Background The ability of Mycobacterium tuberculosis to survive and replicate in macrophages is crucial for the mycobacterium's ability to infect the host and cause tuberculosis. To identify Mycobacterium tuberculosis genes involved in survival in macrophages, a library of non-pathogenic Mycobacterium smegmatis bacteria, each carrying an individual integrated cosmid containing M. tuberculosis H37Rv genomic DNA, was passed through THP-1 human macrophages three times. Results Two of the clones recovered from this enrichment process, sur2 and sur3, exhibited significantly increased survival relative to wild-type bacteria. In coinfection experiments, the ratio of sur2 colonies to wild-type colonies was 1:1 at 0 hours but increased to 20:1 at 24 hours post phagocytosis. The ratio of sur3 colonies to wild-type colonies was 1:1 at 0 hours and 5:1 at 24 hours. The M. tuberculosis ORFs responsible for increased survival were shown to be Rv0365c for the sur2 clone and Rv2235 for the sur3 clone. These ORFs encode proteins with as-of-yet unknown functions. Conclusions We identified two M. tuberculosis ORFs which may be involved in the ability of tubercle bacilli to survive in macrophages.

H. verticillata samples collected from numerous watersheds from 2014 to 2020 were screened for the presence of A. hydrillicola. After isolation of A. hydrillicola from environmental samples and adaptation to laboratory conditions, the cyanobacterium was cultivated in BG11 medium with or without the addition of potassium bromide. H. verticillata leaves colonized with A. hydrillicola were analyzed using fluorescence microscopy as well as AP-MALDI-MSI (9-AA as matrix in negative-ionization mode; lateral resolution, 10 μm). Environmental bromide and bromine concentrations in H. verticillata, sediment, and water samples were analyzed by x-ray fluorescence spectroscopy and ion chromatography. The structure of AETX was elucidated by NMR spectroscopy, high-resolution tandem mass spectrometry, infrared spectroscopy, and x-ray crystallography after isolation of the compound using flash chromatography, semipreparative HPLC, and recrystallization. The genome of A. hydrillicola was amplified from single filaments using multiple displacement amplification and then sequenced using the Illumina MiSeq platform. The putative AETX biosynthetic gene cluster was identified by BLASTp searches for bacterial halogenases against the A. hydrillicola genome. The halogenase AetF was heterologously expressed in E. coli and then purified. Biochemical assays to characterize its activity used tryptophans and indoles as substrates. Reaction products were structurally characterized by HPLC-MS and NMR spectroscopy. A. hydrillicola–H. verticillata extract fractions and pure AETX were tested for activity on C. dubia, D. rerio, C. elegans, and G. gallus. Bioassays on D. rerio and G. gallus were performed in accordance with the National Insitutes of Health Guide for the Care and Use of Laboratory Animals and followed protocol A2017 11-007-Y1-A0, which was reviewed, approved, and overseen by the University of Georgia Institutional Animal Care and Use Committee. VM occurrence in treated birds was confirmed by analysis of the white matter of their optic lobe using light microscopy and transmission electron microscopy. Tissues of deceased wild birds were extracted and analyzed for AETX by HPLC-MS. A full description of the materials and methods used in this study is provided in the supplementary materials. Portions of this dataset are inaccessible because: Data belongs to coauthors at Martin-Luther-University Halle-Wittenberg, Halle (Saale), University of Georgia, Czech Academy of Sciences, and Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), They can be accessed through the following means: NMR and MS raw data are available at Figshare (https://doi.org/10.6084/m9.figshare.6025748.v1). X-ray data and models are available at the Cambridge Crystallographic Data Centre under accession no. CCDC-2018827. (https://doi.org/10.5517/ccdc.csd.cc25rrg5) The whole-genome assemblies (Whole Genome Shotgun projects) of two A. hydrillicola strains, CCALA 1050 and Thurmond2011, have been deposited at DDBJ/ENA/GenBank under the accession nos. JAALHA000000000 and JAAKGC000000000, respectively. The versions described in this paper are JAALHA010000000 and JAAKGC010000000. The sequence of the putative AETX biosynthetic gene cluster can be found at DDBJ/ENA/GenBank under the accession no. MT225528. All other data are available in the main text or the supplementary materials of the journal article. Format: data are in several formats, see accessed information for more information. This dataset is associated with the following publication: Breinlinger, S., T. Phillips, B. Haram, J. Mareš, J. Martínez Yerena, P. Hrouzek, R. Sobotka, W. Henderson, P. Schmieder, S.M. Williams, J.D. Lauderdale, H.D. Wilde, W. Gerrin, A. Kust, J. Washington, C. Wagner, M. Liebeke, H. Enke, T. Niedermeyer, and S. Wilde. Hunting the eagle killer: A cyanobacterial neurotoxin causes vacuolar myelinopathy. SCIENCE. American Association for the Advancement of Science (AAAS), Washington, DC, USA, 371(6536):

