Jose Carlos Huguet-Tapia

Virulence properties are predicted from genome sequence data, in particular the transcription activator-like effector (TALE) gene content, of the rice pathogen Xanthomonas oryzae pv. oryzae (Xoo). Here, we report the genome assembly, annotation, and phylogeny of four legacy isolates of Xoo from Indian rice fields, which clustered closely phylogenetically to previously studied Indian and Nepal Xoo isolates. The TALE genes of these isolates showed both resemblance and unique features when compared with documented Xoo isolates from India, potentially providing insights into possible evolutionary changes in TALE gene profiles. Copyright © 2026 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

We present draft genome sequences of the peanut leaf spot pathogens Passalora arachidicola isolate Ca1604 and Nothopassalora personata isolate Cp1602 that were generated using PacBio sequencing. The assemblies comprise 194 and 571 contigs with total lengths of approximately 37.75 and 49.04 Mb, respectively.

Traditional experimental approaches have greatly advanced our understanding of protein-protein interactions (PPIs) that govern host susceptibility or resistance. Nonetheless, the molecular characterization of microbial effectors and their cognate host targets remains challenging in many economically important plant pathosystems. AlphaFold3 (AF3) has transformed structural biology by achieving near-experimental accuracy in protein structure and complex prediction. A bioinformatics pipeline SPIN (Screening Pathogenicity-Related Host/Pathogen Protein INteractions) was developed for large-scale prediction of interactions between the host and pathogen-secreted proteins. SPIN integrates three core modules: pathogen and host protein preprocessing using bioinformatics tools (SignalP6.0, OrthoFinder, CD-HIT), AF3-based interaction modeling, supported by automated input generation and output filtering. However, AF3 training bias toward mammalian proteins necessitated careful evaluation in plant systems. Benchmarking against experimentally validated plant PPIs revealed that derivative metrics emphasizing interfacial geometry and residue-level contact (ipSAE and pDockQ) provide superior discrimination compared to native global confidence measures (pTM and ipTM), particularly for proteins with intrinsically disordered regions. A multi-metric confidence scoring framework combining pLDDT, PAE, ipSAE, and pDockQ, improved prediction reliability by enhancing recall and reduced false positives through robust assessment of structural confidence and interface quality. For proof-of-concept, SPIN was applied to examine the molecular landscape underlying two economically important diseases of citrus (Citrus L.) caused by Candidatus Liberibacter asiaticus and Ca. Phytoplasma citri. Both pathogens are phloem-limited and cause distinct symptoms, citrus greening and witches' broom, respectively. AF3-predicted interactome data revealed conserved host colonization strategies alongside disease-specific molecular mechanisms, demonstrating the utility of SPIN for dissecting and supporting mechanistic studies in plant-pathogen interactions.

Bacterial blight (BB) of rice, caused by Xanthomonas oryzae pv. oryzae (Xoo), is one of the major drivers of yield losses in Africa and Asia. Xoo secretes TAL‐effectors (TALe) that induce host SWEET sucrose uniporter by binding to the effector binding element (EBE) of SWEET promoters, likely required for Xoo reproduction and virulence. We had multiplex edited the EBEs of three SWEET genes to prevent TALe binding, producing genome‐edited (GE'd) rice mega‐varieties (IR64, Ciherang‐Sub1 for Asia and Komboka for Africa) that were resistant to a wide spectrum of Xoo strains. Here, we report comprehensive analyses of the GE'd lines, including evaluation of agronomic performance in multi‐location multi‐season experimental field plots under different fertilisation regimes and tests for the presence/absence of foreign DNA/transgene in the offspring of GE'd lines (IR64‐BC1T6, Ciherang‐Sub1‐BC1T5, Komboka‐T3). Various strategies were evaluated, including herbicide tolerance, PCR, DNA gel blotting, whole genome sequencing (WGS), and specific tests stipulated by country‐specific biosafety guidelines. Different WGS technologies were evaluated and also used to identify the heritability of the edits, single‐nucleotide polymorphisms (SNPs), and insertions/deletions (indels) that might have resulted from somaclonal variation and potential GE‐induced off‐target mutations. Complete genome reference sequences for the parental lines IR64, Ciherang‐Sub1, and Komboka are provided. In the field experiments, the GE'd lines did not show performance defects. Together, the results indicate that select GE'd lines do not contain foreign DNA or transgene fragments and fulfil the requirements for treatment equivalent to classical breeding lines in countries such as India and Kenya.

