Metabolomic Insights of Biosurfactant Activity from Bacillus niabensis against Planktonic Cells and Biofilm of Pseudomonas stutzeri Involved in Marine Biofouling.

In marine environments, biofilm can cause negative impacts, including the biofouling process. In the search for new non-toxic formulations that inhibit biofilm, biosurfactants (BS) produced by the genus Bacillus have demonstrated considerable potential. To elucidate the changes that BS from B. niabensis promote in growth inhibition and biofilm formation, this research performed a nuclear magnetic resonance (NMR) metabolomic profile analysis to compare the metabolic differences between planktonic cells and biofilms of Pseudomonas stutzeri, a pioneer fouling bacteria. The multivariate analysis showed a clear separation between groups with a higher concentration of metabolites in the biofilm than in planktonic cells of P. stutzeri. When planktonic and biofilm stages were treated with BS, some differences were found among them. In planktonic cells, the addition of BS had a minor effect on growth inhibition, but at a metabolic level, NADP+, trehalose, acetone, glucose, and betaine were up-regulated in response to osmotic stress. When the biofilm was treated with the BS, a clear inhibition was observed and metabolites such as glucose, acetic acid, histidine, lactic acid, phenylalanine, uracil, and NADP+ were also up-regulated, while trehalose and histamine were down-regulated in response to the antibacterial effect of the BS.

Exploring Antifouling Activity of Biosurfactants Producing Marine Bacteria Isolated from Gulf of California.

Biofouling causes major problems and economic losses to marine and shipping industries. In the search for new antifouling agents, marine bacteria with biosurfactants production capability can be an excellent option, due to the amphipathic surface-active characteristic that confers antimicrobial and antibiofilm activities. The aim of this study was to evaluate the antifouling activity of biosurfactants producing marine bacteria from the Gulf of California. The cell free culture supernatant (CFCS) of Bacillus niabensis (S-69), Ralstonia sp. (S-74) (isolated from marine sediment) and of B. niabensis (My-30) (bacteria associated to the sponge Mycale ramulosa) were screened for production of biosurfactants (using hemolysis and drop collapse test, oil displacement and emulsifying activity). The toxicity and antifouling activity were evaluated against biofoulers (bacteria forming biofilm and macrofoulers) both in laboratory and field assays. The results indicate that all bacteria were biosurfactant producers, but the higher capability was shown by B. niabensis (My-30) with high emulsifying properties (E24) of 71%. The CFCS showed moderate toxicity but were considered non-toxic against Artemia franciscana at low concentrations. In the antifouling assay, the CFCS of both strains of B. niabensis showed the best results for the reduction of the biofilm formation (up 50%) against all Gram-positive bacteria and most Gram-negative bacteria with low concentrations. In the field assay, the CFCS of B. niabensis (My-30) led to the reduction of 30% of biofouling compared to the control. The results indicate that the biosurfactant produced by B. niabensis (My-30) has promising antifouling activity.

Exploring the chemical diversity of Capsicum chinense cultivars using NMR-based metabolomics and machine learning methods.

The habanero pepper (Capsicum chinense) is a prominent spicy fruit integral to the historical, social, cultural, and economic fabric of the Yucatan peninsula in Mexico. This study leverages the power of 1H NMR spectroscopy coupled with machine learning algorithms to dissect the metabolomic profile of eleven C. chinense cultivars, including those grown by INIFAP (Habanero-Jaguar, Antillano-HRA 1-1, Antillano-HRA 7-1, Habanero-HAm-18A, Habanero-HC-23C, and Jolokia-NJolokia-22) and commercial hybrids (Habanero-Rey Votán, Habanero-Kabal, Balam, USAPR10117, and Rey Pakal). A total of fifty metabolites, encompassing sugars, amino acids, short-chain organic acids, and nucleosides, were identified from the 1H NMR spectra. The optimized machine learning model proficiently predicted the similarity percentage between the INIFAP-grown cultivars and commercial hybrids, thereby facilitating a comprehensive comparison. Biomarkers unique to each cultivar were delineated, revealing that the Habanero-Rey Votán cultivar is characterized by the highest concentration of sugars. In contrast, the Balam cultivar is rich in amino acids and short-chain organic acids, sharing a similar metabolomic profile with the Jolokia-NJolokia-22 cultivar. The findings of this study underscore the efficacy and reliability of NMR-based metabolomics as a robust tool for differentiating C. chinense cultivars based on their intricate chemical profiles. This approach not only contributes to the scientific understanding of the metabolomic diversity among habanero peppers but also holds potential implications for food science, agriculture, and the culinary arts.

