Marine Chemical Ecology - Chapter 18 (end) doc

Metabolites of Free-Living, Commensal, and Symbiotic Benthic Marine Microorganisms Valerie S.. Metabolites of Free-Living, Commensal, and Symbiotic Benthic Marine Microorganisms 569Stru

Trang 1

Metabolites of Free-Living, Commensal, and Symbiotic Benthic Marine

Microorganisms

Valerie S Bernan

CONTENTS

I Introduction 567

II Marine Sediments 568

A Shallow Marine Sediments 568

B Deep-Sea Marine Sediments 574

III Commensal Marine Microorganisms 577

IV Symbiotic Marine Microorganisms 583

V Summary 589

References 589

I INTRODUCTION

This chapter describes the natural product chemistry that has been identified or associated with marine microorganisms from the benthos and focuses on the marine eubacteria Most of the compounds described in this chapter resulted from a detailed search of the literature for micro-bially derived natural products This approach is biased in many respects, since it only describes bioactive compounds and small molecules However, due to the growing demand for new

comparatively little research has been directed toward the study of natural products from marine

systems of signaling and territorial marking It is estimated that less than 1% of potentially useful chemicals have been discovered from the marine environment, with microbial products

antibiotic, anticancer, anti-inflammatory, and other pharmacological activites.2,3,4 Hopefully a greater understanding of microbial metabolic diversity, coupled with ecological information, will yield a greater understanding of the complexity of marine environments This chapter reviews metabolites of microorganisms that occur in sediments, as well as those in commensal associations and symbiotic relationships

18

9064_ch18/fm Page 567 Tuesday, April 24, 2001 5:30 AM

Trang 2

568 Marine Chemical Ecology

II MARINE SEDIMENTS

Marine sediments are composed of organic debris resulting from ongoing, seasonal, or catastrophicdie-offs of macrobiotic and microbial populations.2 This sedimenting debris provides energy forthe benthic organisms on the sea floor Organic mineral aggregates that are rich in carbohydratesare produced by deposit-feeding animals such as bivalves, which rework the sediments as theyfeed This extensive reworking of the sediments makes them unstable so that the organic aggregatesare resuspended by the tidal flow to produce turbid estuarine waters which are important inrecycling Consumption of resuspended organic debris and phytoplankton by bivalve molluscs andother benthic invertebrates produces rich deposits of feces and pseudofeces that coat mineralfragments These mineral fragments yield additional organic mineral aggregates, which in turn areheavily colonized by microorganisms Other sources of organic matter include those derived fromcordgrass and from beds of seagrasses, such as eelgrass and turtle grass as well as mangroves Asthese plants decompose, they are reworked into smaller fragments by deposit feeders, which cansupport increased populations of microbiota

Within interstitial habitats of sandy beaches, particles are trapped in the upper 5-cm surfacelayer and give rise to a bacterial–protozoan community.5 Below this level, a bacterial flora attached

to sand grains removes some of the dissolved organic carbon while supporting a meiofaunacommunity comprised of nematods and copepods The biotic community of these intertidal sandflats

is supplemented by the production of organic matter via benthic diatoms which migrate verticallywith the tides

Since terrestrial actinomycetes have been such prolific producers of bioactive molecules, it wasnatural to investigate marine species of the same group Therefore, it was no surprise that actino-mycetes isolated from the marine sediments have proven to be one of the most prolific sources ofbioactive secondary compounds.6,7 Their distribution in sediments varies depending on the depthfrom which the sample was collected In several studies, Streptomyces predominated in near-shoremarine sediments, but decreased dramatically past the sublittoral zone In contrast, actinoplanetesare found in greater numbers as the distance from the shoreline increases

