National Committee on Marine Sciences (NCMS)
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- Multifaceted assessment of wastewater-based epidemiology for SARS-CoV-2 in selected urban communities in Davao City, Philippines: A pilot studyOtero, Maria Catherine B.; Murao, Lyre Anni E.; Limen, Mary Antoinette G.; Caalim, Daniel Rev A.; Gaite, Paul Lorenzo A.; Bacus, Michael G.; Acaso, Joan T.; Miguel, Refeim M.; Corazo, Kahlil; Knot, Ineke E.; Sajonia, Homer; de los Reyes, Francis L.; Jaraula, Caroline Marie B.; Baja, Emmanuel S.; Del Mundo, Dann Marie N. (MDPI, 2022-07-19)Over 60 countries have integrated wastewater-based epidemiology (WBE) in their COVID-19 surveillance programs, focusing on wastewater treatment plants (WWTP). In this paper, we piloted the assessment of SARS-CoV-2 WBE as a complementary public health surveillance method in susceptible communities in a highly urbanized city without WWTP in the Philippines by exploring the extraction and detection methods, evaluating the contribution of physico-chemical–anthropogenic factors, and attempting whole-genome sequencing (WGS). Weekly wastewater samples were collected from sewer pipes or creeks in six communities with moderate-to-high risk of COVID-19 transmission, as categorized by the City Government of Davao from November to December 2020. Physico-chemical properties of the wastewater and anthropogenic conditions of the sites were noted. Samples were concentrated using a PEG-NaCl precipitation method and analyzed by RT-PCR to detect the SARS-CoV-2 N, RdRP, and E genes. A subset of nine samples were subjected to WGS using the Minion sequencing platform. SARS-CoV-2 RNA was detected in twenty-two samples (91.7%) regardless of the presence of new cases. Cycle threshold values correlated with RNA concentration and attack rate. The lack of a sewershed map in the sampled areas highlights the need to integrate this in the WBE planning. A combined analysis of wastewater physico-chemical parameters such as flow rate, surface water temperature, salinity, dissolved oxygen, and total dissolved solids provided insights on the ideal sampling location, time, and method for WBE, and their impact on RNA recovery. The contribution of fecal matter in the wastewater may also be assessed through the coliform count and in the context of anthropogenic conditions in the area. Finally, our attempt on WGS detected single-nucleotide polymorphisms (SNPs) in wastewater which included clinically reported and newly identified mutations in the Philippines. This exploratory report provides a contextualized framework for applying WBE surveillance in low-sanitation areas.The authors thank the Davao City Health Office, the local government units under the City Government of Davao, the partner hospitals for their support and assistance, and Diana Aga for discussions on sample collection and analyses. The authors would also like to thank the members of the Bortz Virology Laboratory at the University of Alaska Anchorage- Ralf Dagdag and Matthew Redlinger, Amanda Warr from the Roslin Institute, Nicole Wheeler from the University of Birmingham, Lara Urban, co-founder of PuntSeq, and Joe Russell from MRI Global for their expert advice in nanopore and wastewater sequencing.
- Anti-inflammatory activity of monosubstituted xestoquinone analogues from the marine sponge Neopetrosia compactaSusana, Shalice R.; Salvador-Reyes, Lilibeth A. (MDPI, 2022-03-22)Chronic inflammation is recognized as a contributor to multiple chronic diseases, such as cancer, cardiovascular, and autoimmune disorders. Here, a natural products-initiated discovery of anti-inflammatory agents from marine sponges was undertaken. From the screening of 231 crude extracts, a total of 30 extracts showed anti-inflammatory activity with no direct cytotoxic effects at 50 μg/mL on RAW 264.7 (ATCC®TIB-71™) murine macrophage cells stimulated with 1 μg/mL lipopolysaccharide (LPS). Bioactivity-guided purification of the anti-inflammatory extract from the sponge Neopetrosia compacta led to the isolation of xestoquinone (1), adociaquinone B (2), adociaquinone A (3), 14-hydroxymethylxestoquinone (4), 15-hydroxymethylxestoquinone (5), and an inseparable 2:1 mixture of 14-methoxyxestoquinone and 15-methoxyxestoquinone (6). Compounds 1–6 caused a concentration-dependent reduction of nitric oxide (NO) production in LPS-stimulated RAW 264.7 cells, with 4–6 having low micromolar IC50 and acceptable selectivity index. Gene expression analysis using qRT-PCR showed that 1, 5, and 6 downregulated Il1b and Nos2 expression by 2.1- to 14.8-fold relative to the solvent control at 10 μM. Xestoquinone (1) and monosubstituted analogues (4–6), but not the disubstituted adociaquinones (2 and 3), caused Nrf2 activation in a luciferase reporter MCF7 stable cells. Compounds 5 and 6 caused a modest increase in Nqo1 gene expression at 10 μM. The anti-inflammatory activity of xestoquinone (1) and monosubstituted analogues (4–6) may, in part, be mediated by Nrf2 activation, leading to attenuation of inflammatory mediators such as IL-1β and NOS2.Samples were collected under gratuitous permit numbers GP-0084-15 and GP-0123-17, issued by the Department of Agriculture of the Philippines. We thank the municipalities of Bolinao, Pangasinan, and Puerto Galera, Oriental Mindoro for permission for sample collection. We acknowledge assistance from Z. L. Malto and DDHP chemical ecology group in obtaining the mass spectrometric data and sample collection, respectively.
