Macrocyclic colibactin induces DNA double-strand breaks via copper-mediated oxidative cleavage.

Colibactin is an assumed human gut bacterial genotoxin, whose biosynthesis is linked to the clb genomic island that has a widespread distribution in pathogenic and commensal human enterobacteria. Colibactin-producing gut microbes promote colon tumour formation and enhance the progression of colorectal cancer via cellular senescence and death induced by DNA double-strand breaks (DSBs); however, the chemical basis that contributes to the pathogenesis at the molecular level has not been fully characterized. Here, we report the discovery of colibactin-645, a macrocyclic colibactin metabolite that recapitulates the previously assumed genotoxicity and cytotoxicity. Colibactin-645 shows strong DNA DSB activity in vitro and in human cell cultures via a unique copper-mediated oxidative mechanism. We also delineate a complete biosynthetic model for colibactin-645, which highlights a unique fate of the aminomalonate-building monomer in forming the C-terminal 5-hydroxy-4-oxazolecarboxylic acid moiety through the activities of both the polyketide synthase ClbO and the amidase ClbL. This work thus provides a molecular basis for colibactin's DNA DSB activity and facilitates further mechanistic study of colibactin-related colorectal cancer incidence and prevention

From insect to man: Photorhabdus sheds light on the emergence of human pathogenicity

Photorhabdus are highly effective insect pathogenic bacteria that exist in a mutualistic relationship with Heterorhabditid nematodes. Unlike other members of the genus, Photorhabdus asymbiotica can also infect humans. Most Photorhabdus cannot replicate above 34°C, limiting their host-range to poikilothermic invertebrates. In contrast, P. asymbiotica must necessarily be able to replicate at 37°C or above. Many well-studied mammalian pathogens use the elevated temperature of their host as a signal to regulate the necessary changes in gene expression required for infection. Here we use RNA-seq, proteomics and phenotype microarrays to examine temperature dependent differences in transcription, translation and phenotype of P. asymbiotica at 28°C versus 37°C, relevant to the insect or human hosts respectively. Our findings reveal relatively few temperature dependant differences in gene expression. There is however a striking difference in metabolism at 37°C, with a significant reduction in the range of carbon and nitrogen sources that otherwise support respiration at 28°C. We propose that the key adaptation that enables P. asymbiotica to infect humans is to aggressively acquire amino acids, peptides and other nutrients from the human host, employing a so called “nutritional virulence” strategy. This would simultaneously cripple the host immune response while providing nutrients sufficient for reproduction. This might explain the severity of ulcerated lesions observed in clinical cases of Photorhabdosis. Furthermore, while P. asymbiotica can invade mammalian cells they must also resist immediate killing by humoral immunity components in serum. We observed an increase in the production of the insect Phenol-oxidase inhibitor Rhabduscin normally deployed to inhibit the melanisation immune cascade. Crucially we demonstrated this molecule also facilitates protection against killing by the alternative human complement pathway

Successful validation of a larval dispersal model using genetic parentage data

Larval dispersal is a critically important yet enigmatic process in marine ecology, evolution, and conservation. Determining the distance and direction that tiny larvae travel in the open ocean continues to be a challenge. Our current understanding of larval dispersal patterns at management-relevant scales is principally and separately informed by genetic parentage data and biological-oceanographic (biophysical) models. Parentage datasets provide clear evidence of individual larval dispersal events, but their findings are spatially and temporally limited. Biophysical models offer a more complete picture of dispersal patterns at regional scales but are of uncertain accuracy. Here, we develop statistical techniques that integrate these two important sources of information on larval dispersal. We then apply these methods to an extensive genetic parentage dataset to successfully validate a high-resolution biophysical model for the economically important reef fish species Plectropomus maculatus in the southern Great Barrier Reef. Our results demonstrate that biophysical models can provide accurate descriptions of larval dispersal at spatial and temporal scales that are relevant to management. They also show that genetic parentage datasets provide enough statistical power to exclude poor biophysical models. Biophysical models that included species-specific larval behaviour provided markedly better fits to the parentage data than assuming passive behaviour, but incorrect behavioural assumptions led to worse predictions than ignoring behaviour altogether. Our approach capitalises on the complementary strengths of genetic parentage datasets and high-resolution biophysical models to produce an accurate picture of larval dispersal patterns at regional scales. The results provide essential empirical support for the use of accurately parameterised biophysical larval dispersal models in marine spatial planning and management

Gene probing reveals the widespread distribution, diversity and abundance of isoprene-degrading bacteria in the environment