Depth effect on the prokaryotic community assemblage associated with sponges from different rocky reefs.

Background: Sponge microbiomes are essential for the function and survival of their host and produce biologically active metabolites, therefore, they are ideal candidates for ecological, pharmacologic and clinical research. Next-generation sequencing (NGS) has revealed that many factors, including the environment and host, determine the composition and structure of these symbiotic communities but the controls of this variation are not well described. This study assessed the microbial communities associated with two marine sponges of the genera Aplysina (Nardo, 1834) and Ircinia (Nardo, 1833) in rocky reefs from Punta Arena de la Ventana (Gulf of California) and Pichilingue (La Paz Bay) in the coast of Baja California Sur, México to determine the relative importance of environment and host in structuring the microbiome of sponges. Methods: Specimens of Aplysina sp were collected by scuba diving at 10 m and 2 m; Ircinia sp samples were collected at 2 m. DNA of sponge-associated prokaryotes was extracted from 1 cm3 of tissue, purified and sent for 16S amplicon sequencing. Primer trimmed pair-ended microbial 16S rDNA gene sequences were merged using Ribosomal Database Project (RDP) Paired-end Reads Assembler. Chao1, Shannon and Simpson (alpha) biodiversity indices were estimated, as well permutational analysis of variance (PERMANOVA), and Bray-Curtis distances. Results: The most abundant phyla differed between hosts. Those phyla were: Proteobacteria, Acidobacteria, Cyanobacteria, Chloroflexi, Actinobacteria, Bacteroidetes, and Planctomycetes. In Ircinia sp the dominant phylum was Acidobacteria. Depth was the main factor influencing the microbial community, as analysis of similarities (ANOSIM) showed a significant difference between the microbial communities from different depths. Conclusion: Microbial diversity analysis showed that depth was more important than host in structuring the Aplysina sp and Ircinia sp microbiome. This observation contrast with previous reports that the sponge microbiome is highly host specific.

Bean cultivars (Phaseolus vulgaris L.) under the spotlight of NMR metabolomics.

The seeds of Phaseolus vulgaris are a rich source of protein consumed around the world and are considered as the most important source of proteins and antioxidants in the Mexican diet. This work reports on the 1H NMR metabolomics profiling of the cultivars Peruano (FPe), Pinto (FPi), Flor de mayo (FM), Negro (FN) and Flor de junio (FJ). Total phenolics, total flavonoids and total protein contents were determined to complement the nutritional facts in seeds and leaves. According to our results, the metabolomics fingerprint of beans seeds and leaves were very similar, showing the presence of 52 metabolites, 46 in seeds and 48 in leaves, including 8 sugars, 17 amino acids, 15 organic acids, 5 nucleosides and 7 miscellaneous compounds. In seeds, free amino acids were detected in higher concentrations than in the leaves, whereas organic acids were more abundant in leaves than in seeds. With multivariate and cluster analysis it was possible to rank the cultivars according to their nutritional properties according to NMR profiling, then a machine learning algorithm was used to reveal the most important differential metabolites which are the key for correct classification. The results coincide in highlighting the FN seeds and FPe leaves for the best nutritional facts. Finally, in terms of cultivars, FN and FM present the best nutritional properties, with high protein and flavonoids content, as well as, a high concentration of amino acids and nucleosides.

Marine Sediment Recovered Salinispora sp. Inhibits the Growth of Emerging Bacterial Pathogens and other Multi-Drug-Resistant Bacteria.

Marine obligate actinobacteria produce a wide variety of secondary metabolites with biological activity, notably those with antibiotic activity urgently needed against multi-drug-resistant bacteria. Seventy-five marine actinobacteria were isolated from a marine sediment sample collected in Punta Arena de La Ventana, Baja California Sur, Mexico. The 16S rRNA gene identification, Multi Locus Sequence Analysis, and the marine salt requirement for growth assigned seventy-one isolates as members of the genus Salinispora, grouped apart but related to the main Salinispora arenicola species clade. The ability of salinisporae to inhibit bacterial growth of Staphylococcus epidermidis, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacer baumannii, Pseudomonas aeruginosa, and Enterobacter spp. was evaluated by cross-streaking plate and supernatant inhibition tests. Ten supernatants inhibited the growth of eight strains of S. epidermidis from patients suffering from ocular infections, two out of the eight showed growth inhibition on ten S. epidermidis strains from prosthetic joint infections. Also, it inhibited the growth of the remaining six multi-drug-resistant bacteria tested. These results showed that some Salinispora strains could produce antibacterial compounds to combat bacteria of clinical importance and prove that studying different geographical sites uncovers untapped microorganisms with metabolic potential.

