Publications

4547

ABSTRACT Ubiquitin‐proteasome is a conserved mechanism that regulates cellular responses and disease resistance in plants. However, the regulatory role of ubiquitin‐proteasome in the pathogenicity of “ Candidatus Liberibacter asiaticus” ( C Las), the causal agent of citrus huanglongbing (HLB), one of the most serious citrus diseases, remains poorly defined. In this study, we demonstrated that CLIBASIA_04250 (SDE4250) suppresses 26S proteasome activity in citrus, leading to a reduction in salicylic acid (SA) levels and downregulation of SA‐responsive genes. Further investigation revealed that SDE4250 specifically targets the citrus 26S proteasome non‐ATPase regulatory subunit 4 homologue (CsRPN10) and enhances its degradation via the 26S proteasome pathway. Moreover, overexpression of CsRPN10 inhibited C Las proliferation in citrus hairy roots, which was accompanied by a significant accumulation of SA content and upregulation of related defence genes. Collectively, our findings indicate that SDE4250 targets the citrus 26S proteasome to disrupt SA‐mediated citrus immunity.

Sweet potato witches’ broom disease caused by phytoplasmas threatens crop productivity in Fujian, China. Here, we report a complete phytoplasma genome consisting of a single circular chromosome (680,179 bp with a G + C content of 29.44%) via the next-generation sequencing of sweet potato witches’ broom-infected samples. A total of 730 genes were predicted, and two 16S rRNAs were revealed to be identical to sequences in ‘ Candidatus Phytoplasma australasiaticum’. Following the latest guidelines for the description of phytoplasma species, we have named the strain ‘ Candidatus Phytoplasma australasiaticum’ SPWBPT2024. The genome contained 33 transporters, 35 putative effectors (including SAP11 and SAP54 with multiple copies), and 2 potential mobile units. Analyses of published peanut and sweet potato phytoplasma genomes revealed similarities between their effectors and the effectors in this phytoplasma genome. A motif analysis was performed to clarify the difference between the two SAP54 copies in the obtained phytoplasma genome, as well as the trends in conserved motifs in other strains. In addition, metabolic pathway-related genes in three genomes were examined, revealing deficiencies in the glycolysis and pyrimidine synthesis pathways. A pangenome analysis revealed that the phytoplasma genome is an open genome. To gain further insights into whole-cell metabolic processes, we reconstructed draft genome-scale metabolic models of the genome. This study provides valuable genomic insights into this phytoplasma, characterizing conserved and strain-specific SAP effectors, as well as distinct metabolic deficiencies that may help advance research to combat phytoplasma infections, including accurate diagnostics, vector transmission control, and antimicrobial interventions.

Abstract Magnetotactic bacteria form a highly diverse group of microorganisms, yet early exploration of their diversity was largely centered on the Pseudomonadota. More recently, metagenomic studies have revealed that magnetotaxis, a form of chemotaxis guided by Earth's magnetic field, is widespread in other deep-branching phyla for which little to no ecological or biological information is available beyond that inferred from their genomes. For most of them, the morphology, ultrastructure and magnetosome chain characteristics responsible for the magnetic guidance remain unknown. While screening extreme environments for novel magnetotactic species, we observed magnetotactic Bdellovibrionota in the anoxic and ferruginous sediments of the Fontaine Goyon spring (France). We characterized their cell morphology and ultrastructure using magnetic enrichment, a single-cell sorting approach, and high-resolution electron microscopy. Cells display the morphology typical of the few predatory bacteria described in this phylum, and biomineralize, on average, five irregularly faceted, bullet-shaped magnetite magnetosomes along the concave side of the cell. Metagenomic analysis of approximately one hundred cells revealed a potentially predatory and heterotrophic lifestyle adapted to low-O2 conditions. It also suggests a flexible respiratory metabolism under varying redox conditions, using iron as an alternative terminal electron acceptor. Exploring the diversity of Bdellovibrionota in public databases, we found 21 metagenome-assembled-genomes containing magnetosome genes. None of them harbor the canonical mamK actin-like gene implicated in aligning magnetosomes in described magnetotactic models. Affiliated to an undescribed class, we propose a classification scheme for the magnetotactic Bdellovibrionota species representing the class Bdellonasia class nov., for which no species had been formally described.

