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1
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Lorv JSH, McConkey BJ. Kastor: a reference-based comparative approach for assessment and correction of gene-fragmenting errors in long-read assemblies of small genomes. BMC Genomics 2025; 26:388. [PMID: 40251490 PMCID: PMC12007338 DOI: 10.1186/s12864-025-11569-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2024] [Accepted: 04/04/2025] [Indexed: 04/20/2025] Open
Abstract
Long read sequencing technologies provide an efficient approach to generating highly contiguous and informative assemblies. However, higher relative error rates can introduce frameshifts and premature stop codons that pseudogenize genes, hindering downstream analyses. We developed a software tool that detects gene-fragmenting errors in draft assemblies of small genomes through comparison with a curated set of reference genome sequences and raw read information. In our presented example, detected errors represent less than 0.05% of the genome, but when corrected reduced the rate of pseudogenes from 23.3 to 5.6% in example long read assemblies, comparable to the rate of pseudogenes in short read assemblies. We demonstrate that this software can detect assembly errors in long read assemblies generated from small genomes and correct them to de-fragment genes.
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Affiliation(s)
- Janet S H Lorv
- Department of Biology, University of Waterloo, Waterloo, ON, Canada
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2
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Li J, Guo C, Xie M, Wang K, Wang X, Zou B, Hou F, Ran C, Bi S, Xu Y, Hua Y. Genomic signatures of sensory adaptation and evolution in pangolins. BMC Genomics 2024; 25:1176. [PMID: 39633301 PMCID: PMC11616205 DOI: 10.1186/s12864-024-11063-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2024] [Accepted: 11/18/2024] [Indexed: 12/07/2024] Open
Abstract
BACKGROUND Pangolin is one of the most endangered mammals with many peculiar characteristics, yet the understanding of its sensory systems is still superficial. Studying the genomic basis of adaptation and evolution of pangolin's sensory system is expected to provide further potential assistance for their conservation in the future. RESULTS In this study, we performed a comprehensive comparative genomic analysis to explore the signature of sensory adaptation and evolution in pangolins. By comparing with the aardvark, Cape golden mole, and short-beaked echidna, 124 and 152 expanded gene families were detected in the genome of the Chinese and Malayan pangolins, respectively. The enrichment analyses showed olfactory-related genomic convergence among five concerned mammals. We found 769 and 733 intact OR genes, and 704 and 475 OR pseudogenes in the Chinese and Malayan pangolin species, respectively. Compared to other mammals, far more intact members of OR6 and OR14 were identified in pangolins, particularly for four genes with large copy numbers (OR6C2, OR14A2, OR14C36, and OR14L1). On the genome-wide scale, 1,523, 1,887, 1,110, and 2,732 genes were detected under positive selection (PSGs), intensified selection (ISGs), rapid evolution (REGs), and relaxed selection (RSGs) in pangolins. GO terms associated with visual perception were enriched in PSGs, ISGs, and REGs. Those related to rhythm and sound perception were enriched in both ISGs and REGs, ear development and morphogenesis were enriched in ISGs, and mechanical stimulus and temperature adaptation were enriched in RSGs. The convergence of two vision-related PSGs (OPN4 and ATXN7), with more than one parallel substituted site, was detected among five concerned mammals. Additionally, the absence of intact genes of PKD1L3, PKD2L1, and TAS1R2 and just six single-copy TAS2Rs (TAS2R1, TAS2R4, TAS2R7, TAS2R38, TAS2R40, and TAS2R46) were found in pangolins. Interestingly, we found two large insertions in TAS1R3, distributed in the N-terminal ectodomain, just in pangolins. CONCLUSIONS We found new features related to the adaptation and evolution of pangolin-specific sensory characteristics across the genome. These are expected to provide valuable and useful genome-wide genetic information for the future breeding and conservation of pangolins.
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Affiliation(s)
- Jun Li
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China
- College of Wildlife and Protected Area, Northeast Forestry University, Harbin, 150040, China
| | - Ce Guo
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China
- College of Wildlife and Protected Area, Northeast Forestry University, Harbin, 150040, China
| | - Meiling Xie
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China
- College of Wildlife and Protected Area, Northeast Forestry University, Harbin, 150040, China
| | - Kai Wang
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China
| | - Xianghe Wang
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China
| | - Bishan Zou
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China
| | - Fanghui Hou
- Guangdong Wildlife Rescue Monitoring Center, Guangzhou, 510520, China
- Pangolin Conservation Research Center of National Forestry and Grassland Administration, Guangzhou, 510520, China
| | - Chongyang Ran
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China
| | - Shiman Bi
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China
- College of Wildlife and Protected Area, Northeast Forestry University, Harbin, 150040, China
| | - Yanchun Xu
- College of Wildlife and Protected Area, Northeast Forestry University, Harbin, 150040, China.
| | - Yan Hua
- Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou, 510520, China.
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Poddar BJ, Khardenavis AA. Genomic Insights into the Landfill Microbial Community: Denitrifying Activity Supporting One-Carbon Utilization. Appl Biochem Biotechnol 2024; 196:8866-8891. [PMID: 38980659 DOI: 10.1007/s12010-024-04980-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 06/05/2024] [Indexed: 07/10/2024]
Abstract
In spite of the developments in understanding of denitrifying methylotrophy in the recent years, challenges still exist in unravelling the overall biochemistry of nitrate-dependent methane oxidation in novel or poorly characterized/not-yet-cultured bacteria. In the present study, landfill site was mined for novel C1-carbon-metabolizing bacteria which can use nitrate/nitrite as an electron acceptor. A high-throughput rapid plate assay identified three bacterial isolates with eminent ability for nitrate-dependent methane metabolism under anaerobic conditions. Taxonomic identification by whole-genome sequence-based overall genome relatedness indices accurately assigned the isolates AAK_M13, AAK_M29, and AAK_M39 at the species level to Enterobacter cloacae, Bacillus subtilis, and Bacillus halotolerans, respectively. Several genes encoding sub-components involved in alcohol utilization and denitrification pathways, such as adh, fdh, fdo, nar, nir, and nor, were identified in all the genomes. Though no gene clusters encoding MMO/AMO were annotated, sequencing of PCR amplicons revealed similarity with pMMO/AMO gene using translated nucleotide sequence of strains AAK_M29 and AAK_M39, while strain AAK_M13 showed similarity with XRE family transcriptional regulator. This suggests the horizontal gene transfer and/or presence of a truncated version of a housekeeping enzyme encoded by genes exhibiting partial sequence similarity with pMMO genes that mimicked its function at greenhouse gas emission sites. Owing to lack of conclusive evidence for presence of methane metabolism genes in the selected isolates, further experiment was performed to validate their nitrate-dependent methane oxidation capacities. Bacillus subtilis AAK_M29, Bacillus halotolerans AAK_M39, and Enterobacter cloacae AAK_M13 could oxidize 60%, 75%, and 85% of the added methane respectively accompanied by high nitrate reduction (56-62%) thus supporting the correlation between these two activities. The remarkable ability of these isolates for nitrate-dependent methane metabolism has highlighted their role in ecological contribution and biotechnological potential to serve as methane and nitrate sinks in the landfill sites.
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Affiliation(s)
- Bhagyashri J Poddar
- Environmental Biotechnology and Genomics Division (EBGD), CSIR-National Environmental Engineering Research Institute (NEERI), Nagpur, 440020, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India
| | - Anshuman A Khardenavis
- Environmental Biotechnology and Genomics Division (EBGD), CSIR-National Environmental Engineering Research Institute (NEERI), Nagpur, 440020, India.
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India.
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4
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Fenske L, Jelonek L, Goesmann A, Schwengers O. BakRep - a searchable large-scale web repository for bacterial genomes, characterizations and metadata. Microb Genom 2024; 10:001305. [PMID: 39475723 PMCID: PMC11524574 DOI: 10.1099/mgen.0.001305] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2024] [Accepted: 09/19/2024] [Indexed: 11/02/2024] Open
Abstract
Bacteria are fascinating research objects in many disciplines for countless reasons, and whole-genome sequencing (WGS) has become the paramount methodology to advance our microbiological understanding. Meanwhile, access to cost-effective sequencing platforms has accelerated bacterial WGS to unprecedented levels, introducing new challenges in terms of data accessibility, computational demands, heterogeneity of analysis workflows and, thus, ultimately its scientific usability. To this end, a previous study released a uniformly processed set of 661 405 bacterial genome assemblies obtained from the European Nucleotide Archive as of November 2018. Building on these accomplishments, we conducted further genome-based analyses like taxonomic classification, multilocus sequence typing and annotation of all genomes. Here, we present BakRep, a searchable large-scale web repository of these genomes enriched with consistent genome characterizations and original metadata. The platform provides a flexible search engine combining taxonomic, genomic and metadata information, as well as interactive elements to visualize genomic features. Furthermore, all results can be downloaded for offline analyses via an accompanying command line tool. The web repository is accessible via https://bakrep.computational.bio.
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Affiliation(s)
- Linda Fenske
- Bioinformatics and Systems Biology, Justus Liebig University Giessen, Giessen, Germany
| | - Lukas Jelonek
- Bioinformatics and Systems Biology, Justus Liebig University Giessen, Giessen, Germany
| | - Alexander Goesmann
- Bioinformatics and Systems Biology, Justus Liebig University Giessen, Giessen, Germany
| | - Oliver Schwengers
- Bioinformatics and Systems Biology, Justus Liebig University Giessen, Giessen, Germany
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Bonnici V, Chicco D. Seven quick tips for gene-focused computational pangenomic analysis. BioData Min 2024; 17:28. [PMID: 39227987 PMCID: PMC11370085 DOI: 10.1186/s13040-024-00380-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2024] [Accepted: 08/12/2024] [Indexed: 09/05/2024] Open
Abstract
Pangenomics is a relatively new scientific field which investigates the union of all the genomes of a clade. The word pan means everything in ancient Greek; the term pangenomics originally regarded genomes of bacteria and was later intended to refer to human genomes as well. Modern bioinformatics offers several tools to analyze pangenomics data, paving the way to an emerging field that we can call computational pangenomics. Current computational power available for the bioinformatics community has made computational pangenomic analyses easy to perform, but this higher accessibility to pangenomics analysis also increases the chances to make mistakes and to produce misleading or inflated results, especially by beginners. To handle this problem, we present here a few quick tips for efficient and correct computational pangenomic analyses with a focus on bacterial pangenomics, by describing common mistakes to avoid and experienced best practices to follow in this field. We believe our recommendations can help the readers perform more robust and sound pangenomic analyses and to generate more reliable results.
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Affiliation(s)
- Vincenzo Bonnici
- Dipartimento di Scienze Matematiche Fisiche e Informatiche, Università di Parma, Parma, Italy.
| | - Davide Chicco
- Dipartimento di Informatica Sistemistica e Comunicazione, Università di Milano-Bicocca, Milan, Italy.
