A giant virus infecting the amoeboflagellate Naegleria.

Giant viruses (Nucleocytoviricota) are significant lethality agents of various eukaryotic hosts. Although metagenomics indicates their ubiquitous distribution, available giant virus isolates are restricted to a very small number of protist and algal hosts. Here we report on the first viral isolate that replicates in the amoeboflagellate Naegleria. This genus comprises the notorious human pathogen Naegleria fowleri, the causative agent of the rare but fatal primary amoebic meningoencephalitis. We have elucidated the structure and infection cycle of this giant virus, Catovirus naegleriensis (a.k.a. Naegleriavirus, NiV), and show its unique adaptations to its Naegleria host using fluorescence in situ hybridization, electron microscopy, genomics, and proteomics. Naegleriavirus is only the fourth isolate of the highly diverse subfamily Klosneuvirinae, and like its relatives the NiV genome contains a large number of translation genes, but lacks transfer RNAs (tRNAs). NiV has acquired genes from its Naegleria host, which code for heat shock proteins and apoptosis inhibiting factors, presumably for host interactions. Notably, NiV infection was lethal to all Naegleria species tested, including the human pathogen N. fowleri. This study expands our experimental framework for investigating giant viruses and may help to better understand the basic biology of the human pathogen N. fowleri.

Preadmission course and management of severe pediatric group A streptococcal infections during the 2022-2023 outbreak: a single-center experience.

PURPOSE: The massive increase of infections with Group A Streptococcus (GAS) in 2022-2023 coincided in Switzerland with a change of the recommendations for the management of GAS pharyngitis. Therefore, the objective of the present study was to investigate whether the clinical manifestations and management before hospitalization for GAS infection differed in 2022-2023 compared with 2013-2022. METHODS: Retrospective study of GAS infections requiring hospitalization in patients below 16 years. Preadmission illness (modified McIsaac score), oral antibiotic use, and outcome in 2022-2023 were compared with 2013-2022. Time series were compared with surveillance data for respiratory viruses. RESULTS: In 2022-2023, the median modified McIsaac score was lower (2 [IQR 2-3] vs. 3 [IQR 2-4], p = < 0.0001) and the duration of preadmission illness was longer (4 days [3-7] vs. 3 [2-6], p = 0.004) than in 2013-2022. In both periods, withholding of preadmission oral antibiotics despite a modified McIsaac score ≥ 3 (12% vs. 18%, n.s.) or ≥ 4 (2.4% vs. 10.0%, p = 0.027) was rare. Respiratory disease, skeletal/muscle infection, and invasive GAS disease were significantly more frequent in 2022-2023, but there were no differences in clinical outcome. The time course of GAS cases in 2022-2023 coincided with the activity of influenza A/B. CONCLUSION: We found no evidence supporting the hypothesis that the 2022-2023 GAS outbreak was associated with a change in preadmission management possibly induced by the new recommendation for GAS pharyngitis. However, clinical manifestations before admission and comparative examination of time-series strongly suggest that viral co-circulation played an important role in this outbreak.

Comparative genomics of a vertically transmitted thiotrophic bacterial ectosymbiont and its close free-living relative.

Thiotrophic symbioses between sulphur-oxidizing bacteria and various unicellular and metazoan eukaryotes are widespread in reducing marine environments. The giant colonial ciliate Zoothamnium niveum, however, is the only host of thioautotrophic symbionts that has been cultivated along with its symbiont, the vertically transmitted ectosymbiont Candidatus Thiobius zoothamnicola (short Thiobius). Because theoretical predictions posit a smaller genome in vertically transmitted endosymbionts compared to free-living relatives, we investigated whether this is true also for an ectosymbiont. We used metagenomics to recover the high-quality draft genome of this bacterial symbiont. For comparison we have also sequenced a closely related free-living cultured but not formally described strain Milos ODIII6 (short ODIII6). We then performed comparative genomics to assess the functional capabilities at gene, metabolic pathway and trait level. 16S rRNA gene trees and average amino acid identity confirmed the close phylogenetic relationship of both bacteria. Indeed, Thiobius has about a third smaller genome than its free-living relative ODIII6, with reduced metabolic capabilities and fewer functional traits. The functional capabilities of Thiobius were a subset of those of the more versatile ODIII6, which possessed additional genes for oxygen, sulphur and hydrogen utilization and for the acquisition of phosphorus illustrating features that may be adaptive for the unstable environmental conditions at hydrothermal vents. In contrast, Thiobius possesses genes potentially enabling it to utilize lactate and acetate heterotrophically, compounds that may be provided as byproducts by the host. The present study illustrates the effect of strict host-dependence of a bacterial ectosymbiont on genome evolution and host adaptation.

