The enumeration and characterization of bacteria and fungi associated with marine wood-boring isopods,and the ability of these microorganisms to digest cellulose and wood

Bacteria and fungi associated with the marine wood-boring isopods Limnoria lignorum and Sphaeroma serratum, and with their wood burrows and seawater were investigated. Plate counts using nutrient agar and Czapex Dox agar media in natural and in artificial seawater were used. The diversity and number of bacteria associated with the two isopods were much higher than those associated with wood or seawater, whereas the opposite was true for fungi. The hydrolytic activity of the predominant genera on carboxymethylcellulose and on different types of wood was investigated. The magnitude of fungal cellulase activity exceeded that of bacteria. Activities of bacterial extracellulases exceeded those of intracellulases, while the opposite was observed for fungi. Cellulolytic activity of the predominant isolates biodegraded labkh, peach, white oak and mahogany in decreasing order of activity. Evidence provided by the cellulolytic activities, detected in woodcontaining cultures, suggests that the predominant micro-organisms isolated may play an important role in the woodboring process. Specimens used in the present study were collected from Port Fouad at Port Said Harbour, Egypt.     

Enzyme activities of marine bacteria involved in Laminaria-thallus decomposition and the resulting sugar release

The extracellular decomposing enzyme activities of marine bacteria, including 60 and 16 strains of Laminaria-thallus decomposing bacteria (LDB) and non-LDB, respectively, were determined against several algal polysaccharides. A number of LDB decomposed alginate, fucoidan, and cellulose, but not laminarin. Clear decomposing activity was not observed in the culture supernatant from nutrient broth, but was detected in that from a Laminaria-thallus medium, suggesting that decomposing enzymes were induced. During the incubation of LDB in Laminaria-thallus medium, depolymerized sugar materials with a wide range of molecular size were released into the culture fluid. Total sugars released into the culture fluid after 4 d of incubation amounted to a maximum of 95% of the initial content in the thallus tissues. These findings suggest that microbial decomposition of Laminaria thallus in sea water could be a good source of carbohydrates which may support the heterotrophic growth of marine organisms.     

The trophic role of glycolic acid in coastal seawater. I. Heterotrophic metabolism in seawater and bacterial cultures

Glycolic acid, a known algal excretory product, represents a potentially important energy source for heterotrophic bacteria in marine waters. Measurements of heterotrophic uptake and mineralization by the natural microbes in the plankton indicate quantitative use of glycolic acid comparable to other common microbial substrates. This activity shows vertical and horizontal variations that correlate in a general way with primary productivity. Glycolic acid appears to be present at higher concentrations than other low molecular weight substrates, and it also shows a lower rate of turnover. It is mineralized (respired) to a greater extent than other recorded substrates, an average of 70% of total uptake. Two-thirds of a total of 141 colonies of bacteria cultured from seawater on marine agar proved capable oftaking up and respiring glycolic acid. These same bacteria, however, were unable to utilize glycolic acid for growth when it was the sole carbon source. Glycolate may therefore occur at higher concentrations and show erratic fluctuations in natural waters because microbial use is not tied to glycolate production but to the presence of other substrates. The fact that so many bacteria are able to metabolize it indicates a possibly important trophic role as an energy source. It is suggested that glycolic acid may be a major source of energy for active transport of other substrates by marine bacteria.     

Larval settlement in the serpulid polychaete Hydroides elegans in response to bacterial films: an investigation of the nature of putative larval settlement cue