- Institute of Health Policy Management and Evaluation, University of Toronto, Toronto, Ontario, Canada.
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6
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Griffiths EJ, Mendes I, Maguire F, Guthrie JL, Wee BA, Schmedes S, Holt K, Yadav C, Cameron R, Barclay C, Dooley D, MacCannell D, Chindelevitch L, Karsch-Mizrachi I, Waheed Z, Katz L, Petit III R, Dave M, Oluniyi P, Nasar MI, Raphenya A, Hsiao WWL, Timme RE. PHA4GE quality control contextual data tags: standardized annotations for sharing public health sequence datasets with known quality issues to facilitate testing and training. Microb Genom 2024; 10:001260. [PMID: 38860884 PMCID: PMC11261899 DOI: 10.1099/mgen.0.001260] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2024] [Accepted: 05/22/2024] [Indexed: 06/12/2024] Open
Abstract
As public health laboratories expand their genomic sequencing and bioinformatics capacity for the surveillance of different pathogens, labs must carry out robust validation, training, and optimization of wet- and dry-lab procedures. Achieving these goals for algorithms, pipelines and instruments often requires that lower quality datasets be made available for analysis and comparison alongside those of higher quality. This range of data quality in reference sets can complicate the sharing of sub-optimal datasets that are vital for the community and for the reproducibility of assays. Sharing of useful, but sub-optimal datasets requires careful annotation and documentation of known issues to enable appropriate interpretation, avoid being mistaken for better quality information, and for these data (and their derivatives) to be easily identifiable in repositories. Unfortunately, there are currently no standardized attributes or mechanisms for tagging poor-quality datasets, or datasets generated for a specific purpose, to maximize their utility, searchability, accessibility and reuse. The Public Health Alliance for Genomic Epidemiology (PHA4GE) is an international community of scientists from public health, industry and academia focused on improving the reproducibility, interoperability, portability, and openness of public health bioinformatic software, skills, tools and data. To address the challenges of sharing lower quality datasets, PHA4GE has developed a set of standardized contextual data tags, namely fields and terms, that can be included in public repository submissions as a means of flagging pathogen sequence data with known quality issues, increasing their discoverability. The contextual data tags were developed through consultations with the community including input from the International Nucleotide Sequence Data Collaboration (INSDC), and have been standardized using ontologies - community-based resources for defining the tag properties and the relationships between them. The standardized tags are agnostic to the organism and the sequencing technique used and thus can be applied to data generated from any pathogen using an array of sequencing techniques. The tags can also be applied to synthetic (lab created) data. The list of standardized tags is maintained by PHA4GE and can be found at https://github.com/pha4ge/contextual_data_QC_tags. Definitions, ontology IDs, examples of use, as well as a JSON representation, are provided. The PHA4GE QC tags were tested, and are now implemented, by the FDA's GenomeTrakr laboratory network as part of its routine submission process for SARS-CoV-2 wastewater surveillance. We hope that these simple, standardized tags will help improve communication regarding quality control in public repositories, in addition to making datasets of variable quality more easily identifiable. Suggestions for additional tags can be submitted to PHA4GE via the New Term Request Form in the GitHub repository. By providing a mechanism for feedback and suggestions, we also expect that the tags will evolve with the needs of the community.
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Affiliation(s)
- Emma J. Griffiths
- Centre for Infectious Disease Genomics and One Health, Faculty of Health Sciences, Simon Fraser University, Burnaby, British Columbia, Canada
| | - Inês Mendes
- Theiagen Genomics, LLC, Highlands Ranch, Colorado, USALLC, Highlands Ranch, Colorado, USA
| | - Finlay Maguire
- Department of Community Health & Epidemiology, Faculty of Medicine, Dalhousie University, Halifax, Nova Scotia, Canada, and Faculty of Computer Science, Dalhousie University, Halifax, Nova Scotia, Canada
| | - Jennifer L. Guthrie
- Department of Microbiology & Immunology, Western University, London, Ontario, Canada
| | - Bryan A. Wee
- The Roslin Institute, University of Edinburgh, Edinburgh, UK
| | - Sarah Schmedes
- National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, Georgia, USA
| | - Kathryn Holt
- National Microbiology Laboratory, Public health Agency of Canada, Winnipeg, MB, Canada
| | - Chanchal Yadav
- National Microbiology Laboratory, Public health Agency of Canada, Winnipeg, MB, Canada
| | - Rhiannon Cameron
- Centre for Infectious Disease Genomics and One Health, Faculty of Health Sciences, Simon Fraser University, Burnaby, British Columbia, Canada
| | - Charlotte Barclay
- Centre for Infectious Disease Genomics and One Health, Faculty of Health Sciences, Simon Fraser University, Burnaby, British Columbia, Canada
| | - Damion Dooley
- Centre for Infectious Disease Genomics and One Health, Faculty of Health Sciences, Simon Fraser University, Burnaby, British Columbia, Canada
| | - Duncan MacCannell
- National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, Georgia, USA
| | - Leonid Chindelevitch
- Department of Infection Biology, London School of Hygiene and Tropical Medicine, London, UK
- MRC Centre for Global Infectious Disease Analysis, School of Public Health, Imperial College London, London, UK
| | - Ilene Karsch-Mizrachi
- National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD, USA
| | - Zahra Waheed
- European Bioinformatics Institute, Wellcome Genome Campus, Hinxton, UK
| | - Lee Katz
- Center for Food Safety, University of Georgia, Georgia, USA
| | | | - Mugdha Dave
- McMaster University, Hamilton, Ontario, Canada
| | | | - Muhammad Ibtisam Nasar
- Department of Biology, College of Science, United Arab Emirates University- AL Ain, Abu Dhabi, UAE
| | - Amogelang Raphenya
- Michael G. DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario, Canada
| | - William W. L. Hsiao
- Centre for Infectious Disease Genomics and One Health, Faculty of Health Sciences, Simon Fraser University, Burnaby, British Columbia, Canada
| | - Ruth E. Timme
- Center for Food Safety and Applied Nutrition, U.S. Food and Drug Administration, College Park, Maryland, USA
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Ángeles-Argáiz RE, Aguirre-Beltrán LFL, Hernández-Oaxaca D, Quintero-Corrales C, Trujillo-Roldán MA, Castillo-Ramírez S, Garibay-Orijel R. Assembly collapsing versus heterozygosity oversizing: detection of homokaryotic and heterokaryotic Laccaria trichodermophora strains by hybrid genome assembly. Microb Genom 2024; 10:001218. [PMID: 38529901 PMCID: PMC10995626 DOI: 10.1099/mgen.0.001218] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2023] [Accepted: 03/01/2024] [Indexed: 03/27/2024] Open
Abstract
Genome assembly and annotation using short-paired reads is challenging for eukaryotic organisms due to their large size, variable ploidy and large number of repetitive elements. However, the use of single-molecule long reads improves assembly quality (completeness and contiguity), but haplotype duplications still pose assembly challenges. To address the effect of read length on genome assembly quality, gene prediction and annotation, we compared genome assemblers and sequencing technologies with four strains of the ectomycorrhizal fungus Laccaria trichodermophora. By analysing the predicted repertoire of carbohydrate enzymes, we investigated the effects of assembly quality on functional inferences. Libraries were generated using three different sequencing platforms (Illumina Next-Seq, Mi-Seq and PacBio Sequel), and genomes were assembled using single and hybrid assemblies/libraries. Long reads or hybrid assemby resolved the collapsing of repeated regions, but the nuclear heterozygous versions remained unresolved. In dikaryotic fungi, each cell includes two nuclei and each nucleus has differences not only in allelic gene version but also in gene composition and synteny. These heterokaryotic cells produce fragmentation and size overestimation of the genome assembly of each nucleus. Hybrid assembly revealed a wider functional diversity of genomes. Here, several predicted oxidizing activities on glycosyl residues of oligosaccharides and several chitooligosaccharide acetylase activities would have passed unnoticed in short-read assemblies. Also, the size and fragmentation of the genome assembly, in combination with heterozygosity analysis, allowed us to distinguish homokaryotic and heterokaryotic strains isolated from L. trichodermophora fruit bodies.
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Affiliation(s)
- Rodolfo Enrique Ángeles-Argáiz
- Posgrado en Ciencias Biológicas, Universidad Nacional Autónoma de México, Circuito de los Posgrados s/n, Ciudad Universitaria, Delegación Coyoacán, Ciudad de México, México, C.P. 04510, Mexico
- Instituto de Biología, Universidad Nacional Autónoma de México, Tercer Circuito s/n, Ciudad Universitaria, Delegación Coyoacán, Ciudad de México, México, C.P. 04510, Mexico
- Red de Manejo Biotecnológico de Recursos, Instituto de Ecología A. C. Carretera antigua a Coatepec 351, Col. El Haya, Xalapa, Veracruz, México, C.P. 91612, Mexico
| | - Luis Fernando Lozano Aguirre-Beltrán
- Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México, Avenida Universidad s/n, Universidad Autónoma del Estado de Morelos, Cuernavaca, Morelos, México, C.P. 62210, Mexico
| | - Diana Hernández-Oaxaca
- Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México, Avenida Universidad s/n, Universidad Autónoma del Estado de Morelos, Cuernavaca, Morelos, México, C.P. 62210, Mexico
- Red de Biodiversidad y Sistemática, Instituto de Ecología A. C. Carretera antigua a Coatepec 351, Col. El Haya, Xalapa, Veracruz, México, C.P. 91073, Mexico
| | - Christian Quintero-Corrales
- Posgrado en Ciencias Biológicas, Universidad Nacional Autónoma de México, Circuito de los Posgrados s/n, Ciudad Universitaria, Delegación Coyoacán, Ciudad de México, México, C.P. 04510, Mexico
- Instituto de Biología, Universidad Nacional Autónoma de México, Tercer Circuito s/n, Ciudad Universitaria, Delegación Coyoacán, Ciudad de México, México, C.P. 04510, Mexico
| | - Mauricio A. Trujillo-Roldán
- Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Tercer Circuito s/n, Ciudad Universitaria, Delegación Coyoacán, Ciudad de México, México, C.P. 04510, Mexico
- Centro de Nanociencias y Nanotecnología, Universidad Nacional Autónoma de México, Km 107 carretera Tijuana-Ensenada, Ensenada, Baja California, Mexico, C.P. 22860, Mexico
| | - Santiago Castillo-Ramírez
- Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México, Avenida Universidad s/n, Universidad Autónoma del Estado de Morelos, Cuernavaca, Morelos, México, C.P. 62210, Mexico
| | - Roberto Garibay-Orijel
- Instituto de Biología, Universidad Nacional Autónoma de México, Tercer Circuito s/n, Ciudad Universitaria, Delegación Coyoacán, Ciudad de México, México, C.P. 04510, Mexico
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Chen J, Wang S, Wang D, Chiu Y, Yang N, Lian X, Zhao Z, Wei Q. Uncovering rearrangements in the Tibetan antelope via population-derived genome refinement and comparative analysis with homologous species. Front Genet 2024; 15:1302554. [PMID: 38425715 PMCID: PMC10902437 DOI: 10.3389/fgene.2024.1302554] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2023] [Accepted: 01/24/2024] [Indexed: 03/02/2024] Open
Abstract
Introduction: The Tibetan antelope (Pantholops hodgsonii) is a remarkable mammal thriving in the extreme Qinghai-Tibet Plateau conditions. Despite the availability of its genome sequence, limitations in the scaffold-level assembly have hindered a comprehensive understanding of its genomics. Moreover, comparative analyses with other Bovidae species are lacking, along with insights into genome rearrangements in the Tibetan antelope. Methods: Addressing these gaps, we present a multifaceted approach by refining the Tibetan Antelope genome through linkage disequilibrium analysis with data from 15 newly sequenced samples. Results: The scaffold N50 of the refined reference is 3.2 Mbp, surpassing the previous version by 1.15-fold. Our annotation analysis resulted in 50,750 genes, encompassing 29,324 novel genes not previously study. Comparative analyses reveal 182 unique rearrangements within the scaffolds, contributing to our understanding of evolutionary dynamics and species-specific adaptations. Furthermore, by conducting detailed genomic comparisons and reconstructing rearrangements, we have successfully pioneered the reconstruction of the X-chromosome in the Tibetan antelope. Discussion: This effort enhances our comprehension of the genomic landscape of this species.
