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The clinical and phenotypic features of the uncultivated agent of lacaziosis/lobomycosis in dolphins suggested that this pathogen was the same organism as Lacazia loboi, which causes skin keloidal-like lesions in [...]

Host-specific microbial communities thrive within sponge tissues and this association between sponge and associated microbiota may be driven by the organohalogen chemistry of the sponge animal. Several sponge species produce diverse organobromine secondary metabolites (e.g. brominated phenolics, indoles, and pyrroles) that may function as a chemical defense against microbial fouling, infection or predation. In this study, anaerobic cultures prepared from marine sponges were amended with 2,6-dibromophenol as the electron acceptor and short chain organic acids as electron donors. We observed reductive dehalogenation from diverse sponge species collected at disparate temperate and tropical waters suggesting that biogenic organohalides appear to enrich for populations of dehalogenating microorganisms in the sponge animal. Further enrichment by successive transfers with 2,6-dibromophenol as the sole electron acceptor demonstrated the presence of dehalogenating bacteria in over 20 sponge species collected from temperate and tropical ecoregions in the Atlantic and Pacific Oceans and the Mediterranean Sea. The enriched dehalogenating strains were closely related to Desulfoluna spongiiphila and Desulfoluna butyratoxydans , suggesting a cosmopolitan association between Desulfoluna spp. and various marine sponges. In vivo reductive dehalogenation in intact sponges was also demonstrated. Organobromide-rich sponges may thus provide a specialized habitat for organohalide-respiring microbes and D. spongiiphila and/or its close relatives are responsible for reductive dehalogenation in geographically widely distributed sponge species. [ABSTRACT FROM AUTHOR]

The genus Pseudoblepharisma is currently comprised of only one species, P. tenue, and one variant, P. tenue var. viride, both described as free-living ciliates thriving in oxygen depleted freshwater habitats of Europe. Here we report on this genus being discovered from subtropical freshwaters of Florida, USA. The Florida strain diverges from the P. tenue by being much larger, as well as having a far higher density of intracellular symbiotic green algae, resembling P. tenue var. viride. Unlike its European counterparts, the North American strain was observed to build a lorica, likely useful for protection and feeding; this has not been previously described for this genus. In contrast to P. tenue, the Florida strain does not have endosymbiotic purple bacteria. Despite large distances between sampling zones, the species is a close match at a morphological level to Pseudoblepharisma tenue var. viride. We provide the first 18S rRNA gene sequence for this species, allowing future investigations into the biogeography of this genus. As intensive sampling efforts continue to increase, cryptic microbial species will continue to be recorded from diverse freshwater habitats at a global scale. [ABSTRACT FROM AUTHOR]

The Indian River Lagoon, located on the east coast of Florida, USA, is an Estuary of National Significance and an important economic and ecological resource. The Indian River Lagoon faces several environmental pressures, including freshwater discharges through the St. Lucie Estuary; accumulation of anoxic, fine-grained, organic-rich sediment; and metal contamination from agriculture and marinas. Although the Indian River Lagoon has been well-studied, little is known about its microbial communities; thus, a two-year 16S amplicon sequencing study was conducted to assess the spatiotemporal changes of the sediment bacterial and archaeal groups. In general, the Indian River Lagoon exhibited a prokaryotic community that was consistent with other estuarine studies. Statistically different communities were found between the Indian River Lagoon and St. Lucie Estuary due to changes in porewater salinity causing microbes that require salts for growth to be higher in the Indian River Lagoon. The St. Lucie Estuary exhibited more obvious prokaryotic seasonality, such as a higher relative abundance of Betaproteobacteriales in wet season and a higher relative abundance of Flavobacteriales in dry season samples. Distance-based linear models revealed these communities were more affected by changes in total organic matter and copper than changes in temperature. Anaerobic prokaryotes, such as Campylobacterales, were more associated with high total organic matter and copper samples while aerobic prokaryotes, such as Nitrosopumilales, were more associated with low total organic matter and copper samples. This initial study fills the knowledge gap on the Indian River Lagoon bacterial and archaeal communities and serves as important data for future studies to compare to determine possible future changes due to human impacts or environmental changes. [ABSTRACT FROM AUTHOR]