Larval settlement in the marine polychaete Hydroides elegans (Haswell) is induced by certain bacteria in marine biofilms. The exact nature of the settlement cue that larvae of H. elegans receive from these bacteria remains unknown. In this study, we revealed some properties of the bacterially derived larval settlement cue by investigating the larval settlement inductive activity of two bacterial strains after various treatments. These two bacterial strains, Roseobacter sp. and an α-subclass Proteobacteria, are highly inductive to larval settlement of H. elegans. The larvae responded similarly to Roseobacter and Proteobacteria in all the larval settlement bioassays, suggesting that the larval settlement-inducing substances produced by these bacteria may share common characteristics. First of all, the larvae did not settle in the seawater conditioned by the bacteria attached as a film or by the bacteria that were freely suspended in seawater. The results suggest that the putative larval settlement cue is not released into seawater and, therefore, should be associated with the surface of the bacteria. Secondly, formaldehyde treatment entirely eliminated the larval settlement induction activity of the bacterial films, and streptomycin treatment reduced the percentage of larval settlement on the bacterial films in a concentration-dependent manner. Since both treatments can kill bacteria with little damage to the surface chemistry of bacterial cells, the decline in larval settlement is suggested be due to a reduction of the viable bacterial population in the bacterial films. In fact, the reduction of larval settlement in the streptomycin treatments coincided with the decrease in viable bacterial populations in broth cultures containing respective concentrations of streptomycin. These results suggest that the viability of Roseobacter and Proteobacteria is important to their settlement induction effect. Since the larval settlement induction activity of the bacterial strains appears to correlate with their viability, we suggest that the putative larval settlement cue is derived from a metabolic pathway in the bacteria and that the cue is exported to and concentrated at the extracellular polymer matrix of the bacterial cell, at which the larvae establish contact with the bacteria. The larval settlement cue may be highly susceptible to degradation so that a metabolically active bacterial film is needed to maintain the putative cue at a concentration that surpasses the threshold for induction of larval settlement. Received: 14 October 1998 / Accepted: 5 September 2000     

不同补给水源的城市人工湖中异养菌耐药性及耐药菌种属分布研究

为了阐明不同补给水源的城市人工湖中异养菌耐药状况和耐药菌种属分布特征,选取分别以地表水和再生水为补水来源的XQ湖和FQ湖为代表进行研究。从2018年4—11月逐月采集水样,考察了各水样中对氨苄西林(AMP)、磺胺甲恶唑(SMZ)和四环素(TET)这3种不同种类抗生素具有耐药性的异养菌含量,并对分离菌株的耐药表型、耐药菌株的种属分布以及水质理化指标进行了分析。结果表明,2处人工湖中的AMP耐药菌和SMZ耐药菌含量约为10~2～10~4CFU·(100 m L)~(-1),而TET耐药菌含量则约10~1～10~3CFU·(100 m L)~(-1)。分离出的84株耐药菌归属于19个种,其中蜡状芽孢杆菌、大肠埃希氏菌、维氏气单胞菌、豚鼠气单胞菌和鸟氨酸拉乌尔菌为2个湖共有耐药菌。71.4%的耐药菌都是对AMP单一耐药,以蜡状芽孢杆菌为主。由于具有固有耐药性的细菌在分离出的耐药菌中占比很低,获得性耐药很可能在城市人工湖中异养菌耐药性的发展上发挥了主要贡献作用。地表水补水的XQ湖和再生水补水的FQ湖在总异养菌含量、耐药菌含量和检出率上均无显著差异。分离来源对16株耐药性气单胞菌的聚类无明显影响。     

The marine bacterium Leucothrix mucor as an algal epiphyte

Leucothrix mucor, a filamentous marine heterotrophic bacterium, is found in nature primarily as an algal epiphyte. Its distinctive characteristics make it easily recognizable by dir ect light-microscopic examination, and a method was develop ed to quantify natural populations of L. mucor on algae by direct microscopic counts. L. mucor was found to be widely distributed in the intertidal environment, especially in the mid-to-upper intertidal regions. Populations were most abundant on members of Rhodophyta, particularly on the alga Bangia juscopurpurea. The nature of the L. mucor-algal relationship was investigated by choosing an artificial plastic substrate to which L. mucor would attach. The use of the plastic substrate, both alone, and in conjunction with a natural algal host, gave results supporting the conclusion that L. mucor obtains its nutrients for growth from the alga rather than from seawater. Algae in the upper intertidal region may also protect attached L. mucor from desiccation.     

Study on phosphate uptake of the marine cyanophyte Synechococcus sp. NIBB 1071 in relation to oligotrophic environments in the open ocean