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Affiliation(s)
- Jiarui Chen
- College of Eco‐Environmental Engineering, Qinghai University, Xining, Qinghai, China
- State Key Laboratory of Plateau Ecology and Agriculture, Qinghai University, Xining, Qinghai, China
| | - Shuwen Wang
- College of Eco‐Environmental Engineering, Qinghai University, Xining, Qinghai, China
| | - Dong Wang
- Key Laboratory of Adaptation and Evolution of Plateau Biota, Northwest Institute of Plateau Biology, Chinese Academy of Sciences, Xining, Qinghai, China
- School of Geographical Science, Qinghai Normal University, Xining, Qinghai, China
| | - Yunkang Chiu
- Shenzhen Byoryn Technology Co., Ltd., Shenzhen, China
| | - Nan Yang
- National Genomics Data Center & CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China
| | - Xinming Lian
- Key Laboratory of Adaptation and Evolution of Plateau Biota, Northwest Institute of Plateau Biology, Chinese Academy of Sciences, Xining, Qinghai, China
- Qinghai Provincial Key Laboratory of Animal Ecological Genomics, Xining, Qinghai, China
| | - Zicheng Zhao
- Shenzhen Byoryn Technology Co., Ltd., Shenzhen, China
| | - Qing Wei
- College of Eco‐Environmental Engineering, Qinghai University, Xining, Qinghai, China
- State Key Laboratory of Plateau Ecology and Agriculture, Qinghai University, Xining, Qinghai, China
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9
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Maitreya A, Qureshi A. Genomic and phenotypic characterisation of Enterococcus mundtii AM_AQ_BC8 for its anti-biofilm, antimicrobial and probiotic potential. Arch Microbiol 2024; 206:84. [PMID: 38296886 DOI: 10.1007/s00203-023-03816-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2023] [Revised: 12/15/2023] [Accepted: 12/25/2023] [Indexed: 02/02/2024]
Abstract
Enterococcus mundtii AM_AQ_BC8 isolated from biofouled filtration membrane was characterised as a potential probiotic bacterium showing strong L-lactic acid-producing capability. Experimental studies revealed that E. mundtii AM_AQ_BC8 possess antibiofilm and antimicrobial ability too, as tested against strong biofilm-forming bacteria like Pseudomonas spp. The present study has evaluated the genetic potential of E. mundtii AM_AQ_BC8 through genome sequencing. Whole genome analysis revealed the presence of key genes like ldh_1 and ldh_2 responsible for lactic acid production along with genes encoding probiotic features such as acid and bile salt resistance (dnaK, dnaJ, argS), fatty acid synthesis (fabD, fabE) and lactose utilisation (lacG, lacD). The phylogenomic analysis based on OrthoANI (99.85%) and dDDH (96.8%) values revealed that the strain AM_AQ_BC8 shared the highest homology with E. mundtii. The genome sequence of strain AM_AQ_BC8 has been deposited to NCBI and released with GenBank accession no. SAMN32531201. The study primarily demonstrated the probiotic potential of E. mundtii AM_AQ_BC8 isolate, for L-lactate synthesis in high concentration (8.98 g/L/day), which also showed anti-biofilm and antimicrobial activities.
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Affiliation(s)
- Anuja Maitreya
- Environmental Biotechnology and Genomics Division, CSIR-National Environmental Engineering Research Institute Nehru Marg, Nagpur, 440020, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India
| | - Asifa Qureshi
- Environmental Biotechnology and Genomics Division, CSIR-National Environmental Engineering Research Institute Nehru Marg, Nagpur, 440020, India.
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India.
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10
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Schäfer L, Jehle JA, Kleespies RG, Wennmann JT. A practical guide and Galaxy workflow to avoid inter-plasmidic repeat collapse and false gene loss in Unicycler's hybrid assemblies. Microb Genom 2024; 10:001173. [PMID: 38197876 PMCID: PMC10868617 DOI: 10.1099/mgen.0.001173] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/07/2023] [Accepted: 12/18/2023] [Indexed: 01/11/2024] Open
Abstract
Generating complete, high-quality genome assemblies is key for any downstream analysis, such as comparative genomics. For bacterial genome assembly, various algorithms and fully automated pipelines exist, which are free-of-charge and easily accessible. However, these assembly tools often cannot unambiguously resolve a bacterial genome, for example due to the presence of sequence repeat structures on the chromosome or on plasmids. Then, a more sophisticated approach and/or manual curation is needed. Such modifications can be challenging, especially for non-bioinformaticians, because they are generally not considered as a straightforward process. In this study, we propose a standardized approach for manual genome completion focusing on the popular hybrid assembly pipeline Unicycler. The provided Galaxy workflow addresses two weaknesses in Unicycler's hybrid assemblies: (i) collapse of inter-plasmidic repeats and (ii) false loss of single-copy sequences. To demonstrate and validate how to detect and resolve these assembly errors, we use two genomes from the Bacillus cereus group. By applying the proposed pipeline following an automated assembly, the genome sequence quality can be significantly improved.
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Affiliation(s)
- Lea Schäfer
- Julius Kühn Institute (JKI) – Federal Research Centre for Cultivated Plants, Institute for Biological Control, Schwabenheimer Str. 101, 69221 Dossenheim, Germany
| | - Johannes A. Jehle
- Julius Kühn Institute (JKI) – Federal Research Centre for Cultivated Plants, Institute for Biological Control, Schwabenheimer Str. 101, 69221 Dossenheim, Germany
| | - Regina G. Kleespies
- Julius Kühn Institute (JKI) – Federal Research Centre for Cultivated Plants, Institute for Biological Control, Schwabenheimer Str. 101, 69221 Dossenheim, Germany
| | - Jörg T. Wennmann
- Julius Kühn Institute (JKI) – Federal Research Centre for Cultivated Plants, Institute for Biological Control, Schwabenheimer Str. 101, 69221 Dossenheim, Germany
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11
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Hodgeman R, Mann R, Djitro N, Savin K, Rochfort S, Rodoni B. The pan-genome of Mycobacterium avium subsp. paratuberculosis (Map) confirms ancestral lineage and reveals gene rearrangements within Map Type S. BMC Genomics 2023; 24:656. [PMID: 37907856 PMCID: PMC10619280 DOI: 10.1186/s12864-023-09752-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2023] [Accepted: 10/18/2023] [Indexed: 11/02/2023] Open
Abstract
BACKGROUND To date genomic studies on Map have concentrated on Type C strains with only a few Type S strains included for comparison. In this study the entire pan-genome of 261 Map genomes (205 Type C, 52 Type S and 4 Type B) and 7 Mycobacterium avium complex (Mac) genomes were analysed to identify genomic similarities and differences between the strains and provide more insight into the evolutionary relationship within this Mycobacterial species. RESULTS Our analysis of the core genome of all the Map isolates identified two distinct lineages, Type S and Type C Map that is consistent with previous phylogenetic studies of Map. Pan-genome analysis revealed that Map has a larger accessory genome than Mycobacterium avium subsp. avium (Maa) and Type C Map has a larger accessory genome than Type S Map. In addition, we found large rearrangements within Type S strains of Map and little to none in Type C and Type B strains. There were 50 core genes identified that were unique to Type S Map and there were no unique core genes identified between Type B and Type C Map strains. In Type C Map we identified an additional CE10 CAZyme class which was identified as an alpha/beta hydrolase and an additional polyketide and non-ribosomal peptide synthetase cluster. Consistent with previous analysis no plasmids and only incomplete prophages were identified in the genomes of Map. There were 45 hypothetical CRISPR elements identified with no associated cas genes. CONCLUSION This is the most comprehensive comparison of the genomic content of Map isolates to date and included the closing of eight Map genomes. The analysis revealed that there is greater variation in gene synteny within Type S strains when compared to Type C indicating that the Type C Map strain emerged after Type S. Further analysis of Type C and Type B genomes revealed that they are structurally similar with little to no genetic variation and that Type B Map may be a distinct clade within Type C Map and not a different strain type of Map. The evolutionary lineage of Maa and Map was confirmed as emerging after M. hominissuis.
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Affiliation(s)
- Rachel Hodgeman
- Agriculture Victoria, AgriBio, La Trobe University, Bundoora, VIC, Australia.
