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A Polysaccharide-Degrading Marine Bacterium Flammeovirga Sp. MY04 and Its Extracellular Agarase System

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J. Ocean Univ. China (Oceanic and Coastal Sea Research)

DOI 10.1007/s11802-012-1929-3 ISSN 1672-5182, 2012 11 (3): 375-382

http://www.ouc.edu.cn/xbywb/ E-mail:[email protected]

A Polysaccharide-Degrading Marine Bacterium Flammeovirga sp. MY04 and Its Extracellular Agarase System

HAN Wenjun1), 2), 3), GU Jingyan1), 3), YAN Qiujie2), LI Jungang2), WU Zhihong3), * GU Qianqun1), *, and LI Yuezhong3)
,

1) Key Laboratory of Marine Drugs, Chinese Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao 266003, P . R . China
2) School of Life Science and Biotechnology, Mianyang Normal University, Mianyang 621000, P . R . China 3) State Key Laboratory of Microbial T e chnology, School of Life Science, Shandong University, Jinan 250100, P . R . China

(Received February 2, 2012; revised April 8, 2012; accepted May 29, 2012) © Ocean University of China, Science Press and Springer-Verlag Berlin Heidelberg 2012

Abstract Bacteria of the genus Flammeovirga can digest complex polysaccharides (CPs), but no details have been reported regarding the CP depolymerases of these bacteria. MY04, an agarolytic marine bacterium isolated from coastal sediments, has been identified as a new member of the genus Flammeovirga. The MY04 strain is able to utilize multiple CPs as a sole carbon source and grows well on agarose, mannan, or xylan. This strain produces high concentrations of extracellular proteins (490mgL-1 ±18.2mgL-1 liquid culture) that exhibit efficient and extensive degradation activities on various polysaccharides, especially agarose. These proteins have an activity of 310Umg-1 ±9.6Umg-1 proteins. The extracellular agarase system (EAS) in the crude extracellular enzymes contains at least four agarose depolymerases, which are with molecular masses of approximately 30–70kDa. The EAS is stable at a wide range of pH values (6.0–11.0), temperatures (0–50℃), and sodium chloride (NaCl) concentrations (0–0.9mol L-1). Two major degradation products generated from agarose by the EAS are identified to be neoagarotetraose and neoagarohexaose, suggesting that β-agarases are the major constituents of the MY04 EAS. These results suggest that the Flammeovirga strain MY04 and its polysaccharide-degradation system hold great promise in industrial applications.

Key words Flammeovirga; polysaccharide degradation; extracellular agarase system; neoagaro-oligosaccharide

while only two α-agarases have been reported, one from

Thalassomonas sp. JAMB-A33 and one from Alteromo- nas agarlyticus GJ1B (Ohta et al., 2005b; Potin et al.,

1 Introduction

Complex polysaccharides (CPs), which are composed of repeated units of complex sugars, are widely present in plants, animals, and microorganisms, in which they serve as essential structural and functional components. Agar, composed of agarose and agaropectin, is the main component of the cell wall of red algae (Rhodophyta) (Rochas et al., 1986). Agarose is a polymer composed of 3,6-anhydro-L-galactopyranose-α -1,3-D-galactopyranose units that are joined by β1-4 bonds (Rees, 1969). Agarose can be cleaved by either α-agarase (E.C. 3.2.1.158) at the α1-3 linkage (Rochas et al., 1994) or β-agarase (E.C. 3.2.1.81) at the β1-4 linkage (Morrice et al., 1983a and 1983b), producing a series of oligosaccharides that have reducing ends. The oligomers yielded from the degradation of agarose by α-agarase and β-agarase are termed agaro- and neoagaro-oligosaccharides, respectively. There have been many studies of agarases and their degradation of agarose. Most of the reported agarases are β-agarases,
1993).
Agarases are useful in the preparation of algal protoplasts (Gupta et al., 2010; Yeong et al., 2008) and the recovery of DNA from agarose gels (Cole and Åkerman, 2000) , and have potential in the production of functional oligosaccharides (Hatada et al., 2006; Hu et al., 2006; Wang et al., 2004; Wu et al., 2005). Although the agarose degradation abilities of many microorganisms isolated from marine environment, fresh water, or soil have been investigated and the corresponding enzymes have been characterised, only β-agarase I from Pseudoalteromonas atlantica has been industrially applied for the gel recovery of DNA (Fu and Kim, 2010). There are many limitations in the applications of agarases. For example, the heterologous expression levels of agarases are usually low and the use of these enzymes for some purposes, such as the enzymatic preparation of functional oligosaccharides, is expensive. The use of native agarase-producers or their extracellular agarase systems (EASs) may provide an alternative.

