Publications

4619

Citrus huanglongbing (HLB), associated with the bacterium ‘Candidatus Liberibacter asiaticus’ and spread by the Asian citrus psyllid (Diaphorina citri; ACP), poses a significant threat to California’s citrus industry. First identified in Los Angeles in 2012, HLB has since spread through residential areas across Southern California. A risk-based survey (RBS) model has been developed to improve HLB surveillance and intervention. Within this framework, model components change as HLB dynamics shift, requiring regular updates to maintain data accuracy and model reliability. Disease spread is influenced by natural factors, such as ACP establishment and confirmed HLB locations, as well as human-mediated factors like global mobility (travel introduction from HLB-infected countries), transportation of citrus materials, nurseries, packinghouses, farmers’ markets, and proximity to private or otherwise inaccessible lands. Human-mediated risk factors account for approximately 26.3% (18.4 to 38.4%) of HLB incidence across different years, and natural causes predominantly explain the remaining 73.7% (61.6 to 81.7%). Notably, global mobility was crucial for early HLB detection in new areas, whereas ACP density strongly correlated with disease spread once established. A retrospective analysis from 2015 to 2022 evaluated the RBS model’s performance, showing a predictive power of 88 to 97%, which confirms its validity for developing targeted interventions and early detection strategies in California.

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.

ABSTRACT Citrus huanglongbing (HLB), caused by the phloem‐limited bacterium ‘ Candidatus Liberibacter asiaticus’ ( C Las), is one of the most destructive diseases in global citrus production. Here, we report the functional characterization of a core C Las effector, CLIBASIA_00525 named SDE525, that manipulates host immunity by targeting the citrus nascent polypeptide‐associated complex alpha subunit (CsNACα). We confirmed that SDE525 is a secreted protein that localizes to the nucleus, cytoplasm, and plasma membrane. Notably, the effect of SDE525 exhibits host‐specific context dependence: while its transient expression in the non‐host Nicotiana benthamiana unexpectedly triggered immune responses, in its native host, sweet orange ( Citrus sinensis ), SDE525 interacted with CsNACα to modulate defence, as demonstrated by multiple independent assays. Integrated transcriptomic and metabolomic analyses revealed that this interaction reprograms the host's hormonal landscape, suppressing the jasmonic acid (JA) signalling pathway while simultaneously activating the salicylic acid (SA) pathway. Importantly, under transient overexpression conditions in citrus, this immune reprogramming enhanced resistance against the bacterial canker pathogen Xanthomonas citri subsp. citri —a phenomenon distinct from the susceptibility observed during authentic C Las infection. Our findings uncover a sophisticated mechanism by which a C Las effector rewires host defence signalling, providing fundamental insights into its pathogenesis with potential long‐term implications for future HLB management.

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 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.

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 ).

‘Candidatus Phytoplasma rubi’ (elm yellows group, 16SrV-E phytoplasma subgroup) is the causal agent of rubus stunt disease, a disorder affecting economically important plants—raspberries and blackberries. Although this phytoplasma has been extensively studied in cultivated Rubus crops, its occurrence and molecular identity in wild Rubus species populations in North-Eastern Europe remain poorly documented. In this study, phytoplasmas associated with symptomatic wild Rubus idaeus L. and Rubus nessensis Hall plants were investigated in natural forest ecosystems of the Curonian Spit, Lithuania. A total of 65 symptomatic plants were surveyed, and phytoplasma infection was detected in 30 samples by nested PCR targeting the 16S rRNA gene. Positive samples were characterized using a multilocus molecular approach based on sequence analysis of the additional cpn60 and secA genes. All strains showed high nucleotide sequence similarity across the analysed loci and consistently clustered with reference strains of ‘Candidatus Phytoplasma rubi’. Virtual RFLP profiles derived from the 16S rRNA and cpn60 genes were nearly identical to those of established 16SrV-E phytoplasma subgroup reference strains and clearly distinct from other 16SrV phytoplasma subgroups. These results provide not only the first detailed multilocus molecular characterization of ‘Candidatus Phytoplasma rubi’-related strains infecting wild Rubus species in Lithuania but also represent the first report of this phytoplasma in naturally occurring R. idaeus and R. nessensis plants in the country, thereby extending the known geographical occurrence of this pathogen and documenting its presence in wild Rubus hosts from unmanaged forest habitats in the Eastern Baltic region of Northern Europe.

Ocean warming, disease, and pollution have contributed to global declines in coral abundances and diversity. In the Caribbean, corals previously dominated reefs, providing an architectural framework for diverse ecological habitats, but have significantly declined due to infectious disease and anthropogenic climate change. Key species like the coral Acropora cervicornis are critically endangered, prompting researchers to focus on scientific endeavors to identify factors influencing coral disease resistance and resilience. We previously showed that disease susceptibility, growth rates, and bleaching risk were all associated with the abundance of a single bacterial parasite, Candidatus Aquirickettsia rohweri which proliferates in vivo under nutrient enrichment. Yet how nutrients influence parasite physiology in vivo remains unknown. Here, we analyzed parasite gene expression from a disease-susceptible A. cervicornis genotype exposed to ambient or nutrient enrichment conditions. Electron microscopy showed that Ca. A. rohweri was abundant in coral tissue and densely packed in mucocytes prior to nutrient enrichment. Under ambient conditions, the parasite upregulated genes involved in translation, protein maintenance, and cell envelope integrity, consistent with a conserve-and-maintain strategy. Nutrient enrichment induced expression of genes associated with central metabolism, nutrient import, stress response, host interaction, and two-component systems. Together, these results indicate that nutrient enrichment activates a growth-and-exploitation strategy, likely exacerbating parasitic pressure on A. cervicornis .