Outlining the Hidden Curriculum: Perspectives on Successfully Navigating Scientific Conferences.

Scientific conferences and meetings are valuable opportunities for researchers to network, communicate, and develop knowledge. For early career scientists, conferences can also be intimidating, confusing, and overwhelming, especially without having adequate preparation or experience. In this Perspective, we provide advice based on previous experiences navigating scientific meetings and conferences. These guidelines outline parts of the hidden curriculum around preparing for and attending meetings, navigating conference sessions, networking with other scientists, and participating in social activities while upholding a recommended code of conduct.

Antimicrobial Isoflavans and Other Metabolites of Dalea jamesii.

The phytochemical investigation of extracts of Dalea jamesii root and aerial portions led to the isolation of ten phenolic compounds. Six previously undescribed prenylated isoflavans, summarily named ormegans A - F (1 - 6: ), were characterized, along with two new arylbenzofurans (7, 8: ), a known flavone (9: ), and a known chroman (10: ). The structures of the new compounds were deduced by NMR spectroscopy, supported by HRESI mass spectrometry. The absolute configurations of 1 - 6: were determined by circular dichroism spectroscopy. Compounds 1 - 9: exhibited in vitro antimicrobial activities, causing 98% or greater growth inhibition at concentrations as low as 2.5 - 5.1 µM against methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus faecalis, and Cryptococcus neoformans. Interestingly, the most active compound was the dimeric arylbenzofuran 8: (> 90% growth inhibition at 2.5 µM) against both methicillin-resistant S. aureus and vancomycin-resistant E. faecalis, tenfold more active than its corresponding monomer (7: ).

Reinvestigation of the structure-activity relationships of isoniazid.

Isoniazid (INH) remains a cornerstone for treatment of drug susceptible tuberculosis (TB), yet the quantitative structure-activity relationships for INH are not well documented in the literature. In this paper, we have evaluated a systematic series of INH analogs against contemporary Mycobacterium tuberculosis strains from different lineages and a few non-tuberculous mycobacteria (NTM). Deletion of the pyridyl nitrogen atom, isomerization of the pyridine nitrogen to other positions, replacement of the pyridine ring with isosteric heterocycles, and modification of the hydrazide moiety of INH abolishes antitubercular activity. Similarly, substitution of the pyridine ring at the 3-position is not tolerated while substitution at the 2-position is permitted with 2-methyl-INH 9 displaying antimycobacterial activity comparable to INH. To assess the specific activity of this series of INH analogs against mycobacteria, we assayed them against a panel of gram-positive and gram-negative bacteria, as well as a few fungi. As expected INH and its analogs display a narrow spectrum of activity and are inactive against all non-mycobacterial strains evaluated, except for 4, which has modest inhibitory activity against Cryptococcus neoformans. Our findings provide an updated analysis of the structure-activity relationship of INH that we hope will serve as useful resource for the community.

A nontoxic fungal natural product modulates fin regeneration in zebrafish larvae upstream of FGF-WNT developmental signaling.

BACKGROUND: The regeneration of larvae zebrafish fin emerged as a new model of regeneration in the last decade. In contrast to genetic tools to study fin regeneration, chemical probes to modulate and interrogate regeneration processes are not well developed. RESULTS: We set up a zebrafish larvae fin regeneration assay system and tested activities of natural product compounds and extracts, prepared from various microbes. Colomitide C, a recently isolated product from a fungus obtained from Antarctica, inhibited larvae fin regeneration. Using fluorescent reporter transgenic lines, we show that colomitide C inhibited fibroblast growth factor (FGF) signaling and WNT/ß-catenin signaling, which were activated after larvae fin amputation. By using the endothelial cell reporter line and immunofluorescence, we showed that colomitide C did not affect migration of the blood vessel and nerve into the injured larvae fin. Colomitide C did not show any cytotoxic activities when tested against FGF receptor-amplified human cancer cell lines. CONCLUSION: Colomitide C, a natural product, modulated larvae fin regeneration likely acting upstream of FGF and WNT signaling. Colomitide C may serve as a template for developing new chemical probes to study regeneration and other biological processes.

Diverse subterranean fungi of an underground iron ore mine.