The global prevalence of drug resistance has created an urgent need for the discovery of novel anti-infective drugs. The major source of antibiotics in current clinical practice is terrestrial actinobacteria; the less-exploited deep-sea actinobacteria may serve as an unprecedented source of novel natural products. In this study, we evaluated 50 actinobacteria strains derived from diverse deep water sponges and environmental niches for their anti-microbial activities against a panel of pathogens including Candida albicans, Clostridium difficile, Staphylococcus aureus, and methicillin-resistant S. aureus (MRSA), and Pseudomonas aeruginosa. More than half of the tested strains (27) were identified as active in at least one assay. The rare earth salt lanthanum chloride (LaCl3) was shown to be as an effective elicitor. Among the 27 strains, the antimicrobial activity of 15 were induced or enhanced by the addition of LaCl3. This part of study focused on one strain R818, in which potent antifungal activity was induced by the addition of LaCl3. We found that the LaCl3-activated metabolites in R818 are likely antimycin-type compounds. One of them, compound 1, has been purified. Spectroscopic analyses including HR-MS and 1D NMR indicated that this compound is urauchimycin D. The antifungal activity of compound 1 was confirmed with a minimal inhibitory concentration (MIC) of 25 µg/mL; the purified compound also showed a moderate activity against C. difficile. Additional notable strains are: strain N217 which showed both antifungal and antibacterial (including P. aeruginosa) activities and strain M864 which showed potent activity against C. difficile with an MIC value (0.125 µg/mL) lower than those of vancomycin and metronidazole. Our preliminary studies show that deep-sea actinobacteria is a promising source of anti-infective natural products. [ABSTRACT FROM AUTHOR]

A deep-water sponge of the genus Spongosorites has yielded a bis-indole alkaloid which we have named dragmacidin G. Dragmacidin G was first reported by us in the patent literature and has recently been reported by Hitora et al. from a sponge of the genus Lipastrotheya. Dragmacidin G is the first in this series of compounds to have a pyrazine ring linking the two indole rings. It also has a rare N-(2-mercaptoethyl)-guanidine side chain. Dragmacidin G shows a broad spectrum of biological activity including inhibition of methicillin-resistant Staphylococcus aureus, Mycobacterium tuberculosis, Plasmodium falciparum, and a panel of pancreatic cancer cell lines. [ABSTRACT FROM AUTHOR]

A novel cdc25A inhibitor was isolated from the marine sponge Dysidea etheria de Laubenfels as the search for new protein phosphatase inhibitors from marine organisms continues. Named dysidiolide, the inhibitor is a sesterterpene gamma-hydroxybutenolide, whose structure represents an unusual cyclization of a C(sub 25) isoprenoid. Dysidiolide inhibited the dephosphorylation of p-nitrophenol phosphate by cdc25A with an IC(sub 50) of 9.4 (mu)M. This represents the first account of a cdc25A active compound isolated from a natural source.

Lobomycosis (lacaziosis) is a chronic, granulomatous, fungal infection of the skin and subcutaneous tissues of humans and dolphins. To date, the causative agent, the yeastlike organism Lacazia loboi, has not been grown in the laboratory, and there have been no recent reports describing attempts to culture the organism. As a result, studies on the efficacy of therapeutics and potential environmental reservoirs have not been conducted. Therefore, the objective of the current study was to utilize both classical and novel microbiological methods in order to stimulate growth of Lacazia cells collected from dolphin lesions. This included the experimental inoculation of novel media, cell culture, and the use of artificial skin matrices. Although unsuccessful, the methods and results of this study provide important insight into new approaches that could be utilized in future investigations of this elusive organism. [ABSTRACT FROM AUTHOR]

Abstract: NPAS4 is a brain restricted, activity-induced transcription factor which regulates the expression of inhibitory synapse genes to control homeostatic excitatory/inhibitory balance in neurons. NPAS4 is required for normal social interaction and contextual memory formation in mice. Protein and mRNA expression of NPAS4 is tightly coupled to neuronal depolarization and most prevalent in the cortical and hippocampal regions in the brain, however the precise mechanisms by which the NPAS4 gene is controlled remain unexplored. Here we show that expression of NPAS4 mRNA is actively repressed by RE-1 silencing transcription factor/neuron-restrictive silencer factor (REST/NRSF) in embryonic stem cells and non-neuronal cells by binding multiple sites within the promoter and Intron I of NPAS4. Repression by REST also appears to correlate with the binding of the zinc finger DNA binding protein CTCF within Intron I of NPAS4. In addition, we show that the 3′ untranslated region (3′UTR) of NPAS4 can be targeted by two microRNAs, miR-203 and miR-224 to further regulate its expression. miR-224 is a midbrain/hypothalamus enriched microRNA which is expressed from an intron within the GABAA receptor epsilon (GABRE) gene and may further regionalize NPAS4 expression. Our results reveal REST and microRNA dependent mechanisms that restrict NPAS4 expression to the brain. [Copyright &y& Elsevier]