Long-term pulmonary outcome of children with congenital diaphragmatic hernia: functional lung MRI using matrix-pencil decomposition enables side-specific assessment of lung function.

OBJECTIVES: In patients with congenital diaphragmatic hernia (CDH) the exact functional outcome of the affected lung side is still unknown, mainly due to the lack of spatially resolved diagnostic tools. Functional matrix-pencil decomposition (MP-) lung MRI fills this gap as it measures side-specific ventilation and perfusion. We aimed to assess the overall and side-specific pulmonary long-term outcomes of patients with CDH using lung function tests and MP-MRI. METHODS: Thirteen school-aged children with CDH (seven with small and six with large defect-sized CDH, defined as > 50% of the chest wall circumference being devoid of diaphragm tissue) and thirteen healthy matched controls underwent spirometry, multiple-breath washout, and MP-MRI. The main outcomes were forced expiratory volume in 1 second (FEV1), lung clearance index (LCI2.5), ventilation defect percentage (VDP), and perfusion defect percentage (QDP). RESULTS: Patients with a large CDH showed significantly reduced overall lung function compared to healthy controls (mean difference [95%-CIadjusted]: FEV1 (z-score) -4.26 [-5.61, -2.92], FVC (z-score) -3.97 [-5.68, -2.26], LCI2.5 (TO) 1.12 [0.47, 1.76], VDP (%) 8.59 [3.58, 13.60], QDP (%) 17.22 [13.16, 21.27]) and to patients with a small CDH. Side-specific examination by MP-MRI revealed particularly reduced ipsilateral ventilation and perfusion in patients with a large CDH (mean difference to contralateral side [95%-CIadjusted]: VDP (%) 14.80 [10.50, 19.00], QDP (%) 23.50 [1.75, 45.20]). CONCLUSIONS: Data indicate impaired overall lung function with particular limitation of the ipsilateral side in patients with a large CDH. MP-MRI is a promising tool to provide valuable side-specific functional information in the follow-up of patients with CDH. CLINICAL RELEVANCE STATEMENT: In patients with congenital diaphragmatic hernia, easily applicable MP-MRI allows specific examination of the lung side affected by the hernia and provides valuable information on ventilation and perfusion with implications for clinical practice, making it a promising tool for routine follow-up. KEY POINTS: • Functional matrix pencil decomposition (MP) MRI data from a small sample indicate reduced ipsilateral pulmonary ventilation and perfusion in children with large congenital diaphragmatic hernia (CDH). • Easily applicable pencil decomposition MRI provides valuable side-specific diagnostic information on lung ventilation and perfusion. This is a clear advantage over conventional lung function tests, helping to comprehensively follow up patients with congenital diaphragmatic hernia and monitor therapy effects.

PSMA-PET improves deep learning-based automated CT kidney segmentation.