Background: Approximately 500 Tg of isoprene are emitted to the atmosphere annually, an amount similar to that of methane, and despite its significant effects on the climate, very little is known about the biological degradation of isoprene in the environment. Isolation and characterisation of isoprene degraders at the molecular level has allowed the development of probes targeting isoA encoding the α-subunit of the isoprene monooxygenase. This enzyme belongs to the soluble diiron centre monooxygenase family and catalyses the first step in the isoprene degradation pathway. The use of probes targeting key metabolic genes is a successful approach in molecular ecology to study specific groups of bacteria in complex environments. Here, we developed and tested a novel isoA PCR primer set to study the distribution, abundance, and diversity of isoprene degraders in a wide range of environments. Results: The new isoA probes specifically amplified isoA genes from taxonomically diverse isoprene-degrading bacteria including members of the genera Rhodococcus, Variovorax, and Sphingopyxis. There was no cross-reactivity with genes encoding related oxygenases from non-isoprene degraders. Sequencing of isoA amplicons from DNA extracted from environmental samples enriched with isoprene revealed that most environments tested harboured a considerable variety of isoA sequences, with poplar leaf enrichments containing more phylogenetically diverse isoA genes. Quantification by qPCR using these isoA probes revealed that isoprene degraders are widespread in the phyllosphere, terrestrial, freshwater and marine environments. Specifically, soils in the vicinity of high isoprene-emitting trees contained the highest number of isoprene-degrading bacteria. Conclusion: This study provides the molecular ecology tools to broaden our knowledge of the distribution, abundance and diversity of isoprene degraders in the environment, which is a fundamental step necessary to assess the impact that microbes have in mitigating the effects of this important climate-active gas

Liverpool telescope 2: a new robotic facility for rapid transient follow-up

The Liverpool Telescope is one of the world's premier facilities for time domain astronomy. The time domain landscape is set to radically change in the coming decade, with surveys such as LSST providing huge numbers of transient detections on a nightly basis; transient detections across the electromagnetic spectrum from other facilities such as SVOM, SKA and CTA; and the era of `multi-messenger astronomy', wherein events are detected via non-electromagnetic means, such as gravitational wave emission. We describe here our plans for Liverpool Telescope 2: a new robotic telescope designed to capitalise on this new era of time domain astronomy. LT2 will be a 4-metre class facility co-located with the LT at the Observatorio del Roque de Los Muchachos on the Canary island of La Palma. The telescope will be designed for extremely rapid response: the aim is that the telescope will take data within 30 seconds of the receipt of a trigger from another facility. The motivation for this is twofold: firstly it will make it a world-leading facility for the study of fast fading transients and explosive phenomena discovered at early times. Secondly, it will enable large-scale programmes of low-to-intermediate resolution spectral classification of transients to be performed with great efficiency. In the target-rich environment of the LSST era, minimising acquisition overheads will be key to maximising the science gains from any follow-up programme. The telescope will have a diverse instrument suite which is simultaneously mounted for automatic changes, but it is envisaged that the primary instrument will be an intermediate resolution, optical/infrared spectrograph for scientific exploitation of transients discovered with the next generation of synoptic survey facilities. In this paper we outline the core science drivers for the telescope, and the requirements for the optical and mechanical design

Statistical Optimization of Process Variables for Antibiotic Activity of Xenorhabdus bovienii

The production of secondary metabolites with antibiotic properties is a common characteristic to entomopathogenic bacteria Xenorhabdus spp. These metabolites not only have diverse chemical structures but also have a wide range of bioactivities of medicinal and agricultural interests. Culture variables are critical to the production of secondary metabolites of microorganisms. Manipulating culture process variables can promote secondary metabolite biosynthesis and thus facilitate the discovery of novel natural products. This work was conducted to evaluate the effects of five process variables (initial pH, medium volume, rotary speed, temperature, and inoculation volume) on the antibiotic production of Xenorhabdus bovienii YL002 using response surface methodology. A 25–1 factorial central composite design was chosen to determine the combined effects of the five variables, and to design a minimum number of experiments. The experimental and predicted antibiotic activity of X. bovienii YL002 was in close agreement. Statistical analysis of the results showed that initial pH, medium volume, rotary speed and temperature had a significant effect (P<0.05) on the antibiotic production of X. bovienii YL002 at their individual level; medium volume and rotary speed showed a significant effect at a combined level and was most significant at an individual level. The maximum antibiotic activity (287.5 U/mL) was achieved at the initial pH of 8.24, medium volume of 54 mL in 250 mL flask, rotary speed of 208 rpm, temperature of 32.0°C and inoculation volume of 13.8%. After optimization, the antibiotic activity was improved by 23.02% as compared with that of unoptimized conditions

Characterization of the Partitioning System of Myxococcus Plasmid pMF1

pMF1 is the only autonomously replicating plasmid that has been recently identified in myxobacteria. This study characterized the partitioning (par) system of this plasmid. The fragment that significantly increased the retaining stability of plasmids in Myxococcus cells in the absence of selective antibiotics contained three open reading frames (ORFs) pMF1.21-pMF1.23 (parCAB). The pMF1.22 ORF (parA) is homologous to members of the parA ATPase family, with the highest similarity (56%) to the Sphingobium japonicum ParA-like protein, while the other two ORFs had no homologs in GenBank. DNase I footprinting and electrophoretic mobility shift assays showed that the pMF1.23 (parB) product is a DNA-binding protein of iteron DNA sequences, while the product of pMF1.21 (parC) has no binding activity but is able to enhance the DNA-binding activity of ParB to iterons. The ParB protein autogenously repressed the expression of the par genes, consistent with the type Ib par pattern, while the ParC protein has less repressive activity. The ParB-binding iteron sequences are distributed not only near the partitioning gene loci but also along pMF1. These results indicate that the pMF1 par system has novel structural and functional characteristics