* Corresponding authors. Tel: 0086-532-82032065 E-mail: [email protected]; [email protected]

Flammeovirga is a newly defined bacterial genus be-

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376

HAN et al. / J. Ocean Univ. China (Oceanic and Coastal Sea Research) 2012 11 (3): 375-382

longing to the family Flammeovirgaceae of the class α-Proteobacteria. Five species have been reported in this

genus: F . a prica (Nakagawa et al., 1997), F . a renaria, F . yaeyamensis (Takahashi et al., 2006), F . k amogawensis (Hosoya and Yokota, 2007), and F. pacifica (Xu et al.,

2011). All of the type strains have a potent ability to degrade marine CPs, including agarose. Yang et al. have described the gene cloning and biochemical characteristic

of an agarase, AgaYT, from the F . y aeyamensis strain YT

(Yang et al., 2011). Although the heterologously expressed enzyme exhibited a high degradation activity on agarose, when the protein was expressed in E. coli, it was mostly present in the form of inclusion bodies. To date, we know few of the CP-degradation characteristics of Flammeovirga strains or their extracellular agarase systems, although these strains/systems have been suggested to be relevant for biotechnological applications. glucose. After incubation at 30℃ for 16h with agitation, the turbidity of the broth was measured at 600 nm to evaluate the cellular growth. The initial pH of the medium was adjusted to 5.0–11.0 before autoclaving. Culture temperatures were between 4℃ and 45℃. The cellular tolerance to sodium chloride was assayed over a concentration range of 0–5.0%. The requirement of metal ions for growth was analysed by growing a series of cultures in which a single metal salt had been removed from the medium.

2.3 Polysaccharide-Utilizing Abilities of the MY04
Strain

To evaluate the ability of the bacterial isolate to utilize different polysaccharides, the sole carbon source in the BM broth was supplemented with various CPs, including algae-derived polysaccharides (alginate, agar, agarose, carrageenan, and ι-carrageenan), plant-derived polysaccharides (cellulose, carboxymethylcellulose (CMC), crude mannan, superfine mannan, microcrystalline cellulose (MC), pectate, starch, and xylan), and crustacean polysaccharides (chitin and chitosan). Polysaccharides were added to the media at a final concentration of 0.10%. After the initial culture in the BM broth supplemented with 0.2% glucose, the MY04 bacteria were collected by centrifugation when the culture reached an OD600 of 0.8 and were washed twice using sterilised seawater. The washed bacterial sludge was resuspended in BM broth, and an aliquot was transferred into each restricted broth at a starting OD600 of 0.08. The cellular growth of MY04 was evaluated as described above.
Herein, we report the polysaccharide-degradation characteristics of a marine isolate, Flammeovirga sp. MY04, and its extracellular agarase system.

2 Materials and Methods

2.1 Isolation of Agarose-Degrading Bacteria

Coastal sediments were collected from a laver farm near Ganyu City in Jiangsu Province, China. A basal medium (BM) composed of 3.0% NaCl, 0.75% KCl, 0.11% CaCl2, 0.72% Mg2SO4, 0.15% NH4Cl, and 1.5% agar (pH 7.0) was used to isolate agarolytic bacteria. After incubation at 30℃ for 72h, the colonies that formed deep craters in the agar were transferred to a fresh BM plate for further purification. To evaluate the agar-digestion ability of the isolates, the BM plates were stained with an iodine solution (Hodgson and Chater, 1981) following a 16-h incubation at 30℃.