- School of Applied Systems Biology, AgriBio, La Trobe University, Bundoora, VIC, Australia.
| | - Rachel Mann
- Agriculture Victoria, AgriBio, La Trobe University, Bundoora, VIC, Australia
| | - Noel Djitro
- School of Applied Systems Biology, AgriBio, La Trobe University, Bundoora, VIC, Australia
| | - Keith Savin
- Agriculture Victoria, AgriBio, La Trobe University, Bundoora, VIC, Australia
| | - Simone Rochfort
- Agriculture Victoria, AgriBio, La Trobe University, Bundoora, VIC, Australia
- School of Applied Systems Biology, AgriBio, La Trobe University, Bundoora, VIC, Australia
| | - Brendan Rodoni
- Agriculture Victoria, AgriBio, La Trobe University, Bundoora, VIC, Australia
- School of Applied Systems Biology, AgriBio, La Trobe University, Bundoora, VIC, Australia
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Ramakrishnan P, Ariyan M, Rangasamy A, Rajasekaran R, Ramasamy K, Murugaiyan S, Janahiraman V. Draft Genome Sequence of Enterobacter cloacae S23 a Plant Growth-promoting Passenger Endophytic Bacterium Isolated from Groundnut Nodule Possesses Stress Tolerance Traits. Curr Genomics 2023; 24:36-47. [PMID: 37920731 PMCID: PMC10334703 DOI: 10.2174/1389202924666230403123208] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2022] [Revised: 01/25/2023] [Accepted: 03/03/2023] [Indexed: 04/07/2023] Open
Abstract
Aim This study aims to reveal the passenger endophytic bacterium Enterobacter cloacae S23 isolated from groundnut nodules and to underpin the molecular mechanism and genes responsible for abiotic stress tolerance. Background A variety of microorganisms that contribute to nodulation and encourage plant development activity in addition to the nodulating Rhizobium. Passenger endophytes (PE) are endophytes that accidentally penetrate the plant without any selective pressure keeping them in the interior tissue of the plant. PE possesses characteristics that encourage plant development and boost output while reducing pathogen infection and improving biotic and abiotic stress tolerance. However, there is a lack of molecular evidence on the passenger endophyte-mediated alleviation of abiotic stresses. Objective This study was formulated to reveal the draft genome sequence of Enterobacter cloacae S23, as well as genes and characteristics involved in plant growth promotion and stress tolerance. Method The data were submitted to PATRIC and the TORMES-1.0 Unicyclker tools were used to conduct a complete genome study of Enterobacter cloacae S23. The TORMES-1.0 platform was used to process the reads. RAST tool kit (RASTtk) was used to annotate the S23 sequence. The plant growth-promoting traits such as indole acetic acid production, siderophore secretion, production of extracellular polysaccharides, biofilm formation, phosphate solubilization, and accumulation of osmolytes were examined under normal, 7% NaCl and 30% polyethylene glycol amended conditions to determine their ability to withstand salt and moisture stressed conditions, respectively. Result We report the size of Enterobacter cloacae S23 is 4.82Mb which contains 4511 protein-coding sequences, 71 transfer RNA genes, and 3 ribosomal RNA with a G+C content of DNA is 55.10%. Functional analysis revealed that most of the genes are involved in the metabolism of amino acids, cofactors, vitamins, stress response, nutrient solubilization (kdp, pho, pst), biofilm formation (pga) IAA production (trp), siderophore production (luc, fhu, fep, ent, ybd), defense, and virulence. The result revealed that E. cloacae S23 exhibited multiple plant growth-promoting traits under abiotic stress conditions. Conclusion Our research suggested that the discovery of anticipated genes and metabolic pathways might characterise this bacterium as an environmentally friendly bioresource to support groundnut growth through several mechanisms of action under multi-stresses.
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Affiliation(s)
- Pavithra Ramakrishnan
- Department of Agricultural Microbiology, Tamil Nadu Agricultural University, Coimbatore, 641003, India
| | - Manikandan Ariyan
- Department of Agricultural Microbiology, Tamil Nadu Agricultural University, Coimbatore, 641003, India
- Institute of Ecology and Earth Sciences, University of Tartu, Tartu, Estonia
| | - Anandham Rangasamy
- Department of Agricultural Microbiology, Tamil Nadu Agricultural University, Coimbatore, 641003, India
| | - Raghu Rajasekaran
- Department of Agricultural Microbiology, Tamil Nadu Agricultural University, Coimbatore, 641003, India
| | - Krishnamoorthy Ramasamy
- Department of Crop Management, Vanavarayar Institute of Agriculture, Pollachi, Tamil Nadu, India
| | - SenthilKumar Murugaiyan
- Agricultural College and Research Institute, Tamil Nadu Agricultural University, Eachangkottai, India
| | - Veeranan Janahiraman
- Department of Agricultural Microbiology, Tamil Nadu Agricultural University, Coimbatore, 641003, India
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Lovelace AH, Dorhmi S, Hulin MT, Li Y, Mansfield JW, Ma W. Effector Identification in Plant Pathogens. PHYTOPATHOLOGY 2023; 113:637-650. [PMID: 37126080 DOI: 10.1094/phyto-09-22-0337-kd] [Citation(s) in RCA: 21] [Impact Index Per Article: 10.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/13/2023]
Abstract
Effectors play a central role in determining the outcome of plant-pathogen interactions. As key virulence proteins, effectors are collectively indispensable for disease development. By understanding the virulence mechanisms of effectors, fundamental knowledge of microbial pathogenesis and disease resistance have been revealed. Effectors are also considered double-edged swords because some of them activate immunity in disease resistant plants after being recognized by specific immune receptors, which evolved to monitor pathogen presence or activity. Characterization of effector recognition by their cognate immune receptors and the downstream immune signaling pathways is instrumental in implementing resistance. Over the past decades, substantial research effort has focused on effector biology, especially concerning their interactions with virulence targets or immune receptors in plant cells. A foundation of this research is robust identification of the effector repertoire from a given pathogen, which depends heavily on bioinformatic prediction. In this review, we summarize methodologies that have been used for effector mining in various microbial pathogens which use different effector delivery mechanisms. We also discuss current limitations and provide perspectives on how recently developed analytic tools and technologies may facilitate effector identification and hence generation of a more complete vision of host-pathogen interactions. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.
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Affiliation(s)
| | - Sara Dorhmi
- The Sainsbury Laboratory, Norwich, NR4 7UH, U.K
- Department of Microbiology and Plant Pathology, University of California Riverside, CA 92521, U.S.A
| | | | - Yufei Li
- The Sainsbury Laboratory, Norwich, NR4 7UH, U.K
| | - John W Mansfield
- Faculty of Natural Sciences, Imperial College London, London, SW7 2BX, U.K
| | - Wenbo Ma
- The Sainsbury Laboratory, Norwich, NR4 7UH, U.K
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Schäfer L, Volk F, Kleespies RG, Jehle JA, Wennmann JT. Elucidating the genomic history of commercially used Bacillus thuringiensis subsp. tenebrionis strain NB176. Front Cell Infect Microbiol 2023; 13:1129177. [PMID: 37021121 PMCID: PMC10067926 DOI: 10.3389/fcimb.2023.1129177] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2022] [Accepted: 02/17/2023] [Indexed: 04/07/2023] Open
Abstract
Bacillus thuringiensis subsp. tenebrionis (Btt) produces a coleopteran-specific crystal protoxin protein (Cry3Aa δ-endotoxin). After its discovery in 1982, the strain NB125 (DSM 5526) was eventually registered in 1990 to control the Colorado potato beetle (Leptinotarsa decemlineata). Gamma-irradiation of NB125 resulted in strain NB176-1 (DSM 5480) that exhibited higher cry3Aa production and became the active ingredient of the plant protection product Novodor® FC. Here, we report a comparative genome analysis of the parental strain NB125, its derivative NB176-1 and the current commercial production strain NB176. The entire genome sequences of the parental and derivative strains were deciphered by a hybrid de novo approach using short (Illumina) and long (Nanopore) read sequencing techniques. Genome assembly revealed a chromosome of 5.4 to 5.6 Mbp and six plasmids with a size range from 14.9 to 250.5 kbp for each strain. The major differences among the original NB125 and the derivative strains NB176-1 and NB176 were an additional copy of the cry3Aa gene, which translocated to another plasmid as well as a chromosomal deletion (~ 178 kbp) in NB176. The assembled genome sequences were further analyzed in silico for the presence of virulence and antimicrobial resistance (AMR) genes.
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Affiliation(s)
- Lea Schäfer
- Julius Kühn Institute (JKI) - Federal Research Centre for Cultivated Plants, Institute for Biological Control, Dossenheim, Germany
| | | | - Regina G. Kleespies
- Julius Kühn Institute (JKI) - Federal Research Centre for Cultivated Plants, Institute for Biological Control, Dossenheim, Germany
| | - Johannes A. Jehle
- Julius Kühn Institute (JKI) - Federal Research Centre for Cultivated Plants, Institute for Biological Control, Dossenheim, Germany
| | - Jörg T. Wennmann
- Julius Kühn Institute (JKI) - Federal Research Centre for Cultivated Plants, Institute for Biological Control, Dossenheim, Germany
- *Correspondence: Jörg T. Wennmann,
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Chhetri G, Kim I, Kim J, So Y, Park S, Jung Y, Seo T. Paraburkholderia tagetis sp. nov., a novel species isolated from roots of Tagetes patula enhances the growth and yield of Solanum lycopersicum L. (tomato). Front Microbiol 2023; 14:1140484. [PMID: 37082173 PMCID: PMC10110911 DOI: 10.3389/fmicb.2023.1140484] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2023] [Accepted: 03/21/2023] [Indexed: 04/22/2023] Open
Abstract
A multifunctional, Gram-stain-negative, aerobic, motile by flagella, short-rod shaped bacteria, designated strain RG36T was isolated from roots of marigold plant (Tagetes patula) sampled at Dongguk University, Republic of Korea. A 16S rRNA sequences indicated that the closest phylogenetic neighbors were Paraburkholderia acidiphila 7Q-K02T (99.0%) and Paraburkholderia sacchari IPT101T (98.9%) of the family Burkholderiaceae. The draft genome size was 8.52 Mb (63.7% GC). The genome contained 7,381 coding sequences. Digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values of strain RG36T with its most closely related species were only 83.1-88.7 and 27.6-36.7%, respectively. Strain RG36T contained Q-8 as the major respiratory quinone and its main fatty acids (>10%) were C16:0, C17:0 cyclo, C19:0 cyclo ω8c, and summed feature 8 (comprising C18:1 ω7c and/or C18:1 ω6c). Strain RG36T accumulates polyhydroxybutyrates (PHB) and exhibits multiple plant growth-promoting properties including production of indole-3-acetic acid (IAA), siderophores, protease, phosphate solubilization, and harboring gene clusters for its multifunctional properties. A pot experiment was conducted to evaluate the effect of PGPR on the growth of Solanum lycopersicum L. (Tomato). Result also confirmed the ability of strain RG36T to promote tomato plant growth, especially it increases the yield of tomatoes. Structural assessment of the bioplastic by Fourier transform infrared (FTIR), nuclear magnetic resonance (NMR), and GC-MS spectroscopy, which confirmed the structure of the polymer as PHB. Our study revealed the potential of strain RG36T to promote the growth of tomato plant and fruit yield by stimulating the various phytohormones, which could be use as bio-fertilizers to reduce the use of chemical fertilizers and promotes sustainable agricultural production. The phenotypic, chemotaxonomic and phylogenetic data, and genome analysis showed that strain RG36T represents a novel species of the genus Paraburkholderia, for which the name Paraburkholderia tagetis sp. nov. is proposed. The type strain is RG36T (=KACC 22685T = TBRC 15696T).