Mines and caves are unusual ecosystems containing unique fungi and are greatly understudied compared to other environments. The Soudan Mine in Tower, MN, an iron ore mine that closed in 1963 after operating for 80 years, was sampled to explore fungal diversity and to investigate taxa that tolerate heavy metals for potential bioprocessing technologies or as sources of bioactive molecules for drug discovery and possible biocontrol for white-nose syndrome (WNS) of bats. The mine is 714 m deep, has 18 levels and contains large quantities of wooden timbers, in contrast to many other oligotrophic subterranean environments. Fungi were cultured from samples and the ITS region was sequenced for identification and phylogenetic analysis. Results show Ascomycota are the dominant fungi followed by Basidiomycota and Mucoromycota. Out of 164 identified taxa, 108 belong to the Ascomycota and 26 and 31 to Basidiomycota and Mucoromycota, respectively. There are also 46 taxa that do not match (<97% BLAST GenBank identity) sequenced fungal species. Examples of the most commonly isolated Ascomycota include Scytalidium sp., Mariannaea comptospora, Hypocrea pachybasidioides, Oidiodendron griseum and Pochonia bulbillosa; Basidiomycota include Postia sp., Sistotrema brinkmannii, Calocera sp., Amylocorticiellum sp.; Mucoromycota include Mortierella parvispora, M. gamsii, M. hyaline, M. basiparvispora and Mortierella sp. Unusual growth forms were also found including large quantities of black rhizomorphs of Armillaria sinapina and white mycelial cords of Postia sp. mycelium, as well as Pseudogymnoascus species growing over large areas of mine walls and ceiling. The mine environment is a relatively extreme environment for fungi, with the presence of high levels of heavy metals, complete darkness and poor nutrient availability. Several genera are similar to those isolated in other extreme environments but phylogenetic analyses show differences in species between these environments. Results indicate this subterranean environment hosts a wide diversity of fungi, many of them not found in above ground environments.

Discovery of Antifungal and Biofilm Preventative Compounds from Mycelial Cultures of a Unique North American Hericium sp. Fungus.

Edible mushrooms are an important source of nutraceuticals and for the discovery of bioactive metabolites as pharmaceuticals. In this work, the OSMAC (One Strain, Many Active Compounds) approach was used to isolate two new compounds (1 and 2) along with seven known compounds (3-9) from a mycelial culture of a unique North American edible mushroom Hericium sp. The fruiting body was collected in Marine on St. Croix, Minnesota (USA), and mycelial cultures were grown on four different solid and liquid media. Extracts from the mycelial cultures were screened for antimicrobial activity and only the extract from the Cheerios substrate culture exhibited antifungal activity. Bioassay guided fractionation and HPLC analysis were used to isolate nine pure compounds and the structures of the known compounds were established by analysis of the NMR and mass spectrometry data and comparison to published reports. Compound 1 is a new erinacerin alkaloid and 2 is an aldehyde derivative of 4-hydroxy chroman. Four chlorinated orcinol derivatives (3-6), a pyran (7), erinaceolactone (8), and erinacine (9) were identified. Compound 4 showed antifungal activity against C. albicans and C. neoformans (MIC = 31.3-62.5 µg/mL, respectively). Compound 4 also inhibited biofilm formation of C. albicans and C. neoformans at 7.8 µg/mL. These results suggest that mycelial cultures of edible fungi may provide useful, bioactive compounds.

Antifungal Norditerpene Oidiolactones from the Fungus Oidiodendron truncatum, a Potential Biocontrol Agent for White-Nose Syndrome in Bats.

White-nose syndrome (WNS) is a devastating disease of hibernating bats caused by the fungus Pseudogymnoascus destructans. We obtained 383 fungal and bacterial isolates from the Soudan Iron Mine, an important bat hibernaculum in Minnesota, then screened this library for antifungal activity to develop biological control treatments for WNS. An extract from the fungus Oidiodendron truncatum was subjected to bioassay-guided fractionation, which led to the isolation of 14 norditerpene and three anthraquinone metabolites. Ten of these compounds were previously described in the literature, and here we present the structures of seven new norditerpene analogues. Additionally, this is the first report of 4-chlorophyscion from a natural source, previously identified as a semisynthetic product. The compounds PR 1388 and LL-Z1271α were the only inhibitors of P. destructans (MIC = 7.5 and 15 µg/mL, respectively). Compounds were tested for cytotoxicity against fibroblast cell cultures obtained from Myotis septentrionalis (northern long eared bat) and M. grisescens (gray bat) using a standard MTT viability assay. The most active antifungal compound, PR 1388, was nontoxic toward cells from both bat species (IC50 > 100 µM). We discuss the implications of these results in the context of the challenges and logistics of developing a substrate treatment or prophylactic for WNS.

