The resuscitation-promoting factor (Rpf) from Micrococcus luteus and its putative reaction product 1,6-anhydro-MurNAc increase culturability of environmental bacteria.

Dormant bacterial cells do not divide and are not immediately culturable, but they persist in a state of low metabolic activity, a physiological state having clinical relevance, for instance in latent tuberculosis. Resuscitation-promoting factors (Rpfs) are proteins that act as signalling molecules mediating growth and replication. In this study we aimed to test the effect of Rpfs from Micrococcus luteus on the number and diversity of cultured bacteria using insect and soil samples, and to examine if the increase in culturability could be reproduced with the putative reaction product of Rpf, 1,6-anhydro-N-acetylmuramic acid (1,6-anhydro-MurNAc). The rpf gene from Micrococcus luteus was amplified and cloned into a pET21b expression vector and the protein was expressed in Escherichia coli BL21(DE3) cells and purified by affinity chromatography using a hexa-histidine tag. 1,6-Anhydro-MurNAc was prepared using reported chemical synthesis methods. Recombinant Rpf protein or 1,6-anhydro-MurNAc were added to R2A cultivation media, and their effect on the culturability of bacteria from eight environmental samples including four cockroach guts and four soils was examined. Colony-forming units, 16S rRNA gene copies and Illumina amplicon sequencing of the 16S rRNA gene were measured for all eight samples subjected to three different treatments: Rpf, 1,6-anhydro-MurNAc or blank control. Both Rpf and 1,6-anhydro-MurNAc increased the number of colony-forming units and of 16S rRNA gene copies across the samples although the protein was more effective. The Rpf and 1,6-anhydro-MurNAc promoted the cultivation of a diverse set of bacteria and in particular certain clades of the phyla Actinomycetota and Bacillota . This study opens the path for improved cultivation strategies aiming to isolate and study yet undescribed living bacterial organisms.

Insect Collections as an Untapped Source of Bioactive Compounds-Fireflies (Coleoptera: Lampyridae) and Cardiotonic Steroids as a Proof of Concept.

Natural history collections provide an invaluable basis for systematics, ecology, and conservation. Besides being an important source of DNA, museum specimens may also contain a plethora of natural products. Especially, dried insect collections represent a global repository with billions of inventoried vouchers. Due to their vast diversity, insects possess a great variety of defensive compounds, which they either produce autogenously or derive from the environment. Here, we present a case study on fireflies (Coleoptera: Lampyridae), which produce bufadienolides as a defense against predators. These toxins belong to the cardiotonic steroids, which are used for the treatment of cardiac diseases and specifically inhibit the animal enzyme Na+/K+-ATPase. Bufadienolides have been reported from only seven out of approximately 2000 described firefly species. Using a non-destructive approach, we screened 72 dry coleopteran specimens for bufadienolides using HPLC-DAD and HPLC-MS. We found bufadienolides including five novel compounds in 21 species of the subfamily Lampyrinae. The absence of bufadienolides in the phylogenetically related net-winged beetles (Lycidae) and the lampyrid subfamilies Luciolinae and Lamprohizinae indicates a phylogenetic pattern of bufadienolide synthesis. Our results emphasize the value of natural history collections as an archive of chemical information for ecological and evolutionary basic research and as an untapped source for novel bioactive compounds.

Sequences of Tolypins, Insecticidal Efrapeptin-Type Peptaibiotics from Species of the Fungal Genus Tolypocladium.

A peptide mixture named tolypin, originally isolated from species of the fungal genus Tolypocladium, was structurally characterised and sequences compared to those reported for efrapeptins isolated from strains of Tolypocladium inflatum. Chiral amino acid analysis, direct infusion, and online HPLC electrospray ionization tandem mass spectrometry provided composition, molecular weights of peptides, and series of diagnostic fragment ions. Sequences deduced from ESI-MS revealed that tolypins C-G are identical to efrapeptins C-G. The results were corroborated by ESI-MS and HPLC of an authentic efrapeptin sample from Eli Lilly Research Laboratories (USA). Comparison of the HPLC elution profiles of efrapeptin and tolypin indicated a pronounced microheterogeneity of the former. A high-resolution HPLC of authentic efrapeptin has not been published before. Close relationship and partial identity of sequences of tolypins and efrapeptins, which had previously been postulated, were definitely proven. The geographical origin of the two most important T. inflatum strains used for sequencing of efrapeptins/tolypins could unambiguously be clarified. A new minor compound, designated tolypin H1, was sequenced. High proportions of helicogenic Aib (α-aminoisobutyric acid) and l-isovaline, N-terminal acetyl-l-pipecolic acid and the unusual, amide-bound C-terminal residue, named (S)-2-amino-1-(1,5-diazabicyclo[4.3.0]non-5-ene-5-ylium)-4-methylpentane corresponding to 1-[(2S)-2-amino-4-methylpentyl]-2,3,4,6,7,8-hexahydropyrrolo[1,2-a]pyrimidin-1-ium, define these peptides as linear, cationic peptaibiotics.