For dosimetry of radiopharmaceutical therapies, it is essential to determine the volume of relevant structures exposed to therapeutic radiation. For many radiopharmaceuticals, the kidneys represent an important organ-at-risk. To reduce the time required for kidney segmentation, which is often still performed manually, numerous approaches have been presented in recent years to apply deep learning-based methods for CT-based automated segmentation. While the automatic segmentation methods presented so far have been based solely on CT information, the aim of this work is to examine the added value of incorporating PSMA-PET data in the automatic kidney segmentation. METHODS: A total of 108 PET/CT examinations (53 [68Ga]Ga-PSMA-I&T and 55 [18F]F-PSMA-1007 examinations) were grouped to create a reference data set of manual segmentations of the kidney. These segmentations were performed by a human examiner. For each subject, two segmentations were carried out: one CT-based (detailed) segmentation and one PET-based (coarser) segmentation. Five different u-net based approaches were applied to the data set to perform an automated segmentation of the kidney: CT images only, PET images only (coarse segmentation), a combination of CT and PET images, a combination of CT images and a PET-based coarse mask, and a CT image, which had been pre-segmented using a PET-based coarse mask. A quantitative assessment of these approaches was performed based on a test data set of 20 patients, including Dice score, volume deviation and average Hausdorff distance between automated and manual segmentations. Additionally, a visual evaluation of automated segmentations for 100 additional (i.e., exclusively automatically segmented) patients was performed by a nuclear physician. RESULTS: Out of all approaches, the best results were achieved by using CT images which had been pre-segmented using a PET-based coarse mask as input. In addition, this method performed significantly better than the segmentation based solely on CT, which was supported by the visual examination of the additional segmentations. In 80% of the cases, the segmentations created by exploiting the PET-based pre-segmentation were preferred by the nuclear physician. CONCLUSION: This study shows that deep-learning based kidney segmentation can be significantly improved through the addition of a PET-based pre-segmentation. The presented method was shown to be especially beneficial for kidneys with cysts or kidneys that are closely adjacent to other organs such as the spleen, liver or pancreas. In the future, this could lead to a considerable reduction in the time required for dosimetry calculations as well as an improvement in the results.

Measuring environmental awareness: An analysis using google search data.

Environmental awareness is usually measured using surveys. This paper aims to offer an alternative measure: an Environmental Awareness Index (EAI) constructed using Google search data provided by Google Trends. The benefits of using Google search data over surveys are that (i) they are less costly to obtain, (ii) they are available at high frequency, and (iii) they cover countries where no surveys are available. To test the validity of the proposed EAI, this study empirically assesses the impact of the computed index on individuals' pro-environmental behaviors using the Special Eurobarometer: Attitudes of European citizens towards the Environment data. Results show that the EAI is positively related to pro-environmental behaviors with a statistical significance at the one percent level. This finding stays robust in pooled OLS as well as in panel regression analysis when GDP, mean years of schooling, and population are included as control variables and when time-fixed effects are introduced. Further, the results confirm that environmental awareness is not stable over time and underline the importance of having a timely measure of environmental awareness at hand. Finally, the findings offer several practical implications for managers and policymakers, who will be able to use a timely measure of environmental awareness, assess and measure the impact of their policies aiming to raise environmental awareness as well as depict the type of behavior influenced by their policies.

The Fish Pathogen "Candidatus Clavichlamydia salmonicola"-A Missing Link in the Evolution of Chlamydial Pathogens of Humans.

Chlamydiae like Chlamydia trachomatis and Chlamydia psittaci are well-known human and animal pathogens. Yet, the chlamydiae are a much larger group of evolutionary ancient obligate intracellular bacteria that includes predominantly symbionts of protists and diverse animals. This makes them ideal model organisms to study evolutionary transitions from symbionts in microbial eukaryotes to pathogens of humans. To this end, comparative genome analysis has served as an important tool. Genome sequence data for many chlamydial lineages are, however, still lacking, hampering our understanding of their evolutionary history. Here, we determined the first high-quality draft genome sequence of the fish pathogen "Candidatus Clavichlamydia salmonicola", representing a separate genus within the human and animal pathogenic Chlamydiaceae. The "Ca. Clavichlamydia salmonicola" genome harbors genes that so far have been exclusively found in Chlamydia species suggesting that basic mechanisms important for the interaction with chordate hosts have evolved stepwise in the history of chlamydiae. Thus, the genome sequence of "Ca. Clavichlamydia salmonicola" allows to constrain candidate genes to further understand the evolution of chlamydial virulence mechanisms required to infect mammals.

Genome Dynamics and Temperature Adaptation During Experimental Evolution of Obligate Intracellular Bacteria.