Experimental races of Capsicum annuum cv. jalapeño: Chemical characterization and classification by 1H NMR/machine learning.

This work reports on the metabolic fingerprinting of ten new races of Capsicum annuum cv. jalapeño using 1H NMR based metabolomics coupled to machine learning projections. Ten races were classified and evaluated according to their differential metabolites, variables of commercial interest and by multivariate data analysis/machine learning algorithm. According to our results, experimental races of jalapeño peppers exhibited differences in carbohydrate, amino acid, nucleotide and organic acid contents. Forty-eight metabolites were identified by 1D and 2D NMR and the differential metabolites were quantified by qNMR. Principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) separated the studied races into two groups. The group A included the races Colosus, Emperador, Fundador and Rayo whereas the group B included the races Don Benito, SMJ 1416, SMJ 1417, SMJ 1423, SMJ 145 and STAM J0904. OPLS-DA revealed that levels of citric acid in group A were higher than in group B, while the levels of asparagine, fumaric acid, GABA, glucose, malic acid, pyruvic, quinic acid, sucrose and tryptophan were higher in the group B. Remarkably, ascorbic acid was exclusively found in the race Colosus. Random forest model revealed the diversity of the experimental races and the similarity rate with the well-established races. The most relevant variables used to generate a model were length, weight, yield, width, xylose content and organic acids content.

Allelopathic effects of Margalefidinium polykrikoides and Gymnodinium impudicum in the growth of Gymnodinium catenatum.

Harmful algae blooms (HABs) are characterized for the coexistence of phytoplankton species with dynamic and complex biotic interactions (e.g., competition, symbiosis, predation, parasitism, allelopathy), that occur at fine temporal and spatial scales, and are relevant to understand the role that different species of phytoplankton play in the regulation of HABs. In this work the allelopathic effects of Margalefidinium polykrikoides (=Cochlodinium polykrikoides) and Gymnodinium impudicum on Gymnodinium catenatum were evaluated. The allelopathic abilities of M. polykrikoides and G. impudicum were investigated in bi-algal culture experiments and in trials in which target species were co-cultured, separated by a 10 µm membrane to prevent a direct cell-to-cell contact; and also by the addition of different volumes of culture media without cells. For all trials, cells of each species were harvested during exponential phase and cultured together by triplicate at three relative abundances: 1:1 (200 Cells mL-1 of each species, G. catenatum and M. polykrikoides or G. impudicum), 2:1 (400 Cells mL-1 of G. catenatum and 200 Cells mL-1 of M. polykrikoides or G. impudicum), and 1:2 (200 cells mL-1 of G. catenatum and 400 Cells mL-1 of M. polykrikoides or G. impudicum). All bioassays were carried out by triplicate in 250 mL Erlenmeyer flasks with 150 mL of modified GSe medium with an initial inoculum of 200 or 400 Cells mL-1. During experiments G. catenatum abundances were enumerated daily. In bi-algal culture experiments mortalities of G. catenatum were from 50% to 100% after 48 h of cell contact with M. polykrikoides or G. impudicum. In the case of culture media without cells, only M. polykrikoides caused a decrease in the cell abundance and growth rate of G. catenatum. Morphological changes occurred in G. catenatum when in contact with M. polykrikoides and G. impudicum, such as membrane shedding, prominent nucleus, loss of flagella, cell lysis, as well as the separation of long chains into individual cells. These results suggest that in the natural environment M. polykrikoides and G. impudicum have allelopathic interactions in G. catenatum, which could negatively affect its growth and survival, indicating that these species could displace blooms of G. catenatum.

Amended definitions for Aplysinidae and Aplysina (Porifera, Demospongiae, Verongiida): on three new species from a remarkable population in the Gulf of California.

