Open access
Research Article
6 December 2017

Molecular Mechanisms for Microbe Recognition and Defense by the Red Seaweed Laurencia dendroidea


The ability to recognize and respond to the presence of microbes is an essential strategy for seaweeds to survive in the marine environment, but understanding of molecular seaweed-microbe interactions is limited. Laurencia dendroidea clones were inoculated with the marine bacterium Vibrio madracius. The seaweed RNA was sequenced, providing an unprecedentedly high coverage of the transcriptome of Laurencia, and the gene expression levels were compared between control and inoculated samples after 24, 48, and 72 h. Transcriptomic changes in L. dendroidea in the presence of V. madracius include the upregulation of genes that participate in signaling pathways described here for the first time as a response of seaweeds to microbes. Genes coding for defense-related transcription activators, reactive oxygen species metabolism, terpene biosynthesis, and energy conversion pathways were upregulated in inoculated samples of L. dendroidea, indicating an integrated defensive system in seaweeds. This report contributes significantly to the current knowledge about the molecular mechanisms involved in the highly dynamic seaweed-bacterium interactions.
IMPORTANCE Marine bacteria are part of the healthy microbiota associated with seaweeds, but some species, such as Vibrio spp., are frequently associated with disease outbreaks, especially in economically valuable cultures. In this context, the ability of seaweeds to recognize microbes and, when necessary, activate defense mechanisms is essential for their survival. However, studies dedicated to understanding the molecular components of the immune response in seaweeds are rare and restricted to indirect stimulus. This work provides an unprecedentedly large-scale evaluation of the transcriptional changes involved in microbe recognition, cellular signaling, and defense in the red seaweed Laurencia dendroidea in response to the marine bacterium Vibrio madracius. By expanding knowledge about seaweed-bacterium interactions and about the integrated defensive system in seaweeds, this work offers the basis for the development of tools to increase the resistance of cultured seaweeds to bacterial infections.


Seaweeds are extremely susceptible to microbial colonization due to the release of large amounts of carbon compounds that act as chemical attractants and nutrient sources for bacteria (1). The microbial community associated with seaweeds tends to be species specific and different from that associated with seawater (2). The microbiome associated with healthy individuals of the red seaweed Laurencia dendroidea can fix nitrogen and provide relevant amino acids and vitamins to the seaweed (3). The tight association between seaweeds and their epiphytic microbes led to the establishment of a holobiont concept that is analogous to that corresponding to the well-described microbe-coral relationship (4). However, potential pathogens were also previously detected on seaweed thalli and include microorganisms capable of degrading cell wall polysaccharides (57). Diseases can significantly impact host populations by promoting a decrease of individual fitness and negatively affecting the ability of seaweeds to defend against herbivores (8). Besides, the occurrence of disease outbreaks in valuable reared seaweeds, such as Porphyra (nori) cultures, causes significant economic losses due to a reduction of annual production (9).
Seaweed’s defense against microbes involves a multilevel strategy that starts with the recognition of microbe-associated molecular patterns (MAMPs) or pathogen-induced molecular patterns (PIMPs). Overall, MAMPs include conserved molecules that are characteristic of microbes but are absent in hosts, e.g., bacterial cell wall components (peptidoglycans, lipoteichoic acid, and lipopolysaccharides) or flagellin (10), while PIMPs are the products of the microbial degradation of seaweed cell wall matrix, including oligoagars and oligoguluronates (11). Following the recognition of microbes, evidence has emerged for the occurrence and significant role of innate immunity processes as the first line of defense in seaweeds, similarly to that observed in vascular plants and metazoans (1013), including transient production of reactive oxygen species (ROS) (1416). Besides being directly toxic to microbes (17), ROS participate in intracellular signaling mechanisms leading to the activation of other defense responses (18), such as the expression of genes related to the biosynthesis of secondary metabolites (19). Despite being part of the defensive strategy of seaweeds against fouling (20), the presence of ROS can damage the seaweed cell structures, so the oxidative burst must be tightly regulated through the activation of antioxidant enzymes (21).
Molecular studies in seaweeds have had mixed results regarding the potential costs involved in defense. For example, an increase in the expression of genes involved in cellular energy was detected through suppression subtractive hybridization (SSH) following the exposure of Laminaria digitata to oligoguluronates (19). In contrast, the downregulation of genes involved in energy conversion was detected, through a microarray, after the exposure of Chondrus crispus to methyl jasmonate (22). The conflicting results could be attributed to intrinsic biological differences between the two species or to the relatively small number of sequences analyzed.
Laurencia is a red seaweed genus widely distributed around the world, recognized for the biosynthesis of diverse halogenated secondary metabolites, especially terpenes, with relevant ecological (23, 24) and pharmacological (2529) activities. Some of these halogenated compounds are able to prevent the growth of marine bacteria (3032). Vairappan et al. (32) reported the dominance of L. majuscula during a disease outbreak; its dominance was attributed to the synthesis of secondary metabolites with antibiotic effects. Accordingly, disease symptoms were not observed in natural populations of L. dendroidea. The halogenated metabolites in L. dendroidea are stored inside vacuolar cell structures called corps en cerise (CC) (33), and they are released to the cell surface through regulated vesicle trafficking (34), which can be induced by microbes (35). The compartmentation of secondary metabolites in vacuoles, possibly to avoid autotoxicity, was previously observed in plants and other seaweeds (36, 37). However, the genes involved in this cellular process are still largely unknown. Although a large array of genes responsible for the biosynthesis of terpenes was recently characterized in L. endroidea (38), the molecular mechanisms involved in the response of Laurencia species to bacteria are still largely unknown.
Vibrio is a genus of Gram-negative bacteria associated with ice-ice disease in several red seaweeds, such as Kappaphycus alvarezii and Eucheuma denticulatum (5), and also with hole-rotten disease in the brown seaweed Laminaria japonica (39). Vibrio madracius is phylogenetically close to the V. mediterranei species (40), previously reported to cause bleaching in corals (4143). Additionally, V. madracius is associated with bleached coral (Madracis decactis) (40), indicating that this bacterial species would have a deleterious effect on the symbiotic algae. Oxidative stress resistance proteins are necessary in the pathogenic marine Vibrio species for the progression of virulence (44). Because V. madracius is oxidase and catalase positive, it may tolerate ROS defense responses and colonize algae.
Current knowledge about seaweed-microbe interactions at the molecular level is limited, because studies evaluating seaweed resistance to pathogens have been based on the use of indirect stimulus through the application of MAMPs (16), PIMPS (15, 19, 45, 46), and signaling molecules (e.g., arachidonic acid, linolenic acid, and methyl jasmonate) (22, 47, 48). Recently, an initial attempt to understand the global effects of microbes on a seaweed transcriptome was indirectly made using an agarolytic enzyme (49). Nonetheless, the direct effects of microorganisms on seaweed gene expression have rarely been evaluated and have relied on real-time PCR techniques, monitoring a limited number of genes (49, 50). The dynamic nature of seaweed’s molecular response to microbes implicates temporal complexity and metabolic shifts. Our aim was to identify the major transcriptional responses of L. endroidea in the presence of V. madracius.