Antibiotic-Producing Beneficial Bacteria in the Gut of the Burying Beetle Nicrophorus vespilloides.

The increasing prevalence of antibiotic-resistant human pathogens is a growing public concern and there is intense pressure to identify new antibacterial compounds that can be developed into antibiotics with novel mode of action. Evolutionary theory predicts that insects that have evolved to occupy sophisticated ecological niches by feeding and reproducing on carcasses will depend on their gut microbiome to prevent colonization by invading pathogens taken up with the diet. This inspired our hypothesis that the complex interactions between the core microbiome and the more flexible microbial communities dependent on the environment may promote the outsourcing of antibiotic synthesis to beneficial microbes. We tested this hypothesis by cultivating and characterizing bacteria isolated from the gut of the burying beetle Nicrophorus vespilloides, which feeds and reproduces on small vertebrate carcasses buried in the soil to avoid competitors such as fly maggots. The extracts of isolated bacteria were screened for activity against human pathogens such as Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Candida albicans. More than 400 strains were isolated, among which the crude extract of Serratia marcescens 2MH3-2 displayed promising activity against Staphylococcus aureus. Bioactivity-guided fractionation enabled purification of the primary antimicrobial compound of the extract. By LC-MS and NMR experiments, it was identified as serrawettin W2 (C38H61N5O9), the antibacterial and nematostatic activity of which was corroborated in our study. We postulate that this antibiotic could contribute to the control of both bacteria and phoretic nematodes in the gut, which compete for food when transferred to the carcass. Our study shows that the gut microbiome of N. vespilloides is a promising resource for the screening of antibiotic-producing bacteria.

Sequences of stilboflavin C: towards the peptaibiome of the filamentous fungus Stilbella (= Trichoderma) flavipes.

Filamentous fungi of the genus Stilbella are recognized as an abundant source of naturally occurring α-aminoisobutyric acid-containing peptides. The culture broth of Stilbella (Trichoderma) flavipes CBS 146.81 yielded a mixture of peptides named stilboflavins (SF), and these were isolated and separated by preparative TLC into groups named SF-A, SF-B, and SF-C. Although all three of these groups resolved as single spots on thin-layer chromatograms, HPLC analysis revealed that each of the groups represents very microheterogeneous mixtures of closely related peptides. Here, we report on the sequence analysis of SF-C peptides, formerly isolated by preparative TLC. HPLC coupled to QqTOF-ESI-HRMS provided the sequences of 10 16-residue peptides and five 19-residue peptides, all of which were N-terminally acetylated. In contrast to the previously described SF-A and SF-B peptaibols, SF-C peptaibols contain Ser-Alaol or Ser-Leuol, which are rarely found as C-termini, and repetitive Leu-Aib-Gly sequences, which have not been detected in peptaibols before. Taking the previously determined sequences of SF-A and SF-B into account, the entirety of peptides produced by S. flavipes (the 'peptaibiome') approaches or exceeds 100 non-ribosomally biosynthesized peptaibiotics. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

Metabolites from nematophagous fungi and nematicidal natural products from fungi as alternatives for biological control. Part II: metabolites from nematophagous basidiomycetes and non-nematophagous fungi.

In this second section of a two-part mini-review article, we introduce 101 further nematicidal and non-nematicidal secondary metabolites biosynthesized by nematophagous basidiomycetes or non-nematophagous ascomycetes and basidiomycetes. Several of these compounds have promising nematicidal activity and deserve further and more detailed analysis. Thermolides A and B, omphalotins, ophiobolins, bursaphelocides A and B, illinitone A, pseudohalonectrins A and B, dichomitin B, and caryopsomycins A-C are excellent candidates or lead compounds for the development of biocontrol strategies for phytopathogenic nematodes. Paraherquamides, clonostachydiol, and nafuredins offer promising leads for the development of formulations against the intestinal nematodes of ruminants.