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Rasool Kamli M, Malik A, S M Sabir J, Ahmad Rather I, Kim CB. Insights into the biodegradation and heavy metal resistance potential of the genus Brevibacillus through comparative genome analyses. Gene 2022; 846:146853. [PMID: 36070852 DOI: 10.1016/j.gene.2022.146853] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2022] [Revised: 08/24/2022] [Accepted: 08/26/2022] [Indexed: 11/04/2022]
Abstract
Members of the genus Brevibacillus belonging to the familyPaenibacillaceae are Gram-positive/variable, endospore-forming, and rod-shaped bacteria that dwell in various environmental habitats. Brevibacillus spp. have a wide range of enzyme activities such as degradation of various carbohydrates, plastics, and they possess resistance against heavy metals. These characteristics make them encouraging contenders for biotechnological applications.In this work, we analyzed the reference genomes of 19Brevibacillusspecies, focusing on discovering the biodegradation and heavy metal resistance capabilities of this little studied genus from genomic data. The results indicate that several strain specific traits were identified. For example Brevibacillus halotolerans s-14, and Brevibacillus laterosporus DSM 25 have more glycoside hydrolases (GHs) compared to other carbohydrate-active enzymes, and therefore might be more suitable for biodegradation of carbohydrates. In contrast, strains such as Brevibacillus antibioticus TGS2-1, with a higher number of glycosyltransfereases (GTs) may aid in the biosynthesis of complex carbohydrates. Our results also suggest some correlation between heavy metal resistance and polyurethane degradation, thus indicating that heavy metal resistance strains (e.g. Brevibacillus reuszeri J31TS6) can be a promising source of enzymes for polyurethane degradation. These strain specific features make the members of this bacterial group potential candidates for further investigations with industrial implications. This work also represents the first exhaustive study of Brevibacillus at the genome scale.
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Affiliation(s)
- Majid Rasool Kamli
- Department of Biological Sciences, Faculty of Science, King Abdulaziz University, Jeddah 21589, Saudi Arabia; Center of excellence in Bionanoscience Research, King Abdulaziz University, Jeddah 21589, Saudi Arabia.
| | - Adeel Malik
- Institute of Intelligence Informatics Technology, Sangmyung University, Seoul 03016, Republic of Korea
| | - Jamal S M Sabir
- Department of Biological Sciences, Faculty of Science, King Abdulaziz University, Jeddah 21589, Saudi Arabia; Center of excellence in Bionanoscience Research, King Abdulaziz University, Jeddah 21589, Saudi Arabia
| | - Irfan Ahmad Rather
- Department of Biological Sciences, Faculty of Science, King Abdulaziz University, Jeddah 21589, Saudi Arabia; Center of excellence in Bionanoscience Research, King Abdulaziz University, Jeddah 21589, Saudi Arabia
| | - Chang-Bae Kim
- Department of Biotechnology, Sangmyung University, Seoul 03016, Republic of Korea.
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Afzal M, Vijay AK, Stapleton F, Willcox M. The Relationship between Ciprofloxacin Resistance and Genotypic Changes in S. aureus Ocular Isolates. Pathogens 2022; 11:1354. [PMID: 36422605 PMCID: PMC9695201 DOI: 10.3390/pathogens11111354] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2022] [Revised: 10/26/2022] [Accepted: 11/10/2022] [Indexed: 07/28/2023] Open
Abstract
Staphylococcus aureus (S. aureus) is a frequent cause of eye infections with some isolates exhibiting increased antimicrobial resistance to commonly prescribed antibiotics. The increasing resistance of ocular S. aureus to ciprofloxacin is a serious concern as it is a commonly used as a first line antibiotic to treat S. aureus keratitis. This study aimed to analyse genetic mutations in the genomes of 25 S. aureus isolates from infections or non-infectious ocular conditions from the USA and Australia and their relationship to ciprofloxacin resistance. Overall, 14/25 isolates were phenotypically resistant to ciprofloxacin. All isolates were analyzed for mutations in their quinolone resistance-determining regions (QRDRs) and efflux pump genes. Of the fourteen resistant isolates, 9/14 had ciprofloxacin resistance mutations within their QRDRs, at codons 80 or 84 within the parC subunit and codon 84 within the gyrA subunit of DNA gyrase. The highest resistance (MIC = 2560 μg/mL) was associated with two SNPs in both gyrA and parC. Other resistant isolates (3/14) had mutations within norB. Mutations in genes of other efflux pumps and their regulator (norA, norC, mepA, mdeA, sepA, sdrM, mepR, arlR, and arlS) or the DNA mismatch repair (MMR) system (mutL and mutS) were not associated with increased resistance to ciprofloxacin. The functional mutations associated with ciprofloxacin resistance in QRDRs (gyrA and parC) and norB suggests that these are the most common reasons for ciprofloxacin resistance in ocular isolates. Novel SNPs of gyrA Glu-88-Leu, Asn-860-Thr and Thr-845-Ala and IIe-855-Met, identified in this study, need further gene knock out/in studies to better understand their effect on ciprofloxacin resistance.
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Sulja A, Pothier JF, Blom J, Moretti C, Buonaurio R, Rezzonico F, Smits THM. Comparative genomics to examine the endophytic potential of Pantoea agglomerans DAPP-PG 734. BMC Genomics 2022; 23:742. [DOI: 10.1186/s12864-022-08966-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2022] [Accepted: 10/25/2022] [Indexed: 11/09/2022] Open
Abstract
AbstractPantoea agglomerans DAPP-PG 734 was isolated as endophyte from knots (tumors) caused by Pseudomonas savastanoi pv. savastanoi DAPP-PG 722 in olive trees. To understand the plant pathogen-endophyte interaction on a genomic level, the whole genome of P. agglomerans DAPP-PG 734 was sequenced and annotated. The complete genome had a total size of 5′396′424 bp, containing one circular chromosome and four large circular plasmids. The aim of this study was to identify genomic features that could play a potential role in the interaction between P. agglomerans DAPP-PG 734 and P. savastanoi pv. savastanoi DAPP-PG 722. For this purpose, a comparative genomic analysis between the genome of P. agglomerans DAPP-PG 734 and those of related Pantoea spp. was carried out. In P. agglomerans DAPP-PG 734, gene clusters for the synthesis of the Hrp-1 type III secretion system (T3SS), type VI secretion systems (T6SS) and autoinducer, which could play an important role in a plant-pathogenic community enhancing knot formation in olive trees, were identified. Additional gene clusters for the biosynthesis of two different antibiotics, namely dapdiamide E and antibiotic B025670, which were found in regions between integrative conjugative elements (ICE), were observed. The in-depth analysis of the whole genome suggested a characterization of the P. agglomerans DAPP-PG 734 isolate as endophytic bacterium with biocontrol activity rather than as a plant pathogen.
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Purushothaman S, Meola M, Egli A. Combination of Whole Genome Sequencing and Metagenomics for Microbiological Diagnostics. Int J Mol Sci 2022; 23:9834. [PMID: 36077231 PMCID: PMC9456280 DOI: 10.3390/ijms23179834] [Citation(s) in RCA: 40] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2022] [Revised: 08/24/2022] [Accepted: 08/26/2022] [Indexed: 12/21/2022] Open
Abstract
Whole genome sequencing (WGS) provides the highest resolution for genome-based species identification and can provide insight into the antimicrobial resistance and virulence potential of a single microbiological isolate during the diagnostic process. In contrast, metagenomic sequencing allows the analysis of DNA segments from multiple microorganisms within a community, either using an amplicon- or shotgun-based approach. However, WGS and shotgun metagenomic data are rarely combined, although such an approach may generate additive or synergistic information, critical for, e.g., patient management, infection control, and pathogen surveillance. To produce a combined workflow with actionable outputs, we need to understand the pre-to-post analytical process of both technologies. This will require specific databases storing interlinked sequencing and metadata, and also involves customized bioinformatic analytical pipelines. This review article will provide an overview of the critical steps and potential clinical application of combining WGS and metagenomics together for microbiological diagnosis.
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Affiliation(s)
- Srinithi Purushothaman
- Applied Microbiology Research, Department of Biomedicine, University of Basel, 4031 Basel, Switzerland
- Institute of Medical Microbiology, University of Zurich, 8006 Zurich, Switzerland
| | - Marco Meola
- Applied Microbiology Research, Department of Biomedicine, University of Basel, 4031 Basel, Switzerland
- Institute of Medical Microbiology, University of Zurich, 8006 Zurich, Switzerland
- Swiss Institute of Bioinformatics, University of Basel, 4031 Basel, Switzerland
| | - Adrian Egli
- Applied Microbiology Research, Department of Biomedicine, University of Basel, 4031 Basel, Switzerland
- Institute of Medical Microbiology, University of Zurich, 8006 Zurich, Switzerland
- Clinical Bacteriology and Mycology, University Hospital Basel, 4031 Basel, Switzerland
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Singh AK, Gupta RK, Purohit HJ, Khardenavis AA. Genomic characterization of denitrifying methylotrophic Pseudomonas aeruginosa strain AAK/M5 isolated from municipal solid waste landfill soil. World J Microbiol Biotechnol 2022; 38:140. [PMID: 35705700 DOI: 10.1007/s11274-022-03311-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2021] [Accepted: 05/15/2022] [Indexed: 11/26/2022]
Abstract
Municipal landfills are known for methane production and a source of nitrate pollution leading to various environmental issues. Therefore, this niche was selected for the isolation of one-carbon (C1) utilizing bacteria with denitrifying capacities using anaerobic enrichment on nitrate mineral salt medium supplemented with methanol as carbon source. Eight axenic cultures were isolated of which, isolate AAK/M5 demonstrated the highest methanol removal (73.28%) in terms of soluble chemical oxygen demand and methane removal (41.27%) at the expense of total nitrate removal of 100% and 33% respectively. The whole genome characterization with phylogenomic approach suggested that the strain AAK/M5 could be assigned to Pseudomonas aeruginosa with close neighbours as type strains DVT779, AES1M, W60856, and LES400. The circular genome annotation showed the presence of complete set of genes essential for methanol utilization and complete denitrification process. The study demonstrates the potential of P. aeruginosa strain AAK/M5 in catalysing methane oxidation thus serving as a methane sink vis-à-vis utilization of nitrate. Considering the existence of such bacteria at landfill site, the study highlights the need to develop strategies for their enrichment and designing of efficient catabolic activity for such environments.