The transcriptome sequencing of L. dendroidea 24 h, 48 h, and 72 h after V. madracius inoculation in the culture medium resulted in 12.58 Gbp, which represents approximately 190-fold coverage of the transcriptome of L. endroidea, considering a genome size estimate of 833 Mbp (51) and that 8% of the genetic material codes for proteins (as described for Chondrus crispus by Collén et al. [52]). After the preprocessing step, the sequences were de novo assembled, resulting in 151,740 sequences that were grouped into 53,677 clusters, which are referred to here as genes (Table 1). A total of 36.28% of the genes were shared among all of the control samples regardless of the time that had elapsed since the beginning of the experiment, while 3.79% of the genes were shared among all of the inoculated samples (see Fig. S1 in the supplemental material). We detected in both the control (uninoculated) samples and the samples of L. dendroidea inoculated with V. madracius the expression of genes coding for leucine-rich repeat receptor-like serine/threonine-protein kinase (LRR-RLK) (Fig. S2).
TABLE 1 Characteristics of the cDNA sequences from Laurencia dendroidea after preprocessing and assemblya
Ctrl. 24 h
(n = 2)
InOC. 24 h
(n = 3)
Ctrl. 48 h
(n = 2)
InOC. 48 h
(n = 2)
Ctrl. 72 h
(n = 3)
InOC. 72 h
(n = 3)
Total nucleotides (Mbp)1,9813,8562,6062,4922,1772,32891.46
No. of sequences6,016,98012,040,1248,266,3327,941,10612,588,31013,655,346151,740
Avg sequence size (bp) ± SD172.5 ± 63.4168.8 ± 59.1164.1 ± 59.5163.2 ± 58.3172.6 ± 49.4170.1 ± 49.9602 ± 674
Ctrl., uninoculated samples; InOC., inoculated samples 24 h, 48 h, and 72 h after inoculation with V. madracius; SD, standard deviation.
The number of differentially expressed genes in the seaweed L. endroidea was maximal 24 h after V. madracius inoculation, and the transcriptomic profile tended to be similar to that seen with the control condition 72 h postinoculation (hpi). The concentration of V. madracius in the culture medium was reduced progressively after 72 h in the presence of L. dendroidea (Fig. 1). Plating the seaweed tissue homogenate on thiosulfate-citrate-bile salts-sucrose (TCBS) media did not result in bacterial growth, suggesting that this reduction was not due to bacterial attachment to L. dendroidea thalli.
FIG 1 Concentration of Vibrio madracius in the culture medium in the presence (2 replicates [T1 and T2]) and absence (2 replicates [CV1 and CV2]) of Laurencia dendroidea. The concentration of V. madracius is presented as the number of colony-forming units per milliliter of culture medium as measured for 144 h after bacterial inoculation (average ± standard error).
The comparative analysis of control and inoculated specimens of L. dendroidea revealed the change in the gene expression profile in response to V. madracius. Most of the genes differentially expressed were upregulated in the inoculated samples of L. dendroidea, especially 24 and 48 hpi, while we verified a significant reduction in the number of genes differentially expressed in L. dendroidea 72 h after V. madracius inoculation. Overall, at 24 hpi, we observed in L. dendroidea the upregulation of 675 genes, of which 75.8% were annotated and 6 (16.7% annotated) were downregulated (Fig. 2A). In addition, 48 h after V. madracius inoculation, 299 genes were upregulated, of which 82.3% were annotated and 4 (annotated as encoding hypothetical proteins) were downregulated (Fig. 2B). Finally, 72 h after the introduction of V. madracius in the culture medium, the expression level of 5 genes increased, but none of them were identified through Blast, and 5 genes were repressed, of which 60% were annotated at the protein family level at least (Fig. 2C).
FIG 2 Heat map of expression values (Z score) for differentially expressed genes in Laurencia dendroidea 24 h (A), 48 h (B), and 72 h (C) after Vibrio madracius inoculation. Both annotated and nonannotated genes are represented. The analysis was based on the following numbers of replicates: control 24 h = 2, inoculated 24 h = 3, control 48 h = 2, inoculated 48 h = 2, control 72 h = 3, inoculated 72 h = 3.
The gene coding for NADPH oxidase (NADPH ox), which is responsible for transient production of ROS, was upregulated in L. dendroidea 24 hpi (Fig. 3a). At 24 and 48 hpi, we also observed the upregulation of the genes coding for antioxidant enzymes, such as thioredoxin (TRX), peroxiredoxin (PRX), glutathione S-transferase (GST), and superoxide dismutase (SOD) (Fig. 3a), and of genes associated with protein folding (Fig. S4a).
FIG 3 Relevant differentially expressed genes in Laurencia dendroidea 24, 48, and 72 h after inoculation with Vibrio madracius (data represent logFC values considering “inoculated” versus “control” samples). (a) Products of genes involved in oxidative burst and antioxidant mechanisms are indicated as follows: NADPH oxidase, NADPH ox; thioredoxin, TRX; peroxiredoxin, PRX; glutathione S-transferase, GST; superoxide dismutase, SOD. (b) Products of genes involved in the MAPK cascade and small GTPase-mediated signaling and transcription factors are indicated as follows: mitogen-activated protein kinase kinase, MAPKK; Rho-related protein, Rac1; transcription factor WRKY; transcription factor MYB; ethylene-responsive transcription factor, ERF; heat stress transcription factor, HSF. (c) Products of genes related to phosphoinositide and calcium-dependent signaling are indicated as follows: phosphatidylinositol 4-phosphate 5-kinase, PIP5K; myo-inositol 1-phosphate synthase, MIPS; type II inositol 1,4,5-trisphosphate 5-phosphatase, 5PTase; phosphatidylinositol 4-kinase, P4K; calmodulin, CaM; calcium calmodulin-dependent protein kinase, CDPK; Snf1-related protein kinase, SnRK. (d) Products of genes that participate in the biosynthesis of terpenes are indicated as follows: acetyl-CoA C-acetyltransferase, ACAT; (+)-neomenthol dehydrogenase, NMD; (-)-isopiperitenol dehydrogenase, ISPD. (e) Products of genes involved in vesicle trafficking are indicated as follows: Rab GTPase, Rab; ADP-ribosylation factor, Arf; coatomer, coat α-2; clathrin, CLT; actin, tubulin. (f) Products of genes involved in glycolysis are indicated as follows: glucose-6-phosphate isomerase, G6PI; fructose-bisphosphate aldolase, FBA; glyceraldehyde-3-phosphate dehydrogenase, GAPDH; phosphopyruvate hydratase, PPH. (g) Products of genes involved in tricarboxylic acid cycle and oxidative phosphorylation are indicated as follows: succinyl-CoA ligase, SCS; succinate dehydrogenase, SDH; citrate synthase, CIT; cytochrome c oxidase, Cox; NADH-ubiquinone oxidoreductase, complex I; electron transfer flavoprotein, ETF; ATP synthase. (h) Products of genes related to fatty acid oxidation and branched-chain amino acid catabolism are indicated as follows: enoyl-CoA hydratase, ECH; 3-ketoacyl-CoA thiolase, 3KCT; isovaleryl-CoA dehydrogenase, IVD; 3-hydroxyisobutyryl-CoA hydrolase, HIBCH; propionyl-CoA carboxylase, PCC. Open circles indicate values of logFC that were not statistically significant (P value = >0.001; logFC = <|2.0|). Numbers of replicates were as follows: control 24 h = 2, inoculated 24 h = 3, control 48 h = 2, inoculated 48 h = 2, control 72 h = 3, inoculated 72 h = 3.
The gene coding for a mitogen-activated protein kinase kinase (MAPKK) was upregulated 24 hpi (Fig. 3b). Another relevant biological process overrepresented at 24 and 48 h after V. madracius inoculation was “small GTPase-mediated signal transduction” (Fig. S3), which included Rho-related protein rac1 (Fig. 3b). Additionally, genes coding for phosphatidylinositol 4-phosphate 5-kinase (PIP5K), myo-inositol 1-phosphate synthase (MIPS), type II inositol 1,4,5-trisphosphate 5-phosphatase (5PTase), phosphatidylinositol 4-kinase (P4K), calmodulin (CaM), calcium calmodulin-dependent protein kinase (CDPK), and Snf1-related protein kinase (SnRK) were upregulated mainly 24 hpi (Fig. 3c). The genes coding for WRKY, MYB, ethylene-responsive transcription factor (ERF), and heat stress transcription factor (HSF) were upregulated 24 h and 48 hpi (Fig. 3b).
Several genes related to the biosynthesis of terpenes were also upregulated in L. dendroidea in response to V. madracius, such as the genes coding for acetyl-CoA C-acetyltransferase (ACAT) 24 and 48 h after inoculation and genes homologous to those coding for plant (-)-isopiperitenol dehydrogenase (ISPD) 24 hpi and (+)-neomenthol dehydrogenase (NMD) 24 and 48 hpi (Fig. 3d). Twelve genes involved in the biosynthesis of terpenoid backbones and 10 genes involved in the biosynthesis of monoterpenes (C10), sesquiterpenes (C15), diterpenes (C20), and triterpenes (C30) were detected for the first time in Laurencia (Table 2). Genes coding for the Ras-related protein Rab, ADP-ribosylation factor (Arf), coatomer (coat α-2), and clathrin (CLT) were distributed in the categories “transport” and “intracellular protein transport” and upregulated in the seaweed in response to V. madracius (Fig. 3e, Fig. S3).
TABLE 2 Genes related to the biosynthesis of terpenes characterized for the first time in Laurencia dendroidea with their EC number, Blast E value, identity, and similarity and the metabolic pathway in which they participate
Gene productEC no.Blast e-value% identity% similarityBiosynthetic pathway
Hydroxymethylglutaryl-CoA synthase2. e−483656Terpenoid backbone
Hydroxymethylglutaryl-CoA reductase1.1.1.34/ e−617387Terpenoid backbone
Phosphomevalonate kinase2. e−593244Terpenoid backbone
Diphosphomevalonate decarboxylase4.1.1.333.00 e−995569Terpenoid backbone
Isopentenyl phosphate kinase2. e−202649Terpenoid backbone
(2Z,6E)-farnesyl diphosphate synthase2.5.1.683.00 e−804463Terpenoid backbone
(2E,6E)-farnesyl diphosphate synthase2. e−954765Terpenoid backbone
Prenylcysteine oxidase1. e−933756Terpenoid backbone
Hexaprenyl diphosphate synthase
(geranylgeranyl-diphosphate specific) e−604359Terpenoid backbone
Heptaprenyl diphosphate synthase2.5.1.302.00 e−144363Terpenoid backbone
Undecaprenyl diphosphate synthetase2.5.1.311.00 e−434964Terpenoid backbone
All-trans-octaprenyl-diphosphate synthase2.5.1.902.00 e−283859Terpenoid backbone
Linalool 8-monooxygenase1.14.13.1514.00 e−133558Monoterpenoid
(-)-Isopiperitenol dehydrogenase1.1.1.2239.00 e−243350Monoterpenoid
(+)-Menthofuran synthase1.14.13.1045.00 e−193047Monoterpenoid
(+)-Neomenthol dehydrogenase1.1.1.2084.00 e−233149Monoterpenoid
Germacrene a hydroxylase1.14.13.1231.00 e−292850Sesquiterpenoid
Ent-cassa-12,15-diene 11-hydroxylase1.14.13.1455.00 e−103857Diterpenoid
Ent-kaurene oxidase1.14.13.784.00 e−224053Diterpenoid
Ent-kaurenoic acid oxidase1.14.13.793.00E-63859Diterpenoid
Squalene monooxygenase1. e−855067Triterpenoid
11-Oxo-beta-amyrin 30-oxidase1.14.13.1736.00 e−382743Triterpenoid
Functional categories associated with energy conversion, such as the glycolytic process, including glucose-6-phosphate isomerase (G6PI), fructose-bisphosphate aldolase (FBA), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and phosphopyruvate hydratase (PPH), were overrepresented in the transcriptome of L. dendroidea 24 h and 48 h after the introduction of V. madracius in the culture medium (Fig. 3f). Further, genes related to the tricarboxylic acid cycle and oxidative phosphorylation, e.g., those coding for succinyl-CoA ligase (SCS), succinate dehydrogenase (SDH), citrate synthase (CIT), cytochrome c oxidase (Cox), NADH-ubiquinone oxidoreductase (complex I), electron transfer flavoprotein (ETF), and ATP synthase, were upregulated in L. dendroidea 24 and 48 hpi (Fig. 3g, Fig. S4b and c). Finally, at 24 and 48 hpi, we detected the upregulation of genes related to fatty acid oxidation and the catabolism of leucine, isoleucine, and valine, such as those coding for enoyl-CoA hydratase (ECH), 3-ketoacyl-CoA thiolase (3KCT), isovaleryl-CoA dehydrogenase (IVD), 3-hydroxyisobutyryl-CoA hydrolase (HIBCH), and propionyl-CoA carboxylase (PCC) (Fig. 3h, Fig. S4d).