The synthesis of six thiadiazole nucleoside analogs is reported: 5-diacetylamino-1,2,4-thiadiazol-3-one (1), 5-amino-2-(tetrahydrofuran-2-yl)-1,2,4-thiadiazol-3-one (2), 5-amino-3-[(2'-hydroxyethoxy)methyl]-1,3,4-thiadiazol-2-one (3), 5-amino-3-(4'-hydroxy-2'-hydroxymethyl-butyl)-1,3,4-thiadiazole-2-thione (4), (R)-5-amino-3-(2', 3'-dihydroxypropyl)-1,3,4-thiadiazole- 2-thione (5), and (S)-5-amino-3-(2', 3'-dihydroxypropyl)-1,3,4-thiadiazole-2-thione (6). The synthesis, characterization, and properties of these new synthesized thiadiazole derivatives are discussed. A dimerization of 5-amino-3H-1,3,4-thiadiazole-2- thione (14) by sodium nitrite resulting in di-(5-amino-1,3,4-thiadiazol-2-yl) disulfide (19) is also reported. The preliminary in vitro evaluation of these newly synthesized compounds is discussed. [ABSTRACT FROM AUTHOR]

The article discusses a study on the health of brush-tailed rock-wallabies prior and after being released into a natural reserve. The study was conducted on the 41 wallabies that were sent into the Grampians National Park, Victoria. The wallabies were examined in terms of health factors including blood content, weight, and physical fitness. The brush-tailed rock-wallabies had radio collars attached to them to monitor their health since they live in the park. Results show that 70 percent of reintroduced wallabies had signs of macropod herpes virus, although the study did not reveal cases of oral cavity disease on the animals.

Bacillus strains with >99.7% 16S rRNA gene sequence similarity were characterized with DNA:DNA hybridization, cellular fatty acid (CFA) analysis, and testing of 100 phenotypic traits.When paired with the most closely related type strain, percent DNA:DNA similarities (% S) for six Bacillus strains were all far below the recommended 70% threshold value for species circumscription with Bacillus nealsonii. An apparent genomic group of four Bacillus strain pairings with 94%-70% S was contradicted by the failure of the strains to cluster in CFA- and phenotype-based dendrograms as well as by their differentiation with 9-13 species level discriminators such as nitrate reduction, temperature range, and acid production from carbohydrates. The novel Bacillus strains were monophyletic and very closely related based on 16S rRNA gene sequence. Coherent genomic groups were not however supported by similarly organized phenotypic clusters. Therefore, the strains were not effectively circumscribed within the taxonomic species definition. [ABSTRACT FROM AUTHOR]

Vibrio is a diverse genus of marine-associated bacteria with at least 74 species and more expected as additional marine ecospheres are interrogated. This report describes a phylogenetic reconstruction of Vibrio isolates derived from one such unique ecosystem, marine sponges (Phylum Porifera) collected from depths of 150 to 1242 feet. 16S rRNA gene sequencing along with molecular typing of 16S-23S rRNA intergenic spacer regions clustered many sponge-associated Vibrio (spp) with current known species. That is, several benthic Vibrio species commensal with Porifera sponges seemed genetically linked to vibrios associated with coastal or shallow-water communities, signalling a panmictic population structure among seemingly ecologically disparate strains. Conversely, phylogenetic analysis provided evidence for at least two novel Vibrio speciation events within this specific sponge microcosm. Collectively, these findings earmark this still relatively unknown environment as a bastion of taxonomic and phylogenetic variability for the genus and probably other bacterial taxa. [ABSTRACT FROM AUTHOR]