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Affiliation(s)
- Ashish Kumar Singh
- Environmental Biotechnology and Genomics Division, CSIR-National Environmental Engineering Research Institute, Nehru Marg, Nagpur, Maharashtra, 440020, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India
| | - Rakesh Kumar Gupta
- Environmental Biotechnology and Genomics Division, CSIR-National Environmental Engineering Research Institute, Nehru Marg, Nagpur, Maharashtra, 440020, India
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India
| | - Hemant J Purohit
- Environmental Biotechnology and Genomics Division, CSIR-National Environmental Engineering Research Institute, Nehru Marg, Nagpur, Maharashtra, 440020, India
| | - Anshuman Arun Khardenavis
- Environmental Biotechnology and Genomics Division, CSIR-National Environmental Engineering Research Institute, Nehru Marg, Nagpur, Maharashtra, 440020, India.
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India.
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Abstract
The availability of public genomics data has become essential for modern life sciences research, yet the quality, traceability, and curation of these data have significant impacts on a broad range of microbial genomics research. While microbial genome databases such as NCBI’s RefSeq database leverage the scalability of crowd sourcing for growth, genomics data provenance and authenticity of the source materials used to produce data are not strict requirements. Here, we describe the de novo assembly of 1,113 bacterial genome references produced from authenticated materials sourced from the American Type Culture Collection (ATCC), each with full genomics data provenance relating to bioinformatics methods, quality control, and passage history. Comparative genomics analysis of ATCC standard reference genomes (ASRGs) revealed significant issues with regard to NCBI’s RefSeq bacterial genome assemblies related to completeness, mutations, structure, strain metadata, and gaps in traceability to the original biological source materials. Nearly half of RefSeq assemblies lack details on sample source information, sequencing technology, or bioinformatics methods. Deep curation of these records is not within the scope of NCBI’s core mission in supporting open science, which aims to collect sequence records that are submitted by the public. Nonetheless, we propose that gaps in metadata accuracy and data provenance represent an “elephant in the room” for microbial genomics research. Effectively addressing these issues will require raising the level of accountability for data depositors and acknowledging the need for higher expectations of quality among the researchers whose research depends on accurate and attributable reference genome data. IMPORTANCE The traceability of microbial genomics data to authenticated physical biological materials is not a requirement for depositing these data into public genome databases. This creates significant risks for the reliability and data provenance of these important genomics research resources, the impact of which is not well understood. We sought to investigate this by carrying out a comparative genomics study of 1,113 ATCC standard reference genomes (ASRGs) produced by ATCC from authenticated and traceable materials using the latest sequencing technologies. We found widespread discrepancies in genome assembly quality, genetic variability, and the quality and completeness of the associated metadata among hundreds of reference genomes for ATCC strains found in NCBI’s RefSeq database. We present a comparative analysis of de novo-assembled ASRGs, their respective metadata, and variant analysis using RefSeq genomes as a reference. Although assembly quality in RefSeq has generally improved over time, we found that significant quality issues remain, especially as related to genomic data and metadata provenance. Our work highlights the importance of data authentication and provenance for the microbial genomics community, and underscores the risks of ignoring this issue in the future.
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Smits THM, Arend LNVS, Cardew S, Tång-Hallbäck E, Mira MT, Moore ERB, Sampaio JLM, Rezzonico F, Pillonetto M. Resolving taxonomic confusion: establishing the genus Phytobacter on the list of clinically relevant Enterobacteriaceae. Eur J Clin Microbiol Infect Dis 2022; 41:547-558. [PMID: 35169969 PMCID: PMC8934334 DOI: 10.1007/s10096-022-04413-8] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2021] [Accepted: 01/25/2022] [Indexed: 11/30/2022]
Abstract
Although many clinically significant strains belonging to the family Enterobacteriaceae fall into a restricted number of genera and species, there is still a substantial number of isolates that elude this classification and for which proper identification remains challenging. With the current improvements in the field of genomics, it is not only possible to generate high-quality data to accurately identify individual nosocomial isolates at the species level and understand their pathogenic potential but also to analyse retrospectively the genome sequence databases to identify past recurrences of a specific organism, particularly those originally published under an incorrect or outdated taxonomy. We propose a general use of this approach to classify further clinically relevant taxa, i.e., Phytobacter spp., that have so far gone unrecognised due to unsatisfactory identification procedures in clinical diagnostics. Here, we present a genomics and literature-based approach to establish the importance of the genus Phytobacter as a clinically relevant member of the Enterobacteriaceae family.
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Affiliation(s)
- Theo H M Smits
- Environmental Genomics and Systems Biology Research Group, Institute of Natural Resource Sciences (IUNR), Zurich University of Applied Sciences ZHAW, Wädenswil, Switzerland.
| | - Lavinia N V S Arend
- Central Public Health Laboratory - State of Paraná - LACEN/PR, Molecular Bacteriology Division, São José Dos Pinhais, PR, Brazil
| | - Sofia Cardew
- Culture Collection University of Gothenburg (CCUG), Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland, Gothenburg, Sweden
| | - Erika Tång-Hallbäck
- Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland, Gothenburg, Sweden
| | - Marcelo T Mira
- Core for Advanced Molecular Investigation, Graduate Program in Health Sciences, School of Medicine, Pontifícia Universidade Católica Do Paraná, Curitiba, PR, Brazil
| | - Edward R B Moore
- Culture Collection University of Gothenburg (CCUG), Department of Clinical Microbiology, Sahlgrenska University Hospital, Region Västra Götaland, Gothenburg, Sweden.,Department of Infectious Disease, Institute for Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden
| | - Jorge L M Sampaio
- Faculdade de Ciências Farmacêuticas - University of São Paulo and Fleury Medicina Diagnóstica, São Paulo, SP, Brazil
| | - Fabio Rezzonico
- Environmental Genomics and Systems Biology Research Group, Institute of Natural Resource Sciences (IUNR), Zurich University of Applied Sciences ZHAW, Wädenswil, Switzerland
| | - Marcelo Pillonetto
- Central Public Health Laboratory - State of Paraná - LACEN/PR, Molecular Bacteriology Division, São José Dos Pinhais, PR, Brazil. .,Core for Advanced Molecular Investigation, Graduate Program in Health Sciences, School of Medicine, Pontifícia Universidade Católica Do Paraná, Curitiba, PR, Brazil.
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23
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Sharma P, Singh SP. Identification and profiling of microbial community from industrial sludge. Arch Microbiol 2022; 204:234. [PMID: 35362813 DOI: 10.1007/s00203-022-02831-y] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2021] [Revised: 03/05/2022] [Accepted: 03/06/2022] [Indexed: 02/03/2023]
Abstract
The purpose of this study is to identify microbial communities in pulp and paper industry sludge and their metagenomic profiling on the basis of; phylum, class, order, family, genus and species level. Results revealed that the dominant phyla in 16S rRNA Illumina Miseq analysis inside sludge were Anaerolinea, Pseudomonas, Clostridia, Bacteriodia, Gammaproteobacteria, Spirochetia, Deltaproteobacteria, Spirochaetaceae, Prolixibacteraceae and some unknown microbial strains are also dominant. Metagenomics is a molecular biology-based technology that uses bioinformatics to evaluate huge gene sequences extracted from environmental samples to assess the composition and function of microbiota. The results of metabarcoding of the V3-V4 16S rRNA regions acquired from paired-end Illumina MiSeq sequencing were used to analyze bacterial communities and structure. The present work demonstrates the potential approach to sludge treatment in the open environment via the naturally adapted microorganism, which could be an essential addition to the disposal site. In summary, these investigations indicate that the indigenous microbial community is an acceptable bioresource for remediation or detoxification following secondary treatment. This research aims at understanding the structure of microbial communities and their diversity (%) in highly contaminated sludge to perform in situ bioremediation.
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Affiliation(s)
- Pooja Sharma
- Department of Environmental Microbiology, School for Environmental Sciences, Babasaheb Bhimrao Ambedkar (A Central) University, Lucknow, 226 025, Uttar Pradesh, India.
| | - Surendra Pratap Singh
- Plant Molecular Biology Laboratory, Department of Botany, Dayanand Anglo-Vedic (PG) College, Chhatrapati Shahu Ji Maharaj University, Kanpur, 208 001, India.
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Khullar G, Det-udom R, Prombutar P, Prakitchaiwattana C. Probiogenomic analysis and safety assessment of Bacillus isolates using Omics approach in combination with In-vitro. Lebensm Wiss Technol 2022. [DOI: 10.1016/j.lwt.2022.113216] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
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Comparative Genomics across Three Ensifer Species Using a New Complete Genome Sequence of the Medicago Symbiont Sinorhizobium ( Ensifer) meliloti WSM1022. Microorganisms 2021; 9:microorganisms9122428. [PMID: 34946030 PMCID: PMC8706082 DOI: 10.3390/microorganisms9122428] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2021] [Revised: 11/11/2021] [Accepted: 11/18/2021] [Indexed: 11/17/2022] Open
Abstract
Here, we report an improved and complete genome sequence of Sinorhizobium (Ensifer) meliloti strain WSM1022, a microsymbiont of Medicago species, revealing its tripartite structure. This improved genome sequence was generated combining Illumina and Oxford nanopore sequencing technologies to better understand the symbiotic properties of the bacterium. The 6.75 Mb WSM1022 genome consists of three scaffolds, corresponding to a chromosome (3.70 Mb) and the pSymA (1.38 Mb) and pSymB (1.66 Mb) megaplasmids. The assembly has an average GC content of 62.2% and a mean coverage of 77X. Genome annotation of WSM1022 predicted 6058 protein coding sequences (CDSs), 202 pseudogenes, 9 rRNAs (3 each of 5S, 16S, and 23S), 55 tRNAs, and 4 ncRNAs. We compared the genome of WSM1022 to two other rhizobial strains, closely related Sinorhizobium (Ensifer) meliloti Sm1021 and Sinorhizobium (Ensifer) medicae WSM419. Both WSM1022 and WSM419 species are high-efficiency rhizobial strains when in symbiosis with Medicago truncatula, whereas Sm1021 is ineffective. Our findings report significant genomic differences across the three strains with some similarities between the meliloti strains and some others between the high efficiency strains WSM1022 and WSM419. The addition of this high-quality rhizobial genome sequence in conjunction with comparative analyses will help to unravel the features that make a rhizobial symbiont highly efficient for nitrogen fixation.
