesstoalaboratoryculture.Unfortunately,manyfamiliesof
bacteria are known only from sequence data generated from natural assemblages
and we do not have any cultured representatives of many of the phyla that are
widely distributed and abundant in the ocean. We may know quite a lot about
bacterial assemblages from sequence data, but we know very little of the basic
biology of these organisms; phenotype cannot be adequately determined from
sequence data. Access to laboratory cultures would be immensely beneficial in
answering important questions about the role of microbes in the sea.
One very contentious issue is what constitutes a bacterial species. It is very
common to approach the problem of phylogeny by using the 16S rRNA gene (or
another genetic locus) to define an operational taxonomic unit (OTU). Although a
pragmatic solutionto the problem could describe what may be present ina natural
population, 16S sequence does not encapsulate species information. It does not
answerthebasicquestionofwhatisthisorganism;whatconstitutesthisorganism;
and how has evolution resulted in this particular biological entity that is the center
ofourinterest?Thatentityistheresultofmanyprocesses;mutationofthenuclear
genome, acquisition and mutation of plasmids, what has been acquired among
others by horizontal gene transfer, pathogenicity and/or genomic islands.
One vivid example of the difficulties faced in describing a bacterial species
comesfrommedicalmicrobiologyandastudyofpathogenicEscherichiacoliusing
comparative genomic analysis (Rasko et al. (2008). E. coli is probably the
best-studiedbacteriumeverandisthedefaultorganismforbiochemicalandgenetic
studies.ProbablyallmicrobiologistswouldrecognizeE.coliasasinglespecies.As
such,itmightbeexpectedthatitcouldbedefinedpreciselyonthebasisofgenomic
similarity.Raskoetal.comparedthegenomesofatotalof17cultures,includinga
number of pathogenic strains, and found a great diversity. The mean genome size
ofthe17isolateswas5,020±446genes, andofthattotallessthan halfthegenes
(2, 344 ± 43) could be considered as to comprise a “conserved core”—genes that
were highly conserved in all 17 isolates. A significant number of genes (ca. 300)
were unique to one isolate, i.e., were present in one, but none of the other 17
genomes. Given such great genetic diversity in our best-studied bacterial species,
what is the likelihood that marine species can be described purely on the basis of
Foreword vii
metagenomic data? At present, it is probable that laboratory cultures are the only
sure way to completely describe a bacterium. It is true that genomic analyses of
singlecells,aswellasthetechnologyofmanipulatingsinglecellsfromthenatural
environment, are making significant progress; but, for those whose research inter-
ests are phylogeny, evolution, and identification, the most cost-effective approach
remains isolation and laboratory culture.
Cultures are also essential for commercially important research. Terrestrial
microorganisms,particularlythosederivedfromsoils,havebeenasourceofmany
secondary metabolites, enormously profitable for pharmaceutical companies, and
responsible for huge advances in human health. Microbes from the oceans have
been less important to date, not because marine microbes offer fewer possibilities
for biodiscovery, but because they have not received the same attention. The case
for focusing on marine microbes is strong; the seas offer a wide range of unusual
environments in which microbes have evolved. Unusual environments have unu-
sual microbes that do unusual things—many of which are likely to offer real
opportunities for biodiscovery of commercially important products. The pathway
into these new fields is through isolation and culturing of these novel microbes.
Clearly, with tens of thousands of different species of microorganisms, bacteria
and archaea—as well as protists—in seas, we cannot isolate everything. The
challenge is to prioritize, beginning with microbial families that are known to be
abundant intheoceanbut for which we haveno cultured representative. Sequence
data is clearly an essential tool in that prioritization process. Traditional microbi-
ologicalapproaches,suchasdilutiontoextinction,willcontinuetobecost-effective
but new culture media need to be devised. My personal belief is that we should
move away from traditional media that are based on yeast extract and peptone,
towards media that better mimic the composition of organic matter in the sea.
Progress may depend on the marine chemists developing better ways to quantify
and describe those organic compounds that are likely to be the major substrates
used by natural assemblages (but which will be present at vanishing low concen-
trations).Evolvingtechnologies,suchasthemanipulationofsinglecell,willclearly
be an important aid to successful culturing.
There is no doubt that this is an exciting time for marine microbiology. DNA
sequencinghasshownwhichorganismsarepresentintheseas—andthediversityis
huge.Thechallengenowistoascribefunctiontothisdiversity—toknowwhodoes
what. Isolation and culturing will continue to be very important tools in meeting
thesechallenges—andthisbookisanexcellentdescriptionoftherealprogressthat
is being made.
Ian Joint
The Marine Biological Association
The Laboratory, Citadel Hill
Plymouth, UK
viii Foreword
Reference
RaskoDA,RosovitzMJ,MyersGSAetal.(2008).ThepangenomestructureofEscherichiacoli:
comparative genomic analysis of E. coli commensal and pathogenic isolates. J Bacteriol
190:6881–6893,doi:10.1128/JB.00619-08
Preface
When we accepted the invitation by Springer to edit a book on “marine microbi-
ology,” the next thing was to think about the contents of such book and about
possible authors. There have been a fair number of books published on the topic
and we did not want to duplicate any of them. The lucky coincidence was that we
wereleadingalargeEuropeanconsortium“MaCuMBA,”whichstandsfor“Marine
Microorganisms: Cultivation Methods for Improving their Biotechnological
Applications,” a 4-year one (2012–2016), with 22 partners from 11 European
countries. MaCuMBA is a consortium of industrial and academic partners with
great variety inexpertise inmarinemicrobiology.Hence,itseemedobvioustoask
colleagues and principal investigators of the MaCuMBA consortium to contribute
to this book, for which we gave the title: The marine microbiome—an untold
resource of biodiversity and biotechnological potential.