Microbial recognition and ROS production.

The concentration of V. madracius reduced progressively after 72 h in the presence of L. dendroidea. Because this reduction could not be attributed to biofilm formation, we hypothesized that it should have been due to responses of or defense strategies activated in L. dendroidea in the first 72 h after V. madracius inoculation, as a consequence of recognizing the bacteria through specific membrane receptors. Pattern recognition receptors are largely unknown in seaweeds. A recent study demonstrated the occurrence of genes coding for LRR kinases in the brown seaweed Ectocarpus siliculosus that, due to their molecular structure, were considered to represent candidate pathogen receptors (53). Here, we detected, in both control and inoculated samples, the expression of genes coding for LRR-RLKs, representing a major class of receptors involved in microbe detection in plants through the recognition of MAMPs (54), suggesting that these genes are constitutively expressed in the red seaweed L. dendroidea.
Moreover, at 24 hpi, we verified the upregulation of the gene coding for NADPH oxidase, the major gene for ROS production in seaweeds, in L. dendroidea (14, 47, 55). Because ROS can react with essential host molecules, the activity of antioxidant enzymes is important to limit the oxidative burst. In this work, we report the upregulation of several antioxidant enzymes, especially TRX, PRX, GST, and SOD, 24 and 48 hpi. Accordingly, the expression of TRX, PRX, and GST increased in Laminaria digitata in response to oligoguluronates (19, 56) and the activity of SOD increased in Saccharina japonica as elicited with flg22, a MAMP (57).

Activation of defense-related intracellular signaling cascades and transcription factors.