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Staphylococcus aureus Genomes Harbor Only MpsAB-Like Bicarbonate Transporter but Not Carbonic Anhydrase as Dissolved Inorganic Carbon Supply System. Microbiol Spectr 2021; 9:e0097021. [PMID: 34730408 PMCID: PMC8567241 DOI: 10.1128/spectrum.00970-21] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
In recent years, it became apparent that not only autotrophic but also most other bacteria require CO2 or bicarbonate for growth. Two systems are available for the acquisition of dissolved inorganic carbon supply (DICS): the cytoplasmic localized carbonic anhydrase (CA) and the more recently described bicarbonate transporter MpsAB (membrane potential generating system). In the pathogenic species Staphylococcus aureus, there are contradictions in the literature regarding the presence of a CA or MpsAB. Here, we address these contradictions in detail. We could demonstrate by careful BLASTp analyses with 259 finished and 4,590 unfinished S. aureus genomes that S. aureus does not contain CA and that the bicarbonate transporter MpsAB is the only DICS system in this species. This finding is further supported by two further pieces of evidence: (i) mpsAB deletion mutants in four different S. aureus strains failed to grow under atmospheric air, which should not be the case if they possess CAs, since we have previously shown that both CA and MpsAB can substitute for each other, and (ii) S. aureus is completely resistant to CA inhibitors, whereas Staphylococcus carnosus, which has been shown to have only CA, was inhibited by ethoxyzolamide (EZA). Taken together, we demonstrate beyond doubt that the species S. aureus possesses only the bicarbonate transporter MpsAB as its sole DICS system. IMPORTANCE The discrepancies in the current literature and even in NCBI database, which listed some protein sequences annotated as Staphylococcus aureus carbonic anhydrase (CA), are misleading. One of the existing problems in publicly available sequence databases is the presence of incorrectly annotated genes, especially if they originated from unfinished genomes. Here, we demonstrate that some of these unfinished genomes are of poor quality and should be interpreted with caution. In the present study, we aimed to address these discrepancies and correct the current literature about S. aureus CA, considering the medical relevance of S. aureus. If left unchecked, these misleading studies and wrongly annotated genes might lead to a continual propagation of wrong annotation and, consequently, wrong interpretations and wasted time. In addition, we also show that bicarbonate transporter MpsAB-harboring bacteria are resistant to CA inhibitor, suggesting that pathogens possessing both MpsAB and CA are not treatable with CA inhibitors.
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Moretti C, Rezzonico F, Orfei B, Cortese C, Moreno‐Pérez A, van den Burg HA, Onofri A, Firrao G, Ramos C, Smits THM, Buonaurio R. Synergistic interaction between the type III secretion system of the endophytic bacterium Pantoea agglomerans DAPP-PG 734 and the virulence of the causal agent of olive knot Pseudomonas savastanoi pv. savastanoi DAPP-PG 722. MOLECULAR PLANT PATHOLOGY 2021; 22:1209-1225. [PMID: 34268839 PMCID: PMC8435235 DOI: 10.1111/mpp.13105] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/13/2021] [Revised: 06/09/2021] [Accepted: 06/16/2021] [Indexed: 05/29/2023]
Abstract
The endophytic bacterium Pantoea agglomerans DAPP-PG 734 was previously isolated from olive knots caused by infection with Pseudomonas savastanoi pv. savastanoi DAPP-PG 722. Whole-genome analysis of this P. agglomerans strain revealed the presence of a Hypersensitive response and pathogenicity (Hrp) type III secretion system (T3SS). To assess the role of the P. agglomerans T3SS in the interaction with P. savastanoi pv. savastanoi, we generated independent knockout mutants in three Hrp genes of the P. agglomerans DAPP-PG 734 T3SS (hrpJ, hrpN, and hrpY). In contrast to the wildtype control, all three mutants failed to cause a hypersensitive response when infiltrated in tobacco leaves, suggesting that P. agglomerans T3SS is functional and injects effector proteins in plant cells. In contrast to P. savastanoi pv. savastanoi DAPP-PG 722, the wildtype strain P. agglomerans DAPP-PG 734 and its Hrp T3SS mutants did not cause olive knot disease in 1-year-old olive plants. Coinoculation of P. savastanoi pv. savastanoi with P. agglomerans wildtype strains did not significantly change the knot size, while the DAPP-PG 734 hrpY mutant induced a significant decrease in knot size, which could be complemented by providing hrpY on a plasmid. By epifluorescence microscopy and confocal laser scanning microscopy, we found that the localization patterns in knots were nonoverlapping for P. savastanoi pv. savastanoi and P. agglomerans when coinoculated. Our results suggest that suppression of olive plant defences mediated by the Hrp T3SS of P. agglomerans DAPP-PG 734 positively impacts the virulence of P. savastanoi pv. savastanoi DAPP-PG 722.
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Affiliation(s)
- Chiaraluce Moretti
- Dipartimento di Scienze Agrarie, Alimentari e AmbientaliUniversità degli Studi di PerugiaPerugiaItaly
| | - Fabio Rezzonico
- Environmental Genomics and Systems Biology Research GroupInstitute of Natural Resource SciencesZurich University of Applied Sciences ZHAWWädenswilSwitzerland
| | - Benedetta Orfei
- Dipartimento di Scienze Agrarie, Alimentari e AmbientaliUniversità degli Studi di PerugiaPerugiaItaly
| | - Chiara Cortese
- Dipartimento di Scienze Agrarie, Alimentari e AmbientaliUniversità degli Studi di PerugiaPerugiaItaly
| | - Alba Moreno‐Pérez
- Área de GenéticaFacultad de CienciasUniversidad de MálagaMálagaSpain
- Instituto de Hortofruticultura Subtropical y Mediterránea “La Mayora”Universidad de Málaga‐Consejo Superior de Investigaciones CientíficasMálagaSpain
| | - Harrold A. van den Burg
- Molecular Plant PathologySwammerdam Institute for Life SciencesUniversity of AmsterdamAmsterdamNetherlands
| | - Andrea Onofri
- Dipartimento di Scienze Agrarie, Alimentari e AmbientaliUniversità degli Studi di PerugiaPerugiaItaly
| | - Giuseppe Firrao
- Dipartimento di Scienze Agroalimentati Ambientali e AnimaliUniversità degli Studi di UdineUdineItaly
| | - Cayo Ramos
- Área de GenéticaFacultad de CienciasUniversidad de MálagaMálagaSpain
- Instituto de Hortofruticultura Subtropical y Mediterránea “La Mayora”Universidad de Málaga‐Consejo Superior de Investigaciones CientíficasMálagaSpain
| | - Theo H. M. Smits
- Environmental Genomics and Systems Biology Research GroupInstitute of Natural Resource SciencesZurich University of Applied Sciences ZHAWWädenswilSwitzerland
| | - Roberto Buonaurio
- Dipartimento di Scienze Agrarie, Alimentari e AmbientaliUniversità degli Studi di PerugiaPerugiaItaly
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28
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Hahn EE, Alexander MR, Grealy A, Stiller J, Gardiner DM, Holleley CE. Unlocking inaccessible historical genomes preserved in formalin. Mol Ecol Resour 2021; 22:2130-2147. [PMID: 34549888 DOI: 10.1111/1755-0998.13505] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2021] [Revised: 09/08/2021] [Accepted: 09/10/2021] [Indexed: 11/27/2022]
Abstract
Museum specimens represent an unparalleled record of historical genomic data. However, the widespread practice of formalin preservation has thus far impeded genomic analysis of a large proportion of specimens. Limited DNA sequencing from formalin-preserved specimens has yielded low genomic coverage with unpredictable success. We set out to refine sample processing methods and to identify specimen characteristics predictive of sequencing success. With a set of taxonomically diverse specimens collected between 1962 and 2006 and ranging in preservation quality, we compared the efficacy of several end-to-end whole genome sequencing workflows alongside a k-mer-based trimming-free read alignment approach to maximize mapping of endogenous sequence. We recovered complete mitochondrial genomes and up to 3× nuclear genome coverage from formalin-preserved tissues. Hot alkaline lysis coupled with phenol-chloroform extraction out-performed proteinase K digestion in recovering DNA, while library preparation method had little impact on sequencing success. The strongest predictor of DNA yield was overall specimen condition, which additively interacts with preservation conditions to accelerate DNA degradation. Here, we demonstrate a significant advance in capability beyond limited recovery of a small number of loci via PCR or target-capture sequencing. To facilitate strategic selection of suitable specimens for genomic sequencing, we present a decision-making framework that utilizes independent and nondestructive assessment criteria. Sequencing of formalin-preserved specimens will contribute to a greater understanding of temporal trends in genetic adaptation, including those associated with a changing climate. Our work enhances the value of museum collections worldwide by unlocking genomes of specimens that have been disregarded as a valid molecular resource.
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Affiliation(s)
- Erin E Hahn
- National Research Collections Australia, Commonwealth Scientific Industrial Research Organisation, Canberra, ACT, Australia
| | - Marina R Alexander
- National Research Collections Australia, Commonwealth Scientific Industrial Research Organisation, Canberra, ACT, Australia
| | - Alicia Grealy
- National Research Collections Australia, Commonwealth Scientific Industrial Research Organisation, Canberra, ACT, Australia
| | - Jiri Stiller
- Agriculture and Food, Commonwealth Scientific Industrial Research Organisation, St Lucia, Qld, Australia
| | - Donald M Gardiner
- Agriculture and Food, Commonwealth Scientific Industrial Research Organisation, St Lucia, Qld, Australia
| | - Clare E Holleley
- National Research Collections Australia, Commonwealth Scientific Industrial Research Organisation, Canberra, ACT, Australia
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29
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Soldatou S, Eldjárn GH, Ramsay A, van der Hooft JJJ, Hughes AH, Rogers S, Duncan KR. Comparative Metabologenomics Analysis of Polar Actinomycetes. Mar Drugs 2021; 19:103. [PMID: 33578887 PMCID: PMC7916644 DOI: 10.3390/md19020103] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/21/2021] [Revised: 02/05/2021] [Accepted: 02/08/2021] [Indexed: 12/16/2022] Open
Abstract
Biosynthetic and chemical datasets are the two major pillars for microbial drug discovery in the omics era. Despite the advancement of analysis tools and platforms for multi-strain metabolomics and genomics, linking these information sources remains a considerable bottleneck in strain prioritisation and natural product discovery. In this study, molecular networking of the 100 metabolite extracts derived from applying the OSMAC approach to 25 Polar bacterial strains, showed growth media specificity and potential chemical novelty was suggested. Moreover, the metabolite extracts were screened for antibacterial activity and promising selective bioactivity against drug-persistent pathogens such as Klebsiella pneumoniae and Acinetobacter baumannii was observed. Genome sequencing data were combined with metabolomics experiments in the recently developed computational approach, NPLinker, which was used to link BGC and molecular features to prioritise strains for further investigation based on biosynthetic and chemical information. Herein, we putatively identified the known metabolites ectoine and chrloramphenicol which, through NPLinker, were linked to their associated BGCs. The metabologenomics approach followed in this study can potentially be applied to any large microbial datasets for accelerating the discovery of new (bioactive) specialised metabolites.
