@phdthesis{Abdelmohsen2010,
  author    = {Abdelmohsen, Usama Ramadan},
  title     = {Antimicrobial Activities from Plant Cell Cultures and Marine Sponge-Associated Actinomycetes},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-51483},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2010},
  abstract  = {This thesis is divided into three parts with the main goal allocating novel antimicrobial compounds that could be used as future antibiotics. The first part aimed to evaluate the potential of plant suspension cultures for the production of antimicrobial proteins. The extracellular, intracellular and cell wall bound fractions of seven heterotrophic and photomixotrophic plant cell suspension cultures treated with nine different elicitors were tested for the elicitor dependent production of antimicrobial proteins. Bioactivities were tested against a selected panel of human isolates including Gram-positive and Gram-negative bacteria as well as fungi using the disc diffusion assay. The intracellular fractions of elicited cell cultures were more active than extracellular fractions while the cell wall bound fractions showed lowest activities. Among the 21 fractions tested, the intracellular fraction of Lavendula angustifolia elicited with DC3000 was most active against Candida maltosa. The second most active fraction was the intracellular fraction of Arabidopsis thaliana elicited with salicylic acid which was moreover active against all test strains. The antimicrobial activity of elicited Arabidopsis thaliana cell cultures was tested by bioautography to locate the antimicrobial proteins in the crude extract. The intracellular fraction of photomixotrophic Arabidopsis thaliana cells elicited with salicylic acid was selected for further gel filtration chromatography on S-200 column leading to the purification of one 19 kDa antimicrobially active protein, designated, AtAMP. Our findings suggest that elicited plant cell cultures may present a new promising alternative source of antimicrobial proteins. The second part comprises the isolation of actinomycetes associated with marine sponges and testing the bioactivities of new species for further investigations. Actinobacterial communities of eleven taxonomically different sponges that had been collected from offshore Ras Mohamed (Egypt) and from Rovinj (Croatia) were investigated by a culture-based approach using different standard media for isolation of actinomycetes and media enriched with aqueous sponge extract to target rare and new actinomycete species. Phylogenetic characterization of 52 representative isolates out of 90 based on almost complete sequences of genes encoding 16S rRNA supported their assignment to 18 different actinomycete genera. Altogether 14 putatively new species were identified based on sequence similarity values below 98.2\% to other strains in the NCBI database. The use of M1 agar amended with aqueous sponge extract yielded a putative new genus related to Rubrobacter which highlighting the need for innovative cultivation protocols. Biological activity testing showed that five isolates were active against Gram-positives only, one isolate was active against Candida albicans only and one isolate showed activity against both groups of pathogens. Moreover, the antiparasistic activity was documented for four isolates. These results showed a high diversity of actinomycetes associated with marine sponges as well as highlighted their potential to produce anti-infective agents. The third part of the thesis focused on the isolation and structure elucidation of new bioactive compounds. Streptomyces strain RV15 recovered from sponge Dysidea tupha, was selected for further chemical analysis by virtue of the fact that it exhibited the greatest antimicrobial potential against Staphylococcus aureus as well as Candida albicans among the all tested strains. Moreover, members of the genus Streptomyces are well known as prolific producers of interesting pharmacologically active metabolites. Chemical analysis of the methanolic crude extract using different chromatographic tools yielded four new compounds. The structures of the new compounds were spectroscopically elucidated to be four new cyclic peptides, namely, cyclodysidins A-D. Their bioactivity was tested against different proteases, bacteria and Candida as well as tumor cell lines. The compounds did not show any significant activities at this point.},
  subject      = {Antimikrobieller Wirkstoff},
  language  = {en}
}
@phdthesis{PimentelElardo2008,
  author    = {Pimentel Elardo, Sheila Marie},
  title     = {Novel anti-infective secondary metabolites and biosynthetic gene clusters from actinomycetes associated with marine sponges},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-40463},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2008},
  abstract  = {Marine sponges (Porifera) harbor diverse microbial communities within their mesohyl, among them representatives of the phylum Actinobacteria, commonly known as actinomycetes. Actinomycetes are prolific producers of pharmacologically important compounds and are responsible for producing the majority of antibiotics. The main aim of this Ph.D. study was to investigate the metabolic potential of the sponge-associated actinomycetes to produce novel anti-infective agents. The first aim was to cultivate actinomycetes derived from different marine sponges. 16S rDNA sequencing revealed that the strains belonged to diverse actinomycete genera such as Gordonia, Isoptericola, Micromonospora, Nocardiopsis, Saccharopolyspora and Streptomyces. Phylogenetic analyses and polyphasic characterization further revealed that two of these strains represent new species, namely Saccharopolyspora cebuensis strain SPE 10-1T (Pimentel-Elardo et al. 2008a) and Streptomyces axinellae strain Pol001T (Pimentel-Elardo et al. 2008b). Furthermore, secondary metabolite production of the actinomycete strains was investigated. The metabolites were isolated using a bioassay-guided purification scheme followed by structure elucidation using spectroscopic methods and subjected to an elaborate anti-infective screening panel. Several interesting compounds were isolated namely, the novel polyketides cebulactam A1 and A2 (Pimentel-Elardo et al. 2008c), a family of tetromycin compounds including novel derivatives, cyclodepsipeptide valinomycin, indolocarbazole staurosporine, diketopiperazine cycloisoleucylprolyl and butenolide. These compounds exhibited significant anti-parasitic as well as protease inhibitory activities. The third aim of this Ph.D. study was to identify biosynthetic gene clusters encoding for nonribosomal peptide synthetases (NRPS) and polyketide synthases (PKS) present in the actinomycete strains. Genomic library construction and sequencing revealed insights into the metabolic potential and biosynthetic pathways of selected strains. An interesting NRPS system detected in Streptomyces sp. strain Aer003 was found to be widely distributed in several sponge species, in an ascidian and in seawater and is postulated to encode for a large peptide molecule. Sequencing of the PKS gene cluster of Saccharopolyspora cebuensis strain SPE 10-1T allowed the prediction of the cebulactam biosynthetic pathway which utilizes 3-amino-5-hydroxybenzoic acid as the starter unit followed by successive condensation steps involving methylmalonyl extender units and auxiliary domains responsible for the polyketide assembly. In conclusion, this Ph.D. study has shown that diverse actinomycete genera are associated with marine sponges. The strains, two of them novel species, produced diverse chemical structures with interesting anti-infective properties. Lastly, the presence of biosynthetic gene clusters identified in this study substantiates the biosynthetic potential of actinomycetes to produce exploitable natural products and hopefully provides a sustainable supply of anti-infective compounds.},
  subject      = {Meeresschw{\"a}mme},
  language  = {en}
}