2.4 Preparation of the Crude Extracellular
Enzymes

To prepare the crude extracellular enzymes, the MY04 strain was cultured at 30℃ for 72h in 1L of BM broth supplemented with 0.4% tryptone and 0.25% yeast extract. The following preparation was performed at 4℃. The cultures were centrifuged at 12000\× g for 15 min, and ammonium sulphate was added into the supernatant to reach 80% saturation. After a 4-h incubation to precipitate the proteins, the mixture was centrifuged at 15000×g for 30min. The pellet was dissolved in 50mL of 50mmolL-1 HEPES buffer (pH 7.5) containing 0.5 mmol L-1 EDTA and 5% glycerol, which was dialysed three times against the same buffer for 6h. The dialysed enzyme solution was stored in aliquots at −20℃ before use.

2.2 Identification of the MY04 Isolate

Genomic DNA was prepared using a genomic DNA extraction kit (TianGen Inc., Beijing, China). For PCR amplification of the 16S ribosomal RNA (rRNA) gene sequence, the bacterial universal primer pair 27f (5’-GAGTTTGATCCTGGCTCAG-3’) and 1492r (5’-AA- GGAGGTGATCCAGC C-3’) (Weisburg et al., 1991) were used. The PCR products were gel-purified with a DNA extraction kit (TAKARA Inc., Dalian, China) according to the manufacturer’s instructions and cloned into the pMD18-T vector (TAKARA Inc., Dalian, China) for sequencing. The sequence was analysed against the GenBank database using the on-line BLAST program (Altschul et al., 1990) to search for the most similar sequences.
For transmission electron microscopy (TEM), MY04 cells that were cultured for 16h at 30℃ were mounted on a Formvar-coated copper grid and negatively stained with 1.0% aqueous uranyl acetate. The cell-mounted grid was observed using a JEOL1010 transmission electron microscope (TEM, JEOL) operated at 100kV.

2.5 Polysaccharide-Degradation Activities of the
Crude Extracellular Enzymes

The polysaccharides described above for cellular utilization were further tested as substrates for the degradation-activity assay of the crude enzyme extract. The 3, 5-dinitrosalicylic acid method (Miller, 1959), with minor modifications, was employed to assay the yielded reducing sugar. A 75-µL aliquot of the crude enzyme extract was mixed with 425μL of 50 mmolL-1 HEPES buffer (pH 7.5) that contained an individual polysaccharide at a final concentration of 0.20%. Following a 30-min incubation at
To determine the growth characteristics, strain MY04 was cultivated in BM broth supplemented with 0.20%

HAN et al. / J. Ocean Univ. China (Oceanic and Coastal Sea Research) 2012 11 (3): 375-382

377

40℃, 500μL of 3, 5-dinitrosalicylic acid was added to the reaction mixture and boiled for 10 min until red colour developed, which was measured at 540nm. The red color was derived from the reaction of reducing sugars and 3, 5-dinitrosalicylic acids. A control was performed by boiling the enzyme solution for 5min before adding the solution to the reaction mixture. All of the assays were performed in triplicate. D-Galactose was used as the standard reducing sugar. The enzyme activity (U) was defined as the amount of enzyme that produced 1μmol of D-galactose per minute. The concentration of protein was determined using the Bradford assay (Bradford, 1976) with bovine serum albumin as a standard. were optimised if necessary. The relative activity was defined as the percentage of the residual activity relative to that under the standard conditions described previously.