Inorganic phosphate (Pi) uptake by the marine cyanophyte Synechococcus sp. NIBB 1071 was studied using cells grown in an artificial seawater medium. The phosphate uptake was markedly enhanced in cells grown in the medium of low phosphate concentrations (phosphate-limited cells) than in cells grown in the phosphate-rich medium (phosphate-replete cells). The diagnosis of kinetics of instantaneous phosphate-uptake showed that V _max of the former was more than two orders of magnitude greater than that of the latter, and the k _m of the former was about 1/20 of that of the latter. The enhancement of the phosphate uptake was completed after a 40-h incubation of phosphate-replete cells in the phosphate-free medium. The activation was suppressed by chloramphenicol, an inhibitor of protein synthesis. The uptake developed in phosphate-limited cells was energy dependent and susceptive to osmotic shock, which suggests the involvement of a periplasmic phosphate-binding protein, analogous to that found in heterotrophic gram-negative eubacterial cells. The relationship between phosphate quota and growth rate, together with the kinetical data for phosphate uptake, predicted that ambient phosphate as low as 0.5 nM could support cell growth at a rate of one division per day. Results indicate that cells can grow rapidly even at phosphate concentrations as low as nanomolar levels. A possible regulatory mechanism of phosphate uptake in marine Synechococcus spp. is discussed in relation to a wide distribution of this picophytoplankton in the ocean environment. Received: 19 March 1997 / Accepted: 2 April 1997     

Impact of conditions of cultivation and adsorption on antimicrobial activity of marine bacteria

The potency of free-living and animal-associated marine bacteria to produce antimicrobial substances has been studied in 491 strains isolated from northern and southern parts of the Pacific Ocean. A total of 26% (126 out of 491) of the strains examined produced antimicrobial compounds against 11 test bacterial strains (TBS) including the fish pathogens Aeromonas hydrophila and Vibrio anquillarum. Antimicrobial substances (AS) produced by marine bacteria were especially active against Staphylococcus epidermidis, Proteus vulgaris, Enterococcus faecalis, and Candida albicans. Twelve strains, isolated from different sources, were chosen as promising candidates, producing a number of AS. Production of AS varied within 24 to 72 h, increasing in a culture medium based on natural sea water with Br-ions, and after attachment to polymeric surfaces. In order to study the influence of adsorption, selected strains with a high potential for antimicrobial production were cultivated on polymeric surfaces with different hydrophobicities and chemical functionalities. These parameters of the surface hydrophobicity (measured by means of water contact angles) and chemical functionality of the surfaces were manipulated using the photo- and thermochemistry of a polymeric system (diazo-naphto-quinone/novolak) commonly used as a photoresistant material in semiconducto-manufacturing. The highest antimicrobial activities occurred on hydrophilic surfaces (standard exposed photoresistant films), whereas the number of attached cells was greater on hydrophobic surfaces, characterized as unexposed resistant films. These results suggest that the chemical nature of induced hydrophilicity may also be a major factor in controlling antimicrobial activity of adsorbed bacteria. Received: 5 March 1997 / Accepted: 24 August 1997     

Comparison of growth and lipid accumulation properties of two oleaginous microalgae under different nutrient conditions

This study compared the growth and lipid accumulation properties of two oleaginous microalgae, namely, Scenedesmus sp. LX1 and Chlorella sp. HQ, under different nutrient conditions. Both algal species obtained the highest biomass, lipid content and lipid yield under low-nutrient conditions （mBGll medium）. The biomass, lipid content and lipid yield of Scenedesmus sp. LX1 were 0.42g·L^-1, 22.5% and 93.8mg·L^-1, respectively. These values were relatively higher than those of Chlorella sp. HQ （0.30g·L^-1, 17.1% and 51.3mg·L^-1, respectively）. These algae were then cultivated in an SE medium that contained more nutrients; as a result, the biomass and lipid yield of Scenedesmus sp. LX1 reduced more significantly than those of Chlorella sp. HQ. Opposite results were observed in lipid and triacylglycerols （TAGs） contents. The cell sizes of both algal species under low-nutrient conditions were larger than those under high-nutrient conditions. Chlorella sp. HQ cells did not aggregate, but Scenedesmus sp. LX1 cells flocculated easily, particularly under low-nutrient conditions. In summary, low-nutrient conditions favour the growth and lipid production of both algae, but Scenedesmus sp. LX1 outperforms Chlorella sp. HQ.     