Signaling cascades that modulate the innate immune response have been well described in plants but are still unknown in seaweeds. Despite indirect evidence for the occurrence of mitogen-activated protein kinase (MAPK) cascades in seaweeds (49), the involvement of this pathway in the response to bacteria was not previously investigated. Here, we detected the upregulation in L. dendroidea of a gene coding for a MAPKK 24 hpi, indicating that a MAPK cascade was induced during the response of this seaweed to V. madracius. The MAPK cascade transduces extracellular stimuli into intracellular responses during plant defense against pathogens and can induce the expression of defense-related genes through the phosphorylation of transcription factors, such as ERF (58).
Further, we observed the upregulation of L. dendroidea genes coding for small GTPases, such as Rac, a member of the Rho family considered to be a key regulator in plant immunity, 24 hpi (59). The Rac1 homolog of rice is a regulator of ROS production and induces the expression of defense-related genes promoting resistance against pathogenic bacteria (60). Genes involved in PI signaling were also upregulated in L. dendroidea 24 hpi. Phosphoinositide-mediated signaling affects Ca2+ release and the expression of defense-related genes in plants (61). Indeed, we detected the upregulation of genes coding for CaM and CDPK 24 and 48 hpi which are required for sensing and decoding Ca2+ signals. Pathogenesis-related activation of CDPK was detected in plants (62), and this protein kinase regulates the production of ROS by NADPH ox (63). Another gene coding for a protein kinase upregulated in L. dendroidea 24 hpi was the Snf1-related protein kinase, whose expression in plants is induced by pathogenic bacteria (64). Further, a relevant role was attributed to Snf1-related protein kinases as global regulators of gene expression, inducing catabolic pathways that provide alternative sources of energy and controlling genes that encode signal transduction components and transcription regulators (65). Our work suggests that well-known mechanisms acting on the plant innate immunity response are also present in seaweeds (Fig. 4).
FIG 4 Hypothetical model representing bacterium recognition (through microbe-associated molecular pattern [MAMP]) and some relevant metabolic processes overrepresented in the transcriptomic profile of Laurencia dendroidea in response to Vibrio madracius. LRR (RLK), leucine-rich repeat receptor-like serine/threonine-protein kinase; ROS, reactive oxygen species; PIs, phosphatidylinositol signaling; Rac, Rho family GTPase Rac; CaM, calmodulin; CDPK, calcium calmodulin-dependent protein kinase; TF, transcription factors; Ran, nuclear protein Ran; TI, translation initiation factors; CC, corps en cerise; Arf, ADP-ribosylation factor; Rab, Rab GTPase; TCA, tricarboxylic acid. Note that the figure is not drawn to scale.
WRKY and MYB were upregulated in L. dendroidea 24 h after the inoculation of V. madracius, reinforcing the role of these transcriptional activators positively regulating genes related to immunity (66, 67). Similarly, both transcription factors were upregulated 12 h after peach leaves were inoculated with a pathogenic bacterial species (68) and MYB expression was induced 24 h after the inoculation of Arabidopsis with a pathogenic fungus (69). Another important transcriptional regulatory element upregulated in L. dendroidea 24 hpi was heat stress transcription factor (HSF), associated mainly with defense gene activation, pathogen-induced systemic acquired resistance (86), and transcriptional reprogramming in plants as a consequence of redirecting energy resources from growth to defense mechanisms (87).
Additionally, at 24 hpi, we detected the upregulation of an ERF that has diverse functions in plant defense and responds to jasmonic acid (JA) and ethylene (ET) (70, 71). There is evidence that JA, or a structurally similar compound(s), is also involved in defense signals in macroalgae, as this substance induced the expression of stress-related genes in C. crispus (22), increased the biosynthesis of phlorotannins in Fucus vesiculosus (72), and activated oxidative cascades in Laminaria digitata (47) and C. crispus (12). Although the role of ET signaling in seaweeds was not demonstrated, the ability to synthesize and respond to this plant hormone was previously detected in Enteromorpha intestinalis (73) and Pterocladiella capillacea (74). The present report contributes to evidence indicating the presence of a mechanism in seaweeds similar to plant hormone-regulated defense against microbes.

Energy balance.

Diverse evidence suggests that fighting against microbes is energetically demanding in vascular plants (75). By using high-throughput transcriptome sequencing, we verified the transient upregulation, in response to V. madracius, of L. dendroidea genes involved in energy conversion, especially in relation to glycolysis, the tricarboxylic acid (TCA) cycle, and oxidative phosphorylation (Fig. 4). Further, we observed the upregulation of genes involved in the catabolism of branched-chain amino acids and in the β-oxidation of fatty acids, which provide alternative sources of respiratory substrates for the TCA cycle, especially during severe plant stress and in response to infection (76, 77).

Secondary metabolites and defense.

The expression level of several genes involved in the biosynthesis of terpenes in L. dendroidea increased significantly 24 and 48 h after V. madracius inoculation. Terpenoid compounds are recognized as important secondary metabolites acting to defend Laurencia species against bacterial colonization (30). Indeed, acetyl-CoA C-acetyltransferase (overexpressed 24 and 48 hpi) catalyzes the first step in the biosynthesis of terpenoid backbones through the mevalonate pathway and was suggested to be a regulatory enzyme in isoprenoid biosynthesis during plant abiotic stress adaptation (78). Moreover, the upregulation of genes involved in monoterpene biosynthesis was detected in L. dendroidea in response to V. madracius and offers a possible explanation for the reduction in the concentration of these bacteria in the culture medium in the presence of the seaweed.
Genes relevant for vesicle trafficking—including those coding for Rab, which participates in intracellular membrane trafficking by regulating the movement of vesicles along cytoskeletal filaments (79); actin, which composes the structure of the connections linking the CC to the cell periphery in L. dendroidea; and tubulin, which is responsible for the positioning of the vesicles toward exocytosis sites—were upregulated in this seaweed in response to the inoculation of V. madracius, mainly 24 hpi (34). These findings may corroborate the occurrence of increased vesicle transport in Laurencia as a response to microbes (35).
The present report shows that even though V. madracius cannot be considered a pathogen of L. dendroidea, this seaweed is able to recognize and respond to the microbe through a temporal series of complex metabolic changes. Although we might expect the upregulation of genes related to microbe recognition, signaling, and oxidative burst to precede the upregulation of genes involved in terpene biosynthesis, future studies are needed to explore the timing of the expression of gene groups in shorter time periods (in the window of 24 h after bacterial inoculation). It is also necessary to determine if the measured differences represented a generic response of Laurencia to bacteria or a response to a specific potential pathogen.


The response of L. dendroidea to V. madracius involves transcriptomic reprogramming, especially 24 and 48 h after bacterial inoculation. The upregulation of genes coding for NADPH oxidase and antioxidant enzymes suggests the occurrence of an oxidative burst. Intracellular signaling mediated by a MAPK cascade, small GTPases, phosphatidylinositol, and calcium calmodulin-dependent protein kinases was observed as a seaweed response to bacteria. Further, the upregulation of genes related to the biosynthesis of terpenes, along with the overexpression of genes involved in vesicular transport, suggests increased release of terpenes by L. dendroidea. Finally, we verified the upregulation of genes associated with energy metabolism, indicating that the defense mechanisms in L. dendroidea might involve an energy cost. The upregulation of the genes involved in ROS production and in the biosynthesis of terpenes reveals a previously unknown integrated defensive system in seaweeds. The present study provided novel insights into the complexity of seaweed-microbe interactions and the defensive strategies of L. dendroidea at the molecular level.


