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The Caenorhabditis elegans natural microbiota isolates Pseudomonas lurida MYb11 and Pseudomonas fluorescens MYb115 protect the host against pathogens through distinct mechanisms. While P. lurida produces an antimicrobial compound and directly inhibits pathogen growth, P. fluorescens MYb115 protects the host without affecting pathogen growth. It is unknown how these two protective microbes affect host biological processes. We used a proteomics approach to elucidate the C. elegans response to MYb11 and MYb115. We found that both Pseudomonas isolates increase vitellogenin protein production in young adults, which confirms previous findings on the effect of microbiota on C. elegans reproductive timing. Moreover, the C. elegans responses to MYb11 and MYb115 exhibit common signatures with the response to other vitamin B 12 -producing bacteria, emphasizing the importance of vitamin B 12 in C. elegans -microbe metabolic interactions. We further analyzed signatures in the C. elegans response specific to MYb11 or MYb115. We provide evidence for distinct modifications in lipid metabolism by both symbiotic microbes. We could identify the activation of host-pathogen defense responses as an MYb11-specific proteome signature and provide evidence that the intermediate filament protein IFB-2 is required for MYb115-mediated protection. These results indicate that MYb11 not only produces an antimicrobial compound but also activates host antimicrobial defenses, which together might increase resistance to infection. In contrast, MYb115 affects host processes such as lipid metabolism and cytoskeleton dynamics, which might increase host tolerance to infection. Overall, this study pinpoints proteins of interest that form the basis for additional exploration into the mechanisms underlying C. elegans microbiota-mediated protection from pathogen infection and other microbiota-mediated traits.IMPORTANCESymbiotic bacteria can defend their host against pathogen infection. While some protective symbionts directly interact with pathogenic bacteria, other protective symbionts elicit a response in the host that improves its own pathogen defenses. To better understand how a host responds to protective symbionts, we examined which host proteins are affected by two protective Pseudomonas bacteria in the model nematode Caenorhabditis elegans . We found that the C. elegans response to its protective symbionts is manifold, which was reflected in changes in proteins that are involved in metabolism, the immune system, and cell structure. This study provides a foundation for exploring the contribution of the host response to symbiont-mediated protection from pathogen infection., Competing Interests: The authors declare no conflict of interest.

Spectral similarity networks, also known as molecular networks, are crucial in non-targeted metabolomics to aid identification of unknowns aiming to establish a potential structural relation between different metabolite features. However, too extensive differences in compound structures can lead to separate clusters, complicating annotation. To address this challenge, we developed an automated Annotation Propagation through multiple EXperimental Networks (APEX) workflow, which integrates spectral similarity networks with mass difference networks and homologous series. The incorporation of multiple network tools improved annotation quality, as evidenced by high matching rates of the molecular formula derived by SIRIUS. The selection of manual annotations as the Seed Nodes Set (SNS) significantly influenced APEX annotations, with a higher number of seed nodes enhancing the annotation process. We applied APEX to different Caenorhabditis elegans metabolomics data sets as a proof-of-principle for the effective and comprehensive annotation of glycerophospho N -acyl ethanolamides (GPNAEs) and their glyco-variants. Furthermore, we demonstrated the workflow's applicability to two other, well-described metabolite classes in C. elegans , specifically ascarosides and modular glycosides (MOGLs), using an additional publicly available data set. In summary, the APEX workflow presents a powerful approach for metabolite annotation and identification by leveraging multiple experimental networks. By refining the SNS selection and integrating diverse networks, APEX holds promise for comprehensive annotation in metabolomics research, enabling a deeper understanding of the metabolome.

The Caenorhabditis elegans natural microbiota was described only recently. Thus, our understanding of its effects on nematode physiology is still in its infancy. We previously showed that the C. elegans natural microbiota isolates Pseudomonas lurida MYb11 and P. fluorescens MYb115 protect the worm against pathogens such as Bacillus thuringiensis (Bt). However, the overall effects of the protective microbiota on worm physiology are incompletely understood. Here, we investigated how MYb11 and MYb115 affect C. elegans lifespan, fertility, and intestinal colonization. We further studied the capacity of MYb11 and MYb115 to protect the worm against purified Bt toxins. We show that while MYb115 and MYb11 affect reproductive timing and increase early reproduction only MYb11 reduces worm lifespan. Moreover, MYb11 aggravates killing upon toxin exposure. We conclude that MYb11 has a pathogenic potential in some contexts. This work thus highlights that certain C. elegans microbiota members can be beneficial and costly to the host in a context-dependent manner, blurring the line between good and bad., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Kissoyan, Peters, Giez, Michels, Pees, Hamerich, Schulenburg and Dierking.)