2.9 Identification of the Agarose-Derived
Oligosaccharides

Oligosaccharides derived from agarose were prepared using the method described previously by Li et al. (Li et al., 2007) with some modifications. Briefly, 100 mg of agarose was dissolved in 100 mL of buffer (50 mmol L-1 Tris-Cl, 150 mmolL-1 NaCl, 10mmolL-1 CaCl2, pH 7.5). After sterilisation, the substrate was placed in a 45℃ water bath to prevent gelling. The solution was mixed with 10mL of the crude extracellular enzyme extracts. After a 48-h incubation at 45℃, the reaction was stopped by heating the mixture in a boiling water bath for 10 min. The mixture was subsequently cooled to 4℃ and centrifuged at 12000×g for 30min. The supernatant was concentrated by rotary evaporation at 45℃. A 2-mL aliquot of the solution of the enzymatic products was loaded onto a Sephadex G25 column (superfine, GE-Healthcare, USA) or a Bio-gel P6 column (fine, Bio-Rad Laboratories, USA) for separation using 0.1molL-1 NH4HCO3 as the elution solution. The classic phenol-H2SO4 method was performed to analyse the oligosaccharide fractions. The NH4HCO3 was removed by repeated evaporation under reduced pressure at 45℃ for further component identification.

2.6 Zymogrammatic Analysis of the EAS

The extracellular agarase system (EAS) in the crude enzyme extracts was analysed to identify the agarase components. An aliquot of 10µL of the crude extracellular enzyme extract was mixed with an equal volume of loading buffer and separated using 13.2% non-denaturing polyacrylamide gel electrophoresis (PAGE). After electrophoresis, the gel was washed three times using 50 mmolL-1 HEPES buffer (pH 7.5) for a total of 30min. The gel was then overlaid onto a 1.5% agarose sheet prepared with 50 mmolL-1 HEPES (pH 7.5) and incubated overnight at 37℃. The zymogram was developed using iodine staining.

2.7 Effects of pH and Temperature on the EAS
Activity and Stability

The crude enzymatic products of agarose and the main products from the above preparation, two oligosaccharide fractions (F2 and F3), were applied to silica gel 60 (F254) plates (Merck KGaA Darmstadt, Germany) for TLC analysis. The plates were developed with the solvent n-butanol/ ethanol/water (3:1:1), sprayed with a staining solution (1% diphenylamine and aniline in acetone) (Zhang et al., 2010), and heated at 110℃ for 10min. For MALDI-TOF mass spectrometry, the oligosaccharide fractions F2 and F3 were individually observed as pseudo-molecular ions [M+Na]+ and [M+H]+. The mass spectrometry was performed on a Biflex III instrument (Bruker Daltonics, Billerica, USA) that was set to the positive-ion mode, with formic acid as the matrix. For 13C spectroscopy, 10mg of each oligosaccharide fraction was dissolved in 0.5 mL of D2O in 5-mm NMR tubes. The spectra were recorded on a JNM-ECP600 (JEOL, Japan) apparatus set at 150 MHz, with acetone-d6 as the internal standard.
The optimal temperature of the EAS was determined by carrying out the enzyme activity assay at different temperatures (15–70℃) in 50 mmol L-1 HEPES buffer (pH 7.0) for 30min. The optimal pH of the EAS was assayed at 45℃ in preheated buffers with different pH values, including 50mmolL-1 acetate buffer (pH 4.0–6.0), 50 mmol L-1 HEPES buffer (pH 7.0–8.0), and 50 mmolL-1 glycine-NaOH buffer (pH 8.0–12.0). In addition, the thermostability of the EAS was evaluated by measuring the residual activity of the crude enzyme extract after a preincubation at different temperatures for 1h. The pH stability of the EAS was determined by pre-incubating the crude enzyme extract in each solution (1:20 dilution) of varying pH (4.0–12.0) at 45℃ for 1h and then assaying the residual activity. All of the assays were performed in triplicate. The relative activity was defined as the percentage of activity with respect to the maximum agarase activity.