Cadopherone and colomitide polyketides from Cadophora wood-rot fungi associated with historic expedition huts in Antarctica.

Recent investigations of filamentous fungi isolated from coastal areas and historic wooden structures in the Ross Sea and Peninsula regions of Antarctica have identified the genus Cadophora as one of the most abundant fungal groups, comprising more than 30% of culturable fungi at some locations. A methanol extract of Cadophora luteo-olivacea grown on rice media yielded the known polyketides spiciferone A, spiciferol A, dihydrospiciferone A and dihydrospiciferol A. Additionally, nine related hexaketides were identified, including spiciferone F, two isomers of the known fungal bicyclic ketal colomitide B, cadopherones A-D, similin C, and spicifernin B. HPLC and NMR analysis of extracts from other isolates collected in Antarctica suggests that the spiciferones and colomitides are produced by at least two different Cadophora species. Preliminary precursor feeding experiments provided evidence for the biosynthesis of the colomitides from the same polyketide pathway as the spiciferone phytotoxins, possibly via a type III polyketide synthase (PKS). None of the compounds were active in a panel of anti-bacterial, anti-fungal, and mammalian cytotoxicity assays.

Resource capture and competitive ability of non-pathogenic Pseudogymnoascus spp. and P. destructans, the cause of white-nose syndrome in bats.

White-nose syndrome (WNS) is a devastating fungal disease that has been causing the mass mortality of hibernating bats in North America since 2006 and is caused by the psychrophilic dermatophyte Pseudogymnoascus destructans. Infected bats shed conidia into hibernaculum sediments and surfaces, but it is unknown if P. destructans can form stable, reproductive populations outside its bat hosts. Previous studies have found non-pathogenic Pseudogymnoascus in bat hibernacula, and these fungi may provide insight into the natural history of P. destructans. We compared the relatedness, resource capture, and competitive ability of non-pathogenic Pseudogymnoascus isolates with P. destructans to determine if they have similar adaptations for survival in hibernacula sediment. All non-pathogenic Pseudogymnoascus isolates grew faster, utilized a broader range of substrates with higher efficiency, and were generally more resistant to antifungals compared to P. destructans. All isolates also showed the ability to displace P. destructans in co-culture assays, but only some produced extractible antifungal metabolites. These results suggest that P. destructans would perform poorly in the same environmental niche as non-pathogenic Pseudogymnoascus, and must have an alternative saprophytic survival strategy if it establishes active populations in hibernaculum sediment and non-host surfaces.

Complete Genome Sequence of Streptomyces albus SM254, a Potent Antagonist of Bat White-Nose Syndrome Pathogen Pseudogymnoascus destructans.

We sequenced and annotated the complete 7,170,504-bp genome of a novel secondary metabolite-producingStreptomycesstrain,Streptomyces albusSM254, isolated from copper-rich subsurface fluids at ~220-m depth within the Soudan Iron Mine (Soudan, MN, USA).

Growth inhibition and apoptosis in cancer cells induced by polyphenolic compounds of Acacia hydaspica: Involvement of multiple signal transduction pathways.

Acacia hydaspica R. Parker is known for its medicinal uses in multiple ailments. In this study, we performed bioassay-guided fractionation of cytotoxic compounds from A. hydaspica and investigated their effects on growth and signaling activity in prostate and breast cancer cell lines. Four active polyphenolic compounds were identified as 7-O-galloyl catechin (GC), catechin (C), methyl gallate (MG), and catechin-3-O-gallate (CG). The four compounds inhibited prostate cancer PC-3 cell growth in a dose-dependent manner, whereas CG and MG inhibited breast cancer MDA-MB-231 cell growth. All tested compounds inhibited cell survival and colony growth in both cell lines, and there was evidence of chromatin condensation, cell shrinkage and apoptotic bodies. Further, acridine orange, ethidium bromide, propidium iodide and DAPI staining demonstrated that cell death occurred partly via apoptosis in both PC-3 and MDA-MB-231 cells. In PC-3 cells treatment repressed the expression of anti-apoptotic molecules Bcl-2, Bcl-xL and survivin, coupled with down-regulation of signaling pathways AKT, NFκB, ERK1/2 and JAK/STAT. In MDA-MB-231 cells, treatment induced reduction of CK2α, Bcl-xL, survivin and xIAP protein expression along with suppression of NFκB, JAK/STAT and PI3K pathways. Our findings suggest that certain polyphenolic compounds derived from A. hydaspica may be promising chemopreventive/therapeutic candidates against cancer.