Laurencia dendroidea (Hudson) J. V. Lamouroux was sampled at Castelhanos Beach in Anchieta municipality, Espírito Santo state (20°51′40″S, 40°37′00″W), and was maintained in a laboratory. The unialgal culture of this seaweed was established through successive excision of the apices. Clones were used to prevent intraspecific variations in transcriptomic profiles from masking the effect of bacterial inoculation. These algal clones were treated with 100 µg/ml ampicillin, 120 µg/ml streptomycin, and 60 µg/ml gentamicin, which reduced the levels of bacteria in the culture by more than 95%. The clones were grown in sterile seawater with germanium dioxide (1 mg/liter) and 50% Provasoli solution (enriched seawater medium [ESW]) for 2 days before the experiment. The culture and experimental conditions were as follows: temperature, 22 ± 1°C; salinity, 32 ± 1; irradiance, 80 ± 5 µmol photons ⋅ m−2 ⋅ s−1; 14 h light/10 h dark.
Vibrio madracius was isolated from the coral Madracis decactis sampled in Saint Peter and Saint Paul archipelago (40). The bacteria were grown at 30°C in sterile marine broth to an optical density at 600 nm (OD600) of 0.8, corresponding to 108 CFU ⋅ ml−1, and precipitated for 5 min at 3,000 rpm (5415R centrifuge; Eppendorf). The supernatant was discarded, and the pellet was resuspended in sterile seawater and inoculated in Falcon tubes containing 250 mg of L. dendroidea and 40 ml of ESW (n = 2). The final concentration of V. madracius in the treatment was 107 CFU ⋅ ml−1 (i.e., in the presence of L. dendroidea; replicates T1 and T2). The same quantity of bacteria was inoculated in Falcon tubes containing 40 ml of ESW (n = 2) in the absence of L. dendroidea (CV1 and CV2). The culture medium was plated in TCBS media (n = 3) immediately after bacterial inoculation and 24, 48, 72, 96, and 144 h after bacterial inoculation in the presence and absence of L. dendroidea. Also, 144 h after bacterial inoculation, the seaweed thalli were homogenized in a sterile 3% NaCl solution for 1 h using vortex mixing and this tissue homogenate was plated in TCBS media. The petri dishes were incubated overnight at 30°C, and the colonies of V. madracius were counted when present.
To evaluate the transcriptomic profile of L. dendroidea in the presence and absence of V. madracius, control tubes were set up with 250 mg of L. dendroidea and 40 ml of ESW (n = 3); the inoculated tubes contained 250 mg of L. dendroidea, 40 ml of ESW, and V. madracius at 107 CFU ml−1 (n = 3). After 24, 48, and 72 h, control and inoculated L. dendroidea specimens were frozen and separately ground in liquid nitrogen using a mortar and pestle. Total RNA was extracted using the TRIzol (Life Technologies, Inc.) protocol. Double-strand cDNA libraries were prepared using a TruSeq stranded mRNA LT sample preparation kit (Illumina). Library size distribution was accessed using a model 2100 Bioanalyzer (Agilent) and a High Sensitivity DNA kit (Agilent). The accurate quantification of the libraries was accomplished using model 7500 real-time PCR (Applied Biosystems) and a Kapa library quantification kit (Kapa Biosystems). Paired-end sequencing (2 × 250 bp) was performed on a MiSeq sequencer (Illumina) for the following numbers of replicates: control 24 h = 2, inoculated 24 h = 3, control 48 h = 2, inoculated 48 h = 2, control 72 h = 3, inoculated 72 h = 3.
The sequences were preprocessed to trim poly(A-T) tails that were at least 20 bp long, to remove reads shorter than 35 bp, and to trim sequences with a quality score lower than Phred 30, using Prinseq software (80). The processed sequences from all of the samples were assembled using Trinity software, and sequences larger than 199 bp were used in the downstream analysis. Sequences from each sample were mapped against the assembled reads using Bowtie 2 (81) (with the following parameters: --end-to-end; --no-mixed; --no-discordant; --score-min L,-0.1,-0.1) and were clustered into genes using Corset software (minimum read count = 5) (82). A few bacterial sequences were detected through Blast searches against the NCBI-nr database and were removed from subsequent analysis. Statistically relevant genes differentially expressed between the control and the inoculated samples were identified using the edgeR software package associated with the Fisher exact test and Bonferroni correction for multiple tests, considering the following parameters: corrected P value, ≤0.001; log fold change [logFC] value, ≥2.0 (83). To plot a heat map of gene expression levels comparing control and inoculated samples (Fig. 2), we used Z score analysis, a conventional method of data normalization that calculates the mean expression value for a gene under the different conditions and normalizes the deviations as a function of the mean. The differentially expressed genes were annotated through a Blast search against the NCBI-nr database (E value, <10−5), and GO terms were assigned using the Blast2go tool (84). To identify the transcripts associated with the biosynthesis of terpenoid compounds, we analyzed the transcriptome of L. dendroidea using hidden Markov models generated from the alignment of sequences available in the KEGG database through the use of HMMER 3.0 software (85), following the method previously used by de Oliveira et al. (38). The sequences matching these profiles were annotated through a Blast search against the NCBI-nr, PlantCyc, and Uniprot databases. The functional identifications were manually confirmed.


This paper is part of the DSc requirements of Louisi Souza de Oliveira at the Biodiversity and Evolutionary Biology Graduate Program of the Federal University of Rio de Janeiro.
This research received the financial support of CAPES, CNPq, and FAPERJ. F.L.T. and R.C.P. thank CNPq for their Research Productivity Fellowships. The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
L.S.D.O. performed experiments, RNA extraction, and sequencing and bioinformatics analysis and drafted the manuscript. D.A.T. participated in the bioinformatics analysis and in the discussions and drafting of the manuscript. A.C.R.M.L. participated in the experiments and in the drafting of the manuscript. D.B.S. performed the isolation of clones of Laurencia dendroidea and participated in the discussion of the results and drafting of the manuscript. P.M.M. participated in the bioinformatics analysis and drafting of the manuscript. C.C.T. participated in the acquisition of funding and drafting of the manuscript. R.C.P. participated in the acquisition of funding, work planning, discussion of the results, and drafting of the manuscript. F.L.T. participated in the acquisition of funding, work planning, discussion of the results, and drafting of the manuscript. All of us read and approved the final manuscript.

Supplemental Material

File (sph006172418sf1.eps)
File (sph006172418sf2.tif)
File (sph006172418sf3.jpg)
File (sph006172418sf4.eps)
ASM does not own the copyrights to Supplemental Material that may be linked to, or accessed through, an article. The authors have granted ASM a non-exclusive, world-wide license to publish the Supplemental Material files. Please contact the corresponding author directly for reuse.