3 Results

2.8 Effects of Different Compounds on the
EAS Activity
3.1 Isolation and Identification of the Agarolytic
MY04 Strain

A basal medium (BM), described in the Materials and
Methods, was used for the screening and isolation of bacterial strains that are able to utilize agar as their sole carbon source. The agarolytic activity of the colonies formed on BM plates was detected based on etched agar and iodine staining. The agarolytic strain MY04 was isolated from coastal sediments collected from a laver farm near
Various metal ions, chelators, denaturants, reducing reagents, and a thiol modifier were added to the reaction mixtures to separately assay the effects of these substances on the EAS activity. The reaction was performed in 50mmolL-1 HEPES buffer (pH 7.5) at 45℃ for 30min. The final concentration employed for each reagent was 1, 10 or 100mmolL-1. The concentrations of the metal ions

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HAN et al. / J. Ocean Univ. China (Oceanic and Coastal Sea Research) 2012 11 (3): 375-382

Ganyu City in Jiangsu Province, China, in 2004. On a BM plate, strain MY04 spread and formed large and deeply etched colonies. After the plate was stained with the iodine solution, a clear agar-degradation zone developed around individual colonies (Fig.1a). The solid medium on which MY04 was cultured turned to liquid after 5 d of incubation at 30℃. These culture characteristics suggested that MY04 has potent agarolytic activities.

3.2 Growth Characterisation of MY04

MY04 grew well in the presence of 1.0%−5.0% NaCl, with an optimal concentration of 3.0%, suggesting that this strain is a halophilic bacterium. Among the metal ion

+

components in the BM, NH4 , Mg2+, and Ca2+ were essential for the growth of MY04, whereas K+ was not essential. MY04 grew optimally at 30℃. The tolerable pH range for the growth of MY04 was between 6.0 and 10.0, with optimal growth at pH 7.0.
MY04 was able to grow with most of the tested CPs as the sole carbon source (Fig.2a). Agarose was the best carbon source for the growth of this strain. Some of the other CPs, such as mannan and xylan, were also good carbon sources for MY04 growth. It is interesting to note that, in contrast to the type strains of Flammeovirga, the MY04 strain grew weakly on carboxylmethyl cellulose (CMC). Moreover, the MY04 strain was unable to utilize chitosan.

Fig.1 Phenotypic morphologies of the marine isolate MY04. (A) Colonies of MY04 on a BM plate stained using the iodine method. The regions with yellow or red color denote the presence of reducing sugars produced from the degradation of agar by the extracellular agarase secreted from MY04 colonies, which are in the middle of the yellow regions (the dark dots or lines). (B) Transmission electron micrograph of negatively stained MY04 cells. Scale bar=1μm.

The 16S rRNA gene of MY04 was sequenced, and the sequence was deposited in the GenBank database under accession number AY849869. According to the BLASTn results using the GenBank databases, MY04 is highly similar (99.6% similarity with respect to the 16S rRNA

gene sequence) to F . y aeyamensis NBRC 100898T (Taka-

hashi et al., 2006). However, the MY04 strain had two phenotypic characteristics that were different from the

type strain of F . y aeyamensis. One characteristic is that

the MY04 culture is reddish-orange during exponential growth but changes to a white color during the late stationary phase, which is similar to the colors reported for F . kamogawensis (Hosoya and Yokota, 2007). Additionally, MY04 cells are curved with a size of (0.4–0.7)μm× (4–6) μm (Fig.1b), whereas the reported Flammeovirga members are longer rods.

Fig.2 Utilization and degradation abilities of Flammeo- virga sp. MY04 and its extracellular enzyme extracts. (A) Growth ability of MY04 cells in BM containing various CPs as the sole carbon source. (B) Degradation activities of the crude extracellular enzyme extracts from strain MY04 on CPs. * OD540 = 18.4. The results are means ± S.D. of three different experiments.