Marine sediments and sponges may show steep variations in redox potential, providing niches for both aerobic and anaerobic microorganisms. Geodia spp. and sediment specimens from the Straits of Florida were fixed using paraformaldehyde and 95% ethanol (v/v) for fluorescence in situ hybridization (FISH). In addition, homogenates of sponge and sediment samples were incubated anaerobically on various cysteine supplemented agars. FISH analysis showed a prominent similarity of microbiota in sediments and Geodia spp. samples. Furthermore, the presence of sulfate-reducing and annamox bacteria as well as other obligate anaerobic microorganisms in both Geodia spp. and sediment samples were also confirmed. Anaerobic cultures obtained from the homogenates allowed the isolation of a variety of facultative anaerobes, primarily Bacillus spp. and Vibrio spp. Obligate anaerobes such as Desulfovibrio spp. and Clostridium spp. were also found. We also provide the first evidence for a culturable marine member of the Chloroflexi, which may enter into symbiotic relationships with deep-water sponges such as Geodia spp. Resuspended sediment particles, may provide a source of microorganisms able to associate or form a symbiotic relationship with sponges. [ABSTRACT FROM AUTHOR]

The lithistid sponge Discodermia dissoluta (family Theonellidae), is found in deep-waters throughout the Caribbean sea and is the source of discodermolide, a natural product with potential anticancer properties, and other secondary metabolites. As with other sponges, large numbers of microbes are harbored in the sponge mesohyl. The microbial population of the sponge mesohyl shows an abundance of large filamentous microbes. Fractionation of the dissociated sponge allowed enrichment of this microbe, which was then identified by analysis of the 16S rRNA genes. Its identity was confirmed through the use of fluorescent in situ hybridization. These studies have allowed the identification of this eubacterial microbe as belonging to the genus Entotheonella.The ISME Journal (2008) 2, 335–339; doi:10.1038/ismej.2007.91; published online 7 February 2008 [ABSTRACT FROM AUTHOR]

This study examined the symbiotic microbiota of the hexacoral Cirrhipathes lutkeni using traditional plate culture, fluorescence in situ hybridization (FISH) and 16S rDNA characterization. FISH counts for the whole coral (holobiont) showed a major presence of γ-Proteobacteria (22%) and Actinobacteria (19%), followed by α-Proteobacteria (14%), Firmicutes (9%), Cytophaga-Flavobacterium (7%), β-Proteobacteria (6%) and Chloroflexi (2%). In contrast to the diversity observed by FISH, plate cultures were found to be selective for γ-Proteobacteria (22 cultures) with the exception of an Actinobacterium. The methods employed in this study detected 76% of all microbes estimated by DAPI staining of C. lutkeni homogenates. The absence of zooxanthellae in this particular hexacoral was confirmed by PCR and spectrophotometry using fresh tissue isolated from the holobiont. This is the first study describing the microbial associations of shallow-water hexacorallia, which opens further insight into coral microbial ecology and may enhance the search for novel natural products in the near future.The ISME Journal (2007) 1, 654–659; doi:10.1038/ismej.2007.77; published online 18 October 2007 [ABSTRACT FROM AUTHOR]

Most isolates recovered from marine environments are Gram-negative proteobacteria, even with the use of various media and media additions to enhance recoverability. Cultivation studies with two genera of deep-water sponges yielded nine isolates that demonstrated bulbous branching rod morphology, which is usually associated with microorganisms staining Gram-positive. Gram reactions indicated that the isolates were Gram-negative, which was confirmed by partial 16S rDNA sequencing. All nine isolates were shown to be α-proteobacteria most closely related to other α-proteobacteria isolated from various sponges. [Copyright &y& Elsevier]

Production of sponge-derived bioactive compounds in vitro has been proposed as an alternative to wild harvest, aquaculture, and chemical synthesis to meet the demands of clinical drug development and manufacture. Until recently, this was not possible because there were no marine invertebrate cell lines. Recent breakthroughs in the development of sponge cell lines and rapid cell division in improved nutrient media now make this approach a viable option. We hypothesized that three-dimensional (3-D) cell cultures would better represent how sponges function in nature, including the production of bioactive compounds. We successfully cultured sponge cells in 3-D matrices using FibraCel® disks, thin hydrogel layers, and gel microdroplets (GMDs). For in vitro production of bioactive compounds, the use of GMDs is recommended. Nutrients and sponge products rapidly diffuse into and out of the 3-D matrix, the GMDs may be scaled up in spinner flasks, and cells and/or secreted products can be easily recovered. Research on scale-up and production is in progress in our laboratory. [ABSTRACT FROM AUTHOR]