Verongiid sponges inhabiting the La Paz region, Gulf of California are described herein as new species. Although morphological evidence was sufficient to determine the identity between species, we have confirmed their uniqueness and relationships with molecular (CO1 and ITS1 and 2), and ecological studies. An amended definition of family Aplysinidae and genus Aplysina is presented to highlight a novel skeletal trait for the latter, clearly described herein as a complex of dendritic fibers sustained by anastomosed fibers deep in the choanosome. This novel fiber arrangement combination is a constant trait of Aplysina encarnacionae sp. nov. and A. airapii sp. nov., which otherwise conform to our current concept of Aplysina. The former species has a long tubular morphology, reddish purple color, with the longest dendritic fibers; while A. airapii sp. nov. is a short tubular sponge, yellow with reddish tints, and smaller choanosomal dendritic fibers that depart from a uniplanar anastomosed skeleton. A third species, A. sinuscaliforniensis sp. nov., is characterized by a completely anastomosed skeleton, massive habit with short tubes superimposed one over the other, sympatric with the other new species. Comparisons with Suberea azteca verified that the same novel skeletal architecture described here occurs in the latter, while Aiolochroia thiona examined as well, proved to bear an anastomosed skeleton only, besides sharing more features with Aplysina than with any other genera in Verongiida. Moreover, molecular sequencing recovered "S." azteca nested in the Aplysina clade, prompting us to reallocate it in Aplysina, as originally proposed. The new species described herein are probably endemics within the Gulf of California since they have not been recorded elsewhere along the Mexican Pacific coast.

Allelopathic effect of Chattonella marina var. marina (Raphidophyceae) on Gymnodinium catenatum (Dinophycea).

The allelopathic effect of the raphidophyte Chattonella marina var. marina on the dinoflagellate Gymnodinium catenatum was determined. Both species are harmful algal bloom forming algae, produce toxic metabolites, and can co-exist in the environment. In general, raphidophytes tend to dominate over dinoflagellates, which may indicate an allelopathic effect of the former algae. Strains of C. marina var. marina and G. catenatum isolated from Bahía de La Paz were cultured in bi-algal cultures with and without cell contact. Additionally, cultures of G. catenatum were exposed to cell-free culture filtrates of the raphidophyte to test whether soluble allelopathic molecules are active. During late stationary phase, both species were cultivated in mixed cultures for 72h using the following cell abundance proportions: 20×103cellsL-1: 20×103cellsL-1 (1:1; G. catenatum: C. marina); 10×103cellsL-1: 20×103cellsL-1 (1:2), and 20×103cellsL-1: 10×103cellsL-1 (2:1). Cells of G. catenatum were also exposed to different volumes of cell filtrates of C. marina (10, 20, and 50mL) using the same cell abundance proportions for 24h. Samples were taken daily for cell counts and microscopic observations. Growth inhibition was higher when there was cell contact between both species, however mortality of G. catenatum was also observed without direct cell contact, indicating that toxic metabolites are liberated to the culture medium. Changes in cell morphology of G. catenatum occurred in the presence of cells and filtrates of C. marina, such as loss of flagella and motility, swelling, loss of girdle and sulci, prominent nucleus, rupture of cell membrane, and cell lysis. Induction of temporary cysts was also observed. These results suggest that toxic metabolites are liberated to the medium by C. marina, affecting G. catenatum by inhibiting its growth and causing changes in its life history, providing new insights of interactions between raphidophytes and dinoflagellates that could happen in the natural environment when both species are present.

Protocol for Assessing Antifouling Activities of Macroalgal Extracts.

The development of novel environmentally friendly antifouling (AF) solutions is a very active field in fundamental and applied research. An attractive option in producing such material resides in biomimetic studies: living organisms have evolved well-adapted structures and materials over geological times through natural selection. In this chapter, we explain the experimental procedure to be followed for the preparation of macroalgal extracts and to assess their AF efficiency towards key species. All bioassays described here have the advantage of being fast, reliable, and standardized.

Isothiocyanate sesquiterpenes from a sponge of the genus axinyssa.

The chemical study of a sponge of the genus Axinyssa collected in the Gulf of California has led to the isolation of the new bicyclic sesquiterpenes axinisothiocyanates A-L ( 1- 12) together with the known compounds (1 R,6 S,7 S,10 S)-10-isothiocyanato-4-amorphene ( 13), (4 R*,5 R*,7 S*,10 R*)-4-isocyanoeudesm-11-ene, (-)-epipolasin A, and (+)-aristolone. The structures of the new metabolites have been established by spectroscopic techniques, including the analysis of pyridine-induced 1H NMR chemical shifts. The cytotoxic activity has been tested against three human tumor cell lines.

Cytotoxic dibromotyrosine-derived metabolites from the sponge Aplysina gerardogreeni.

The chemical study of the sponge Aplysina gerardogreeni collected at the Gulf of California has led to the isolation of four new dibromotyrosine-derived metabolites, aplysinones A-D, whose structures were determined by spectroscopic analysis and chemical methods. The new compounds and four semisynthetic analogues prepared in this study have shown cytotoxic activity against human tumor cell lines.