ABSTRACT Acidophilic ammonia-oxidizing bacteria (AOB) have only recently been discovered. These organisms hold great promise for acidic wastewater treatment; however, their physiology remains poorly understood compared to that of neutrophilic AOB. Here, we investigated the physiology of the acidophilic AOB “Candidatus Nitrosacidococcus tergens ” sp. RJ19 across a broad pH range (2.5–7.0) using a specialized bioreactor system. We monitored nitrogen (N) transformations combined with microbial community composition and transcriptomic profiles, focusing on nitrogen metabolism and proton stress responses. Our results show that above pH 6.0, “ Ca . Na. tergens ” performs complete and stoichiometric conversion of ammonium to nitrite, coinciding with isotopic fractionation effects specific for ammonia oxidation and increased expression of key ammonia oxidation genes. The apparent absence of nirK and cycA did not impede ammonia oxidation, suggesting that these genes are non-essential in this context. Below pH 6.0, nitric oxide and nitrate accumulated, and nitrous oxide (N₂O) levels, although negligible compared to the other N-compounds, peaked near pH 4.0. Stable isotope analysis, including the site-specific 15 N-enrichment at the inner (α) and outer (β) nitrogen positions of the N 2 O molecule, indicated nitrifier-denitrification as the source of N₂O, supported by the highest norB expression at this pH. These findings provide new insights into the acid-tolerant physiology of “ Ca . Na. tergens ” and advance its potential application in engineered nitrogen removal systems under acidic conditions. IMPORTANCE The world is facing a climate crisis intensified by human-driven nutrient pollution. Ammonia and the bacteria that oxidize it are central both to the global nitrogen cycle and to wastewater treatment. The acidophilic ammonia oxidizer “ Candidatus Nitrosacidococcus tergens” was previously shown to oxidize ammonia under highly acidic conditions; however, a complete understanding of its metabolism is lacking. Our study now shows that “ Ca . Na. tergens” performs canonical ammonia oxidation across a broad pH range. At pH values below 6, however, a combination of chemical and biological processes leads to the production of nitrate, nitric oxide, and the greenhouse gas nitrous oxide. In addition, we show that these bacteria adapt to proton stress through mechanisms beyond transcriptional mechanisms. Our study highlights the robust metabolism of acidophilic ammonia oxidizers and expands our understanding of nitrification under acidic conditions.

A hydrogenotrophic methanogen, designated strain ACI-7 T , was isolated from an alkaline, saline wetland located in eastern Nebraska, USA. Cells were identified as non-motile rods (0.9–4.2 µm in length and 0.2–0.4 µm in diameter) that occurred singly, in chains, or as twisted chains and aggregates. Strain ACI-7 T utilized H 2 +CO 2 or formate as methanogenic substrates and required growth factors present in yeast extract for continuous cultivation. The strain grew at 20–45 °C (optimum, 40 °C), at pH 6.5–8.5 (optimum, pH 7.3) and with 0–2.5% NaCl (optimum, 0–1%). The genomic G+C content of ACI-7 T was 31.79 mol%. Phylogenetic analysis of the 16S rRNA gene sequence indicated strain ACI-7 T was affiliated with the genus Methanobacterium , most closely related to Methanobacterium oryzae FPi T (97.1% sequence similarity) and Methanobacterium veterum MK4 T (96.9% sequence similarity). Overall genome relatedness indices for strain ACI-7 T compared to other Methanobacterium species ranged from 68.45 to 78.17% for average nucleotide identity and 18.9 to 25.7% for digital DNA–DNA hybridization. Morphological, physiological and genomic characteristics indicate that strain ACI-7 T represents a novel species, for which the name Methanobacterium nebraskense sp. nov. is proposed. The type strain is ACI-7 T (=DSM 118696 T =ATCC TSD-487 T ).

ABSTRACT The oil refinery (OR) consortium is a model methanogenic enrichment culture used to study anaerobic benzene degradation. Over half of the culture’s bacterial community consists of two closely related Desulfobacterota strains, designated ORM2a and ORM2b, whose mechanisms of benzene activation are unknown. Three proteomics data sets were integrated and analyzed using high-quality OR metagenomes and metagenome-assembled genomes (MAGs), including a complete circularized ORM2a MAG, to identify active metabolic pathways and proteins expressed during methanogenic benzene degradation. Among the proteins identified were Bam-like subunits of an ATP-independent benzoyl-CoA degradation pathway, as well as downstream β-oxidation proteins yielding acetate. The most abundant proteins identified mapped to two ORM2a gene clusters of unknown function. Homologous and syntenic gene clusters were identified in the MAGs of ORM2b and a sulfate-reducing Pelotomaculum that also degrades benzene, as well as in nine contigs assembled from hydrothermal vent metagenomes. Extensive homology and structural predictions suggest that the first cluster—termed the “Magic” gene cluster—encodes for enzymes catalyzing the chemically challenging activation of benzene and subsequent transformation steps yielding benzoyl-CoA. The second (“Nanopod”) gene cluster encodes a transmembrane complex that may facilitate benzene transport across the cell membrane. Phylogenomic analyses place ORM2a and ORM2b within a novel genus of strict anaerobes specialized for benzene degradation, which we propose naming “ Candidatus Anaerobenzenivorax.” IMPORTANCE Benzene is a widespread, persistent, and toxic pollutant that can accumulate in anoxic environments such as groundwater and sediments. Benzene can be metabolized in the absence of oxygen; however, despite decades of research, the biochemical mechanisms for benzene activation under anaerobic conditions remain unproven. This study provides strong genetic and proteomic evidence for a new suite of enzymes that initiate anaerobic benzene activation. These findings lay a foundation for future biochemical studies and expand our understanding of how microbes carry out difficult chemical reactions in the absence of oxygen.