Metabolites from nematophagous fungi and nematicidal natural products from fungi as an alternative for biological control. Part I: metabolites from nematophagous ascomycetes.

Plant-parasitic nematodes are estimated to cause global annual losses of more than US$ 100 billion. The number of registered nematicides has declined substantially over the last 25 years due to concerns about their non-specific mechanisms of action and hence their potential toxicity and likelihood to cause environmental damage. Environmentally beneficial and inexpensive alternatives to chemicals, which do not affect vertebrates, crops, and other non-target organisms, are therefore urgently required. Nematophagous fungi are natural antagonists of nematode parasites, and these offer an ecophysiological source of novel biocontrol strategies. In this first section of a two-part review article, we discuss 83 nematicidal and non-nematicidal primary and secondary metabolites found in nematophagous ascomycetes. Some of these substances exhibit nematicidal activities, namely oligosporon, 4',5'-dihydrooligosporon, talathermophilins A and B, phomalactone, aurovertins D and F, paeciloxazine, a pyridine carboxylic acid derivative, and leucinostatins. Blumenol A acts as a nematode attractant. Other substances, such as arthrosporols and paganins, play a decisive role in the life cycle of the producers, regulating the formation of reproductive or trapping organs. We conclude by considering the potential applications of these beneficial organisms in plant protection strategies.

The peptaibiotics database--a comprehensive online resource.

In this work, we present the 'Peptaibiotics Database' (PDB), a comprehensive online resource, which intends to cover all Aib-containing non-ribosomal fungal peptides currently described in scientific literature. This database shall extend and update the recently published 'Comprehensive Peptaibiotics Database' and currently consists of 1,297 peptaibiotic sequences. In a literature survey, a total of 235 peptaibiotic sequences published between January 2013 and June 2014 have been compiled, and added to the list of 1,062 peptides in the recently published 'Comprehensive Peptaibiotics Database'. The presented database is intended as a public resource freely accessible to the scientific community at peptaibiotics-database.boku.ac.at. The search options of the previously published repository and the presentation of sequence motif searches have been extended significantly. All of the available search options can be combined to create complex database queries. As a public repository, the presented database enables the easy upload of new peptaibiotic sequences or the correction of existing informations. In addition, an administrative interface for maintenance of the content of the database has been implemented, and the design of the database can be easily extended to store additional information to accommodate future needs of the 'peptaibiomics community'.

Peptaibol, secondary-metabolite, and hydrophobin pattern of commercial biocontrol agents formulated with species of the Trichoderma harzianum complex.

The production of bioactive polypeptides (peptaibiotics) in vivo is a sophisticated adaptation strategy of both mycoparasitic and saprotrophic Trichoderma species for colonizing and defending their natural habitats. This feature is of major practical importance, as the detection of peptaibiotics in plant-protective Trichoderma species, which are successfully used against economically relevant bacterial and fungal plant pathogens, certainly contributes to a better understanding of these complex antagonistic interactions. We analyzed five commercial biocontrol agents (BCAs), namely Canna(®) , Trichosan(®) , Vitalin(®) , Promot(®) WP, and TrichoMax(®) , formulated with recently described species of the Trichoderma harzianum complex, viz. T. afroharzianum, T. simmonsii, and T. guizhouense. By using the well-established, HPLC/MS-based peptaibiomics approach, it could unequivocally be demonstrated that all of these formulations contained new and recurrent peptaibols, i.e., peptaibiotics carrying an acetylated N-terminus, the C-terminus of which is reduced to a 1,2-amino alcohol. Their chain lengths, including the amino alcohol, were 11, 14, and 18 residues, respectively. Peptaibols were also to be the dominating secondary metabolites in plate cultures of the four strains obtained from four of the Trichoderma- based BCAs, contributing 95% of the UHPLC-UV/VIS peak areas and 99% of the total ion count MS peak area from solid media. Furthermore, species-specific hydrophobins, as well as non-peptaibiotic secondary metabolites, were detected, the latter being known for their antifungal, siderophore, or plant-growth-promoting activities. Notably, none of the isolates produced low-molecular weight mycotoxins.

Systematics of the Trichoderma harzianum species complex and the re-identification of commercial biocontrol strains.