Evolution experiments with free-living microbes have radically improved our understanding of genome evolution and how microorganisms adapt. Yet there is a paucity of such research focusing on strictly host-associated bacteria, even though they are widespread in nature. Here, we used the Acanthamoeba symbiont Protochlamydia amoebophila, a distant relative of the human pathogen Chlamydia trachomatis and representative of a large group of protist-associated environmental chlamydiae, as a model to study how obligate intracellular symbionts evolve and adapt to elevated temperature, a prerequisite for the pivotal evolutionary leap from protist to endothermic animal hosts. We established 12 replicate populations under two temperatures (20 °C, 30 °C) for 510 bacterial generations (38 months). We then used infectivity assays and pooled whole-genome resequencing to identify any evolved phenotypes and the molecular basis of adaptation in these bacteria. We observed an overall reduction in infectivity of the symbionts evolved at 30 °C, and we identified numerous nonsynonymous mutations and small indels in these symbiont populations, with several variants persisting throughout multiple time points and reaching high frequencies. This suggests that many mutations may have been beneficial and played an adaptive role. Mutated genes within the same temperature regime were more similar than those between temperature regimes. Our results provide insights into the molecular evolution of intracellular bacteria under the constraints of strict host dependance and highly structured populations and suggest that for chlamydial symbionts of protists, temperature adaptation was facilitated through attenuation of symbiont infectivity as a tradeoff to reduce host cell burden.

Use of gene sequences as type for naming prokaryotes: Recommendations of the international committee on the taxonomy of chlamydiae.

The International Committee on Systematics of Prokaryotes (ICSP) discussed and rejected in 2020 a proposal to modify the International Code of Nomenclature of Prokaryotes to allow the use of gene sequences as type for naming prokaryotes. An alternative nomenclatural code, the Code of Nomenclature of Prokaryotes Described from Sequence Data (SeqCode), which considers genome sequences as type material for naming species, was published in 2022. Members of the ICSP subcommittee for the taxonomy of the phylum Chlamydiae (Chlamydiota) consider that the use of gene sequences as type would benefit the taxonomy of microorganisms that are difficult to culture such as the chlamydiae and other strictly intracellular bacteria. We recommend the registration of new names of uncultured prokaryotes in the SeqCode registry.

Colocalization and potential interactions of Endozoicomonas and chlamydiae in microbial aggregates of the coral Pocillopora acuta.

Corals are associated with a variety of bacteria, which occur in the surface mucus layer, gastrovascular cavity, skeleton, and tissues. Some tissue-associated bacteria form clusters, termed cell-associated microbial aggregates (CAMAs), which are poorly studied. Here, we provide a comprehensive characterization of CAMAs in the coral Pocillopora acuta. Combining imaging techniques, laser capture microdissection, and amplicon and metagenome sequencing, we show that (i) CAMAs are located in the tentacle tips and may be intracellular; (ii) CAMAs contain Endozoicomonas (Gammaproteobacteria) and Simkania (Chlamydiota) bacteria; (iii) Endozoicomonas may provide vitamins to its host and use secretion systems and/or pili for colonization and aggregation; (iv) Endozoicomonas and Simkania occur in distinct, but adjacent, CAMAs; and (v) Simkania may receive acetate and heme from neighboring Endozoicomonas. Our study provides detailed insight into coral endosymbionts, thereby improving our understanding of coral physiology and health and providing important knowledge for coral reef conservation in the climate change era.

One to host them all: genomics of the diverse bacterial endosymbionts of the spider Oedothorax gibbosus.

Bacterial endosymbionts of the groups Wolbachia, Cardinium and Rickettsiaceae are well known for their diverse effects on their arthropod hosts, ranging from mutualistic relationships to reproductive phenotypes. Here, we analysed a unique system in which the dwarf spider Oedothorax gibbosus is co-infected with up to five different endosymbionts affiliated with Wolbachia, 'Candidatus Tisiphia' (formerly Torix group Rickettsia), Cardinium and Rhabdochlamydia. Using short-read genome sequencing data, we show that the endosymbionts are heterogeneously distributed among O. gibbosus populations and are frequently found co-infecting spider individuals. To study this intricate host-endosymbiont system on a genome-resolved level, we used long-read sequencing to reconstruct closed genomes of the Wolbachia, 'Ca. Tisiphia' and Cardinium endosymbionts. We provide insights into the ecology and evolution of the endosymbionts and shed light on the interactions with their spider host. We detected high quantities of transposable elements in all endosymbiont genomes and provide evidence that ancestors of the Cardinium, 'Ca. Tisiphia' and Wolbachia endosymbionts have co-infected the same hosts in the past. Our findings contribute to broadening our knowledge about endosymbionts infecting one of the largest animal phyla on Earth and show the usefulness of transposable elements as an evolutionary 'contact-tracing' tool.