Haas AF, Nelson CE, Wegley Kelly L, Carlson CA, Rohwer F, Leichter JJ, Wyatt A, and Smith JE. 2011. Effects of coral reef benthic primary producers on dissolved organic carbon and microbial activity. PLoS One6:e27973.
Burke C, Steinberg P, Rusch D, Kjelleberg S, and Thomas T. 2011. Bacterial community assembly based on functional genes rather than species. Proc Natl Acad Sci U S A108:14288–14293.
de Oliveira LS, Gregoracci GB, Silva GGZ, Salgado LT, Filho GA, Alves-Ferreira M, Pereira RC, and Thompson FL. 2012. Transcriptomic analysis of the red seaweed Laurencia dendroidea (Florideophyceae, Rhodophyta) and its microbiome. BMC Genomics13:487.
Egan S, Harder T, Burke C, Steinberg P, Kjelleberg S, and Thomas T. 2013. The seaweed holobiont: understanding seaweed-bacteria interactions. FEMS Microbiol Rev37:462–476.
Largo DB, Fukami K, and Nishijima T. 1995. Occasional pathogenic bacteria promoting ice-ice disease in the carrageenan-producing red algae Kappaphycus alvarezii and Eucheuma denticulatum (Solieriaceae, Gigartinales, Rhodophyta). J Appl Phycol7:545–554.
Jaffray AE, Anderson RJ, and Coyne VE. 1997. Investigation of bacterial epiphytes of the agar-producing red seaweed Gracilaria gracilis (Stackhouse) Steentoft, Irvine et Farnham from Saldanha Bay, South Africa and Lüderitz, Namibia. Bot Mar40:569–576.
Sakai T, Ishizuka K, and Kato I. 2003. Isolation and characterization of a fucoidan-degrading marine bacterium. Mar Biotechnol5:409–416.
Campbell AH, Vergés A, and Steinberg PD. 2014. Demographic consequences of disease in a habitat-forming seaweed and impacts on interactions between natural enemies. Ecology95:142–152.
Ding H and Ma J. 2005. Simultaneous infection by red rot and chytrid diseases in Porphyra yezoensis Ueda. J Appl Phycol17:51–56.
Nürnberger T, Brunner F, Kemmerling B, and Piater L. 2004. Innate immunity in plants and animals: striking similarities and obvious differences. Immunol Rev198:249–266.
Weinberger F. 2007. Pathogen-induced defense and innate immunity in macroalgae. Biol Bull213:290–302.
Bouarab K, Adas F, Gaquerel E, Kloareg B, Salaün JP, and Potin P. 2004. The innate immunity of a marine red alga involves oxylipins from both the eicosanoid and octadecanoid pathways. Plant Physiol135:1838–1848.
Egan S, Fernandes ND, Kumar V, Gardiner M, and Thomas T. 2014. Bacterial pathogens, virulence mechanism and host defence in marine macroalgae. Environ Microbiol16:925–938.
Weinberger F, Leonardi P, Miravalles A, Correa JA, Lion U, Kloareg B, and Potin P. 2005. Dissection of two distinct defense-related responses to agar oligosaccharides in Gracilaria chilensis (Rhodophyta) and Gracilaria conferta (Rhodophyta). J Phycol41:863–873.
Küpper FC, Müller DG, Peters AF, Kloareg B, and Potin P. 2002. Oligoalginate recognition and oxidative burst play a key role in natural and induced resistance of sporophytes of Laminariales. J Chem Ecol28:2057–2081.
Küpper FC, Gaquerel E, Boneberg EM, Morath S, Salaün JP, and Potin P. 2006. Early events in the perception of lipopolysaccharides in the brown alga Laminaria digitata include an oxidative burst and activation of fatty acid oxidation cascades. J Exp Bot57:1991–1999.
Mellersh DG, Foulds IV, Higgins VJ, and Heath MC. 2002. H2O2 plays different roles in determining penetration failure in three diverse plant-fungal interactions. Plant J29:257–268.
Schmitt FJ, Renger G, Friedrich T, Kreslavski VD, Zharmukhamedov SK, Los DA, Kuznetsov VV, and Allakhverdiev SI. 2014. Reactive oxygen species: re-evaluation of generation, monitoring and role in stress-signaling in phototrophic organisms. Biochim Biophys Acta1837:835–848.
Cosse A, Potin P, and Leblanc C. 2009. Patterns of gene expression induced by oligoguluronates reveal conserved and environment-specific molecular defense responses in the brown alga Laminaria digitata. New Phytol182:239–250.
da Gama BAP, Plouguerné E, and Pereira RC. 2014. The antifouling defence mechanisms of marine macroalgae. Adv Bot Res71:413–440.
Dring MJ. 2005. Stress resistance and disease resistance in seaweeds: the role of reactive oxygen metabolism. Adv Bot Res43:175–207.
Collén J, Hervé C, Guisle-Marsollier I, Léger JJ, and Boyen C. 2006. Expression profiling of Chondrus crispus (Rhodophyta) after exposure to methyl jasmonate. J Exp Bot57:3869–3881.
Da Gama BAP, Pereira RC, Carvalho AGV, Coutinho R, and Yoneshigue-Valentin Y. 2002. The effects of seaweed secondary metabolites on biofouling. Biofouling18:13–20.
Pereira RC, Da Gama BA, Teixeira VL, and Yoneshigue-Valentin Y. 2003. Ecological roles of natural products of the Brazilian red seaweed Laurencia obtusa. Braz J Biol63:665–672.
Vairappan CS, Kawamoto T, Miwa H, and Suzuki M. 2004. Potent antibacterial activity of halogenated compounds against antibiotic-resistant bacteria. Planta Med70:1087–1090.
Dos Santos AO, Veiga-Santos P, Ueda-Nakamura T, Filho BPD, Sudatti DB, Bianco EM, Pereira RC, and Nakamura CV. 2010. Effect of elatol, isolated from red seaweed Laurencia dendroidea, on Leishmania amazonensis. Mar Drugs8:2733–2743.
Veiga-Santos P, Pelizzaro-Rocha KJ, Santos AO, Ueda-Nakamura T, Dias Filho BP, Silva SO, Sudatti DB, Bianco EM, Pereira RC, and Nakamura CV. 2010. In vitro anti-trypanosomal activity of elatol isolated from red seaweed Laurencia dendroidea. Parasitology137:1661–1670.
Chatter R, Ben Othman R, Rabhi S, Kladi M, Tarhouni S, Vagias C, Roussis V, Guizani-Tabbane L, and Kharrat R. 2011. In vivo and in vitro anti-inflammatory activity of neorogioltriol, a new diterpene extracted from the red algae Laurencia glandulifera. Mar Drugs9:1293–1306.
Lhullier C, Falkenberg M, Ioannou E, Quesada A, Papazafiri P, Horta PA, Schenkel EP, Vagias C, and Roussis V. 2010. Cytotoxic halogenated metabolites from the Brazilian red alga Laurencia catarinensis. J Nat Prod73:27–32.
Vairappan CS, Daitoh M, Suzuki M, Abe T, and Masuda M. 2001. Antibacterial halogenated metabolites from the Malaysian Laurencia species. Phytochemistry58:291–297.
Vairappan CS. 2003. Potent antibacterial activity of halogenated metabolites from Malaysian red algae, Laurencia majuscula (Rhodomelaceae, Ceramiales). Biomol Eng20:255–259.
Vairappan CS, Anangdan SP, Tan KL, and Matsunaga S. 2010. Role of secondary metabolites as defense chemicals against ice-ice disease bacteria in biofouler at carrageenophyte farms. J Appl Phycol22:305–311.
Salgado LT, Viana NB, Andrade LR, Leal RN, da Gama BAP, Attias M, Pereira RC, and Amado Filho GM. 2008. Intra-cellular storage, transport and exocytosis of halogenated compounds in marine red alga Laurencia obtusa. J Struct Biol162:345–355.
Reis VM, Oliveira LS, Passos RMF, Viana NB, Mermelstein C, Sant’anna C, Pereira RC, Paradas WC, Thompson FL, Amado-Filho GM, and Salgado LT. 2013. Traffic of secondary metabolites to cell surface in the red alga Laurencia dendroidea depends on a two-step transport by the cytoskeleton. PLoS One8:e63929.
Paradas WC, Salgado LT, Sudatti DB, Crapez MA, Fujii MT, Coutinho R, Pereira RC, and Amado Filho GM. 2010. Induction of halogenated vesicle transport in cells of the red seaweed Laurencia obtusa. Biofouling26:277–286.
Paul NA, Cole L, de Nys R, and Steinberg PD. 2006. Ultrastructure of the gland cells of the red alga Asparagopsis armata (Bonnemaisoniaceae). J Phycol42:637–645.
Dworjanyn SA, De Nys R, and Steinberg PD. 1999. Localisation and surface quantification of secondary metabolites in the red alga Delisea pulchra. Mar Biol133:727–736.
de Oliveira LS, Tschoeke DA, de Oliveira AS, Hill LJ, Paradas WC, Salgado LT, Thompson CC, Pereira RC, and Thompson FL. 2015. New insights on the terpenome of the red seaweed Laurencia dendroidea (Florideophyceae, Rhodophyta). Mar Drugs13:879–902.
Wang G, Shuai L, Li Y, Lin W, Zhao X, and Duan D. 2008. Phylogenetic analysis of epiphytic marine bacteria on hole-rotten diseased sporophytes of Laminaria japonica. J Appl Phycol20:403–409.
Moreira APB, Duytschaever G, Tonon LAC, Dias GM, Mesquita M, Cnockaert M, Francini-Filho RB, De Vos P, Thompson CC, and Thompson FL. 2014. Vibrio madracius sp. nov. isolated from Madracis decactis (Scleractinia) in St Peter & St Paul Archipelago, mid-Atlantic Ridge, Brazil. Curr Microbiol69:405–411.
Ben-Haim Y, Banim E, Kushmaro A, Loya Y, and Rosenberg E. 1999. Inhibition of photosynthesis and bleaching of zooxanthellae by the coral pathogen Vibrio shiloi. Environ Microbiol1:223–229.
Kushmaro A, Banin E, Loya Y, Stackebrandt E, and Rosenberg E. 2001. Vibrio shiloi sp. nov., the causative agent of bleaching of the coral Oculina patagonica. Int J Syst Evol Microbiol51:1383–1388.