Trichoderma harzianum is known as a cosmopolitan, ubiquitous species associated with a wide variety of substrates. It is possibly the most commonly used name in agricultural applications involving Trichoderma, including biological control of plant diseases. While various studies have suggested that T. harzianum is a species complex, only a few cryptic species are named. In the present study the taxonomy of the T. harzianum species complex is revised to include at least 14 species. Previously named species included in the complex are T. guizhouense, T. harzianum, and T. inhamatum. Two new combinations are proposed, T. lentiforme and T. lixii. Nine species are described as new, T. afarasin, T. afroharzianum, T. atrobrunneum, T. camerunense, T. endophyticum, T. neotropicale, T. pyramidale, T. rifaii and T. simmonsii. We isolated Trichoderma cultures from four commercial biocontrol products reported to contain T. harzianum. None of the biocontrol strains were identified as T. harzianum s. str. In addition, the widely applied culture 'T. harzianum T22' was determined to be T. afroharzianum. Some species in the T. harzianum complex appear to be exclusively endophytic, while others were only isolated from soil. Sexual states are rare. Descriptions and illustrations are provided. A secondary barcode, nuc translation elongation factor 1-α (TEF1) is needed to identify species in this complex.

Front line defenders of the ecological niche! Screening the structural diversity of peptaibiotics from saprotrophic and fungicolous Trichoderma/Hypocrea species.

Approximately 950 individual sequences of non-ribosomally biosynthesised peptides are produced by the genus Trichoderma/Hypocrea that belong to a perpetually growing class of mostly linear antibiotic oligopeptides, which are rich in the non-proteinogenic α-aminoisobutyric acid (Aib). Thus, they are comprehensively named peptaibiotics. Notably, peptaibiotics represent ca. 80 % of the total inventory of secondary metabolites currently known from Trichoderma/Hypocrea. Their unique membrane-modifying bioactivity results from amphipathicity and helicity, thus making them ideal candidates in assisting both colonisation and defence of the natural habitats by their fungal producers. Despite this, reports on the in vivo-detection of peptaibiotics have scarcely been published in the past. In order to evaluate the significance of peptaibiotic production for a broader range of potential producers, we screened nine specimens belonging to seven hitherto uninvestigated fungicolous or saprotrophic Trichoderma/Hypocrea species by liquid chromatography coupled to electrospray high resolution mass spectrometry. Sequences of peptaibiotics found were independently confirmed by analysing the peptaibiome of pure agar cultures obtained by single-ascospore isolation from the specimens. Of the nine species examined, five were screened positive for peptaibiotics. A total of 78 peptaibiotics were sequenced, 56 (=72 %) of which are new. Notably, dihydroxyphenylalaninol and O-prenylated tyrosinol, two C-terminal residues, which have not been reported for peptaibiotics before, were found as well as new and recurrent sequences carrying the recently described tyrosinol residue at their C-terminus. The majority of peptaibiotics sequenced are 18- or 19-residue peptaibols. Structural homologies with 'classical representatives' of subfamily 1 (SF1)-peptaibiotics argue for the formation of transmembrane ion channels, which are prone to facilitate the producer capture and defence of its substratum.

Homoserine lactones influence the reaction of plants to rhizobia.

Bacterial quorum sensing molecules not only grant the communication within bacterial communities, but also influence eukaryotic hosts. N-acyl-homoserine lactones (AHLs) produced by pathogenic or beneficial bacteria were shown to induce diverse reactions in animals and plants. In plants, the reaction to AHLs depends on the length of the lipid side chain. Here we investigated the impact of two bacteria on Arabidopsis thaliana, which usually enter a close symbiosis with plants from the Fabaceae (legumes) family and produce a long-chain AHL (Sinorhizobium meliloti) or a short-chain AHL (Rhizobium etli). We demonstrate that, similarly to the reaction to pure AHL molecules, the impact, which the inoculation with rhizosphere bacteria has on plants, depends on the type of the produced AHL. The inoculation with oxo-C14-HSL-producing S. meliloti strains enhanced plant resistance towards pathogenic bacteria, whereas the inoculation with an AttM lactonase-expressing S. meliloti strain did not. Inoculation with the oxo-C8-HSL-producing R. etli had no impact on the resistance, which is in agreement with our previous hypothesis. In addition, plants seem to influence the availability of AHLs in the rhizosphere. Taken together, this report provides new insights in the role of N-acyl-homoserine lactones in the inter-kingdom communication at the root surface.

The comprehensive peptaibiotics database.