Mathematical modeling of pneumococcal transmission dynamics in response to PCV13 infant vaccination in Germany predicts increasing IPD burden due to serotypes included in next-generation PCVs.

INTRODUCTION: Two next-generation pneumococcal conjugate vaccines (PCVs), a 15- and a 20-valent PCV (PCV15 and PCV20), have recently been licensed for use in adults, and PCV15 has also been licensed in children. We developed a dynamic transmission model specific for Germany, with the aim to predict carriage prevalence and invasive pneumococcal disease (IPD) burden for serotypes included in these vaccines. METHODS: The model allows to follow serotype distributions longitudinally both in the absence and presence of PCV vaccinations. We considered eight age cohorts and seven serotype groups according to the composition of different pneumococcal vaccines. This comprises the additional serotypes contained in PCV15 and PCV20 but not in PCV13. RESULTS: The model predicted that by continuing the current vaccine policy (standard vaccination with PCV13 in children and with PPSV23 in adults) until 2031, IPD case counts due to any serotype in children <2 years of age will remain unchanged. There will be a continuous decrease of IPD cases in adults aged 16-59y, but a 20% increase in adults ≥60y. Furthermore, there will be a steady decrease of the proportion of carriage and IPD due to serotypes included in PCV7 and PCV13 over the model horizon and a steady rise of non-PCV13 serotypes in carriage and IPD. The highest increase for both pneumococcal carriage and absolute IPD case counts was predicted for serotypes 22F and 33F (included in both PCV15 and PCV20) and serotypes 8, 10A, 11A, 12F, and 15B (included in PCV20 only), particularly in older adults. Between 2022 and 2031, serotypes included in PCV20 only are expected to cause 19.7-25.3% of IPD cases in adults ≥60y. CONCLUSIONS: We conclude that introduction of next-generation PCVs for adults may prevent a substantial and increasing proportion of adult IPDs, with PCV20 having the potential to provide the broadest protection against pneumococcal disease.

The Influence of ESG Ratings On Idiosyncratic Stock Risk: The Unrated, the Good, the Bad, and the Sinners.

This study analyzes whether stocks of companies with environmental social governance (ESG) rating show lower idiosyncratic risk. The main analysis covers 898,757 company-month observations of US stocks in the period from 1991 to 2018 and controls for stocks' exposure to liquidity, mispricing, innovations in volatility risk, investor sentiment, and analysts' forecast divergence. The main finding is that the receipt of an ESG rating decreases idiosyncratic stock risk. The effect is stronger for stocks that receive a higher ESG rating. Nevertheless, even when companies receive a lower ESG rating, they show significantly lower idiosyncratic risk than stocks without an ESG rating. Furthermore, stocks subject to a negative screen show lower idiosyncratic risk during recessions than comparable stocks with an ESG rating but without a negative screen. The results support the notion that the receipt of an ESG rating decreases uncertainty regarding future stock risk and return and show that ESG ratings and negative screens individually influence stock risk and, therefore, should be considered separately.

Gene gain facilitated endosymbiotic evolution of Chlamydiae.

Chlamydiae is a bacterial phylum composed of obligate animal and protist endosymbionts. However, other members of the Planctomycetes-Verrucomicrobia-Chlamydiae superphylum are primarily free living. How Chlamydiae transitioned to an endosymbiotic lifestyle is still largely unresolved. Here we reconstructed Planctomycetes-Verrucomicrobia-Chlamydiae species relationships and modelled superphylum genome evolution. Gene content reconstruction from 11,996 gene families suggests a motile and facultatively anaerobic last common Chlamydiae ancestor that had already gained characteristic endosymbiont genes. Counter to expectations for genome streamlining in strict endosymbionts, we detected substantial gene gain within Chlamydiae. We found that divergence in energy metabolism and aerobiosis observed in extant lineages emerged later during chlamydial evolution. In particular, metabolic and aerobic genes characteristic of the more metabolically versatile protist-infecting chlamydiae were gained, such as respiratory chain complexes. Our results show that metabolic complexity can increase during endosymbiont evolution, adding an additional perspective for understanding symbiont evolutionary trajectories across the tree of life.