Thompson FL, Hoste B, Thompson CC, Huys G, and Swings J. 2001. The coral bleaching Vibrio shiloi Kushmaro et al. 2001 is a later synonym of Vibrio mediterranei Pujalte and Garay 1986. Syst Appl Microbiol24:516–519.
Ma L, Chen J, Liu R, Zhang XH, and Jiang YA. 2009. Mutation of rpoS gene decreased resistance to environmental stresses, synthesis of extracellular products and virulence of Vibrio anguillarum. FEMS Microbiol Ecol70:130–136.
Küpper FC, Kloareg B, Guern J, and Potin P. 2001. Oligoguluronates elicit an oxidative burst in the brown algal kelp Laminaria digitata. Plant Physiol125:278–291.
Tonon T, Rousvoal S, Roeder V, and Boyen C. 2008. Expression profiling of the mannuronan C5-epimerase multigenic family in the brown alga Laminaria digitata (Phaeophyceae) under biotic stress conditions(1). J Phycol44:1250–1256.
Küpper FC, Gaquerel E, Cosse A, Adas F, Peters AF, Müller DG, Kloareg B, Salaün JP, and Potin P. 2009. Free fatty acids and methyl jasmonate trigger defense reactions in Laminaria digitata. Plant Cell Physiol50:789–800.
Zambounis A, Strittmatter M, and Gachon CMM. 2013. Chronic stress and disease resistance in the genome model marine seaweed Ectocarpus siliculosus. Aquat Bot104:147–152.
Lim EL, Siow RS, Abdul Rahim R, and Ho CL. 2016. Global transcriptome analysis of Gracilaria changii (Rhodophyta) in response to agarolytic enzyme and bacterium. Mar Biotechnol18:189–200.
Strittmatter M, Grenville-Briggs LJ, Breithut L, van West P, Gachon CMM, and Küpper FC. 2016. Infection of the brown alga Ectocarpus siliculosus by the oomycete Eurychasma dicksonii induces oxidative stress and halogen metabolism. Plant Cell Environ39:259–271.
Kapraun DF. 2005. Nuclear DNA content estimates in multicellular green, red and brown algae: phylogenetic considerations. Ann Bot95:7–44.
Collén J, Porcel B, Carré W, Ball SG, Chaparro C, Tonon T, Barbeyron T, Michel G, Noel B, Valentin K, Elias M, Artiguenave F, Arun A, Aury JM, Barbosa-Neto JF, Bothwell JH, Bouget FY, Brillet L, Cabello-Hurtado F, Capella-Gutiérrez S, Charrier B, Cladière L, Cock JM, Coelho SM, Colleoni C, Czjzek M, Da Silva C, Delage L, Denoeud F, Deschamps P, Dittami SM, Gabaldón T, Gachon CMM, Groisillier A, Hervé C, Jabbari K, Katinka M, Kloareg B, Kowalczyk N, Labadie K, Leblanc C, Lopez PJ, McLachlan DH, Meslet-Cladiere L, Moustafa A, Nehr Z, Nyvall Collén P, Panaud O, Partensky F, et al. 2013. Genome structure and metabolic features in the red seaweed Chondrus crispus shed light on evolution of the Archaeplastida. Proc Natl Acad Sci U S A110:5247–5252.
Zambounis A, Elias M, Sterck L, Maumus F, and Gachon CMM. 2012. Highly dynamic exon shuffling in candidate pathogen receptors … what if brown algae were capable of adaptive immunity?Mol Biol Evol29:1263–1276.
Nürnberger T and Kemmerling B. 2006. Receptor protein kinases—pattern recognition receptors in plant immunity. Trends Plant Sci11:519–522.
Luo Q, Zhu Z, Yang R, Qian F, Yan X, and Chen H. 2015. Characterization of a respiratory burst oxidase homologue from Pyropia haitanensis with unique molecular phylogeny and rapid stress response. J Appl Phycol27:945–955.
Thomas F, Cosse A, Le Panse S, Kloareg B, Potin P, and Leblanc C. 2014. Kelps feature systemic defense responses: insights into the evolution of innate immunity in multicellular eukaryotes. New Phytol204:567–576.
Wang S, Zhao F, Wei X, Lu B, Duan D, and Wang G. 2013. Preliminary study on flg22-induced defense responses in female gametophytes of Saccharina japonica (Phaeophyta). J Appl Phycol25:1215–1223.
Meng X and Zhang S. 2013. MAPK cascades in plant disease resistance signaling. Annu Rev Phytopathol51:245–266.
Kawano Y, Kaneko-Kawano T, and Shimamoto K. 2014. Rho family GTPase-dependent immunity in plants and animals. Front Plant Sci5:522.
Ono E, Wong HL, Kawasaki T, Hasegawa M, Kodama O, and Shimamoto K. 2001. Essential role of the small GTPase Rac in disease resistance of rice. Proc Natl Acad Sci U S A98:759–764.
Hung CY, Aspesi P, Hunter MR, Lomax AW, and Perera IY. 2014. Phosphoinositide-signaling is one component of a robust plant defense response. Front Plant Sci5:267.
Romeis T, Ludwig AA, Martin R, and Jones JD. 2001. Calcium-dependent protein kinases play an essential role in a plant defence response. EMBO J20:5556–5567.
Kobayashi M, Ohura I, Kawakita K, Yokota N, Fujiwara M, Shimamoto K, Doke N, and Yoshioka H. 2007. Calcium-dependent protein kinases regulate the production of reactive oxygen species by potato NADPH oxidase. Plant Cell19:1065–1080.
Xie C, Zhou X, Deng X, and Guo Y. 2010. PKS5, a SNF1-related kinase, interacts with and phosphorylates NPR1, and modulates expression of WRKY38 and WRKY62. J Genet Genomics37:359–369.
Baena-González E, Rolland F, Thevelein JM, and Sheen J. 2007. A central integrator of transcription networks in plant stress and energy signalling. Nature448:938–942.
Meng Y and Wise RP. 2012. HvWRKY10, HvWRKY19, and HvWRKY28 regulate Mla-triggered immunity and basal defense to barley powdery mildew. Mol Plant Microbe Interact25:1492–1505.
Raffaele S and Rivas S. 2013. Regulate and be regulated: integration of defense and other signals by the AtMYB30 transcription factor. Front Plant Sci4:98.
Socquet-Juglard D, Kamber T, Pothier JF, Christen D, Gessler C, Duffy B, and Patocchi A. 2013. Comparative RNA-seq analysis of early-infected peach leaves by the invasive phytopathogen Xanthomonas arboricola pv. pruni. PLoS One8:e54196.
Mengiste T, Chen X, Salmeron J, and Dietrich R. 2003. The BOTRYTIS SUSCEPTIBLE1 gene encodes an R2R3MYB transcription factor protein that is required for biotic and abiotic stress responses in Arabidopsis. Plant Cell15:2551–2565.
Oñate-Sánchez L and Singh KB. 2002. Identification of Arabidopsis ethylene-responsive element binding factors with distinct induction kinetics after pathogen infection. Plant Physiol128:1313–1322.
Oñate-Sánchez L, Anderson JP, Young J, and Singh KB. 2007. AtERF14, a member of the ERF family of transcription factors, plays a nonredundant role in plant defense. Plant Physiol143:400–409.
Arnold TM, Targett NM, Tanner CE, Hatch WI, and Ferrari KE. 2001. Evidence for methyl jasmonate-induced phlorotannin production in Fucus vesiculosus (Phaeophyceae). J Phycol37:1026–1029.
Plettner I, Steinke M, and Malin G. 2005. Ethene (ethylene) production in the marine macroalga Ulva (Enteromorpha) intestinalis L. (Chlorophyta, Ulvophyceae): effect of light-stress and co-production with dimethyl sulphide. Plant Cell Environ28:1136–1145.
García-Jiménez P and Robaina RR. 2012. Effects of ethylene on tetrasporogenesis in Pterocladiella capillacea (Rhodophyta)(1). J Phycol48:710–715.
Rojas CM, Senthil-Kumar M, Tzin V, and Mysore KS. 2014. Regulation of primary plant metabolism during plant-pathogen interactions and its contribution to plant defense. Front Plant Sci5:17.
Taylor NL, Heazlewood JL, Day DA, and Millar AH. 2004. Lipoic acid-dependent oxidative catabolism of keto acids in mitochondria provides evidence for branched-chain amino acid catabolism in Arabidopsis. Plant Physiol134:838–848.
Bolton MD. 2009. Primary metabolism and plant defense—fuel for the fire. Mol Plant Microbe Interact22:487–497.
Soto G, Stritzler M, Lisi C, Alleva K, Pagano ME, Ardila F, Mozzicafreddo M, Cuccioloni M, Angeletti M, and Ayub ND. 2011. Acetoacetyl-CoA thiolase regulates the mevalonate pathway during abiotic stress adaptation. J Exp Bot62:5699–5711.
Stenmark H and Olkkonen VM. 2001. The Rab GTPase family. Genome Biol2:REVIEWS3007.
Schmieder R and Edwards R. 2011. Quality control and preprocessing of metagenomic datasets. Bioinformatics27:863–864.
Langmead B and Salzberg SL. 2012. Fast gapped-read alignment with Bowtie 2. Nat Methods9:357–359.
Davidson NM and Oshlack A. 2014. Corset: enabling differential gene expression analysis for de novo assembled transcriptomes. Genome Biol15:410.
Robinson MD, McCarthy DJ, and Smyth GK. 2010. edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics26:139–140.
Conesa A, Götz S, García-Gómez JM, Terol J, Talón M, and Robles M. 2005. Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics21:3674–3676.
Finn RD, Clements J, and Eddy SR. 2011. HMMER web server: interactive sequence similarity searching. Nucleic Acids Res39:W29–W37.
Pick T, Jaskiewicz M, Peterhänsel C, and Conrath U. 2012. Heat shock factor HsfB1 primes gene transcription and systemic acquired resistance in Arabidopsis. Plant Physiol159:52–55.
Pajerowska-Mukhtar KM, Wang W, Tada Y, Oka N, Tucker CL, Fonseca JP, and Dong X. 2012. The HSF-like transcription factor TBF1 is a major molecular switch for plant growth-to-defense transition. Curr Biol22:103–112.