Peptaibiotics are nonribosomally biosynthesized peptides, which - according to definition - contain the marker amino acid α-aminoisobutyric acid (Aib) and possess antibiotic properties. Being known since 1958, a constantly increasing number of peptaibiotics have been described and investigated with a particular emphasis on hypocrealean fungi. Starting from the existing online 'Peptaibol Database', first published in 1997, an exhaustive literature survey of all known peptaibiotics was carried out and resulted in a list of 1043 peptaibiotics. The gathered information was compiled and used to create the new 'The Comprehensive Peptaibiotics Database', which is presented here. The database was devised as a software tool based on Microsoft (MS) Access. It is freely available from the internet at http://peptaibiotics-database.boku.ac.at and can easily be installed and operated on any computer offering a Windows XP/7 environment. It provides useful information on characteristic properties of the peptaibiotics included such as peptide category, group name of the microheterogeneous mixture to which the peptide belongs, amino acid sequence, sequence length, producing fungus, peptide subfamily, molecular formula, and monoisotopic mass. All these characteristics can be used and combined for automated search within the database, which makes The Comprehensive Peptaibiotics Database a versatile tool for the retrieval of valuable information about peptaibiotics. Sequence data have been considered as to December 14, 2012.

Screening the biosphere: the fungicolous fungus Trichoderma phellinicola, a prolific source of hypophellins, new 17-, 18-, 19-, and 20-residue peptaibiotics.

To investigate the significance of antibiotics for the producing organism(s) in the natural habitat, we screened a specimen of the fungicolous fungus Trichoderma phellinicola (syn. Hypocrea phellinicola) growing on its natural host Phellinus ferruginosus. Results revealed that a particular group of non-ribosomal antibiotic polypeptides, peptaibiotics, which contain the non-proteinogenic marker amino acid, α-aminoisobutyric acid, was biosynthesized in the natural habitat by the fungicolous producer and, consequently, released into the host. By means of liquid chromatography coupled to electrospray high-resolution time-of-flight mass spectrometry, we detected ten 20-residue peptaibols in the specimen. Sequences of peptaibiotics found in vivo were independently confirmed by analyzing the peptaibiome of an agar plate culture of T. phellinicola CBS 119283 (ex-type) grown under laboratory conditions. Notably, this strain could be identified as a potent producer of 39 new 17-, 18-, and 19-residue peptaibiotics, which display the same building scheme as the 20-residue peptaibols found in the specimen. Two of the 19-residue peptaibols are tentatively assigned to carry tyrosinol, a novel C-terminal residue, as deduced from high-resolution tandem mass-spectrometry data. For the new peptaibiotics produced by T. phellinicola, the name 'hypophellin(s)', based on the teleomorph name, is introduced.

The sequences of the eleven-residue peptaibiotics: suzukacillins-B.

The filamentous fungus designated 'Trichoderma viride' strain 63 C-1 simultaneously produces suzukacillins (SZs), two microheterogeneous groups of peptaibols, under submerged culture conditions. Both groups are readily distinguishable by TLC: the major group is designated SZ-A, whereas the minor group with a higher Rf value is named SZ-B. The peptide mixture was obtained from a MeOH extract of the mycelium. SZ-B was separated from SZ-A by Sephadex LH-20 column chromatography. Although it provided one single spot on silica-gel TLC plates, 15 individual peptides could be separated by C8 reversed-phase (RP) HPLC, and their sequences were determined by HPLC/QqTOF-ESI-HRMS. Fourteen peptides exhibit the C-terminal sequence Pro(6) -Lxx-Lxx-Aib-Pro-Vxxol/Lxxol(11) , which is common for eleven-residue peptaibols. The remaining peptide is tentatively assigned as a ten-residue sequence, in which the C-terminal 1,2-amino alcohol is deleted, thus terminating in free proline. Nine of the peptides carry an Ac-Aib residue at the N-terminus, very frequently found in eleven-residue peptaibols. Four peptides comprise the rare Ac-Ala N-terminus, and for two peptides, N-terminal Ac-D-Iva residues were identified. One peptide contains a C-terminal residue of yet undetermined structure. Comparison with previously reported eleven-residue peptaibol sequences reveals that eight of the peptides represent new sequence analogs.

Hypopulvins, novel peptaibiotics from the polyporicolous fungus Hypocrea pulvinata, are produced during infection of its natural hosts.

