Miniaturized Cultivation of Microbiota for Antimalarial Drug Discovery

Abstract

The ongoing search for effective antiplasmodial agents remains essential in the fight against malaria worldwide. Emerging parasitic drug resistance places an urgent need to explore chemotherapies with novel structures and mechanisms of action. Natural products have historically provided effective antimalarial drug scaffolds. In an effort to search nature's chemical potential for antiplasmodial agents, unconventionally sourced organisms coupled with innovative cultivation techniques were utilized. Approximately 60,000 niche microbes from various habitats (slow-growing terrestrial fungi, Antarctic microbes, and mangrove endophytes) were cultivated on a small-scale, extracted, and used in high-throughput screening to determine antimalarial activity. About 1% of crude extracts were considered active and 6% partially active (≥ 67% inhibition at 5 and 50 μg/mL, respectively). Active extracts (685) were cultivated on a large-scale, fractionated, and screened for both antimalarial activity and cytotoxicity. High interest fractions (397) with an IC50 < 1.11 μg/mL were identified and subjected to chromatographic separation for compound characterization and dereplication. Identifying active compounds with nanomolar antimalarial activity coupled with a selectivity index tenfold higher was accomplished with two of the 52 compounds isolated. This microscale, high-throughput screening project for antiplasmodial agents is discussed in the context of current natural product drug discovery efforts.

Characterizing the Personalized Microbiota Dynamics for Disease Classification by Individual-Specific Edge-Network Analysis

“…There is growing interest in bolstering resistance to infections or diseases by altering the microbiota (Jia et al, 2008; Holmes, 2016; Waterman et al, 2016; Delzenne and Bindels, 2018). Here, we have presented a computational framework, i.e., iENA, to identify the key OTU features to distinguish normal and disease states, by extracting higher-order statistics and dynamic information from 16S rRNA (ribosomal RNA) gene sequencing data in a one-sample manner.…”

Exploitation of Mangrove Endophytic Fungi for Infectious Disease Drug Discovery

“…Tissues from mangroves ( Rhizophora mangle, Avicennia germinans , and Laguncularia racemosa ), associated trees ( Conocarpus erectus , and Coccoloba uvifera ), sediments, and marine invertebrates were collected over the course of several years (2010–2014) at sites around Tampa Bay, the Florida Keys, the Gulf of Mexico and Tapachula, Mexico. As previously reported [ 31 ], small pieces of organic material were surfaced sterilized in bleach and/or isopropyl alcohol and pressed against or transferred onto various solid agar media types meant to target a large scope of bacteria and fungi. Media preparation was as follows: a nutrient medium (e.g., potato dextrose, Sabaurad dextrose, actino, malt, tryptic soy, or glycerol) was combined with agar, salt, and a combination of antibacterial or antifungal small molecules (e.g., nystatin, cycloheximide, or chloramphenicol).…”

“…Generally following methods previously reported [ 30 , 31 ], for each fungal isolate, two 1 cm 2 pieces of fungal material on agar were inoculated into 1.25 mL of each: untreated SDB (Sabouraud dextrose broth), 100 µM sodium butyrate in SDB, and 100 µM 5-azacytadine in SDB and agitated. Each of these aliquots was poured over 3 g autoclaved brown rice in a 20 mL scintillation vial.…”

“…While invertebrates remain a major source of new bioactive marine compounds, developments in microbial isolation, culture, and genome-sequencing techniques continue to expand the natural products frontier of marine microbial sources [ 13 , 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 , 26 ]. Mangrove forests are marine-margin habitats regarded for their microbial, chemical, and biological diversity [ 27 , 28 , 29 , 30 , 31 ]. While endophytic microorganisms are well studied in South-East Asian mangroves [ 27 , 28 , 30 ], the microbial inhabitants of the expansive mangrove forests in the Americas and the Caribbean remain largely unexplored.…”

Healthy collaborations are needed for mangrove land use and mosquito control

“…The mangrove sediment-derived microorganisms may be a valuable reservoir of natural product diversity. The products could be utilized in the exploration of new antibiotics or drugs [ 4 , 5 ] in addition to other benefits. Therefore, a decision to develop urban communities through encroachment of the mangrove ecosystem should be considered with care before it is implemented.…”