The study of the metabolites produced by marine actinomycetes was pioneered by researchers atthe Institute of Microbial Chemistry in Tokyo in the early 1970s One of the first compounds

purpurgensa SS-228 collected in mud samples from Sagami Bay, Japan.8 This antibiotic selectivelyinhibited Gram-positive bacteria and was active against Ehrlich ascites tumor cells in mice It alsoproduced a hypotensive effect in mice, probably due to its inhibition of dopamine hydroxylase inthe pathway of epinephrine biosynthesis Most interestingly, this bacterium only produced theantibiotic when it was fermented in diluted yeast extract containing “Kobu Cha” (the brown seaweed

micro-organisms have nutritional requirements corresponding to nutrients in their natural habitats Thisstudy was also one of the first to report that bacteria from the order Actinomycetales could beisolated from the marine environment Actinomycetes were originally thought to have entered themarine environment via rivers or runoff or to have existed as spores of terrestrial species However,

occurred as indigenous populations, and that populations increased in relative abundance in response

to the availability of certain nutrients Greater abundances of culturable streptomycetes found incoastal environments vs deepwater marine systems may be attributed to higher amounts of organicdetritus, much of which is derived from vascular plants concentrated in shallow marine systems.Unlike the terrestrial actinomycetes, marine actinomycetes have been shown to produce mac-rolides only rarely One example of this class of compounds is the aplasmomycins A–C

Trang 3

Metabolites of Free-Living, Commensal, and Symbiotic Benthic Marine Microorganisms 569

(Structure 18.1a–c) These compounds were isolated from the fermentation of Streptomyces griseus

SS-20 collected from shallow mud in Sagami Bay, Japan The antibiotic was only produced underlow-nutrient media containing “Kobu Cha” and 3% NaCl The aplasmomycins are inhibitory against

aplasmo-mycin is an effective antimalarial agent in Plasmodium berghei-infected mice X-ray phy revealed that the active compound contained a symmetric ring in which boron was coordinated

crystallogra-in the center with a crown ether-like structure.10 Fenical’s group11 from Scripps Institute of ography also isolated a new member of a rare class of macrolides, maduralide (Structure 18.2).Maduralide was isolated from the fermentation of a Maduramycete isolated from shallow sedimentsfrom Bodega Bay, California This compound is a member of a rare 24-membered ring lactonegroup represented by rectilavendomycin

Ocean-Among the alkaloids, the most unusual example is an acaricidal (lethal to arachnids) erpene derivative, altemicidin (Structure 18.3) This novel alkaloid was purified from a marine strain

part of Japan It yielded potent antitumor activity in vitro against L1210 murine leukemia and IMCcarcinoma cell lines, but was toxic in vivo in mice Altemicidin is a novel sulfur- and nitrogen-

derivative of phenylpropionic acid (Structure 18.4) A series of analogs was synthesized and theiranti-algae properties examined These methyl ester analogs were found to be more active againstalgae than the free acids in both agar diffusion and liquid test systems The compounds did notexhibit any antimicrobial activity against Staphylococcus aureus, Escherichia coli, or Mucor miehei

at concentrations up to 200 µg/mL

Christchurch, New Zealand was fermented in a saline medium and found to produce modestantibacterial activity against Bacillus subtilis.14 The isolation of the active compound revealed it to

be an actinoflavoside, a molecule of an unprecedented structure class (Structure 18.5) voside resembles the common plant-derived flavonoid glycosides, but this compound contains an

Actinofla-O O

O

O O

O B

OR

R'O

M+

-18.1a: Aplasmomycin A R = R' = H 18.1b: Aplasmomycin B R = H, R' = Ac 18.1c: Aplasmomycin C R = R' = Ac 9064_ch18/fm Page 569 Tuesday, April 24, 2001 5:30 AM

Trang 4

additional alkylation at C-5 Due to the general conclusion that prokaryotes do not produce thisclass of compounds, its origin via the flavinoid biosythetic pathway seems questionable

A marine actinomycete, Streptomyces virdostaticus spp "litoralis" was isolated from an tidal sample collected in Key West, Florida Fermentation samples exhibited both antibacterial andDNA-damaging activites Although activity was observed in a tap-water based medium, the addition

inter-of 2% NaCl increased the biomass by 33% and increased activity four-fold Purification inter-of theactive materials revealed four related bioxalomycins (Structure 18.6a and b) The β species werefound to be the quinone forms of the corresponding α components The β components are distin-guished from the antibiotic naphthyridinomycin by the presence of a second oxazolidine ring in aregion of the molecule analogous to quinocarcin Bioxalomycin α2 was the most potent antibiotic

of the group, showing MIC values between 0.002–0.25 µg/mL, and also exhibited excellent in vitro

activity against neoplastic cell lines This compound was active in vivo in a mouse P388 leukemiamodel demonstrating an 80% increase in life survival (ILS) Mechanistic studies have shown thatfollowing metabolic reduction of the quinone, bioxalomycin α2 cross-links DNA through alkylation