H. verticillata samples collected from numerous watersheds from 2014 to 2020 were screened for the presence of A. hydrillicola. After isolation of A. hydrillicola from environmental samples and adaptation to laboratory conditions, the cyanobacterium was cultivated in BG11 medium with or without the addition of potassium bromide. H. verticillata leaves colonized with A. hydrillicola were analyzed using fluorescence microscopy as well as AP-MALDI-MSI (9-AA as matrix in negative-ionization mode; lateral resolution, 10 μm). Environmental bromide and bromine concentrations in H. verticillata, sediment, and water samples were analyzed by x-ray fluorescence spectroscopy and ion chromatography. The structure of AETX was elucidated by NMR spectroscopy, high-resolution tandem mass spectrometry, infrared spectroscopy, and x-ray crystallography after isolation of the compound using flash chromatography, semipreparative HPLC, and recrystallization. The genome of A. hydrillicola was amplified from single filaments using multiple displacement amplification and then sequenced using the Illumina MiSeq platform. The putative AETX biosynthetic gene cluster was identified by BLASTp searches for bacterial halogenases against the A. hydrillicola genome. The halogenase AetF was heterologously expressed in E. coli and then purified. Biochemical assays to characterize its activity used tryptophans and indoles as substrates. Reaction products were structurally characterized by HPLC-MS and NMR spectroscopy. A. hydrillicola–H. verticillata extract fractions and pure AETX were tested for activity on C. dubia, D. rerio, C. elegans, and G. gallus. Bioassays on D. rerio and G. gallus were performed in accordance with the National Insitutes of Health Guide for the Care and Use of Laboratory Animals and followed protocol A2017 11-007-Y1-A0, which was reviewed, approved, and overseen by the University of Georgia Institutional Animal Care and Use Committee. VM occurrence in treated birds was confirmed by analysis of the white matter of their optic lobe using light microscopy and transmission electron microscopy. Tissues of deceased wild birds were extracted and analyzed for AETX by HPLC-MS. A full description of the materials and methods used in this study is provided in the supplementary materials. Portions of this dataset are inaccessible because: Data belongs to coauthors at Martin-Luther-University Halle-Wittenberg, Halle (Saale), University of Georgia, Czech Academy of Sciences, and Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), They can be accessed through the following means: NMR and MS raw data are available at Figshare (https://doi.org/10.6084/m9.figshare.6025748.v1). X-ray data and models are available at the Cambridge Crystallographic Data Centre under accession no. CCDC-2018827. (https://doi.org/10.5517/ccdc.csd.cc25rrg5) The whole-genome assemblies (Whole Genome Shotgun projects) of two A. hydrillicola strains, CCALA 1050 and Thurmond2011, have been deposited at DDBJ/ENA/GenBank under the accession nos. JAALHA000000000 and JAAKGC000000000, respectively. The versions described in this paper are JAALHA010000000 and JAAKGC010000000. The sequence of the putative AETX biosynthetic gene cluster can be found at DDBJ/ENA/GenBank under the accession no. MT225528. All other data are available in the main text or the supplementary materials of the journal article. Format: data are in several formats, see accessed information for more information. This dataset is associated with the following publication: Breinlinger, S., T. Phillips, B. Haram, J. Mareš, J. Martínez Yerena, P. Hrouzek, R. Sobotka, W. Henderson, P. Schmieder, S.M. Williams, J.D. Lauderdale, H.D. Wilde, W. Gerrin, A. Kust, J. Washington, C. Wagner, M. Liebeke, H. Enke, T. Niedermeyer, and S. Wilde. Hunting the eagle killer: A cyanobacterial neurotoxin causes vacuolar myelinopathy. SCIENCE. American Association for the Advancement of Science (AAAS), Washington, DC, USA, 371(6536):

Freshwater mussels of the order Unionida are among the most endangered animal groups globally, but the causes of population declines are often enigmatic with little known about the role of disease. In 2018, we collected wild adult pheasantshell (Actinonaias pectorosa) and mucket (Actinonaias ligamentina) during an epidemiologic survey investigating an ongoing mussel mass mortality event in the Clinch River, USA. Histopathology and transmission electron microscopy showed a novel microsporidian parasite primarily infecting the ovary of pheasantshell. Sequencing of the small subunit rRNA gene produced a 1333 bp sequence with greatest similarity to Pseudonosema cristatellae (AF484694.1; 86.36%; e-value = 0), a microsporidium infecting the freshwater bryozoan (Cristatella mucedo). Microsporidia were observed in 65% (17/26) of the examined female pheasantshell (A. pectorosa) and in no (0/2) female mucket (A. ligamentina), and occurred at mortality and non-mortality sites. Our findings indicate that a novel parasite, Microsporidium clinchi n. sp., is present in pheasantshell in the Clinch River, USA, and while likely not a cause of mass mortality, could reduce fecundity and recruitment in this declining population and threaten the success of reintroductions. Surveillance for M. clinchi n. sp. and evaluation of brood stock and their progeny for microsporidia would therefore be prudent.

Co-infection data in the form of colony forming units and amoeba cell counts. This dataset is associated with the following publication: Buse , H., F. Schaefer, and G. Rice. Enhanced survival but not amplification of Francisella spp. in the presence of free-living amoebae. Acta Microbiologica et Immunologica Hungarica. Akademiai Kiado, Budapest, HUNGARY, 64(1): 17-36, (2016).