- Global mass spectrometric analysis reveals chemical diversity of secondary metabolites and 44-Methylgambierone production in Philippine Gambierdiscus strainsMalto, Zabrina Bernice L.; Benico, Garry A.; Batucan, Jeremiah D.; Dela Cruz, James; Romero, Marc Lawrence J.; Azanza, Rhodora V.; Salvador-Reyes, Lilibeth A. (Frontiers Media SA, 2022-02-04)Surveillance and characterization of emerging marine toxins and toxigenic dinoflagellates are warranted to evaluate their associated health risks. Here, we report the occurrence of the ciguatera poisoning-causative dinoflagellate Gambierdiscus balechii in the Philippines. Toxin production and chemical diversity of secondary metabolites in G. balechii GtoxSAM092414, G. balechii Gtox112513, and the recently reported Gambierdiscus carpenteri Gam1BOL080513 were assessed using targeted and untargeted UPLC-MS/MS analysis and radioligand receptor-binding assay (RBA). 44-methylgambierone was produced by all three strains, albeitwith different levels based on RBA and UPLC-HRMS/MS analysis. The fatty acid composition was similar in all strains, while subtle differences in monosaccharide content were observed, related to the collection site rather than the species. Molecular networking using the GNPS database identified 45 clusters belonging to at least ten compound classes, with terpene glycosides, carbohydrate conjugates, polyketides, and macrolides as major convergence points. Species-specific peptides and polyhydroxylated compounds were identified in G. balechii GtoxSAM092414 and G. carpenteri Gam1BOL080513, respectively. These provide a glimpse of the uncharacterized biosynthetic potential of benthic dinoflagellates and highlight the intricate and prolific machinery for secondary metabolites production in these organisms.We would like to thank H. Junio and the Secondary Metabolites Profiling Laboratory of the Institute of Chemistry, University of the Philippines Diliman and K. B. Davis for assistance in the conduct of this study.
- Emerging pharmaceutical contaminants in key aquatic environments of the PhilippinesMariano, Shyrill Mae F.; Angeles, Luisa F.; Aga, Diana S.; Villanoy, Cesar L.; Jaraula, Caroline Marie B. (Frontiers Media SA, 2023-09-13)Pharmaceuticals in natural waters are considered emerging pollutants due to their low concentrations and the negative effects they pose to the environment. Common sources of such pollutants include untreated wastewater from hospitals, residential, industrial, and agricultural sources. Many wastewater treatment methods only remove a subset of all pharmaceuticals from the wastewater; remaining pharmaceuticals are discharged into natural waters, and ultimately drain into coastal areas. Regions without proper wastewater treatment are especially susceptible to such contamination. This study deals with the distribution, sources, and seasonal variability of pharmaceuticals in key aquatic systems in the Philippines. Two watershed continuums (Davao Gulf, Davao City; Macajalar Bay, Cagayan de Oro City); two tourist areas (Boracay Island, Aklan; Mabini, Batangas); and one pristine atoll (Tubbataha Reefs, Palawan)—all with varied prevailing human population pressures—were studied. Samples of hospital wastewater as well as groundwater, surface and bottom water samples from rivers and coastal seas collected during dry and wet seasons were analyzed using solid-phase extraction and liquid chromatography-tandem mass spectrometry. Thirty-four target pharmaceutical residues and antibiotics were extracted and quantified. Acetaminophen was detected at concentrations of up to 289.17 ppb in freshwater samples, and at concentrations of up to 253.39 ppb in seawater samples. Ubiquitous to all the sites was caffeine, reaching 1848.57 ppb. Sulfamethazine, a commonly used veterinary antibiotic, was detected at 764.91 ppb in a river site in Cagayan de Oro. Untreated hospital wastewater contained metformin, iopamidol, sulfamethoxazole, acetylsulfamethoxazole, ciprofloxacin, and azithromycin, but these pharmaceuticals were not detected in other river and coastal waters. Samples collected during the dry season exhibited higher concentrations than those from the wet season, which appears to be related to increase in transient populations from tourism activities as well as dilution. The presence of pharmaceutical residues and antibiotics in these areas and the potential impact on the environment indicate the need for stricter wastewater management measures, particularly in communities located near water bodies. As the results of this study show, such measures might be most beneficial and effective if imposed during dry season and in areas open to tourism.We thank the crew and researchers aboard M/Y Panata expedition PA0421 to Tubbataha in October 2019 and cruise to Boracay December 2019. Our gratitude to Mary Antoinette Limen, Mishel Valery Rañada, Gio Ferson Bautista, and Ernest Guiller Pineda for helping us in the field, and to Lahiruni Halwatura for assisting in the creation of a standard calibration curve for saltwater.
