Publications

4373

Abstract Aim Candidatus Methanoperedens-related archaea have recently been identified as anaerobic methane oxidizers in paddy soils. Fertilization practices, including the application of inorganic and organic fertilizers (e.g. chicken manure), may significantly influence their community dynamics and the associated methane cycling processes. However, the comparative effects of inorganic and chicken manure fertilization on these archaeal community in paddy fields remain unclear. This study aimed to examine the diversity, community composition, and abundance of Methanoperedens-related archaea at three representative soil layers of 0–10, 20–30, and 40–50 cm under three fertilization treatments (no fertilizer, inorganic fertilizer, chicken manure fertilizer). Methods and results High-throughput sequencing revealed significant differences in community composition among treatments, while overall diversity showed minimal changes. Quantitative Polymerase Chain Reaction indicated that archaeal abundance under inorganic (2.3 × 106 copies g−1) and chicken manure fertilization (2.2 × 106 copies g−1) treatments was significantly greater than that under no fertilization (1.8 × 106 copies g−1) in upper 30 cm soils, with no significant difference at 40–50 cm depth. Inorganic fertilization more strongly promoted archaeal abundance, whereas chicken manure had a greater effect on community structure. Soil ammonium, nitrate, and organic carbon contents were significantly correlated with archaeal community patterns. Conclusion Both inorganic and organic fertilization can substantially influence the community structure and abundance of Methanoperedens-related archaea in paddy soils.

AbstractBACKGROUNDAlthough it is known that Candidatus Liberibacter asiaticus (CLas), the agent of citrus Huanglongbing, circulates and multiplies within the insect vector Diaphorina citri, the specific factors enabling CLas transmission remain unclear. Previous studies have shown that ATPSyn‐β facilitates phytoplasma movement in vector insects, and functions as a transport protein in D. citri. In this study, the role of ATPSyn‐β was expected to be unveiled in CLas transmission in D. citri.RESULTSThe interaction between ATPSyn‐β of D. citri and the outer membrane protein A (OmpA) of CLas was identified using glutathione S‐transferase pull‐down and yeast two‐hybrid assays. No interactions existed between OmpA and vitellogenin (Vg). Bioinformatics analysis revealed that ATPSyn‐β contained two conserved domains, ATP‐synt_ab_N and ATP‐synt_ab. Notably, ATP‐synt_ab_N interacted with both OmpA and Vg, indicating a competitive relationship between Vg and OmpA for binding to ATPSyn‐β. After silencing ATPSyn‐β expression using RNAi, the feeding behavior of D. citri decreased, resulting in a lower acquisition of CLas. Subsequently, this inhibited CLas movement within D. citri and reduced the inoculation of CLas to healthy citrus plants.CONCLUSIONATPSyn‐β was involved in the acquisition, movement in vivo, and inoculation of CLas by interacting with OmpA, thus offering a novel target to prevent the spread of citrus Huanglongbing. © 2025 Society of Chemical Industry.

Ticks spread pathogens that affect both human and animal health and often cause significant economic losses to the livestock industry. However, there is limited data on the distribution of ticks and tick-borne pathogens, particularly those affecting cattle in Cameroon. In this study, ticks were collected from cattle in Buea, the capital of the South West Region of Cameroon, to determine their diversity and identify tick-borne pathogens through PCR and sequencing. The tick samples were screened for pathogens using assays that target the rOmpA gene (ompA) of Rickettsia, the ssrRNA gene of Babesia and Theileria and the Ehrlichia genus 16SrRNA gene. A total of 458 ticks were collected with Rhipicephalus annulatus (65.6%) as the predominant species. From 68 tick pools screened, 26 (38.2%) were positive for pathogen/ symbiont DNA. The pathogen identified was Rickettsia africae (22.1%). This study reports the first molecular detection of the symbiont Candidatus Midichloria mitochondrii (22.1%) in the sampled tick species. It was observed that male ticks were significantly more likely to test positive for R. africae (OR = 208, 95% CI = 28.6 - 4553, p<0.001). Animal handlers may face the risk of these zoonotic infections and as such, there is a need to employ protective measures to prevent or reduce pathogen spread.

