Horizontal acquisition of a patchwork Calvin cycle by symbiotic and free-living Campylobacterota (formerly Epsilonproteobacteria).

Assie A, Leisch N, Meier DV, et al. Horizontal acquisition of a patchwork Calvin cycle by symbiotic and free-living Campylobacterota (formerly Epsilonproteobacteria). The ISME journal. 2019;14(1):104-122.Most autotrophs use the Calvin-Benson-Bassham (CBB) cycle for carbon fixation. In contrast, all currently described autotrophs from the Campylobacterota (previously Epsilonproteobacteria) use the reductive tricarboxylic acid cycle (rTCA) instead. We discovered campylobacterotal epibionts ("Candidatus Thiobarba") of deep-sea mussels that have acquired a complete CBB cycle and may have lost most key genes of the rTCA cycle. Intriguingly, the phylogenies of campylobacterotal CBB cyclegenes suggest they were acquired in multiple transfers from Gammaproteobacteria closely related to sulfur-oxidizing endosymbionts associated with the mussels, as well as from Betaproteobacteria. We hypothesize that "Ca. Thiobarba" switched from the rTCA cycle to a fully functional CBB cycle during its evolution, by acquiring genes from multiple sources, including co-occurring symbionts. We also found key CBB cycle genes in free-living Campylobacterota, suggesting that the CBB cycle may be more widespread in this phylum than previously known. Metatranscriptomics and metaproteomics confirmed high expression of CBB cycle genes in mussel-associated "Ca. Thiobarba". Direct stable isotope fingerprinting showed that "Ca. Thiobarba" has typical CBB signatures, suggesting that it uses this cycle for carbon fixation. Our discovery calls into question current assumptions about the distribution of carbon fixation pathways in microbial lineages, and the interpretation of stable isotope measurements in the environment

Microanatomy of the trophosome region of Paracatenula cf. polyhymnia (Catenulida, Platyhelminthes) and its intracellular symbionts

Marine catenulid platyhelminths of the genus Paracatenula lack mouth, pharynx and gut. They live in a symbiosis with intracellular bacteria which are restricted to the body region posterior to the brain. The symbiont-housing cells (bacteriocytes) collectively form the trophosome tissue, which functionally replaces the digestive tract. It constitutes the largest part of the body and is the most important synapomorphy of this group. While some other features of the Paracatenula anatomy have already been analyzed, an in-depth analysis of the trophosome region was missing. Here, we identify and characterize the composition of the trophosome and its surrounding tissue by analyzing series of ultra-thin cross-sections of the species Paracatenula cf. polyhymnia. For the first time, a protonephridium is detected in a Paracatenula species, but it is morphologically reduced and most likely not functional. Cells containing needle-like inclusions in the reference species Paracatenula polyhymnia Sterrer and Rieger, 1974 were thought to be sperm, and the inclusions interpreted as the sperm nucleus. Our analysis of similar cells and their inclusions by EDX and Raman microspectroscopy documents an inorganic spicule consisting of a unique magnesium–phosphate compound. Furthermore, we identify the neoblast stem cells located underneath the epidermis. Except for the modifications due to the symbiotic lifestyle and the enigmatic spicule cells, the organization of Paracatenula cf. polyhymnia conforms to that of the Catenulida in all studied aspects. Therefore, this species represents an excellent model system for further studies of host adaptation to an obligate symbiotic lifestyle

Bacterial Symbiosis Maintenance in the Asexually Reproducing and Regenerating Flatworm Paracatenula galateia

Bacteriocytes set the stage for some of the most intimate interactions between animal and bacterial cells. In all bacteriocyte possessing systems studied so far, de novo formation of bacteriocytes occurs only once in the host development, at the time of symbiosis establishment. Here, we present the free-living symbiotic flatworm Paracatenula galateia and its intracellular, sulfur-oxidizing bacteria as a system with previously undescribed strategies of bacteriocyte formation and bacterial symbiont transmission. Using thymidine analogue S-phase labeling and immunohistochemistry, we show that all somatic cells in adult worms – including bacteriocytes – originate exclusively from aposymbiotic stem cells (neoblasts). The continued bacteriocyte formation from aposymbiotic stem cells in adult animals represents a previously undescribed strategy of symbiosis maintenance and makes P. galateia a unique system to study bacteriocyte differentiation and development. We also provide morphological and immunohistochemical evidence that P. galateia reproduces by asexual fragmentation and regeneration (paratomy) and, thereby, vertically transmits numerous symbiont-containing bacteriocytes to its asexual progeny. Our data support the earlier reported hypothesis that the symbiont population is subjected to reduced bottleneck effects. This would justify both the codiversification between Paracatenula hosts and their Candidatus Riegeria symbionts, and the slow evolutionary rates observed for several symbiont genes