Information & Contributors


Published In

cover image mSphere
Volume 2Number 627 December 2017
eLocator: 10.1128/msphere.00094-17
Editor: Yonghua Li-Beisson, Aix-Marseille University


Received: 3 March 2017
Accepted: 8 November 2017
Published online: 6 December 2017


  1. bacteria
  2. cell signaling
  3. defense
  4. differential expression
  5. seaweed
  6. terpenes



Louisi Souza de Oliveira
Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
Diogo Antonio Tschoeke
Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
Núcleo em Ecologia e Desenvolvimento Sócio-Ambiental de Macaé (NUPEM), Universidade Federal do Rio de Janeiro, Macaé, Rio de Janeiro, Brazil
Ana Carolina Rubem Magalhães Lopes
Departamento de Biologia Marinha, Universidade Federal Fluminense (UFF), Niterói, Rio de Janeiro, Brazil
Daniela Bueno Sudatti
Departamento de Biologia Marinha, Universidade Federal Fluminense (UFF), Niterói, Rio de Janeiro, Brazil
Pedro Milet Meirelles
Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
Cristiane C. Thompson
Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
Renato Crespo Pereira
Departamento de Biologia Marinha, Universidade Federal Fluminense (UFF), Niterói, Rio de Janeiro, Brazil
Fabiano L. Thompson
Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil


Yonghua Li-Beisson
Aix-Marseille University


Address correspondence to Fabiano L. Thompson, [email protected].

Metrics & Citations


Note: There is a 3- to 4-day delay in article usage, so article usage will not appear immediately after publication.

Citation counts come from the Crossref Cited by service.


If you have the appropriate software installed, you can download article citation data to the citation manager of your choice. Simply select your manager software from the list below and click Download.

View Options

Figures and Media






Share the article link

Share with email

Email a colleague

Share on social media

American Society for Microbiology ("ASM") is committed to maintaining your confidence and trust with respect to the information we collect from you on websites owned and operated by ASM ("ASM Web Sites") and other sources. This Privacy Policy sets forth the information we collect about you, how we use this information and the choices you have about how we use such information.
FIND OUT MORE about the privacy policy