Natural Products (NP) are essential for the discovery of novel drugs and products for numerous biotechnological applications. The NP discovery process is expensive and time-consuming, having as major ...hurdles dereplication (early identification of known compounds) and structure elucidation, particularly the determination of the absolute configuration of metabolites with stereogenic centers. This review comprehensively focuses on recent technological and instrumental advances, highlighting the development of methods that alleviate these obstacles, paving the way for accelerating NP discovery towards biotechnological applications. Herein, we emphasize the most innovative high-throughput tools and methods for advancing bioactivity screening, NP chemical analysis, dereplication, metabolite profiling, metabolomics, genome sequencing and/or genomics approaches, databases, bioinformatics, chemoinformatics, and three-dimensional NP structure elucidation.
Although the rise of antibiotic and multidrug resistant bacteria is one of the biggest current threats to human health, our understanding of the mechanisms involved in antibiotic resistance selection ...remains scarce. We performed whole genome sequencing of 21 Pseudomonas strains, previously isolated from an active submarine volcano of Greece, the Kolumbo volcano. Our goal was to identify the genetic basis of the enhanced co-tolerance to antibiotics and acidity of these Pseudomonas strains. Pangenome analysis identified 10,908 Gene Clusters (GCs). It revealed that the numbers of phage-related GCs and sigma factors, which both provide the mechanisms of adaptation to environmental stressors, were much higher in the high tolerant Pseudomonas strains compared to the rest ones. All identified GCs of these strains were associated with antimicrobial and multidrug resistance. The present study provides strong evidence that the CO
-rich seawater of the volcano associated with low pH might be a reservoir of microorganisms carrying multidrug efflux-mediated systems and pumps. We, therefore, suggest further studies of other extreme environments (or ecosystems) and their associated physicochemical parameters (or factors) in the rise of antibiotic resistance.
Background: The distribution of microorganisms, and especially pathogens, over airborne particles of different sizes has been ignored to a large extent, but it could have significant implications ...regarding the dispersion of these microorganisms across the planet, thus affecting human health. Objectives: We examined the microbial quality of the aerosols over the eastern Mediterranean region during an African storm to determine the size distribution of microorganisms in the air. Methods: We used a five-stage cascade impactor for bioaerosol collection in a coastal city on the eastern Mediterranean Sea during a north African dust storm. Bacterial communities associated with aerosol particles of six different size ranges were characterized following molecular culture-independent methods, regardless of the cell culturability (analysis of 16S rRNA genes). Results: All 16S rDNA clone libraries were diverse, including sequences commonly found in soil and marine ecosystems. Spore-forming bacteria such as Firmicutes dominated large particle sizes (> 3.3 micrometers), whereas clones affiliated with Actinobacteria (found commonly in soil) and Bacteroidetes (widely distributed in the environment) gradually increased their abundance in aerosol particles of reduced size (< 3.3 micrometers). A large portion of the clones detected at respiratory particle sizes (< 3.3 micrometers) were phylogenetic neighbors to human pathogens that have been linked to several diseases. Conclusions: The presence of aerosolized bacteria in small size particles may have significant implications to human health via intercontinental transportation of pathogens.
The majority of studies focusing on microbial functioning in various environments are based on DNA or RNA sequencing techniques that have inherent limitations and usually provide a distorted picture ...about the functional status of the studied system. Untargeted proteomics is better suited for that purpose, but it suffers from low efficiency when applied in complex consortia. In practice, the scanning capabilities of the currently employed LC-MS/MS systems provide limited coverage of key-acting proteins, hardly allowing a semiquantitative assessment of the most abundant ones from most prevalent species. When particular biological processes of high importance are under investigation, the analysis of specific proteins using targeted proteomics is a more appropriate strategy as it offers superior sensitivity and comes with the added benefits of increased throughput, dynamic range and selectivity. However, the development of targeted assays requires
a priori
knowledge regarding the optimal peptides to be screened for each protein of interest. In complex, multi-species systems, a specific biochemical process may be driven by a large number of homologous proteins having considerable differences in their amino acid sequence, complicating LC-MS/MS detection. To overcome the complexity of such systems, we have developed an automated pipeline that interrogates UniProt database or user-created protein datasets (e.g. from metagenomic studies) to gather homolog proteins with a defined functional role and extract respective peptide sequences, while it computes several protein/peptide properties and relevant statistics to deduce a small list of the most representative, process-specific and LC-MS/MS-amenable peptides for the microbial enzymatic activity of interest.