Soudanones A-G: Antifungal Isochromanones from the Ascomycetous Fungus Cadophora sp. Isolated from an Iron Mine.

One new isochromane (pseudoanguillosporin C, 2), seven isochromanones (soudanones A-G, 3-9), and six known analogues including 10 and 11 were isolated from a culture of the fungus Cadophora sp. 10-5-2 M, collected from the subterranean 10th level of the Soudan Underground Iron Mine in Minnesota. All of the compounds were tested against a panel of microbial pathogens, and 2, 3, 10, and 11 were found to have activity against Cryptococcus neoformans (MIC = 35, 40, 20, and 30 µg/mL, respectively). Compound 11 was also active against Candida albicans, with an MIC of 40 µg/mL.

Synthesis and evaluation of N-alkyl-9-aminoacridines with antibacterial activity.

A series of 9-alkylaminoacridines were synthesized and evaluated for activity against two strains of methicillin-resistant and one strain of methicillin-sensitive Staphylococcus aureus. Results are presented that show a clear structure activity relationship between the N-alkyl chain length and antibacterial activity with peak MIC99 values of 2-3 µM for alkyl chains ranging from 10 to 14 carbons in length. Although prior work has linked the function of acridine-based compounds to intercalation and topoisomerase inhibition, the present results show that 9-alkylaminoacridines likely function as amphiphilic membrane-active disruptors potentially in a similar manner as quaternary ammonium antimicrobials.

Ultra-High-Throughput Screening of Natural Product Extracts to Identify Proapoptotic Inhibitors of Bcl-2 Family Proteins.

Antiapoptotic Bcl-2 family proteins are validated cancer targets composed of six related proteins. From a drug discovery perspective, these are challenging targets that exert their cellular functions through protein-protein interactions (PPIs). Although several isoform-selective inhibitors have been developed using structure-based design or high-throughput screening (HTS) of synthetic chemical libraries, no large-scale screen of natural product collections has been reported. A competitive displacement fluorescence polarization (FP) screen of nearly 150,000 natural product extracts was conducted against all six antiapoptotic Bcl-2 family proteins using fluorochrome-conjugated peptide ligands that mimic functionally relevant PPIs. The screens were conducted in 1536-well format and displayed satisfactory overall HTS statistics, with Z'-factor values ranging from 0.72 to 0.83 and a hit confirmation rate between 16% and 64%. Confirmed active extracts were orthogonally tested in a luminescent assay for caspase-3/7 activation in tumor cells. Active extracts were resupplied, and effort toward the isolation of pure active components was initiated through iterative bioassay-guided fractionation. Several previously described altertoxins were isolated from a microbial source, and the pure compounds demonstrate activity in both Bcl-2 FP and caspase cellular assays. The studies demonstrate the feasibility of ultra-high-throughput screening using natural product sources and highlight some of the challenges associated with this approach.

Antiproliferative activities of halogenated thieno[3,2-d]pyrimidines.

The in vitro evaluation of thieno[3,2-d]pyrimidines identified halogenated compounds 1 and 2 with antiproliferative activity against three different cancer cell lines. A structure activity relationship study indicated the necessity of the chlorine at the C4-position for biological activity. The two most active compounds 1 and 2 were found to induce apoptosis in the leukemia L1210 cell line. Additionally, the compounds were screened against a variety of other microbial targets and as a result, selective activity against several fungi was also observed. The synthesis and preliminary biological results are reported herein.

Subinhibitory antibiotic concentrations mediate nutrient use and competition among soil streptomyces.