- Total polyphenol content of tropical marine and coastal flora: Potentials for food and nutraceutical applicationsNarvarte, Bienson Ceasar V.; Genovia, Tom Gerald T.; Hinaloc, Lourie Ann R.; Gonzaga, Shienna Mae C.; Tabonda-Nabor, April Mae; Palecpec, Flora Maye R.; Dayao, Helen M.; Roleda, Michael Y. (Springer, 2023-07-08)The marine environment is abundant in natural products that are beneficial to humans. Among these compounds are the polyphenols produced by marine flora as secondary metabolites and used as a defense against stressful environmental conditions. Accordingly, recent pharmacological and biomedical studies showed that polyphenols from marine and coastal floras have several important bioactivities including antioxidant property. In this study, we measured the total polyphenol content (TPC) of 75 species of marine-associated flora. The TPC of their methanolic extracts was measured spectrophotometrically using the Folin-Ciocalteu assay and was expressed both as mg phloroglucinol equivalent per g of dry weight (mg PGE g−1 DW) and as mg gallic acid equivalent per g dry weight (mg GAE g−1 DW). The TPC values are higher when expressed in terms of GAE compared to PGE. Also, the mean TPC of tracheopytes (229 ± 43.0 mg PGE g−1 DW) was higher compared to the mean TPC of macroalgae (69.4 ± 9.59 mg PGE g−1 DW). For macroalgae, ochrophytes (97.9 ± 22.7 mg PGE g−1 DW) had the highest mean TPC followed by chlorophytes (80.0 ± 20.5 mg PGE g−1 DW) and rhodophytes (49.5 ± 8.60 mg PGE g−1 DW). Moreover, our study also showed that TPC varied between young and mature tissues, among different color morphotypes and different parts of the plants. Although the concentrations of total polyphenols varied among species, ages, strains and parts of the plant, our study showed that marine and coastal floras are rich sources of polyphenols that could be further examined for their biological activities and other applications in food industry.
- Total synthesis and bioactivity evaluation of hydrophobic microcionamide‐inspired peptidesInocentes, Carl Rogel V.; Salvador‐Reyes, Lilibeth A.; Villaraza, Aaron Joseph L. (Wiley, 2023-01)In this report, we describe the facile synthesis of four microcionamide-inspired peptides where the atypical 2-phenylethylenamine (2-PEA) functional group in the marine natural product, microcionamide A, was replaced with a similarly-aromatic but more easily incorporated tryptophan (Trp) residue. Compounds 1–4 were synthesized using a standard Fmoc-based solid-phase synthesis strategy followed by iodine-mediated on-resin cyclization for disulfide-bridged compounds 1–3. Compound 1 showed antimicrobial activity against Staphylococcus aureus and Pseudomonas aeruginosa, with minimum inhibitory concentrations (MICs) of 9.1 μM and 15 μM, respectively. The inactivity of alanine analogs 2–4 against these pathogens suggests that the N-terminal Val, the cyclic scaffold, the contiguous Ile residues, and consequently, the hydrophobicity of compound 1 are essential for antibacterial activity. Compound 1 also favorably exhibited minimal cytotoxicity against normal mammalian cell lines. In summary, we have synthesized an analog of microcionamide A where replacement of the 2-PEA moiety with a Trp residue retained the antibacterial activity and with favorably low cytotoxicity.