Given the dramatic increase in the L. sceleratus population in the southeastern Aegean Sea, there is growing interest in assessing the toxicity of this pufferfish and the factors controlling its ...tetrodotoxin (TTX) content. In the present study, liver, gonads, muscle and skin of 37 L. sceleratus specimens collected during May and June 2021 from the island of Rhodes, Greece, were subjected to multi-analyte profiling using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in order to quantitate TTX and evaluate whether this biotoxin interrelates with hormones. TTX and its analogues 4-epiTTX, 11-deoxyTTX, 11-norTTX-6-ol, 4,9-anhydroTTX and 5,11/6,11-dideoxyTTX were detected in all tissue types. Liver and gonads were the most toxic tissues, with the highest TTX concentrations being observed in the ovaries of female specimens. Only 22% of the analyzed muscle samples were non-toxic according to the Japanese toxicity threshold (2.2 μg TTX eq g−1), confirming the high poisoning risk from the inadvertent consumption of this species. Four steroid hormones (i.e., cortisol, testosterone, androstenedione and β-estradiol) and the gonadotropin-releasing hormone (GnRH) were detected in the gonads. Androstenedione dominated in female specimens, while GnRH was more abundant in males. A positive correlation of TTX and its analogues with β-estradiol was observed. However, a model incorporating sex rather than β-estradiol as the independent variable proven to be more efficient in predicting TTX concentration, implying that other sex-related characteristics are more important than specific hormone-regulated processes.
The administration of antibiotics in aquaculture has raised concern about the impact of their overuse in marine ecosystems, seafood safety and consumers' health. This "green consumerism" has forced ...researchers to find new alternatives against fish pathogens. The present study focused on 12 Mediterranean medicinal-aromatic plants as potential antimicrobials and antioxidant agents that could be used in fish aquaculture.
assays showed that the essential oils (EOs) from all studied plants had anti-bacterial and antioxidant properties, with their efficacy being dependent on their chemical composition. More specifically, EOs rich in carvacrol, p-cymene and γ-terpinene exhibited not only the strongest inhibitory activity against the growth of bacterial pathogens (inhibitory concentration: 26-88 μg mL
), but also the greatest total antioxidant capacity (ABTS: 2591-5879 μmole mL
; CUPRAC: 931-2733 μmole mL
). These compounds were mainly found in the EOs from Greek oregano (
subsp.
), Spanish oregano (
) and savoury (
) collected from cultivations in Greece. The specific EOs stand out as promising candidates for the treatment of bacterial diseases and oxidative stress in farmed fish. Further
experiments are needed to fully understand the effects of EO dietary supplementation on fish farming processes.
•We examined the pollution status of deep-sea sediments in the southern Cretan margin.•AHs, UCM, PAHs, PCBs and chlorinated pesticides were ubiquitous in sediments.•AHs and PAHs were of ...terrestrial-biogenic and pyrolytic origin, respectively.•The presence of PCBs and HCHs was attributed to historical emission sources.•An on-going, fresh input of DDT was revealed.
Deep sediments from the southern Cretan margin were analyzed to establish baseline levels for various types of organic pollutants before the anticipated intensification of anthropogenic activities. The total concentration of aliphatic hydrocarbons (ΣAH:326–3758ngg−1, dry weight) was similar to those reported for deep sediments of the western Mediterranean Sea, while considerably lower levels were measured for polycyclic aromatic hydrocarbons (ΣPAH:9–60ngg−1). Source-diagnostic ratios suggested that the aliphatic hydrocarbons in sediments were mainly of terrestrial biogenic origin, while polycyclic aromatic hydrocarbons stemmed from the deposition of long-range transported combustion aerosols. Among the organochlorinated compounds analyzed, β-hexachlorocyclohexane (β-HCH:222–7052pgg−1), 1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (p,p′-DDT:37–2236pgg−1) and polychlorinated biphenyls (ΣPCB:38–1182pgg−1) showed the highest abundance in sediments. The presence of HCHs and PCBs was attributed to historical inputs that have undergone extensive weathering, whereas an ongoing fresh input was suggested for p,p′-DDT. Multiple linear regression analysis revealed that the levels of the various pollutants in sediments were controlled by different factors, but with organic carbon content playing a prominent role in most cases.