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  • Characterization of a Recombinant Thermostable Arylsulfatase from Deep-Sea Bacterium Flammeovirga Pacifica Chao Gao1†, Min Jin2†, Zhiwei Yi2, and Runying Zeng2,3,4*

    Characterization of a Recombinant Thermostable Arylsulfatase from Deep-Sea Bacterium Flammeovirga Pacifica Chao Gao1†, Min Jin2†, Zhiwei Yi2, and Runying Zeng2,3,4*

    J. Microbiol. Biotechnol. (2015), 25(11), 1894–1901 http://dx.doi.org/10.4014/jmb.1504.04028 Research Article Review jmb Characterization of a Recombinant Thermostable Arylsulfatase from Deep-Sea Bacterium Flammeovirga pacifica Chao Gao1†, Min Jin2†, Zhiwei Yi2, and Runying Zeng2,3,4* 1XingHuaLing District Food and Drug Administration, Taiyuan 030000, P.R. China 2State Key Laboratory Breeding Base of Marine Genetic Resource; Third Institute of Oceanography, SOA, Xiamen 361005, P.R. China 3Fujian Collaborative Innovation Center for Exploitation and Utilization of Marine Biological Resources, Xiamen 361005, P.R. China 4South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, Guangzhou 510000, P.R. China Received: April 13, 2015 Revised: August 4, 2015 A novel sulfatase gene, ary423 (1,536 bp ORF), encoding a protein of 511 amino acids with a Accepted: August 8, 2015 calculated molecular mass of 56 kDa, was identified from Flammeovirga pacifica, which was isolated from deep-sea sediments of west Pacific Ocean. Amino acid sequence analysis revealed that Ary423 possessed a conserved C-X-A-X-R motif, which was recognized as the First published online sulfatase signature. Phylogenetic analysis suggested that Ary423 belonged to arylsulfatases. August 13, 2015 After heterologous expression in Escherichia coli cells, the recombinant Ary423 was purified + *Corresponding author with a Ni affinity column, and was shown to be highly active at a broad range of Phone: +86-592-2195323; temperatures from 30° to 70°C, with maximum activity at 40°C. Furthermore, recombinant Fax: +86-592-2195323; Ary423 retained more than 70% and 40% of its maximum activity after 12 h of incubation at E-mail: [email protected] 50°C and 60°C, respectively, exhibiting good thermostability at high temperatures.
  • Carrageenan Catabolism Is Encoded by a Complex Regulon in Marine Heterotrophic Bacteria

    Carrageenan Catabolism Is Encoded by a Complex Regulon in Marine Heterotrophic Bacteria

    ARTICLE DOI: 10.1038/s41467-017-01832-6 OPEN Carrageenan catabolism is encoded by a complex regulon in marine heterotrophic bacteria Elizabeth Ficko-Blean1, Aurélie Préchoux1, François Thomas 1, Tatiana Rochat 2, Robert Larocque1, Yongtao Zhu 3, Mark Stam4, Sabine Génicot1, Murielle Jam1, Alexandra Calteau4, Benjamin Viart4, David Ropartz5, David Pérez-Pascual2, Gaëlle Correc1, Maria Matard-Mann1, Keith A. Stubbs6, Hélène Rogniaux5, Alexandra Jeudy1, Tristan Barbeyron1, Claudine Médigue4, Mirjam Czjzek1, David Vallenet 4, Mark J. McBride3, Eric Duchaud2 & Gurvan Michel 1 1234567890 Macroalgae contribute substantially to primary production in coastal ecosystems. Their biomass, mainly consisting of polysaccharides, is cycled into the environment by marine heterotrophic bacteria using largely uncharacterized mechanisms. Here we describe the complete catabolic pathway for carrageenans, major cell wall polysaccharides of red mac- roalgae, in the marine heterotrophic bacterium Zobellia galactanivorans. Carrageenan cata- bolism relies on a multifaceted carrageenan-induced regulon, including a non-canonical polysaccharide utilization locus (PUL) and genes distal to the PUL, including a susCD-like pair. The carrageenan utilization system is well conserved in marine Bacteroidetes but modified in other phyla of marine heterotrophic bacteria. The core system is completed by additional functions that might be assumed by non-orthologous genes in different species. This complex genetic structure may be the result of multiple evolutionary events including gene duplica- tions and horizontal gene transfers. These results allow for an extension on the definition of bacterial PUL-mediated polysaccharide digestion. 1 Sorbonne Universités, UPMC Univ Paris 06, CNRS, UMR 8227, Integrative Biology of Marine Models, Station Biologique de Roscoff, CS 90074 Roscoff, Bretagne, France.
  • University of Florida Thesis Or Dissertation Formatting

