Publications

4485

Abstract Members of the phylum Deferribacterota inhabit diverse environments, but their symbiosis with protists has never been reported. We discovered an ectosymbiotic clade of Deferribacterota specifically associated with spirotrichonymphid protists in the guts of the termites Reticulitermes speratus and Hodotermopsis sjostedti and trichonymphid protists in the gut of the wood-feeding cockroach Cryptocercus punctulatus. The ectosymbiotic Deferribacterota were spiral shaped and attached to 16%–91% of the host protist cells. These formed a monophyletic cluster within an uncultured insect gut-associated family-level clade, which is sister to the vertebrate gut-associated family Mucispirillaceae. The complete genome of an ectosymbiotic Deferribacterota was obtained from a Trichonympha acuta cell in a C. punctulatus gut and analyzed together with a single-cell amplified genome of another ectosymbiotic Deferribacterota associated with Holomastigotes sp. in the gut of R. speratus. Genome analyses suggest that these Deferribacterota ferment monosaccharides and conduct fumarate and oxidative respiration with H2 as an electron donor. They thus possibly contribute to the removal of hydrogen and oxygen to protect the fermentative activity of the protist hosts. The ectosymbionts possess reduced signal transduction gene repertoires, implying that the association has provided a relatively stable environment for these bacteria. The ectosymbionts likely possess flagella with an unusually expanded number of flagellin variants up to 40, which may reflect an adaptation to their ectosymbiotic lifestyle. We propose a novel genus, Termitispirillum, for these ectosymbionts and a novel family, Termitispirillaceae, for the insect-gut clade, under SeqCode. Our findings provide new insights into the ecology and evolution of Deferribacterota.

An anaerobic, salt-tolerant, rod-shaped bacterium, strain JL216-2 T , was isolated from the shell of the hydrothermal gastropod Alviniconcha marisindica in the Tianxiu hydrothermal field on the Carlsberg Ridge, Indian Ocean. Cells of strain JL216-2 T were motile and 0.5×3–6 µm in size. Strain JL216-2 T grew at 10–32 °C (optimal 28 °C), pH 6.0–9.0 (optimal pH 7.0) and 0.5–12.0% (w/v) sodium chloride(NaCl, optimal 2.5%). Strain JL216-2 T exhibited metabolic versatility, being capable of heterotrophic growth on complex organic compounds. It also performed autotrophic growth by utilizing carbon dioxide (CO₂) fixation or sodium bicarbonate (NaHCO 3 ) as the sole carbon source, driven by hydrogen oxidation coupled with Fe(III) reduction. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the strain was most closely related to Helicovermis profundi S502 T (90.43% sequence similarity). The genome size of strain JL216-2 T was 4.91 Mb, with a G+C content of 39.54 mol%. Phylogenomic analysis and overall genome relatedness indices values indicated a close relatedness of strain JL216-2 T to H. profundi S502 T and Fusibacter spp. The combined genotypic and phenotypic data showed that strain JL216-2 T represents a novel genus within the order Eubacteriales . The type strain is JL216-2 T (=MCCC M30262 T =KACC 23553 T ).

ABSTRACT X‐disease, caused by strains in the 16SrIII‐A subgroup of ‘ Candidatus Phytoplasma pruni’, is a devastating disease of Prunus species (stone fruits). Multiple outbreaks of this disease have occurred across much of North America for more than a century, with the most recent one beginning around 2010 in the Pacific Northwest of the United States, causing severe damage to the stone fruit industry. Sensitive and specific detection of X‐disease is critical to prevent the propagation and spread of infected plant material; however, current PCR‐based detection workflows lack specificity to X‐disease and require one or more PCR assays followed by additional testing to confirm the species identity. In this study, two real‐time PCR assays, one targeting the secY gene and the other targeting the tuf gene, were developed for specific detection of X‐disease in stone fruits. The CPH‐TXD assay targeting the tuf gene provided high‐sensitivity target detection of as few as five target copies with significantly reduced cross‐reactivity compared to current methods. The CPH‐SXD assay targeting the secY gene detected a minimum of 30 target copies and showed no cross‐reaction with closely related strains. The repeatability and reproducibility of both assays were verified with acceptable results. Together, these assays provide efficient and high‐specificity detection methods to combat the growing threat of X‐disease in North America.