Despite their large environmental impact and multiple independent emergences, the processes leading to the evolution of anaerobic methanotrophic archaea (ANME) remain unclear. This work uses comparative metagenomics of a recently evolved but understudied ANME group, “ Candidatus Methanovorans” (ANME-3), to identify evolutionary processes and innovations at work in ANME, which may be obscured in earlier evolved lineages. We identified horizontal transfer of hdrA homologs and convergent evolution in carbon and energy metabolic genes as potential early steps in Methanovorans evolution. We also identified the erosion of genes required for methylotrophic methanogenesis along with horizontal acquisition of multiheme cytochromes and other loci uniquely associated with ANME. The assembly and comparative analysis of multiple Methanovorans genomes offers important functional context for understanding the niche-defining metabolic differences between methane-oxidizing ANME and their methanogen relatives. Furthermore, this work illustrates the multiple evolutionary modes at play in the transition to a globally important metabolic niche.

Abstract The CRISPR–Cas (clustered regularly interspaced short palindromic repeats and CRISPR-associated protein) systems are adaptive immune mechanisms that play critical roles in defending archaea and bacteria against invading entities. These systems can be divided into two classes, with class 2 comprising three types (II, V, and VI). Because of their ability to cleave double-stranded DNA, many class 2 CRISPR–Cas proteins have been harnessed as genome editing tools. Unlike the well-studied type II Cas9 proteins, the structural studies of the type V-B Cas12b proteins are limited, hindering their engineering and broader application. Here, we report four complex structures of ChCas12b, which reveal many unique structural features. The folding of the single guide RNA (sgRNA) of ChCas12b is distinct from that of AacCas12b and BthCas12b. Notably, many of these unique features are involved in ChCas12b–sgRNA interaction, suggesting that they are co-evolved. While ChCas12b shares a conserved two-cation-assisted catalytic mechanism with its homologs, it recognizes a longer guide:target heteroduplex, potentially offering higher fidelity in DNA editing. Altogether, our studies suggested that Cas12b family proteins exhibit significant diversity in their folding, sgRNA and target DNA binding. In the future, it is worth characterizing more representative proteins to identify CRISPR–Cas proteins with higher gene editing ability and fidelity.

The genus Moraxella (Moraxellaceae, Pseudomonadales) comprises a diverse group of bacteria inhabiting human and animal mucosa, as well as environmental niches such as water, soil and food. While some species are clinically significant pathogens, others play ecological or biotechnological roles. Despite previous taxonomic revisions, Moraxella remains polyphyletic, necessitating a refined classification. In this study, we conducted comprehensive taxogenomic analyses integrating core protein phylogeny, average amino acid identity and the percentage of conserved proteins, along with 16S rRNA similarities and inferred phylogeny. Our results revealed three distinct phylogenetic clusters within Moraxella. The core group, comprising Moraxella lacunata NBRC 102154T and closely related species, exhibited strong genomic cohesion. A second cluster, consisting of Moraxella boevrei DSM 14165T, Moraxella osloensis CCUG 350T and Moraxella atlantae NBRC 14588T, showed greater genetic affinity to Faucicola mancuniensis GVCNT2T, supporting their reassignment to the genus Faucicola. The third group, represented by Moraxella lincolnii CCUG 9405T, was phylogenetically distinct, occupying a basal position relative to Psychrobacter, indicating the need for its classification within a novel genus within the family Moraxellaceae, for which we propose the name Lwoffella lincolnii gen. nov., comb. nov. Phenotypic data compiled from the original published descriptions of the respective species, including fatty acid composition and enzymatic profiles, further corroborated these genomic findings. This study refines Moraxella taxonomy by clarifying genus boundaries and evolutionary relationships, with implications for ecology and clinical microbiology.

The actinobacterial family Microbacteriaceae comprises a diverse group of Gram-positive bacteria with high G+C content and complex taxonomic challenges. Traditional polyphasic approaches, based on 16S rRNA phylogeny and phenotypic traits, have in some instances resulted in polyphyletic taxonomic groupings, necessitating genome-wide methodologies to better resolve evolutionary relationships. This study employs a taxogenomic approach – incorporating 16S rRNA gene sequencing, core protein phylogeny, average amino acid identity and percentage of conserved proteins – to reassess the genera Leifsonia, Salinibacterium, Homoserinimonas, Glaciibacter, Antiquaquibacter and Diaminobutyricibacter. The findings reveal significant polyphyly in Leifsonia, Glaciibacter and Salinibacterium, indicating ecological convergence rather than shared ancestry. Proposed taxonomic revisions include the reclassification of Salinibacterium soli and Salinibacterium metalliresistens into Antiquaquibacter, Salinibacterium hongtaonis and Salinibacterium sedimenticola into Homoserinimonas, and Leifsonia psychrotolerans and Leifsonia kafniensis into Glaciibacter. Diaminobutyricibacter tongyongensis should be incorporated into Leifsonia. Additionally, Leifsonia rubra is reassigned to Salinibacterium. Two new genera were also proposed to encompass “Leifsonia flava” and Glaciibacter flavus, named Orlajensenia gen. nov. and another to encompass Leifsonia bigeumensis, named Leifsonella gen. nov. These genome-based insights provide a refined framework for the taxonomy of Microbacteriaceae, enhancing our understanding of their evolutionary and ecological roles.