Synthetic microbial communities (SynComs) are curated microbial groups that can be designed to optimize microbial functions, such as enhancing plant growth or disease resistance. Attaining SynCom stability in the presence of native soil communities remains a key challenge. This study investigated the survival, persistence, and chemical interactions of a SynCom with a native soil microbial community using a transwell system that spatially constrains bacteria while permitting chemical interactions. The SynCom, composed of six compatible Pseudomonas species identified through whole-genome sequencing, was analyzed for antagonistic interactions with native microbes over time and assessed using biomass and viability measurements. Over time, the SynCom exhibited reduced growth in the presence of native soil microbes compared to the SynCom not exposed to the native microbes. Flow cytometry analysis showed an 81% reduction of live cells for the persistent strain in the presence of native microbes and a 78% and 99% increase in dead and unstained cells, respectively. Compared to a non-persistent strain, one persistent SynCom strain showed lower metabolic utilization across five key compound classes: polymers, carboxylic acids, amino acids, amines, and phenols when exposed to the native soil microbes. These findings underscore the importance of understanding complex SynCom-environment interactions to enhance SynCom stability and optimize in situ applications. IMPORTANCE: SynComs are an emerging technology that can augment plant health. Still, their application in situ depends on deciphering the complex interactions between SynCom microbes and native microbial communities. This study provides insight into several Pseudomonas strains displaying persistent characteristics. Understanding the persistent traits of these bacteria is a vital advancement in SynCom technology and an important next step toward implementing SynComs in agricultural systems.

Synthetic microbial communities (SynComs) are curated microbial groups designed to enhance plant growth or disease resistance by augmenting soil microbiomes. Attaining SynCom stability in the presence of native soil communities remains a key challenge. This study investigated the survival, persistence, and chemical interactions of a SynCom with a native soil microbial community using a transwell system that spatially constrains bacteria while permitting chemical interactions. The SynCom, composed of six compatible Pseudomonas species identified through whole-genome sequencing, was analyzed for antagonistic interactions with native microbes over time and assessed using biomass and viability measurements. Over time, the SynCom exhibitedreduced growth in the presence of native soil microbes compared to the SynCom not exposed to the native microbes. Flow cytometry analysis showed an 81% reduction of live cells for the persistent strain in the presence of native microbes and a 78% and 99% increase in dead and unstained cells, respectively. Compared to a non-persistent strain, one persistent SynCom strain showed lower metabolic utilization across five key compound classes: polymers, carboxylic acids, amino acids, amines, and phenols when exposed to the native soil microbes. These findings underscore the importance of understanding complex SynCom-environment interactions to enhance SynCom stability and optimize in situ applications. Importance: Synthetic Microbial Communities, or SynComs, are an emerging technology that can potentially augment plant health. Still, their application in situ depends on deciphering the complex interactions between SynCom microbes and native microbial communities. This study provides insight into several Pseudomonas strains displaying persistent characteristics, which makes these bacteria promising candidates for SynCom stability in implanted environments. Understanding the persistent traits of these bacteria is a vital advancement in SynCom technology, and an important next step toward implementing SynComs in agricultural systems.

We performed genome sequencing and comparative analysis of Pantoea ananatis strains isolated from corn leaves expressing typical bacterial leaf streak (BLS) and maize white spot (MWS) symptoms to confirm bacterial identity and to understand the relationship among these strains and P. ananatis strains isolated from different plant hosts in Brazil. In pathogenicity tests, strains 4.2 and 13.3 isolated from symptomatic BLS leaves were non-pathogenic on corn. In contrast, strain B13 isolated from MWS-diseased leaf tissue caused symptoms typical of MWS. Our comparative analysis revealed that all three strains are very genetically similar. The G+C (%) content of strains 4.2 and 13.3 was 53.5%, while the B13 content was 53.7%. Average nucleotide identity (ANI) analysis showed that strains B13 and 13.3, B13 and 4.2, and 4.2 and 13.3 shared ANIs of 99.17%, 99.15% and 99.99%, respectively. Strains 13.3, B13, and 4.2 shared ~99% ANI with P. ananatis type strain LMG 2665. To the best of our knowledge, these are the first genome sequences of P. ananatis strains isolated from corn in Brazil.