    University of Florida Thesis Or Dissertation Formatting

    CHARACTERIZING THE MOLECULAR MECHANISMS CONTRIBUTING TO BIOLOGICALLY INDUCED CARBONATE MINERALIZATION AND THROMBOLITE FORMATION By ARTEMIS S. LOUYAKIS A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2017 © 2017 Artemis S. Louyakis To my mother, for supporting every single goal I’ve ever had, the memory of my father, for keeping me focused, and my partner, for all he’s done ACKNOWLEDGMENTS I would like to begin by acknowledging and thanking my mentor, Dr. Jamie Foster, for all her guidance throughout this Ph.D. I thank my committee members for all of their advice and support - Drs. Eric Triplett, Julie Maupin, Nian Wang, and Eric McLamore. I’d like to thank the rest of the Department of Microbiology and Cell Science, staff for always keeping my academic life in order, faculty for never turning me away when I came to use equipment or ask for help, especially Drs. K.T. Shanmugan and Wayne Nicholson, as well as Dr. Andy Schuerger from the Dept. of Plant Pathology for his advice over the years. I’d also like to acknowledge those lab members and extended lab members who made themselves readily available to talk through any problems I came up against and celebrate when all went well, including Drs. Rafael Oliveira, Jennifer Mobberley, and Giorgio Casaburi, and Lexi Duscher, Rachelle Banjawo, Maddie Vroom, Hadrien Gourlé, and so many more. I’d also like to profusely thank my family and friends who have never been anything less than completely supportive of me, specifically my partner Nathan Prince, my mother and siblings Denise Louyakis, Bobbi Louyakis, Nick Newman, Cori Sergi, extended parents and siblings Carol Prince, Barry Prince, Aaron Prince, my nieces and nephew Bailey O’Regan, Bella O’Regan, Layla Newman, Colton Prince, and Summer Prince, and my dearest friends Tina Pontbriand, Tom Pontbriand, Karen Chan, Dalal Haouchar, Alexi Casaburi, and Eloise Stikeman.
  • The Influence of Bacteria on the Adaptation to Changing Environments in Ectocarpus : a Systems Biology Approach Hetty Kleinjan

    The Influence of Bacteria on the Adaptation to Changing Environments in Ectocarpus : a Systems Biology Approach Hetty Kleinjan

    The influence of bacteria on the adaptation to changing environments in Ectocarpus : a systems biology approach Hetty Kleinjan To cite this version: Hetty Kleinjan. The influence of bacteria on the adaptation to changing environments in Ectocar- pus : a systems biology approach. Molecular biology. Sorbonne Université, 2018. English. NNT : 2018SORUS267. tel-02868508 HAL Id: tel-02868508 https://tel.archives-ouvertes.fr/tel-02868508 Submitted on 15 Jun 2020 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. Sorbonne Université ED 227 - Sciences de la Nature et de l'Homme : écologie & évolution Laboratoire de Biologie Intégrative des Modèles Marins Equipe Biologie des algues et interactions avec l'environnement The influence of bacteria on the adaptation to changing environments in Ectocarpus: a systems biology approach Par Hetty KleinJan Thèse de doctorat en Biologie Marine Dirigée par Simon Dittami et Catherine Boyen Présentée et soutenue publiquement le 24 septembre 2018 Devant un jury composé de : Pr. Ute Hentschel Humeida, Rapportrice GEOMAR, Kiel, Germany Dr. Suhelen Egan, Rapportrice UNSW Sydney, Australia Dr. Fabrice Not, Examinateur Sorbonne Université – CNRS Dr. David Green, Examinateur SAMS, Oban, UK Dr. Aschwin Engelen, Examinateur CCMAR, Faro, Portugal Dr.
  • Genome-Based Taxonomic Classification Of