Background: The interplay between hosts and their associated microbiome is now recognized as a fundamental basis of the ecology, evolution, and development of both players. These interdependencies inspired a new view of multicellular organisms as "metaorganisms." The goal of the Collaborative Research Center "Origin and Function of Metaorganisms" is to understand why and how microbial communities form long-term associations with hosts from diverse taxonomic groups, ranging from sponges to humans in addition to plants., Methods: In order to optimize the choice of analysis procedures, which may differ according to the host organism and question at hand, we systematically compared the two main technical approaches for profiling microbial communities, 16S rRNA gene amplicon and metagenomic shotgun sequencing across our panel of ten host taxa. This includes two commonly used 16S rRNA gene regions and two amplification procedures, thus totaling five different microbial profiles per host sample., Conclusion: While 16S rRNA gene-based analyses are subject to much skepticism, we demonstrate that many aspects of bacterial community characterization are consistent across methods. The resulting insight facilitates the selection of appropriate methods across a wide range of host taxa. Overall, we recommend single- over multi-step amplification procedures, and although exceptions and trade-offs exist, the V3 V4 over the V1 V2 region of the 16S rRNA gene. Finally, by contrasting taxonomic and functional profiles and performing phylogenetic analysis, we provide important and novel insight into broad evolutionary patterns among metaorganisms, whereby the transition of animals from an aquatic to a terrestrial habitat marks a major event in the evolution of host-associated microbial composition.

Caenorhabditis elegans is associated in nature with a species-rich, distinct microbiota, which was characterized only recently [1]. Thus, our understanding of the relevance of the microbiota for nematode fitness is still at its infancy. One major benefit that the intestinal microbiota can provide to its host is protection against pathogen infection [2]. However, the specific strains conferring the protection and the underlying mechanisms of microbiota-mediated protection are often unclear [3]. Here, we identify natural C. elegans microbiota isolates that increase C. elegans resistance to pathogen infection. We show that isolates of the Pseudomonas fluorescens subgroup provide paramount protection from infection with the natural pathogen Bacillus thuringiensis through distinct mechanisms. We found that the P. lurida isolates MYb11 and MYb12 (members of the P. fluorescens subgroup) protect C. elegans against B. thuringiensis infection by directly inhibiting growth of the pathogen both in vitro and in vivo. Using genomic and biochemical analyses, we further demonstrate that MYb11 and MYb12 produce massetolide E, a cyclic lipopeptide biosurfactant of the viscosin group [4, 5], which is active against pathogenic B. thuringiensis. In contrast to MYb11 and MYb12, P. fluorescens MYb115-mediated protection involves increased resistance without inhibition of pathogen growth and most likely depends on indirect, host-mediated mechanisms. This work provides new insight into the functional significance of the C. elegans natural microbiota and expands our knowledge of bacteria-derived compounds that can influence pathogen colonization in the intestine of an animal., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Introduction: Acinetobacter baumannii has become one of the most feared organisms in hospital-acquired infections during the past decades. Their multi-drug resistant profiles have rendered many broad-spectrum antibiotics ineffective. The purpose of this retrospective study is to describe and compare molecular characteristics of A. baumannii isolated from patients at a tertiary care center in Lebanon from two outbreaks, the first in 2007-2008 as part of a case-controlled study involving Acinetobacter baumannii cases admitted to the ICU and the second in 2013., Methodology: A total of 148 A. baumannii clinical isolates were collected from various clinical specimens during 2007-2008 and 2013. All A. baumannii isolates were subjected to PCR amplification of blaOXA-23-like and blaOXA-51-like genes of carbapenem resistance. Random amplification of polymorphic DNA (RAPD) was also performed to assess their genomic relatedness., Results: There was an increase in the prevalence of blaOXA-23-like and blaOXA-51-like between the two time periods; however, only with 22% genomic relatedness between 2007-2008 and 2013 isolates. Taking 80% as margin of compatibility, 31 distinct clusters containing 2 to 11 strains were observed in both time periods., Conclusion: The presence of numerous clusters accompanied by a predominant increase in the prevalence of blaOXA-23-like gene between 2007 and 2013 suggests a horizontal transmission of the gene within various strains of the species, constituting a primary factor in the continued increase of carbapenem resistance over the years. As such, infection control measures ought to be taken with the highest priority and compliance among all involved healthcare workers is of utmost importance., Competing Interests: No Conflict of Interest is declared, (Copyright (c) 2018 Souha S Kanj, Ralph Tayyar, Marwa Shehab, Bassam El-Hafi, Sari S Rasheed, Kohar Annie Berge Kissoyan, Zeina A Kanafani, Rima Hanna Wakim, Nada Kara Zahreddine, George F Araj, Ghassan Dbaibo, Ghassan M Matar.)

Background: The objective of this study was to determine whether patients infected with extended-spectrum beta-lactamase (ESBL)-producing organisms are colonized at multiple body sites., Methods: This was a prospective cohort study at a tertiary care center in Beirut, Lebanon. Hospitalized patients with infections caused by ESBL-producing organisms were included. Cultures were obtained from the primary site of infection as well as from other sites (skin, nasopharynx, urine, rectum). Molecular analysis was performed on isolates to determine clonal relatedness., Results: One hundred patients were included in the study. Only 22 patients had positive cultures from sites other than the primary site of infection. The most common ESBL gene was CTX-M-15 followed by TEM-1. In 11 of 22 patients, isolates collected from the same patient were 100% genetically related, while in the remaining patients, genomic relatedness ranged from 42.9% to 97.1%., Conclusions: Colonization at sites other than the primary site of infection was not common among our patient population infected with ESBL-producing organisms. The dynamics of transmission of these bacterial strains should be studied in further prospective studies to determine the value of routine active surveillance and the need for expanded precautions in infected and colonized patients.