Differentiation of neural stem cells (NSCs) to neurons requires the activation of genes controlled by the repressor element 1 (RE1) silencing transcription factor (REST)/neuron-restrictive silencer factor (NRSF) protein complex. Important components of REST/NRSF are phosphatases (termed RNA polymerase II C-terminal domain small phosphatases [CTDSPs]) that inhibit RNA polymerase II and suppress neuronal gene expression in NSCs. Activation of genes controlled by CTDSPs is required for neurogenesis, but how this is achieved is not fully understood. Here we show that ctdsp2 is a target of miR-26b, a microRNA that is encoded in an intron of the ctdsp2 primary transcript. This intrinsic negative feedback loop is inactive in NSCs because miR-26b biogenesis is inhibited at the precursor level. Generation of mature miR-26b is activated during neurogenesis, where it suppresses Ctdsp2 protein expression and is required for neuronal cell differentiation in vivo.

New and emerging environmental pathogens pose some of the greatest threats to modern aquaculture, a critical source of food protein globally. As with other intensive farming practices, increasing our understanding of the biology of infections is important to improve animal welfare and husbandry. The gill infection epitheliocystis is increasingly problematic in gilthead seabream (Sparus aurata), a major Mediterranean aquaculture species. Epitheliocystis is generally associated with chlamydial bacteria, yet we were not able to localise chlamydial targets within the major gilthead seabream lesions. Two previously unidentified species within a novel β-proteobacterial genus were instead identified. These co-infecting intracellular bacteria have been characterised using highresolution imaging and genomics, presenting the most comprehensive study on epitheliocystis agents to date. Draft genomes of the two uncultured species, Ca. Ichthyocystis hellenicum and Ca. Ichthyocystis sparus, have been de novo sequenced and annotated from preserved material. Analysis of the genomes shows a compact core indicating a metabolic dependency on the host, and an accessory genome with an unprecedented number of tandemly arrayed gene families. This study represents a critical insight into novel, emerging fish pathogens and will be used to underpin future investigations into the bacterial origins, and to develop diagnostic and treatment strategies.

Aquaculture is a burgeoning industry, requiring diversification into new farmed species, which are often at risk from infectious disease. We used a mesocosm technique to investigate the susceptibility of sharpsnout seabream (Diplodus puntazzo) larvae to potential environmental pathogens in seawater compared to control borehole water. Fish exposed to seawater succumbed to epitheliocystis from 21 days post hatching, causing mortality in a quarter of the hosts. The pathogen responsible was not chlamydial, as is often found in epitheliocystis, but a novel species of the γ-proteobacterial genus Endozoicomonas. Detailed characterisation of this pathogen within the infectious lesions using high resolution fluorescent and electron microscopy showed densely packed rod shaped bacteria. A draft genome sequence of this uncultured bacterium was obtained from preserved material. Comparison with the genome of the Endozoicomonas elysicola type strain shows that the genome of Ca. Endozoicomonas cretensis is undergoing decay through loss of functional genes and insertion sequence expansion, often indicative of adaptation to a new niche or restriction to an alternative lifestyle. These results demonstrate the advantage of mesocosm studies for investigating the effect of environmental bacteria on susceptible hosts and provide an important insight into the genome dynamics of a novel fish pathogen.

Chlamydia are obligate intracellular bacteria and important pathogens of humans and animals. Chlamydia-related bacteria are also major fish pathogens, infecting epithelial cells of the gills and skin to cause the disease epitheliocystis. Given the wide distribution, ancient origins and spectacular diversity of bony fishes, this group offers a rich resource for the identification and isolation of novel Chlamydia. The broad-nosed pipefish (Syngnathus typhle) is a widely distributed and genetically diverse temperate fish species, susceptible to epitheliocystis across much of its range. We describe here a new bacterial species, Candidatus Syngnamydia venezia; epitheliocystis agent of S. typhle and close relative to other chlamydial pathogens which are known to infect diverse hosts ranging from invertebrates to humans.