A new series of phenazines has been isolated from a strain of marine-derived Streptomyces byFenical’s group.16 A study from the shallow sediments of Bodega Bay, California resulted in the

N COOH

HO

NH HO

H

O

18.5: Actinoflavoside 9064_ch18/fm Page 570 Tuesday, April 24, 2001 5:30 AM

Trang 5

antibacterial activity Subsequent fermentation of this isolate in a saltwater based medium produced

was isolated from the fermentation of a Streptomyces sp isolated from a sediment sample collected

50% seawater medium and produced the novel 5,10-dihydrophencomycin methyl ester and theknown microbial metabolites (2-hydroxyphenyl)acetamide, meanquinone MK9, and phencomycin.The new 5,10-dihydrophencomycin methyl ester (Structure 18.8) exhibited less antimicrobial activ-ity than phencomycin, and the dimerization of identical m-C7N units may explain its origin

(Structures 18.9 a–e) were isolated together with the known compounds enterocin and enterocin from the fermentation broth of a new Streptomyces sp.18 The strain was isolated fromthe shallow marine sediments collected at Wailupe beach park on the southeast shore of Oahu,Hawaii The α-pyrone moeity is commonly observed in many antibiotics and toxins Interestingly,enterocin and 5-deoxyenterocin, along with the 5-behenate and 5-arachidate esters of enterocin,

compounds in both prokaryotes and chordates raises the question of whether symbiotic or ciated microorganisms are responsible for the production of the metabolites isolated from somemarine invertebrates

asso-In a study of estuarine microorganisms isolated from Torrey Pines, La Jolla, California, Fenical

fermentation broth of an unidentified actinomycete Both compounds possess a common quinone with rare sesquiterpenoid structural components In addition, marinone possesses a brominesubstituent in the dihydroxybenzene ring, a position typical for bromination in marine metabolites.These new molecules are among a rare group of bacterial metabolites produced via mixed biosyn-thesis involving both acetate and terpene pathways Marinone and debromomarinone exhibit sig-nificant in vitro antibacterial activity against Gram-positive bacteria More recently, the same groupisolated a different unidentified actinomycete, from a sediment sample collected at 1m depth inBatiquitos Lagoon, California, that also produced marinone in addition to several cytotoxic metab-olites related to marinone One compound, neomarinone (Structure 18.11), is a novel metabolitepossesssing a new sesquiterpene- and polyketide-derived carbon skeleton The other two derivativesare isomarinone and methoxydebromomarinone All three compounds are also derived from a mixedbiosynthetic pathway involving polyketide and terpene pathways Connection of the sesquiterpenoidside-chain to the naphthoquinone core occurs on the nonquinone side in neomarinone The origin

naphtho-of the sesquiterpenoid side-chain appears complex; it is possibly derived from a cation-inducedmethyl migration as observed in the trichothecenes All three compounds exhibited moderate in vitro

Trang 6

cytotoxicity in the National Cancer Institute’s 60 cancer cell line panel with a mean IC50 value

of 10 µM.20

As part of the continuing interest in isolating secondary metabolites from marine estuaries, aStreptomycete was isolated from a sediment sample collected in Mission Bay, California Whenthe culture was fermented under saline culture conditions, it was found to produce a family ofnovel cyclic heptapeptides, cyclomarines A, B, and C While the major metabolite, cyclomarine

A (Structure 18.12), is cytotoxic in vitro toward cancer cells, it is more interesting for its significant

in vitro and in vivo inflammatory properties The compound displays significant topical inflammatory activity in the phorbol ester-induced mouse edema assay, showing 92% inhibition

anti-at the standard test dose Cyclomarine A contains three common and four unusual amino acids

well known synthetic building block, but is a rare constituent of natural products, found in only