Determination and properties of actively metabolizing heterotrophic bacteria in the sea,investigated by means of micro-autoradiography

Substrate transformation and microbial biomass production in aquatic ecosystems depend mainly on the total number of actively metabolizing heterotrophic bacteria. The most common methods used concern the determination of either the colony-forming bacteria or the total number of bacteria including autotrophs and inactive organisms a micro-autoradiographic method is presented which enables the substrate uptake of single bacteria by means of ³H-amino-acid mixture and Nuclepore filters to be determined. The standardization procedure revealed the greatest success after 3 h incubation with 10 Ci/ml tritiated amino-acid mixture and an exposure of 14 days to the X-ray film. Preliminary experiments showed inactivation of an active fresh-water population from 100% to 0.6% within 3 h at 28S. With increasing distance from the shore, the number of colony-forming units decreases from 6 to 0.01% of the total number of active heterotrophic bacteria. It is concluded from the results that the fraction of very small heterotrophic bacteria which cannot be cultured on nutrient media is responsible for the continuous breakdown of organic matter in off-shore regions of the sea.     

Effects of nutrient concentration and litter cover on quantitative shoot parameters and belowground biomass of Zizania latifolia (L)

This study investigated the effect of nutrient concentration and litter cover on the development of quantitative shoot parameters and belowground biomass (BGB) production of Zizania latifolia. Zizania latifolia is a common emergent aquatic species in East Asia. Four treatments were done at the study site, and were observed between May 2003 and December 2005. The treatments are namely, high nutrient (HN) with litter cover (HNWL), high nutrient without litter cover (HNNL), low nutrient (LN) with litter cover (LNWL), and low nutrient without litter cover (LNNL). The quantitative shoot parameters and BGB had higher values for treatments with high nutrient (HN) compared to the low nutrient treatments (LN), independent of the presence of litter cover. Furthermore, the life span of the secondary shoots was also higher in HN treatments compared to LN treatments. The BGB productivity was higher in the HNNL treatment compared to the other treatments. The LNWL treatment showed the least developed quantitative shoot parameters, e.g. plant height, and the lowest BGB for Z. latifolia. It was generally observed that the combined effects of low nutrients and litter cover negatively affected shoot development and BGB production.     

The influence of storage on antibacterial activity of sea water. I. Experiments with sea water stored at 18°C

Three sea-water samples of about 25 l each, stored at 18°C in the dark, were examined for antibacterial activity and bacterial content. Serratia marinorubra mainly was used as test bacterium. During storage for 197 to 454 days, bactericidal capacity varied considerably. In raw sea water, extreme log (N ₃–N ₀) values (N: number of colony-forming test bacteria/ml after 0 and 3 days of any test) were higher than-1 and lower than-4. Antibacterial activity of filtersterilized and autoclaved, stored, sea water likewise varied, but to a lesser degree. The number of colony-forming marine bacteria fluctuated about 3 orders of magnitude. Variations in bacterial content and in bactericidal action of stored sea water were positively correlated during the first months of storage; later, this correlation became negative. From these results it is concluded that variations in nutrient concentration — due to successive periods of growth and autolysis of marine bacteria — were the primary reason for the changes observed in antibacterial activity of stored sea-water samples.     

A brominated secondary metabolite synthesized by the cyanobacterial symbiont of a marine sponge and accumulation of the crystalline metabolite in the sponge tissue

The dictyoceratid marine sponge Dysidea herbacea (Keller, 1889) is common in shallow waters of the tropical Pacific Ocean. Polybrominated biphenyl ethers such as 2-(2,4-dibromophenyl)-4,6-dibromophenol (1) are characteristic secondary metabolites of some specimens of this sponge and may represent as much as 12% of the dry weight. We have found 1 to be deposited as conspicuous crystals throughout the sponge tissue. The dominant prokaryotic endosymbiont in the mesohyl of the sponge is a filamentous cyanobacterium (Oscillatoria spongeliae), although a vacuole-containing, heterotrophic bacterium is also present. The cyanobacteria were separated from the sponge cells and heterotrophic bacteria by flow cytometry. Coupled gas chromatography—mass spectrometry and proton nuclear magnetic-resonance spectroscopy revealed that the major brominated Compound 1 isolated from the intact symbiotic association is found in the cyanobacteria and not in the sponge cells or heterotrophic bacteria. This suggests that the production of the compound is due to the cyanobacterium, and not to the sponge or symbiotic heterotrophic bacteria, as had been suggested earlier.     