In order to investigate the significance of antibiotics for the producing organism(s) in the natural habitat, we screened specimens of the polyporicolous fungus Hypocrea pulvinata growing on its natural hosts Piptoporus betulinus and Fomitopsis pinicola. Results showed that a particular group of nonribosomally biosynthesised antibiotic polypeptides, the peptaibiotics, which contain the nonproteinogenic marker amino acid α-aminoisobutyric acid (Aib), was produced in the natural habitat by the fungicolous producer and, consequently, released into the host. Using liquid chromatography coupled to electrospray high-resolution mass spectrometry we detected especially 19-, but also 11-, 18-, and 20-residue peptaibiotics in the five infected specimens analysed. Structures of peptaibiotics found were confirmed by analysing the peptaibiome of pure agar cultures obtained by single-ascospore isolation from the specimens. The 19-residue peptaibols were determined as deletion sequences of the trichosporins B lacking the Aib residue in position 6. Notably, 26 of the 28 peptaibiotics sequenced were novel; therefore the name 'hypopulvins' was introduced. Considering not only the ubiquity of both the two host species but also the highly specific association between H. pulvinata and P. betulinus/F. pinicola, and the abundance of this fungicolous species in north temperate regions of the world, a decisive role for the peptaibiotics detected in this study is predicted, which may act as mediators of the complex interactions between the basidiomycetous host and its fungicolous ascomycete 'partner'. Structural analogies of the hypopulvins, particularly with other 18-, 19-, and 20-residue peptaibiotics, suggest that the hypopulvins are forming transmembrane ion channels and could thus support the hypothesis of a parasitic lifestyle of the fungicolous producer.

The production of multiple small peptaibol families by single 14-module Peptide synthetases in Trichoderma/Hypocrea.

The most common sequences of peptaibiotics are 11-residue peptaibols found widely distributed in the genus Trichoderma/Hypocrea. Frequently associated are 14-residue peptaibols sharing partial sequence identity. Genome sequencing projects of three Trichoderma strains of the major clades reveal the presence of up to three types of nonribosomal peptide synthetases with 7, 14, or 18-20 amino acid-adding modules. Here, we provide evidence that the 14-module NRPS type found in T. virens, T. reesei (teleomorph Hypocrea jecorina), and T. atroviride produces both 11- and 14-residue peptaibols based on the disruption of the respective NRPS gene of T. reesei, and bioinformatic analysis of their amino acid-activating domains and modules. The sequences of these peptides may be predicted from the gene sequences and have been confirmed by analysis of families of 11- and 14-residue peptaibols from the strain 618, termed hypojecorins A (23 sequences determined, 4 new) and B (3 sequences determined, 2 new), and the recently established trichovirins A from T. virens. The distribution of 11- and 14-residue products is strain-specific and depends on growth conditions as well. Possible mechanisms of module skipping are discussed.

Harmonine, a defence compound from the harlequin ladybird, inhibits mycobacterial growth and demonstrates multi-stage antimalarial activity.

The harlequin ladybird beetle Harmonia axyridis has been introduced in many countries as a biological control agent, but has become an invasive species threatening the biodiversity of native ladybirds. Its invasive success has been attributed to its vigorous resistance against diverse pathogens. This study demonstrates that harmonine ((17R,9Z)-1,17-diaminooctadec-9-ene), which is present in H. axyridis haemolymph, displays broad-spectrum antimicrobial activity that includes human pathogens. Antibacterial activity is most pronounced against fast-growing mycobacteria and Mycobacterium tuberculosis, and the growth of both chloroquine-sensitive and -resistant Plasmodium falciparum strains is inhibited. Harmonine displays gametocytocidal activity, and inhibits the exflagellation of microgametocytes and zygote formation. In an Anopheles stephensi mosquito feeding model, harmonine displays transmission-blocking activity.

Secondary metabolites released by the burying beetle Nicrophorus vespilloides: chemical analyses and possible ecological functions.

Burying beetles of the genus Nicrophorus localize small vertebrate carcasses by emitted volatiles. The carcass that serves as reproduction and breeding site is buried in the soil by the beetles. Biparental care for offspring includes both preservation of the carrion and its preparation as diet and nursery. Buried carcasses show no signs of microbial decay, and those experimentally treated with Nicrophorus secretions are known to grow fewer bacteria and fungi. In order to investigate the chemical composition of these secretions, we used GC-MS for analysis of methanolic extracts of anal and oral secretions released by adult N. vespilloides. Furthermore, we analyzed the headspace of adult N. vespilloides by SPME-GC-MS and searched for compounds with known antimicrobial activity. We identified 34 compounds in the headspace, and anal and oral secretions, 26 of which occurred consistently. We discuss the ecological relevance of these compounds with respect to both their antimicrobial activity and ecological relevance.