N H

H N

N N

O O

R R OH

Trang 7

O

O OH

OH HO

O

CH3O O

O O

O O OCH3HO

O

O O

H

HO O O

OCH3

H

O

O O

OH

O

CH3O HO

OH H H O

18.9a: Wailupemycin A 18.9b: Wailupemycin B

18.9c: Wailupemycin C 18.9d: Wailupemycin D

H N

N H O

N O HN O

H N

O HN

O N O O

HO N

HO

CH3O O

Trang 8

These amino acids may be products of unusual biosynthetic pathways, and it will be interesting

to elucidate their biosynthesis.21 Another estuarine Streptomyces was isolated from the sandy

sediment collected near San Diego, California in the San Luis Estuary Cultivation of the

Strep-tomyces sp resulted in the isolation of two new aromatic tetraols, luisols A and B (Structure 18.13a

and b) Luisol A, formally a reduced hydroquinone, appears to be related to the quinones of the

granaticin class The structure of luisol B contains the rare epoxynaphtho[2,3c]furan which is only

found in one other natural product, the fungal metabolite anthrinone.22 Lastly, a Streptomycete sp

was isolated from an estuary near Doheny Beach, California and fermented in a salt-based medium

The culture broth was found to contain a new pentacyclic polyether, arenic acid (Structure 18.14),

which is related to two known polyether antibiotics, K41-A and oxolonomycin The structure of

arenaric acid was established by spectroscopic methods involving comprehensive two-dimensional

Although marine actinomycetes are the most prolific source for bioactive metabolites from

shallow sediments, marine Bacilli spp have also been isolated, and unusual secondary metabolites

substi-tuted with a ketide extended leucine PM-94128 was quite potent with cytoxicity against several

protein synthesis

Many interesting metabolites have also been isolated from deep-sea sediments.2 The energy input

to the sea floor below a depth of 2 km is thought to be less than 10% of the primary productivity

in the euphotic zone At these depths, food consists of a slow rain of fecal pellets and zooplankton

along with the carcasses of larger organisms from the nekton Most of these carcasses are consumed

quickly by fish who scatter the remains in the form of feces over large areas to be utilized by

benthic microorganisms A bacterial isolate from deep-sea mud collected at a depth of 3300 m

off the Aomori coast of the Japan Sea required a seawater medium to grow and produce bioactive

substances Even though the strain was isolated at 700 atm pressure, it appeared to grow well at

surface pressure and temperature The strain was identified as Alteromonas haloplanktis and was

This compound had little cytotoxicity but, when added to a mixed macrophage-tumor cell culture,

induced macrophage-mediated cytolysis of tumor cells Its dimeric structure contains two

hydrox-amates and two amide functionalitites and is similar to other siderophores such as nocardimin

and desferioxamine

While screening for antitumor effects, Fenical et al.26 isolated a deep-sea bacterium from a

sediment sample obtained from a 1000-m depth along the California coast Fermentation of the

slow-growing bacterium in a salt-based medium yielded a series of novel cytotoxic and antiviral

macrolides, the macrolactins A–F This bacterium was an unidentified Gram-positive organism that

produced six macrolides and two open-chain hydroxy acids when fermented in the presence of salt

at atmospheric pressure Macrolactin A (Structure 18.17) was the predominate compound produced,

showing moderate antibacterial activity, yet it was quite potent against B16-F10 murine melanoma

in vitro with an IC50 of 3.5 µg/ml Of potentially greater significance, macrolactin A inhibited

several viruses, including Herpes simplex (IC50 = 5.0 µg/ml) and HIV, the human immunodeficiency

virus (IC50 = 10.0 µg/mL)

Gram-positive bacterium cultured from deep-sea sediments The caprolactams A and B (Structure 18.18a

Trang 9

7-methyloctanoic acid and 6-methyloctanoic acid, respectively Natural products containing acyclized lysine are uncommon and have only been reported in several sponges and fungi In fact,these structures are quite similar to the bengamides, which are sponge-derived caprolactams withoxidized acyl side-chains Thus, one may speculate that the bengamides isolated from sponges maytruly be produced by a symbiotic microorganism The compounds are mildly cytotoxic towardhuman epidermoid carcinoma and colorectal adenocarcinoma cells with MIC values of 10 and

γ-indomycinone (Structure 18.19), from a Streptomyces species isolated from a deep-sea sediment