The trophic role of glycolic acid in coastal seawater. II. Seasonal changes in concentration and heterotrophic use in Ipswich Bay,Massachusetts, USA

Glycolic acid is a known algal excretory product which is found in marine waters and is readily metabolized by marine bacteria. The following parameters were measured over the course of a year in Ipswich Bay: chlorophyll a, temperature, viable bacteria, heterotrophic uptake of glycolate, and glycolate concentrations. The latter two were combined to give estimates of the flux of glycolate for a station 3 km out in the bay and for an inshore station. Pronounced seasonal changes were found for all parameters. Statistically significant correlations between heterotrophic V _max and glycolic acid concentrations and temperature were found, but not between planktonic chlorophyll a and any of the other parameters. Measurements of glycolic acid flux yielded an annual flux of 2.84 g m^-2 for the bay station, which is about 0.5% of the bay phytoplankton production. Glycolic acid turns over roughly 12 times per year in the bay. It contributes perhaps as much to bacterial metabolism as any other single substrate, but is apparently not of dominating importance.     

Pyruvate-utilizing bacteria as potential contributors to the food web in the Arabian Gulf

Water samples from the Kuwaiti coasts of the Arabian Gulf were used for counting and isolating bacteria capable of growth on low molecular weight organic compounds known to be released by picocyanobacteria. The compounds tested were potassium acetate, sodium pyruvate, fumaric acid, succinic acid, sodium citrate and glycerol. For comparison, the bacterial numbers on glucose (a conventional carbon source) and Tween 80 and crude oil (unconventional carbon sources), as sole sources of carbon and energy were also determined. Sodium pyruvate was, in most cases, the carbon and energy source most commonly utilized by the cultivable surface water bacteria. The most common cultivable bacterial genera on the test carbon sources were Pseudoalteromonas, Vibrio, Cobetia and Roseobacter. Less common genera were Rhodococcus, Pseudomonas and Bacillus. Quantification of heterotrophic bacteria associated with cultures of local picocyanobacterial strains, originally isolated from the Gulf surface water, also revealed that the carbon source most commonly utilized by cultivable bacteria was sodium pyruvate. However, a different bacterial composition was identified, with Alcanivorax, Stappia and Marinobacter as the major heterotrophic genera. All heterotrophic bacteria that grew on sodium pyruvate could also grow on β-alanine, as sole sources of carbon and energy. From this study we suggest that the Arabian Gulf bacteria utilizing picocyanobacterial low molecular weight compounds, particularly pyruvate may potentially contribute to the food web in this aquatic system. The experiments comply with the current laws of Kuwait.     

Algal plating as a tool for investigating allelopathy among marine microalgae

Chemical interactions among marine microalgae were studied in cultures of pennate diatoms on agar plates. Nine marine and freshwater pennate diatoms were surveyed as potential bioassay organisms; Cylindrotheca fusiformis was most favourable for assays because of its rapid and even growth on agar. Diatom and bacterial bioassays were used to screen cell and filtrate extracts of 14 microalgal cultures. A number of these algal species, which were grown axenically, produced extractable, intracellular and/or extracellular substances that inhibited the growth of C. fusiformis. Our results suggest that the culturing of pennate diatoms on a solid medium can provide a simple bioassay for screening algal extracts which potentially contain growth inhibitors involved in microalgal allelopathy.     

不同氮浓度下三角褐指藻生长特性和化学组成

三角褐指藻是一类海洋单细胞硅藻,富含多不饱和脂肪酸,可以作为鱼、虾、贝等理想的饵料。而近年该藻曾多次在我国沿海海域发生暴发性增殖,给当地生态环境带来了一定的影响。为了探讨不同氮浓度对三角褐指藻生长特性和化学组成的影响,设置了低氮(44 μ mol.L-1)、中氮(880 μmol.L-1)和高氮(4 400 μmol.L-1)浓度三种处理,着重测定三角褐指藻的细胞密度、比生长率、生物量、可溶性糖、蛋白质含量和叶绿素含量等指标。结果表明,高氮浓度明显地促进了藻细胞的生长繁殖。高氮浓度下的藻细胞密度、比生长率和生物量分别比低氮浓度下的提高了 5.38 倍、0.81 倍和 2.86 倍。藻生长前期,高氮浓度和中氮浓度下的生长曲线相似,呈现一个"S"型的曲线。另外,高氮浓度下的藻细胞可溶性糖、蛋白质和叶绿素a含量分别是低氮浓度下的 2.5 倍、1.5 倍和 15 倍,说明高氮浓度促进了藻细胞化学组成的转化和积累。结果揭示,氮浓度可能是导致三角褐指藻近年在我国沿海海域发生暴发性增殖的重要因素。     