The invasion of the tetrodotoxin (TTX)-bearing silver-cheeked toadfish and potential poisoning due to its consumption (tetrodotoxication) threatens public safety in the Mediterranean Sea. In this ...study, TTX and TTX analogues of
(Gmelin, 1789) were measured using liquid chromatography tandem mass spectrometry (LC-MS/MS) in fish collected off the island of Crete (Southern Mediterranean). We tested the synergistic effect of a suite of factors potentially affecting toxins' levels and tetrodotoxication risk using general and generalized linear models, respectively. The type of tissue, geographic origin (Cretan Sea, Libyan Sea), sex, and fish maturity stage were significant predictors of toxin concentrations. Mean TTX was higher in gonads and lower in muscles, higher in the Libyan Sea and in female fish, and lower in juvenile (virgin) fish. The concentration of TTX was also significantly and positively correlated with the concentration of several TTX analogues (4-epiTTX, 4,9-anhydroTTX, 11-deoxyTTX, 5,11/6,11-dideoxyTTX, 5,6,11-trideoxyTTX, 11-norTTX-6-ol). The analysis showed that fish originating from the Libyan Sea had significantly higher probability to cause tetrodotoxication in case of consumption. The variability explained by the models developed in this study was relatively low, indicating that toxin levels are hard to predict and the consumption of
should therefore be avoided.
Despite progress achieved, there is limited available information about the antibacterial activity of constituents of essential oils (EOs) from different medicinal-aromatic plants (MAPs) against fish ...pathogens and the complex interactions of blended EOs thereof. The present study aimed to investigate possible synergistic antimicrobial effects of EOs from seven Greek MAPs with strong potential against
bv.
a fish pathogen associated with aquaculture disease outbreaks. The main objective was to evaluate whether blending of these EOs can lead to increased antimicrobial activity against the specific microorganism. A total of 127 combinations of EOs were prepared and their effect on
bv.
growth was tested in vitro. We examined both the inhibitory and bactericidal activities of the individual EOs and compared them to those of the blended EOs. The vast majority of the investigated combinations exhibited significant synergistic and additive effects, while antagonistic effects were evident only in a few cases, such as the mixtures containing EOs from rosemary, lemon balm and pennyroyal. The combination of EOs from Greek oregano and wild carrot, as well as the combinations of those two with Spanish oregano or savoury were the most promising ones. Overall, Greek oregano, savoury and Spanish oregano EOs were the most effective ones when applied either in pure form or blended with other EOs.
Tetrodotoxin (TTX) is a potent marine neurotoxin involved in poisoning cases, especially through the consumption of puffer fish. Knowledge of the toxicity equivalency factors (TEFs) of TTX analogues ...is crucial in monitoring programs to estimate the toxicity of samples analyzed with instrumental analysis methods. In this work, TTX analogues were isolated from the liver of a Lagocephalus sceleratus individual caught on South Crete coasts. A cell-based assay (CBA) for TTXs was optimized and applied to the establishment of the TEFs of 5,11-dideoxyTTX, 11-norTTX-6(S)-ol, 11-deoxyTTX and 5,6,11-trideoxyTTX. Results showed that all TTX analogues were less toxic than the parent TTX, their TEFs being in the range of 0.75–0.011. Then, different tissues of three Lagocephalus sceleratus individuals were analyzed with CBA and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The obtained TEFs were applied to the TTX analogues’ concentrations obtained by LC-MS/MS analysis, providing an indication of the overall toxicity of the sample. Information about the TEFs of TTX analogues is valuable for food safety control, allowing the estimation of the risk of fish products to consumers.
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