1-hydroxy-1-methylpropyl side-chain The compound exhibits only mild cytotoxicity against a humancolon cancer cell line HCT-116 but shows a differential cytotoxicity against the Chinese hamsterovary cell lines UV20 (deficient in DNA excision repair) and BR1 (proficient in DNA repair)

Another Streptomycete species was isolated from deep-sea sediments collected at 1500 m inthe sea surrounding Tokyo, Japan When this culture was fermented in the presence of seawater,

O O OH

N H O

H H

O

H H

Trang 10

potential to be developed as antimetastasis or anti-HIV drugs The two inhibitors were

never been isolated from nature but were found in marine organisms, suggesting that they are ofmicrobial origin.29

Simidu et al.30 isolated 49 bacterial strains from deep-sea core sediments collected at a depth

of 4000 m and examined them for the production of tetrodotoxin This study indicates that dotoxin-producing bacteria are not restricted to certain taxonomic groups A variety of groups of

tetro-bacteria, including Bacillus, Micrococcus, Acinetobacter, Aeromonas, Alteromonas, Moraxella, Vibrio, and one unidentified bacterium, all produce tetrotoxin Although the strains are limited in

number, the tetrodotoxin-producing bacteria are quite widespread among various bacterial groups

in marine sediments It has been postulated that the tetrodotoxins are synthesized solely by bacteria

in sediments and subsequently accumulated by benthic organisms, such as fish and crabs, thatacquire them through the food web

Another cyclic peptide, halobacillin, was isolated from a marine-derived Bacillus sp isolated

from a deep-sea sediment core.31 Halobacillin (Structure 18.21) was only produced in based media and is similar in structure to the surfactins and iturins Interestingly, the compoundexhibits cytotoxicity against the human colon tumor cell line HCT-116 but lacks the antibacterialactivity associated with surfactin

sea-water-H N

HO OH

OH

OH O

18.20: D -Glucono-1,5-lactam

N

N H

H N

N OH HO

O O O

O R

Trang 11

III COMMENSAL MARINE MICROORGANISMS

Commensal marine bacteria inhabit surfaces, tissues, and internal spaces of other organisms andplants, but, often, the exact associations are transient and the interactions are not well understood.Since bacteria occur in seawater at concentrations of one million cells per milliliter, marine plantsand animals are constantly exposed to extremely high concentrations of bacteria Many of thesebacteria are motile, chemotactic, and/or pathogenic, and readily colonize a variety of surfaces.Bacteria are the first detectable microorganisms to colonize a surface, which they do by twoprocesses The first process is an instantaneous but reversible adsorption of the bacteria onto a

adsorption, in which different populations develop with time These bacterial films often play avital role in the growth and development of macroscopic marine plants A number of seaweedshave distinctive epibacterial floras that may supply growth factors and contribute to the destruction

of algal autoinhibitory substances Animal surfaces also appear to selectively enhance or inhibitmicrobial colonization Documentation of these coexistances is increasing, but evidence supportingtrue symbiosis rather than a nonobligate association is still lacking Nonetheless, these associationsare significant in the quest for new bioactive natural products because of the potential cooperativerole in the production of novel metabolites These surfaces are richer in nutrients than seawaterand most sediments, thus providing a unique niche for the isolation of many diverse bacteria, asdescribed below.32

Umezawa and co-workers screened fermentation broths containing various bacterial species,isolated from the surface of seaweeds, for the production of polysaccharides.33 One genus, Fla- vobacterium uglignosum, was found to produce marinactan, which is capable of suppressing

sarcoma-180 tumors in mice At daily doses of 10 to 50 mg/Kg in mice, marinactan inhibited75–95% of the growth of these tumors Marinactan is a neutral heteroglycan consisting of fucose,

mannose, and glucose In an antibacterial screening program by Sano et al.,34 a Pseudoalteromonas

sp was found to produce the novel antibiotic korormicin (Structure 18.22) The organism was

isolated from the tropical green alga Halimeda sp., collected in Palau and required a

seawater-based medium for growth and survival Korormicin is a combination of an oxidized fatty acid and

an unusual lactonized amino acid This unusual amino acid is without biosynthetic precedent andposes an interesting question as to its origin Interestingly, korormicin was harmless to terrestrialbacteria and Gram-positive marine bacteria, but was active against 11 marine Gram-negative

bacteria During the course of an anticancer screening program, a new marine bacterium, bacter variabilis, was isolated from the blades of the tropical brown alga Pocockiella variegata.35