No abstract

Enterohemorrhagic Escherichia coli (EHEC) is a foodborne pathogen which can cause diarrhea, vomiting, and, in some cases, severe complications such as kidney failure in humans. Up to 30% of cattle are colonized with EHEC, which can enter the food chain through contaminated meat, dairy, and vegetables. In order to control infections and stop transmission, it is important to understand what factors allow EHEC to colonize its hosts, cause virulence, and aid transmission. Since this cannot be systematically studied in humans, it is important to develop animal models of infection and transmission. We developed a model which allows us to study foodborne infection in zebrafish, a vertebrate host that is transparent and genetically tractable. Our results show that foodborne infection is more efficient than waterborne infection and that the locus of enterocyte effacement is a key virulence determinant in the zebrafish model. It is induced early during infection, and loss of tight LEE regulation leads to a decreased bacterial burden and decreased host mortality. Overall, the zebrafish model allows us to study foodborne infection, including pathogen release from the food vehicle and gene regulation and its context of host-microbe interactions, as well as environmental shedding and transmission to naive hosts.

Bacteria belonging to the genus Cronobacter spp. have been recognized as causative agents of life-threatening systemic infections, primarily in premature, low-birth weight and/or immune-compromised neonates. Knowledge remains scarce regarding the underlying molecular mechanisms of disease development. In this study, we evaluated the use of a zebrafish model to study the pathogenesis of Cronobacter turicensis LMG 23827T, a clinical isolate responsible for two fatal sepsis cases in neonates. Here, the microinjection of approximately 50 colony forming units (CFUs) into the yolk sac resulted in the rapid multiplication of bacteria and dissemination into the blood stream at 24 h post infection (hpi), followed by the development of a severe bacteremia and larval death within 3 days. In contrast, the innate immune response of the embryos was sufficiently developed to control infection after the intravenous injection of up to 104 CFUs of bacteria. Infection studies using an isogenic mutant devoid of surviving and replicating in human macrophages (ΔfkpA) showed that this strain was highly attenuated in its ability to kill the larvae. In addition, the suitability of the zebrafish model system to study the effectiveness of antibiotics to treat Cronobacter infections in zebrafish embryos was examined. Our data indicate that the zebrafish model represents an excellent vertebrate model to study virulence-related aspects of this opportunistic pathogen in vivo.

The Planctomycetes-Verrucomicrobia-Chlamydiae (PVC) bacterial superphylum constitutes a broad range of organisms with an intriguing array of ultrastructural morphologies, including intracellular membranes and compartments and their corresponding complex genomes encoding these forms. The phylum Chlamydiae are all obligate intracellular bacteria and, although much is already known of their genomes from various families and how these regulate the various morphological forms, we know remarkably little about what is likely the deepest rooting clade of this phylum, which has only been found to contain pathogens of marine and fresh water vertebrates. The disease they are associated with is called epitheliocystis; however, analyses of the causative agents is hindered by an inability to cultivate them for refined in vitro experimentation. For this reason, we have developed tools to analyse both the genomes and the ultrastructures of bacteria causing this disease, directly from infected tissues. Here we present structural data for a member of the family Ca. Similichlamydiaceae from this deep-rooted clade, which we have identified using molecular tools, in epitheliocystis lesions of gilthead seabream (Sparus aurata) in Greece. We present evidence that the chlamydial inclusions appear to develop in a perinuclear location, similar to other members of the phylum and that a chlamydial developmental cycle is present, with chlamydial forms similar to reticular bodies (RBs) and elementary bodies (EBs) detected. Division of the RBs appeared to follow a budding process, and larger RBs with multiple condensed nucleoids were detected using both transmission electron microscopy (TEM) and by focused-ion beam, scanning electron microscopy (FIB-SEM). As model hosts, fish offer many advantages for investigation, and we hope by these efforts to encourage others to explore the biology of fish pathogens from the PVC.