The genus Herbaspirillum comprises 13 species, the majority of which are plant colonizers. However, some species are occasionally isolated from environmental sources, including water and polluted soil, while others are opportunistic human pathogens. Four novel bacterial strains were isolated from diseased foliage of tomato and Boston fern in Florida, USA. Phylogenetic analysis based on the 16S rRNA gene sequence placed all strains into the genus Herbaspirillum. The Gram-negative strains produced opaque, creamy white, mucoid colonies, which is typical of the genus Herbaspirillum. Biolog biochemical profiling also identified those strains as members of Herbaspirillum. The strains were subjected to whole-genome sequencing, and their genomes were compared with those of reference strains of Herbaspirillum spp. using average nucleotide identity (ANI). The two strains isolated from Boston fern shared 99% pairwise ANI, as did the two strains isolated from tomato. Among all reference genomes tested, the novel strains shared the highest ANI to Herbaspirillum huttiense subsp. huttiense (G21-1742 and NC 40101, 96.76%; SE1, 97.23%; F1, 97.16%) and to H. huttiense subsp. putei. These values are above the established 95% threshold for species delineation based on ANI. As the ANI between members of the two currently described subspecies of H. huttiense, i.e. huttiense and putei, is also ~97%, it can be inferred that the two groups of novel strains described in this study should be considered as candidates for classification as two new subspecies of H. huttiense, given that the current H. huttiense subspecies also have ~97% with the fern and tomato strains. In silico DNA–DNA hybridization results were consistent with those of ANI; comparison of G21-1742 and NC 40101 with H. huttiense subsp. putei IAM 15032^Tand H. huttiense subsp. huttiense LMG 2199^T produced DNA–DNA hybridization (DDH) values of 66.1 and 73.6 %, respectively. Similarly, SE1 and F1 had DDH values of 68.9 and 68.8% with H. huttiense subsp. putei IAM 15032^T and 77.1 and 76.7% with H. huttiense subsp. huttiense LMG 2199^T, respectively. The genomes of all novel isolates carry genes involved in plant pathogenesis, including those of the type III secretion system, which are not present in other H. huttiense strains. Based on genomic and phenotypic data, we conclude that these strains represent the first phytopathogenic subspecies within H. huttiense and the names proposed are H. huttiense subsp. nephrolepidis for the two strains isolated from Nephrolepis exaltata (designated strain, G21−1742=LMG 33362=NCPPB 4765) and H. huttiense subsp. lycopersici (designated strain, SE1=LMG 3361=NCPPB 4764) for the two strains isolated from Solanum lycopersicum.

Due to the error-prone nature of viral RNA-dependent RNA polymerases, the replication of RNA viruses results in a diversity of viral genomes harboring point mutations, deletions, insertions, and genome rearrangements. Citrus tristeza virus (CTV), a causal agent of diseases of economically important citrus species, shows intrinsic genetic stability. While the virus appears to have some mechanism that limits the accumulation of single-nucleotide variants, the production of defective viral genomes (DVGs) during virus infection has been reported for certain variants of CTV. The intra-host diversity generated during plant infection with variant T36 (CTV-T36) remains unclear. To address this, we analyzed the RNA species accumulated in the initially infected and systemic leaves of Nicotiana benthamiana plants inoculated with an infectious cDNA clone of CTV-T36, which warranted that infection was initiated by a known, well-defined sequence variant of the virus. CTV-T36 limited the accumulation of single-nucleotide mutants during infection. With that, four types of DVGs—deletions, insertions, and copy- and snap-backs—were found in all the samples, with deletions and insertions being the most common types. Hot-spots across the genome for DVG recombination and short direct sequence repeats suggest that sequence complementarity could mediate DVG formation. In conclusion, our study illustrates the formation of diverse DVGs during CTV-T36 infection. To the best of our knowledge, this is the first study that has analyzed the genetic variability and recombination of a well-defined sequence variant of CTV in an herbaceous host.