Pelagio-This Gram-negative, pleomorphic, halophillic bacterium represents a new genus of marine teria Fermentation extracts of this culture were found to contain the known compound, griseoluticacid, and three new phenazine antibiotics, the pelagiomicins A–C Pelagiomicin A (Structure 18.23)demonstrated strong antibacterial activity to several Gram-negative and Gram-positive bacteria and

eubac-inhibited several cancer cell lines in vitro [ID50 values between 0.04 to 0.2 µg/mL]

Interest in finding new antibiotics against multidrug resistant Mycobacterium tuberculosis and

M avium-intracellulare led Andersen et al.36 to isolate one Pseudomonas sp from the surface of

an unidentified leafy red alga collected in Masset Inlet, British Columbia, and another Pseudomonas

sp from an unidentified tube worm collected near Moira Island, British Columbia When cultured

on solid agar in the presence of salt, the two strains produced the novel cyclic depsipeptides,massetolides A–H, and the known compound viscosin Massetolide A (Structure 18.24) was two

to four times more potent than viscosin in its in vitro inhibition of M tuberculosis (MIC = 5–10 µg/mL) and M avium-intracellulare (MIC = 2.5–5.0 µg/mL), and a single intraperitoneal injection

of 10 mg/Kg of massetolide A was found to be nontoxic to mice No activity was observed foreither compound against a panel of other human pathogens A number of unnatural and intriguing

L-cyclopropylalanine, etc.) but the supply was not sufficient for testing The same group also isolated

Trang 12

a family of novel cyclic decapeptide antibiotics, loloatin A–D (Structure 18.25a–d) from a Bacillus

Bacillus sp was cultured on solid agar containing salt, and the extract from the cells was found to

produce related antibiotics that were active against several strains of antibiotic resistant bacteria

Loloatins A–C inhibited the growth of methicillin-resistant S aureus, vancomycin-resistant coccus sp., and penicillin-resistant Streptococcus pneumoniae with MIC of 0.5 to 4 µg/mL Inter-estingly, only loloatin C exhibited antibacterial activity against the Gram-negative bacterium

Entero-Escherichia coli (MIC 1 µg/mL), and loloatin D was four times less active than A–C against positive bacteria These results demonstrate that subtle changes in cyclic decapeptide structure canhave a significant impact on antimicrobial activity Even though the loloatins share structural

Gram-n-C7H15H

N N

H

H N

O

O H N O

NH O NH O HN O

NH O

H N

NH

N HN O

O

O O O

O

COOH

O O H

Trang 13

features with the tyrocidines, the latter have never been reported to demonstrate Gram-negativeantibacterial activity

Shigemori et al.38 isolated a new cyclic alkaloid, alteramide A (Structure 18.26), produced by

an Alteromonas species isolated from the marine sponge Halichondria okadai collected near Nagai,

Kanagawa, Japan Alteramide A is a macrocyclic lactam containing dienone and dienoyltetramicacid functionalities that can undergo a [4 + 4] cycloaddition to generate a hexacyclic derivative.Alteramide A exhibited cytotoxicity against murine leukemia P388 cells, murine lymphoma L1210cells, and human epidermoid carcinoma KB cells Related macrocylic lactams, such as ikaruga-

mycin and discodermolide, have been previously isolated from the terrestrial Streptomyces ochromogenes var ikaruganesis and the sponge Discodermia dissoluta, respectively, but not from

phae-symbiotic bacteria The isolation of alteramide may provide insights into the metabolic origin ofdiscodermolide

Investigations of microorganisms associated with the Antarctic sponge Isodictya setifera led

to the isolation of a strain of Pseudomonas aeruginosa that exhibited Gram-positive antibacterial

activity.39 Fractionation of the culture broth identified a new diketopiperazine, cyclo-(L

alka-loids Investigations of the sponge revealed that neither metabolite was present This suggests

H

O

O NH O N

O N

O N O

HN O

N H

O N H

X=H

R2=

18.25c: Loloatin C R1=

N H

R2=

N H X=H

18.25d: Loloatin D R1=

N H

Định dạng
Số trang	26
Dung lượng	474,4 KB