    Genome-Based Taxonomic Classification Of

    ORIGINAL RESEARCH published: 20 December 2016 doi: 10.3389/fmicb.2016.02003 Genome-Based Taxonomic Classification of Bacteroidetes Richard L. Hahnke 1 †, Jan P. Meier-Kolthoff 1 †, Marina García-López 1, Supratim Mukherjee 2, Marcel Huntemann 2, Natalia N. Ivanova 2, Tanja Woyke 2, Nikos C. Kyrpides 2, 3, Hans-Peter Klenk 4 and Markus Göker 1* 1 Department of Microorganisms, Leibniz Institute DSMZ–German Collection of Microorganisms and Cell Cultures, Braunschweig, Germany, 2 Department of Energy Joint Genome Institute (DOE JGI), Walnut Creek, CA, USA, 3 Department of Biological Sciences, Faculty of Science, King Abdulaziz University, Jeddah, Saudi Arabia, 4 School of Biology, Newcastle University, Newcastle upon Tyne, UK The bacterial phylum Bacteroidetes, characterized by a distinct gliding motility, occurs in a broad variety of ecosystems, habitats, life styles, and physiologies. Accordingly, taxonomic classification of the phylum, based on a limited number of features, proved difficult and controversial in the past, for example, when decisions were based on unresolved phylogenetic trees of the 16S rRNA gene sequence. Here we use a large collection of type-strain genomes from Bacteroidetes and closely related phyla for Edited by: assessing their taxonomy based on the principles of phylogenetic classification and Martin G. Klotz, Queens College, City University of trees inferred from genome-scale data. No significant conflict between 16S rRNA gene New York, USA and whole-genome phylogenetic analysis is found, whereas many but not all of the Reviewed by: involved taxa are supported as monophyletic groups, particularly in the genome-scale Eddie Cytryn, trees. Phenotypic and phylogenomic features support the separation of Balneolaceae Agricultural Research Organization, Israel as new phylum Balneolaeota from Rhodothermaeota and of Saprospiraceae as new John Phillip Bowman, class Saprospiria from Chitinophagia.
  • The Influence of Invasive Jellyfish Blooms on The

    The Influence of Invasive Jellyfish Blooms on The

    Biol Invasions DOI 10.1007/s10530-014-0810-2 MOLECULAR TOOLS The influence of invasive jellyfish blooms on the aquatic microbiome in a coastal lagoon (Varano, SE Italy) detected by an Illumina-based deep sequencing strategy Caterina Manzari • Bruno Fosso • Marinella Marzano • Anita Annese • Rosa Caprioli • Anna Maria D’Erchia • Carmela Gissi • Marianna Intranuovo • Ernesto Picardi • Monica Santamaria • Simonetta Scorrano • Giuseppe Sgaramella • Loredana Stabili • Stefano Piraino • Graziano Pesole Received: 14 November 2013 / Accepted: 31 October 2014 Ó The Author(s) 2014. This article is published with open access at Springerlink.com Abstract The rapid expansion of multicellular microbial taxonomic and functional diversity. Here native and alien species outbreaks in aquatic and we used an effective and highly sensitive experimental terrestrial ecosystems (bioinvasions) may produce strategy, bypassing the efficiency bottleneck of the significant impacts on bacterial community dynamics traditional bacterial isolation and culturing method, to and nutrient pathways with major ecological implica- identify changes of the planktonic microbial commu- tions. In aquatic ecosystems, bioinvasions may cause nity inhabiting a marine coastal lagoon (Varano, adverse effects on the water quality resulting from Adriatic Sea) under the influence of an outbreak- changes in biological, chemical and physical proper- forming alien jellyfish species. Water samples were ties linked to significant transformations of the collected from two areas that differed in their level of confinement inside in the lagoon and jellyfish densities (W, up to 12.4 medusae m-3; E, up to 0.03 medusae Caterina Manzari, Bruno Fosso and Marinella Marzano have -3 contributed equally to this work. m ) to conduct a snapshot microbiome analysis by a metagenomic approach.