The term “microbiome” is fashionable and is used to describe the whole
microbial community, i.e., all microorganisms and their genetic information in a
certain habitat or environment. The marine microbiome refers to the totality of
microorganismslivingintheocean,itsfringingseas,estuaries,andbaysandfjords.
This includes the intertidal areas of the coast but also the seafloor and the
sub-seafloor,thousandsofmetersdowninthebottomofthesea.Italsoincludesthe
microorganisms living on and in marine animals, plants (seagrasses), and
macroalgae, even though each of them forms its own microbiome.
Microorganisms are basically defined by their size and as a rule of thumb we
consideranyorganismamicroorganismwhenitssizeistoosmalltobeobservedin
detail by the naked eye. This would mean everything smaller than 1 mm. In
practice,mostorganismsthatweconsiderasmicroorganismsareinthemicrometer
(lm)(one-thousandthofamillimeter)range,thesmallestmaybeonly0.2lm,but
the biggest can be several hundreds of lm. Microorganisms comprise all three
domainsoflife:Bacteria,Archaea,andEukarya.Whiletheformertwodomainsare
traditionally considered microorganisms (bacteria), all macroorganisms (plants,
animals, macroalgae, and many fungi) are Eukarya. However, what is often for-
gotten that by far most Eukarya are in fact microorganisms (protists).
ix
x Preface
BacteriaandArchaeaareoftenreferredtoas“Prokaryotes,”(organismswithout
anucleus“karyon”intheircells)todistinguish“bacteria”fromtheeukaryotesthat
do have a nucleus. We agree with Norman Pace (‘Time for a change’ Nature 441:
289,2006,and‘It’stimetoretiretheprokaryote’.MicrobiologyToday,May2009,
85–87)whoarguesthataprokaryoteisdefinedbywhatisdoesnothave(anucleus)
andthatthisisnotagoodcriterion.Inthisbookwedecidedtoavoidthisterm,even
thoughsomeauthorswerenotfullyconvinced.However,aswiththeknowledgewe
have today, there is no doubt that Bacteria and Archaea comprise very different
domains of life, even though you cannot tell much from the morphology observed
underthemicroscope.Themorphologyofmanymicroorganismsisanywaydevoid
ofmuchresolution.Thediversityofmicroorganismsisintheirgenomeandinwhat
theydo.Using“prokaryote”isoftensloppy,becausewhenonestartsasking,often
only one of the domains of Bacteria or Archaea is meant. It is therefore more
accurate to name that domain. And in the more rare cases that both domains are
meant, it is not a big deal to name both. That is what we consequently did in this
book.
There is another biological entity that we have not named yet. Viruses are not
representingexactlyadomainoflife(i.e.,notbelongingtoanyofthethreedomains
of life), but they are surely a biological entity that needs to be considered when
talkingaboutanymicrobiome.Virusesarenot“living”simplybythefactthatthey
need a living cell to replicate. But viruses play an extremely important role in
maintaining the biodiversity, maintaining the microbial foodweb, and transferring
genetic information between organisms, perhaps even between domains. Recent
discoveries also show that the border between bacteria and virus is vanishing. The
numberofviruses intheoceanisoverwhelming withanorderofmagnitudelarger
than that of all microorganisms (10 and 1 million per milliliter of seawater,
respectively).
Itisexactly70yearsagowhenClaudeE.ZoBell’sbook“MarineMicrobiology”
appeared(C.E.ZoBell,MarineMicrobiology.Amonographonhydrobacteriology,
Waltham, Massachusetts, USA, 1946, 240 p.). ZoBell was at that time Associate
Professor of Marine Microbiology at Scripps Institute of Oceanography, La Jolla,
California.Hismonographisstillworthreadingandpresentsuswitharemarkably
complete picture of marine microbial life and you could ask yourself how much
more we know today. We hope nevertheless that the present edited volume does
give a taste of what hasbeen achieved inthose 70years and where we stand now.
The marine microbiome is not just interesting from a scientific point of view.
Certainly, with 70 % of the Earth’s surface covered by the ocean and the ocean
probably being the largest continuous habitat, the marine microbiome plays a
prominent role in the biogeochemical cycling of elements, is at the basis of the
marine foodweb, critical for the ecology of the sea, and essential for climate reg-
ulation and counteracting the effects of global change. However, science has also
made great discoveries of bioactive compounds made by marine microorganisms
that have found applications in biotechnology, bioenergy, and pharmacy, and
activities that are displayed by these organisms that find application in bioreme-
diation.Beingawarethatwemightnotknowmostofthemicrobialdiversitythatis