ABSTRACT Ca . Babelota is a phylum of strictly intracellular bacteria whose representatives are commonly detected in various environments through metagenomics, though their presence, ecology, and biology have never been addressed so far. As a group of strict intracellular, we hypothesize that their presence, occurrence, and abundance heavily depend on their hosts, which are known as heterotrophic protists, based on few described isolates. Here, we conducted a sampling campaign allowing to characterize protists and associated bacterial communities, using high-throughput sequencing. In parallel, a systematic enrichment of protists from samples was performed to attempt characterization and isolation of new Ca . Babelota within native hosts. We found that Ca . Babelota are among the most widespread phylum among the rare ones. Protist enrichments are allowed in certain cases to enrich as well for Ca . Babelota, which could be visualized in vivo infecting protist cells. Though cosmopolitan, Ca . Babelota diversity was highly site-specific. Cooccurrence analyses allowed to retrieve well-known as well as new putative associations involving numerous protists of various trophic regimes. The combination of approaches developed in this study enhances our understanding of Ca . Babelota ecology and biology, while paving the way for future isolation of new members of this elusive phylum, which could have huge impact on protists—and ecosystems—functioning. IMPORTANCE Our understanding of microbial diversity surrounding us and colonizing the environment has been dramatically impacted by the advent of DNA-based analyses. Such progress helped shine a new light on numerous lineages of yet-to-be-characterized microbes, whose ecology and biology are basically unknown. Among those uncharacterized clades is the Candidatus Babelota, a bacterial phylum for which parasitism seems to be an ancestral trait. All known Ca. Babelota thrive by infecting phagotrophic protist hosts, thereby impacting this basal link of the trophic chain. The Ca. Babelota constitutes a model that stands out, as phylum-wide conserved parasitism has only been described in one previous occurrence for Bacteria, with the Chlamydiota. Thus, exploring the intricate interplay between Ca. Babelota and their protist hosts will advance our knowledge of bacterial diversity, their ecology, and global impact on ecosystem functioning.

Abstract Methanol, the simplest alcohol, has long been recognized as a key energy and carbon source for soil and plant-associated bacteria and fungi, and is increasingly recognized as an important substrate for marine bacteria. Lanthanide-dependent methanol dehydrogenases (encoded by the gene xoxF) are now recognized as the key catalysts for methylotrophy in many environments, yet the identity of the most transcriptionally active methylotrophs in open ocean waters (“Clade X”) has remained elusive. Here we show that “Clade X” methylotrophs belong to the deep-branching alphaproteobacterial order TMED127, which we propose be renamed ‘Methylaequorales’; ‘methyl’ for ‘methylotrophic metabolism’ and ‘aequor’ for ‘ocean surface’, as these bacteria are most transcriptionally active near the sea surface. TMED127/Methylaequorales are present in surface waters of tropical and subtropical oceans throughout the global ocean. They have small, streamlined genomes (∼1.5 Mb), and appear to be obligate methylotrophs that use the serine cycle for carbon assimilation. They display a diel pattern of xoxF5 and glucose dehydrogenase (gdh) transcription, peaking in the late afternoon, in oligotrophic surface water of the Sargasso Sea. Several other highly transcribed genes of unknown function had no homologs outside of TMED127/Methylaequorales genomes. Our findings illuminate an overlooked marine methylotrophic bacterium and predict a diel cycle of methanol production in surface seawater by an unknown pathway. Importance Methanol metabolism is increasingly recognized as an important process in the marine carbon cycle, yet the identity and metabolism of the microorganisms mediating methylotrophy in the open ocean have remained unknown. This study reveals that bacteria in the TMED127 order of Alphaproteobacteria, proposed new name of “Methylaequorales”, abundantly transcribe the key gene for lanthanide-dependent methylotrophy in oligotrophic surface waters of the world’s oceans. TMED127/Methylaequorales likely require methanol as a carbon and energy source and display a diel pattern of transcription of key genes for methylotrophy that peaks in the late afternoon. These findings motivate future studies on the mechanisms of methanol production in surface seawater.