Citrus huanglongbing (HLB), caused by ‘Candidatus Liberibacter asiaticus’ (CLas), is a destructive disease threatening global citrus industry. Although citrus cultivars differ in HLB sensitivity, how infection alters endophytic bacterial communities in cultivars with contrasting susceptibility remains unclear. Here, we compared endophytic microbiome shifts in leaf and root tissues of HLB-susceptible Shatangju mandarin (Citrus reticulata cv. Shatangju) and HLB-tolerant Shatian pomelo (C. maxima cv. Shatian) at 10 months postgrafting with CLas-infected buds. Infected Shatangju mandarin showed severe symptoms and higher CLas levels in leaves than in roots, while infected Shatian pomelo remained asymptomatic with higher CLas in roots than in leaves. High-throughput 16S rRNA gene sequencing revealed that CLas infection restructured the endophytic microbial community in a tissue-specific manner. CLas-infected Shatian pomelo leaves displayed increased microbial diversity and network complexity, while CLas-infected Shatangju mandarin roots harbored more disease-suppressive taxa like Streptomyces. Across both cultivars, network complexity correlated inversely with CLas abundance. Key genera (e.g., Pseudomonas, Streptomyces, and Prevotella) correlated positively with CLas, suggesting roles in mitigating pathogen effects. Predicted functional profiling indicated activated pathways (amino acid metabolism, terpenoid/polyketide biosynthesis, and xenobiotic biodegradation) potentially supporting host defense in infected tissues. The HLB tolerance of Shatian pomelo appeared linked to tissue-specific bacterial restructuring, particularly recruitment of antibiotic-producing bacteria and enhanced metabolic resilience. These findings elucidated cultivar-specific microbial strategies against CLas, offering insights for microbiome-mediated HLB management.

Background: The Asian Haemaphysalis longicornis is capable of transmitting a wide range of zoonotic pathogens. This study aimed to investigate the prevalence and characteristics of major pathogens in questing H. longicornis ticks collected from two geographically and ecologically distinct regions in China. Methods: A total of 1004 questing H. longicornis ticks were collected from Liaoning Province (Northeast China; n = 882) and Anhui Province (Central and Eastern China; n = 122) and pooled into 670 mixed samples. Tick species were identified by mitochondrial cytochrome oxidase I gene sequencing. Nucleic acid samples from all pools were first screened for Dabie bandavirus using RT-PCR and for bacteria using universal 16S rRNA primers. Bacteria-positive samples were further tested using polygenic seminested PCR for Rickettsia spp. and Coxiella -like endosymbionts (CLEs). Phylogenetic analyses, including polygene sequence analysis, were performed to ensure the accuracy of species identification and to explore genetic relationships. Results: No Dabie bandavirus was detected in any samples. Among the 670 tick pools, 14 were positive for spotted fever group rickettsiae (SFGR), with an overall minimum infection rate (MIR) of 1.4% (14/1004). Notably, the detected SFGR species showed clear geographic segregation: Candidatus Rickettsia jingxinensis was detected only in Liaoning Province (MIR of 1.4%), while Rickettsia heilongjiangensis was found exclusively in Anhui Province (MIR of 1.6%). Additionally, CLEs (overall MIR of 2.0%) were identified in 20 tick pools, with a significantly higher prevalence in Anhui Province (10.7%) than in Liaoning Province (0.8%; p < 0.001). Conclusions: This surveillance revealed a distinct geographical distribution pattern of SFGR species in H. longicornis populations in China and confirmed the presence of Ca . R. jingxinensis in the northeast and R. heilongjiangensis in the central and eastern regions. These findings, supported by polygenic evidence, provide essential epidemiological data for understanding regional tick-borne disease risks and highlight the need for continued region-specific surveillance.