Though traditionally perceived as weapons, antibiotics are also hypothesized to act as microbial signals in natural habitats. However, while subinhibitory concentrations of antibiotics (SICA) are known to shift bacterial gene expression, specific hypotheses as to how SICA influence the ecology of natural populations are scarce. We explored whether antibiotic 'signals', or SICA, have the potential to alter nutrient utilization, niche overlap, and competitive species interactions among Streptomyces populations in soil. For nine diverse Streptomyces isolates, we evaluated nutrient utilization patterns on 95 different nutrient sources in the presence and absence of subinhibitory concentrations of five antibiotics. There were significant changes in nutrient use among Streptomyces isolates, including both increases and decreases in the capacity to use individual nutrients in the presence vs. in the absence of SICA. Isolates varied in their responses to SICA and antibiotics varied in their effects on isolates. Furthermore, for some isolate-isolate-antibiotic combinations, competition-free growth (growth for an isolate on all nutrients that were not utilized by a competing isolate), was increased in the presence of SICA, reducing the potential fitness cost of nutrient competition among those competitors. This suggests that antibiotics may provide a mechanism for bacteria to actively minimize niche overlap among competitors in soil. Thus, in contrast to antagonistic coevolutionary dynamics, antibiotics as signals may mediate coevolutionary displacement among coexisting Streptomyces, thereby hindering the emergence of antibiotic resistant phenotypes. These results contribute to our broad understanding of the ecology and evolutionary biology of antibiotics and microbial signals in nature.

Solphenazines A-F, glycosylated phenazines from Streptomyces sp. strain DL-93.

During a survey of actinobacteria known to suppress the growth of Streptomyces scabies (the causative agent of potato scab disease) in vivo, six new rhamnosylated alkaloids, the solphenazines A-F (1-6), were isolated from a biological control strain of Streptomyces (DL-93). The known rhamnosyl analogue of paraben (9) was also isolated along with a new rhamnosylated derivative of N-methyl-p-aminobenzoic acid (10). None of the compounds exhibited any antibacterial or antifungal activity against a standard panel of microorganisms, but compounds 1, 2, and 6 displayed some cytotoxicity against HCT-116 cancer cells. Additional in vitro testing provided data suggesting that the cytotoxic activity is not due to DNA intercalation or topoisomerase inhibition.

Total synthesis and biological evaluation of transvalencin Z.

The emerging global epidemic of drug-resistant tuberculosis has created an urgent need to identify novel therapeutic approaches for disease treatment. Transvalencin Z (1) is a natural product from Nocardia transvalensis with relatively potent and selective antimycobacterial activity against Mycobacterium smegmatis, making it an attractive target for structure-activity and mechanism of action studies. The total synthesis of the four possible diastereomers of transvalencin Z was completed (1a-d), and the absolute configurations were defined using chemical synthesis, HPLC retention times, and optical rotation measurements. Surprisingly, none of the transvalencin Z diastereomers exhibited any inhibitory activity against a panel of microbial pathogens, including several species of mycobacteria.

Natural products as leads for tuberculosis drug development.

New therapeutics are urgently needed to combat the immense disease burden of tuberculosis and related mycobacterial diseases worldwide. Natural products continue to provide leads for the development of novel drugs to treat the rapidly growing numbers of patients with multi- and extensively-drug resistant tuberculosis. This review presents natural products and synthesized analogues with anti-mycobacterial activity published between 2006 through 2009. Structure activity relationships, synthetic analogues and newly reported activities of known compounds reported during this period are also included.

Pharmacophore and structure-activity relationships of integrase inhibition within a dual inhibitor scaffold of HIV reverse transcriptase and integrase.

Rational design of dually active inhibitors against human immunodeficiency virus (HIV) reverse transcriptase (RT) and integrase (IN) has proved viable with 1-[(2-hydroxyethoxy)methyl]-6-(phenylthio)thymine (HEPT) type of non-nucleoside RT inhibitors (NNRTIs). To establish the pharmacophore and study the structure-activity relationships (SAR) of integrase inhibition within a previously disclosed RT/IN dual inhibitor scaffold, new analogues featuring substitution at different sites of the HEPT ring were designed and synthesized. These studies have revealed an IN inhibition pharmacophore that is merged with the known RT pharmacophore through a shared C-6 benzyl group. Further SAR also demonstrated that optimal IN inhibition within our dual inhibitor scaffold requires a regiospecific (N-1) diketoacid (DKA)-carrying pendant with a certain length.