Antimicrobial activity of Red Sea corals

Scleractinian corals and alcyonacean soft corals are the two most dominant groups of benthic marine organisms inhabiting the coral reefs of the Gulf of Eilat, northern Red Sea. Antimicrobial assays performed with extracts of six dominant Red Sea stony corals and six dominant soft corals against marine bacteria isolated from the seawater surrounding the corals revealed considerable variability in antimicrobial activity. The results demonstrated that, while the majority (83%) of Red Sea alcyonacean soft corals exhibited appreciable antimicrobial activity against marine bacteria isolated from the seawater surrounding the corals, the stony corals had little or no antimicrobial activity. From the active soft coral species examined, Xenia macrospiculata exhibited the highest and most potent antimicrobial activity. Bioassay-directed fractionation indicated that the antimicrobial activity was due to the presence of a range of compounds of different polarities. One of these antibiotic compounds was isolated and identified as desoxyhavannahine, with a minimum inhibitory concentration (MIC) of 48 μg ml⁻¹ against a marine bacterium. The results of the current study suggest that soft and hard corals have developed different means to combat potential microbial infections. Alcyonacean soft corals use chemical defense through the production of antibiotic compounds to combat microbial attack, whereas stony corals seem to rely on other means.     

The culturable microbial community of the Great Barrier Reef sponge Rhopaloeides odorabile is dominated by an α-Proteobacterium

The microbial community cultured from the marine sponge Rhopaloeides odorabile Thompson et al. is dominated by a single bacterium, designated strain NW001. Sequence analysis of 1212 bp of the16S rRNA gene of strain NW001 indicates that it is a member of the α-subgroup of the class Proteobacteria. The association between this bacterium and its host sponge was observed in healthy R. odorabile collected from six different reefs in the Great Barrier Reef representing a geographic distance of 460 km, and in four collections in different seasons in 1997–1998 at Davies Reef (18°49.6′S; 147°34.49′E). The proportion of colonies of strain NW001 in samples from R. odorabile, expressed as a percentage of the total heterotrophic bacterial colony count, showed no significant spatial (range: 81–98%) or temporal differences (range: 81–99%), although colony counts of strain NW001 varied by up to two orders of magnitude between reef sites and sampling periods. The location of strain NW001 within the sponge mesohyl was visualized by in situ hybridization, using fluorescently labeled probes based on the 16S rRNA gene sequence of this strain. Cells of strain NW001 surround the choanocyte chambers, suggesting that these bacteria may play a role in nutrient uptake by the sponge. The absence of strain NW001 from corresponding seawater samples indicates that it has a specific, intimate relationship with R. odorabile and is not being utilized as a food source. A unique cyanobacterium related to the genera Leptolyngbya and Plectonema was also isolated from R. odorabile and characterized by 16S rRNA gene sequencing. Received: 19 May 2000 / Accepted: 18 November 2000     

Antimicrobial activity of extracts of Caribbean gorgonian corals

Extracts of 39 species of Caribbean gorgonians were tested for antimicrobial activity against 15 strains of marine bacteria. The bacteria consisted of three opportunistic pathogens, Vibrio parahaemolyticus, Leucothrix mucor, and Aerococcus viridans, and 12 strains isolated from either healthy or decayed gorgonians. Overall, only 15% (79 out of 544) of the tests resulted in antibacterial activity with 33% (13 out of 39) of the gorgonians inhibiting only one bacterial strain and 23% (9 out of 39) showing no activity. The extracts of four Pseudopterogorgia species showed relatively high levels of activity, inhibiting 43 to 86% of the bacterial strains. The potency of the active Pseudopterogorgia species was variable, however, and three additional Pseudopterogorgia species were inactive against all bacterial strains. With the exception of one sensitive strain, Vibrio species were resistant to gorgonian metabolites. Our results indicate that organic extracts of most Caribbean gorgonians do not possess potent, broad-spectrum antibacterial activity inhibitory to the growth of opportunistic marine pathogens and bacteria associated with healthy and decayed gorgonian surfaces. These findings suggest that the inhibition of bacterial growth is not the primary ecological function of gorgonian secondary metabolites and that bacteria may not be important selective agents in the evolution of gorgonian secondary chemistry.