Interrater variability of ML-based CT-FFR during TAVR-planning: influence of image quality and coronary artery calcifications.

Objective: To compare machine learning (ML)-based CT-derived fractional flow reserve (CT-FFR) in patients before transcatheter aortic valve replacement (TAVR) by observers with differing training and to assess influencing factors. Background: Coronary computed tomography angiography (cCTA) can effectively exclude CAD, e.g. prior to TAVR, but remains limited by its specificity. CT-FFR may mitigate this limitation also in patients prior to TAVR. While a high reliability of CT-FFR is presumed, little is known about the reproducibility of ML-based CT-FFR. Methods: Consecutive patients with obstructive CAD on cCTA were evaluated with ML-based CT-FFR by two observers. Categorization into hemodynamically significant CAD was compared against invasive coronary angiography. The influence of image quality and coronary artery calcium score (CAC) was examined. Results: CT-FFR was successfully performed on 214/272 examinations by both observers. The median difference of CT-FFR between both observers was -0.05(-0.12-0.02) (p < 0.001). Differences showed an inverse correlation to the absolute CT-FFR values. Categorization into CAD was different in 37/214 examinations, resulting in net recategorization of Δ13 (13/214) examinations and a difference in accuracy of Δ6.1%. On patient level, correlation of absolute and categorized values was substantial (0.567 and 0.570, p < 0.001). Categorization into CAD showed no correlation to image quality or CAC (p > 0.13). Conclusion: Differences between CT-FFR values increased in values below the cut-off, having little clinical impact. Categorization into CAD differed in several patients, but ultimately only had a moderate influence on diagnostic accuracy. This was independent of image quality or CAC.

A Novel Widespread MITE Element in the Repeat-Rich Genome of the Cardinium Endosymbiont of the Spider Oedothorax gibbosus.

Free-living bacteria have evolved multiple times to become host-restricted endosymbionts. The transition from a free-living to a host-restricted lifestyle comes with a number of different genomic changes, including a massive loss of genes. In host-restricted endosymbionts, gene inactivation and genome reduction are facilitated by mobile genetic elements, mainly insertion sequences (ISs). ISs are small autonomous mobile elements, and one of, if not the most, abundant transposable elements in bacteria. Proliferation of ISs is common in some facultative endosymbionts, and is likely driven by the transmission bottlenecks, which increase the level of genetic drift. In this study, we present a manually curated genome annotation for a Cardinium endosymbiont of the dwarf spider Oedothorax gibbosus. Cardinium species are host-restricted endosymbionts that, similarly to ColbachiaWolbachia spp., include strains capable of manipulating host reproduction. Through the focus on mobile elements, the annotation revealed a rampant spread of ISs, extending earlier observations in other Cardinium genomes. We found that a large proportion of IS elements are pseudogenized, with many displaying evidence of recent inactivation. Most notably, we describe the lineage-specific emergence and spread of a novel IS-derived Miniature Inverted repeat Transposable Element (MITE), likely being actively maintained by intact copies of its parental IS982-family element. This study highlights the relevance of manual curation of these repeat-rich endosymbiont genomes for the discovery of novel MITEs, as well as the possible role these understudied elements might play in genome streamlining. IMPORTANCE Cardinium bacteria, a widespread symbiont lineage found across insects and nematodes, have been linked to reproductive manipulation of their hosts. However, the study of Cardinium has been hampered by the lack of comprehensive genomic resources. The high content of mobile genetic elements, namely, insertion sequences (ISs), has long complicated the analyses and proper annotations of these genomes. In this study, we present a manually curated annotation of the Cardinium symbiont of the spider Oedothorax gibbosus. Most notably, we describe a novel IS-like element found exclusively in this strain. We show that this mobile element likely evolved from a defective copy of its parental IS and then spread throughout the genome, contributing to the pseudogenization of several other mobile elements. We propose this element is likely being maintained by the intact copies of its parental IS element and that other similar elements in the genome could potentially follow this route.