Comparison of four different fixatives using Mytilus edulis gill tissue

Gill-pieces of the same individual of Mytilus edulis were subjected to fixation with glutaraldehyde, buffered with different buffers (1) FSW= filtered sea water, (2)PBS = phosphate buffer saline, (3) PHEM= PIPES,HEPES,EGTA and MgCl2 and (4)Cacodylate<br

Flexible Generation of E-Learning Exams in R: Moodle Quizzes, OLAT Assessments, and Beyond

The capabilities of the package exams for automatic generation of (statistical) exams in R are extended by adding support for learning management systems: As in earlier versions of the package exam generation is still based on separate Sweave ?les for each exercise but rather than just producing di?erent types of PDF output ?les, the package can now render the same exercises into a wide variety of output formats. These include HTML (with various options for displaying mathematical content) and XML speci?cations for online exams in learning management systems such as Moodle or OLAT. This ?exibility is accomplished by a new modular and extensible design of the package that allows for reading all weaved exercises into R and managing associated supplementary ?les (such as graphics or data ?les). The manuscript discusses the readily available user interfaces, the design of the underlying infrastructure, and how new functionality can be built on top of the existing tools

Coming together—symbiont acquisition and early development in deep-sea bathymodioline mussels

How and when symbionts are acquired by their animal hosts has a profound impact on the ecology and evolution of the symbiosis. Understanding symbiont acquisition is particularly challenging in deep-sea organisms because early life stages are so rarely found. Here, we collected early developmental stages of three deep-sea bathymodioline species from different habitats to identify when these acquire their symbionts and how their body plan adapts to a symbiotic lifestyle. These mussels gain their nutrition from chemosynthetic bacteria, allowing them to thrive at deep-sea vents and seeps worldwide. Correlative imaging analyses using synchrotron-radiation based microtomography together with light, fluorescence and electron microscopy revealed that the pediveliger larvae were aposymbiotic. Symbiont colonization began during metamorphosis from a planktonic to a benthic lifestyle, with the symbionts rapidly colonizing first the gills, the symbiotic organ of adults, followed by all other epithelia of their hosts. Once symbiont densities in plantigrades reached those of adults, the host's intestine changed from the looped anatomy typical for bivalves to a straightened form. Within the Mytilidae, this morphological change appears to be specific to Bathymodiolus and Gigantidas, and is probably linked to the decrease in the importance of filter feeding when these mussels switch to gaining their nutrition largely from their symbionts

Growth in width and FtsZ ring longitudinal positioning in a gammaproteobacterial symbiont

Rod-shaped bacteria usually grow in length and place their FtsZ ring and division site at midcell, perpendicular to their long axis [1,2]. Here, we provide morphometric and immunocytochemical evidence that a nematode-associated gammaproteobacterium [3,4] grows in width, sets a constricting FtsZ ring parallel to its long axis, and divides longitudinally by default. Remarkably, the newly described FtsZ ring appears to be not only 90 degrees shifted with respect to model rods, but also elliptical and discontinuous. This reveals an unexpected versatility of the gammaproteobacterial cytokinetic machinery

Correlative 3D anatomy and spatial chemistry in animal-microbe symbioses: developing sample preparation for phase-contrast synchrotron radiation based micro-computed tomography and mass spectrometry imaging

The three-dimensional (3D) architecture of organs, tissues and cells together with the spatial distribution of specific molecules, both enables and drives the close interactions between hosts and microbes. To image the 3D anatomy and molecular composition of an animal-microbe system at a single cell scale we developed a correlative workflow combining phase-contrast synchrotron radiation based micro-computed tomography (PC-SRμCT) and mass spectrometry imaging (MSI). The key challenge in combining both techniques was the synchronization of sample preparation and imaging conditions. We used PC-SRμCT for high-resolution 3D imaging, which is the most suitable x-ray tomographic technique to image soft tissue samples without contrast enhancing agents at a single-cell level. For method development, we used small invertebrates that live in symbiosis with bacteria. For PC-SRμCT samples were paraformaldehyde-fixed and submersed in a buffer-filled capillary. We tested sample preparation for Matrix-assisted laser desorption ionization (MALDI)-MSI after PC-SRμCT and achieved the best results for PFA-fixed samples, processed under cryogenic conditions. The resulting molecular composition permitted imaging of hundreds of different molecule distributions from single tissue sections on a micrometer resolution. Image analysis revealed organ- and cell-specific metabolite distributions, which we could match to corresponding structures in the anatomical 3D PC-SRμCT model.Our PC-SRμCT-MSI sample preparation- and imaging workflow results in a detailed 3D scenario, which provides an atlas to guide other microscopy or omics approaches towards specific organs and cells. Particularly, for animal-microbe systems this approach presents a powerful tool to visualize anatomical and chemical processes at a microscale, linking structure and function in the symbiosis