'Flagship' ciliates were investigated from soil samples collected in Florida, USA. This was undertaken to determine if species thought to be restricted to a given world region could be uncovered from similar habitats in a novel location, e.g. another continent. Two species of Condylostomides were discovered, and recorded from the North American continent for the first time. Condylostomides etoschensis was known only from Africa, but was found to be thriving in a Florida study site. An 18S rDNA sequence for this species was determined for the first time. Also discovered from the same study site was the ciliate Condylostomides coeruleus , previously known only from Central and South America. These two 'flagship' ciliates were found in the same habitat, from a continent well outside of their previously recorded biogeographies. Molecular sequencing and microscopy investigations were conducted to form the baseline for future work within this genus. Soil ciliates can obtain large population numbers and form cysts and are therefore likely able to disperse globally. These new records provide additional evidence that large distances, even between continents, do not hinder microbes from thriving globally. The absence of these conspicuously-colored gold and blue ciliates from previous studies is likely due to undersampling, rather than to any physical barriers. [ABSTRACT FROM AUTHOR]

Vibrio represents a diverse bacterial genus found in different niches of the marine environment, including numerous genera of marine sponges (phylum Porifera), inhabiting different depths and regions of benthic seas, that are potentially important in driving adaptive change among Vibrio spp. Using 16S rRNA gene sequencing, a previous study showed that sponge-derived (SD) vibrios clustered with their mainstream counterparts present in shallow, coastal ecosystems, suggesting a genetic relatedness between these populations. Sequences from the topA, ftsZ, mreB, rpoD, rctB and toxR genes were used to investigate the degree of relatedness existing between these two separate populations by examining their phylogenetic and genetic disparity. Phylogenies were constructed from the concatenated sequences of the six housekeeping genes using maximum-parsimony, maximum-likelihood and neighbour-joining algorithms. Genetic recombination was evaluated using the incongruence length difference test, Split decomposition and measuring overall compatibility of sites. This combined technical approach provided evidence that SD Vibrio strains are largely genetically homologous to their shallow-water counterparts. Moreover, the analyses conducted support the existence of extensive horizontal gene transfer between these two groups, supporting the idea of a single panmictic population structure among vibrios from two seemingly distinct, marine environments. [ABSTRACT FROM AUTHOR]

Increases in resistance to commonly used antibiotics have been reported globally in isolates from humans, wildlife, and the environment. To date, few studies have examined long-term trends in antibiotic resistance in organisms isolated from marine mammal populations. The objective of this study was to examine temporal trends in resistance to antibiotics among pathogens isolated from common bottlenose dolphins (Tursiops truncatus) between 2003 and 2015. Dolphins were captured and released in the Indian River Lagoon, Florida, an ecosystem with a large coastal human population and significant environmental impacts. Swab samples for microbiology were taken from the blowhole, gastric fluid, and feces and cultured on standard media under aerobic conditions. Isolates were identified using gram stain morphology and growth on selective media. Antibiotic resistance was measured using disc diffusion on Mueller Hinton agar and the Multiple Antibiotic Resistance (MAR) index calculated for each pathogen. A total of 733 isolates was obtained from 171 individual dolphins. The most commonly cultured pathogens included Aeromonas hydrophila, Escherichia coli, Edwardsiella tarda, and Vibrio alginolyticus. The overall prevalence of resistance to at least one antibiotic for the 733 isolates was 88.2%. The MAR index increased significantly between 2003 and 2007 and 2010 and 2015 for Pseudomonas aeruginosa and V. alginolyticus. For all bacterial isolates, resistance to cefotaxime, ceftazidime, and gentamicin increased significantly between sampling periods. This is one of few studies to use the MAR index for bacterial isolates from a marine mammal. The significant increases in resistance for some bacterial species likely reflect shared environmental exposures to antibiotics and transfer of resistance to dolphins from terrestrial sources or from animal or human populations. [ABSTRACT FROM AUTHOR]