Defensive symbiosis against giant viruses in amoebae.

Protists are important regulators of microbial communities and key components in food webs with impact on nutrient cycling and ecosystem functioning. In turn, their activity is shaped by diverse intracellular parasites, including bacterial symbionts and viruses. Yet, bacteria-virus interactions within protists are poorly understood. Here, we studied the role of bacterial symbionts of free-living amoebae in the establishment of infections with nucleocytoplasmic large DNA viruses (Nucleocytoviricota). To investigate these interactions in a system that would also be relevant in nature, we first isolated and characterized a giant virus (Viennavirus, family Marseilleviridae) and a sympatric potential Acanthamoeba host infected with bacterial symbionts. Subsequently, coinfection experiments were carried out, using the fresh environmental isolates as well as additional amoeba laboratory strains. Employing fluorescence in situ hybridization and qPCR, we show that the bacterial symbiont, identified as Parachlamydia acanthamoebae, represses the replication of the sympatric Viennavirus in both recent environmental isolates as well as Acanthamoeba laboratory strains. In the presence of the symbiont, virions are still taken up, but viral factory maturation is inhibited, leading to survival of the amoeba host. The symbiont also suppressed the replication of the more complex Acanthamoeba polyphaga mimivirus and Tupanvirus deep ocean (Mimiviridae). Our work provides an example of an intracellular bacterial symbiont protecting a protist host against virus infections. The impact of virus-symbiont interactions on microbial population dynamics and eventually ecosystem processes requires further attention.

CT295 Is Chlamydia trachomatis' Phosphoglucomutase and a Type 3 Secretion Substrate.

The obligate intracellular bacteria Chlamydia trachomatis store glycogen in the lumen of the vacuoles in which they grow. Glycogen catabolism generates glucose-1-phosphate (Glc1P), while the bacteria can take up only glucose-6-phosphate (Glc6P). We tested whether the conversion of Glc1P into Glc6P could be catalyzed by a phosphoglucomutase (PGM) of host or bacterial origin. We found no evidence for the presence of the host PGM in the vacuole. Two C. trachomatis proteins, CT295 and CT815, are potential PGMs. By reconstituting the reaction using purified proteins, and by complementing PGM deficient fibroblasts, we demonstrated that only CT295 displayed robust PGM activity. Intriguingly, we showed that glycogen accumulation in the lumen of the vacuole of a subset of Chlamydia species (C. trachomatis, C. muridarum, C. suis) correlated with the presence, in CT295 orthologs, of a secretion signal recognized by the type three secretion (T3S) machinery of Shigella. C. caviae and C. pneumoniae do not accumulate glycogen, and their CT295 orthologs lack T3S signals. In conclusion, we established that the conversion of Glc1P into Glc6P was accomplished by a bacterial PGM, through the acquisition of a T3S signal in a "housekeeping" protein. Acquisition of this signal likely contributed to shaping glycogen metabolism within Chlamydiaceae.

Formation of Nano- and Micro-Scale Surface Features Induced by Long-Range Femtosecond Filament Laser Ablation.

In this work, we study the characteristics of femtosecond-filament-laser-matter interactions and laser-induced periodic surface structures (LIPSS) at a beam-propagation distance up to 55 m. The quantification of the periodicity of filament-induced self-organized surface structures was accomplished by SEM and AFM measurements combined with the use of discrete two-dimensional fast Fourier transform (2D-FFT) analysis, at different filament propagation distances. The results show that the size of the nano-scale surface features increased with ongoing laser filament processing and, further, periodic ripples started to form in the ablation-spot center after irradiation with five spatially overlapping pulses. The effective number of irradiating filament pulses per spot area affected the developing surface texture, with the period of the low spatial frequency LIPSS reducing notably at a high pulse number. The high regularity of the filament-induced ripples was verified by the demonstration of the angle-of-incidence-dependent diffraction of sunlight. This work underlines the potential of long-range femtosecond filamentation for energy delivery at remote distances, with suppressed diffraction and long depth focus, which can be used in biomimetic laser surface engineering and remote-sensing applications.