Background: Ticks are significant vectors of pathogens, and their nonpathogenic intracellular bacteria, known as endosymbionts, influence tick physiology and may interact with tick-borne pathogens. Such relationships can affect transmission dynamics and understanding them is essential for designing strategies that control tick-borne diseases and reduce their impact on human and animal health. Aim: This study aimed to determine the prevalence and density of two key endosymbiotic bacteria, Francisella-like endosymbiont (FLE) and Candidatus Midichloria mitochondrii (Ca. M. mitochondrii), and to investigate possible correlations between these symbionts and the protozoan parasite Theileria annulata in Hyalomma anatolicum ticks. Methods: Between January and October 2024, 116 ticks were collected from cattle in the Diwaniyah province, Iraq. The morphological examination was followed by molecular confirmation using mitochondrial 16S rRNA sequencing. Quantitative polymerase chain reaction was applied to detect and quantify FLE, Ca. M. mitochondrii, and T. annulata in H. anatolicum. Results: H. anatolicum was the predominant species, representing 86% of ticks, whereas H. excavatum accounted for 13%. Mitochondrial 16S rRNA sequences confirmed species identity and showed 99.48-100% similarity to reference isolates from China, Pakistan, Turkey, India, and Russia. FLE was detected in 100% of H. anatolicum, with mean loads of 7.9× 103 copies per tick. Ca. M. mitochondrii was detected in 99% of specimens, with mean loads of 3.7×102 copies per tick. FLE densities tended to be higher in females compared to males (9.7 × 10³ copies vs. 1.8 × 10³ copies) but difference was not statistically significant. Ca. M. mitochondrii loads also differed significantly (4.8 × 10² copies vs. 1.3 × 10¹ copies). T. annulata was present in all ticks, with the highest copy numbers in semi-engorged females (1.9 × 104 copies), followed by males (2.8 × 10³ copies). Our findings revealed a statistically significant association (p<0.05) between T. annulata and Ca. M. mitochondrii, whereas no significant correlation was observed between T. annulata and FLE. Conclusion: The presence of FLE and Ca. M. mitochondrii in H. anatolicum from the Diwaniyah province was documented. Evidence indicated associations between symbiont density and T. annulata infection, and the results point toward a possible role of endosymbiotic bacteria in pathogen persistence and the epidemiology of tick-borne diseases.

Abstract BACKGROUND Huanglongbing (HLB) is the most devastating citrus disease worldwide, with no resistant commercial citrus varieties identified and no effective chemical treatments currently available on the market. The main challenge in developing effective strategies for controlling HLB is the inability to culture its associated pathogen, the fastidious Candidatus Liberibacter asiaticus ( C Las), in defined media or axenic cultures. RESULTS In this study, we developed an effective method for screening antimicrobials against C Las using periwinkle as a model plant. Compared with citrus, periwinkle offers notable advantages, including reduced time, increased efficiency, and lower costs. This method consists of two main components: screening exogenously applied antimicrobials and evaluating overexpressed endogenous antimicrobial peptides. Using this method, we demonstrated that bromothalonil and cyproconazole effectively inhibit the proliferation of C Las in hydroponically grown periwinkle. Further analyses revealed that bromothalonil and cyproconazole were also highly toxic to other citrus pathogens, including Xanthomonas citri subsp. citri ( Xcc ), Alternaria alternata tangerine pathotype (Z7), and Penicillium digitatum (KH8). In addition, we established an Agrobacterium rhizogenes ‐mediated hairy root genetic transformation system for periwinkle and demonstrated that the proliferation of C Las was significantly inhibited in transgenic roots overexpressing antimicrobial peptide (SAMP). CONCLUSION An efficient antimicrobial screening system for C Las and several other citrus pathogens has been established. By addressing challenges such as high costs, prolonged experimental periods, and heavy workloads, we anticipate that this developed method will serve as a useful tool for the identification of antimicrobials effective against C Las and other citrus pathogens. © 2025 Society of Chemical Industry.