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Affiliation(s)
- Sylvia Soldatou
- Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow G4 0RE, UK; (S.S.); (A.H.H.)
| | | | - Andrew Ramsay
- School of Computing Science, University of Glasgow, Glasgow G12 8RZ, UK; (G.H.E.); (A.R.); (S.R.)
| | | | - Alison H. Hughes
- Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow G4 0RE, UK; (S.S.); (A.H.H.)
| | - Simon Rogers
- School of Computing Science, University of Glasgow, Glasgow G12 8RZ, UK; (G.H.E.); (A.R.); (S.R.)
| | - Katherine R. Duncan
- Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow G4 0RE, UK; (S.S.); (A.H.H.)
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Amatore Z, Gunn S, Harris LK. An Educational Bioinformatics Project to Improve Genome Annotation. Front Microbiol 2020; 11:577497. [PMID: 33365016 PMCID: PMC7750189 DOI: 10.3389/fmicb.2020.577497] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2020] [Accepted: 10/27/2020] [Indexed: 01/28/2023] Open
Abstract
Scientific advancement is hindered without proper genome annotation because biologists lack a complete understanding of cellular protein functions. In bacterial cells, hypothetical proteins (HPs) are open reading frames with unknown functions. HPs result from either an outdated database or insufficient experimental evidence (i.e., indeterminate annotation). While automated annotation reviews help keep genome annotation up to date, often manual reviews are needed to verify proper annotation. Students can provide the manual review necessary to improve genome annotation. This paper outlines an innovative classroom project that determines if HPs have outdated or indeterminate annotation. The Hypothetical Protein Characterization Project uses multiple well-documented, freely available, web-based, bioinformatics resources that analyze an amino acid sequence to (1) detect sequence similarities to other proteins, (2) identify domains, (3) predict tertiary structure including active site characterization and potential binding ligands, and (4) determine cellular location. Enough evidence can be generated from these analyses to support re-annotation of HPs or prioritize HPs for experimental examinations such as structural determination via X-ray crystallography. Additionally, this paper details several approaches for selecting HPs to characterize using the Hypothetical Protein Characterization Project. These approaches include student- and instructor-directed random selection, selection using differential gene expression from mRNA expression data, and selection based on phylogenetic relations. This paper also provides additional resources to support instructional use of the Hypothetical Protein Characterization Project, such as example assignment instructions with grading rubrics, links to training videos in YouTube, and several step-by-step example projects to demonstrate and interpret the range of achievable results that students might encounter. Educational use of the Hypothetical Protein Characterization Project provides students with an opportunity to learn and apply knowledge of bioinformatic programs to address scientific questions. The project is highly customizable in that HP selection and analysis can be specifically formulated based on the scope and purpose of each student's investigations. Programs used for HP analysis can be easily adapted to course learning objectives. The project can be used in both online and in-seat instruction for a wide variety of undergraduate and graduate classes as well as undergraduate capstone, honor's, and experiential learning projects.
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Affiliation(s)
- Zoie Amatore
- Science Department, Harris Interdisciplinary Research, Davenport University, Lansing, MI, United States
| | - Susan Gunn
- College of Urban Education, Davenport University, Grand Rapids, MI, United States
| | - Laura K. Harris
- Science Department, Harris Interdisciplinary Research, Davenport University, Lansing, MI, United States
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Yu M, Singh J, Khan A, Sundin GW, Zhao Y. Complete Genome Sequence of the Fire Blight Pathogen Strain Erwinia amylovora Ea1189. MOLECULAR PLANT-MICROBE INTERACTIONS : MPMI 2020; 33:1277-1279. [PMID: 32808873 DOI: 10.1094/mpmi-06-20-0158-a] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/08/2023]
Abstract
Erwinia amylovora causes fire blight, the most devastating bacterial disease of apples and pears in the United States and worldwide. The model strain E. amylovora Ea1189 has been extensively used to understand bacterial pathogenesis and molecular mechanisms of bacterial-plant interactions. In this work, we sequenced and assembled the de novo genome of Ea1189, using a combination of long Oxford Nanopore Technologies and short Illumina sequence reads. A complete gapless genome assembly of Ea1189 consists of a 3,797,741-bp circular chromosome and a 28,259-bp plasmid with 3,472 predicted genes, including 78 transfer RNAs, 22 ribosomal RNAs, and 20 noncoding RNAs. A comparison of the Ea1189 genome to previously sequenced E. amylovora complete genomes showed 99.94 to 99.97% sequence similarity with 314 to 946 single nucleotide polymorphisms. We believe that the availability of the complete genome sequence of strain Ea1189 will further support studies to understand evolution, diversity and structural variations of Erwinia strains, as well as the molecular basis of E. amylovora pathogenesis and its interactions with host plants, thus facilitating the development of effective management strategies for this important disease.
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Affiliation(s)
- Menghao Yu
- Department of Crop Sciences, University of Illinois at Urbana-Champaign, IL 61801, U.S.A
| | - Jugpreet Singh
- Plant Pathology and Plant-Microbe Biology Section, Cornell University, Geneva, NY 14456, U.S.A
| | - Awais Khan
- Plant Pathology and Plant-Microbe Biology Section, Cornell University, Geneva, NY 14456, U.S.A
| | - George W Sundin
- Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI 48824, U.S.A
| | - Youfu Zhao
- Department of Crop Sciences, University of Illinois at Urbana-Champaign, IL 61801, U.S.A
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Complete or High-Quality Draft Genome Sequences of Six Xanthomonas hortorum Strains Sequenced with Short- and Long-Read Technologies. Microbiol Resour Announc 2020; 9:9/41/e00828-20. [PMID: 33033126 PMCID: PMC7545280 DOI: 10.1128/mra.00828-20] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2023] Open
Abstract
We report the genome sequences of six Xanthomonas hortorum species-level clade members, X. hortorum pathovars taraxaci, pelargonii, cynarae, and gardneri (complete genome sequences) and X. hortorum pathovars carotae and vitians (high-quality draft genome sequences). Both short- and long-read sequencing technologies were used.
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33
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Khan M, Stapleton F, Summers S, Rice SA, Willcox MDP. Antibiotic Resistance Characteristics of Pseudomonas aeruginosa Isolated from Keratitis in Australia and India. Antibiotics (Basel) 2020; 9:antibiotics9090600. [PMID: 32937932 PMCID: PMC7559795 DOI: 10.3390/antibiotics9090600] [Citation(s) in RCA: 25] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2020] [Revised: 09/08/2020] [Accepted: 09/09/2020] [Indexed: 02/07/2023] Open
Abstract
This study investigated genomic differences in Australian and Indian Pseudomonas aeruginosa isolates from keratitis (infection of the cornea). Overall, the Indian isolates were resistant to more antibiotics, with some of those isolates being multi-drug resistant. Acquired genes were related to resistance to fluoroquinolones, aminoglycosides, beta-lactams, macrolides, sulphonamides, and tetracycline and were more frequent in Indian (96%) than in Australian (35%) isolates (p = 0.02). Indian isolates had large numbers of gene variations (median 50,006, IQR = 26,967-50,600) compared to Australian isolates (median 26,317, IQR = 25,681-33,780). There were a larger number of mutations in the mutL and uvrD genes associated with the mismatch repair (MMR) system in Indian isolates, which may result in strains losing their efficacy for DNA repair. The number of gene variations were greater in isolates carrying MMR system genes or exoU. In the phylogenetic division, the number of core genes were similar in both groups, but Indian isolates had larger numbers of pan genes (median 6518, IQR = 6040-6935). Clones related to three different sequence types-ST308, ST316, and ST491-were found among Indian isolates. Only one clone, ST233, containing two strains was present in Australian isolates. The most striking differences between Australian and Indian isolates were carriage of exoU (that encodes a cytolytic phospholipase) in Indian isolates and exoS (that encodes for GTPase activator activity) in Australian isolates, large number of acquired resistance genes, greater changes to MMR genes, and a larger pan genome as well as increased overall genetic variation in the Indian isolates.
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Affiliation(s)
- Mahjabeen Khan
- School of Optometry and Vision Science, UNSW, Sydney, NSW 2052, Australia; (M.K.); (F.S.)
| | - Fiona Stapleton
- School of Optometry and Vision Science, UNSW, Sydney, NSW 2052, Australia; (M.K.); (F.S.)
| | - Stephen Summers
- The Singapore Centre for Environment Life Sciences Engineering (SCELSE), The School of Biological Sciences, Nanyang Technological University, Singapore 639798, Singapore; (S.S.); (S.A.R.)
| | - Scott A. Rice
- The Singapore Centre for Environment Life Sciences Engineering (SCELSE), The School of Biological Sciences, Nanyang Technological University, Singapore 639798, Singapore; (S.S.); (S.A.R.)
- The ithree Institute, The University of Technology Sydney, Sydney, NSW 2007, Australia
| | - Mark D. P. Willcox
- School of Optometry and Vision Science, UNSW, Sydney, NSW 2052, Australia; (M.K.); (F.S.)
- Correspondence: ; Tel.: +61-2-9385-4164
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Hulin MT, Jackson RW, Harrison RJ, Mansfield JW. Cherry picking by pseudomonads: After a century of research on canker, genomics provides insights into the evolution of pathogenicity towards stone fruits. PLANT PATHOLOGY 2020; 69:962-978. [PMID: 32742023 PMCID: PMC7386918 DOI: 10.1111/ppa.13189] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/04/2019] [Revised: 03/09/2020] [Accepted: 03/23/2020] [Indexed: 05/10/2023]
Abstract
Bacterial canker disease is a major limiting factor in the growing of cherry and other Prunus species worldwide. At least five distinct clades within the bacterial species complex Pseudomonas syringae are known to be causal agents of the disease. The different pathogens commonly coexist in the field. Reducing canker is a challenging prospect as the efficacy of chemical controls and host resistance may vary against each of the diverse clades involved. Genomic analysis has revealed that the pathogens use a variable repertoire of virulence factors to cause the disease. Significantly, strains of P. syringae pv. syringae possess more genes for toxin biosynthesis and fewer encoding type III effector proteins. There is also a shared pool of key effector genes present on mobile elements such as plasmids and prophages that may have roles in virulence. By contrast, there is evidence that absence or truncation of certain effector genes, such as hopAB, is characteristic of cherry pathogens. Here we highlight how recent research, underpinned by the earlier epidemiological studies, is allowing significant progress in our understanding of the canker pathogens. This fundamental knowledge, combined with emerging insights into host genetics, provides the groundwork for development of precise control measures and informed approaches to breed for disease resistance.
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Affiliation(s)
| | - Robert W. Jackson
- Birmingham Institute of Forest Research (BIFoR), University of BirminghamBirminghamUK
- School of Biosciences, University of BirminghamBirminghamUK
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Spacova I, Dodiya HB, Happel AU, Strain C, Vandenheuvel D, Wang X, Reid G. Future of Probiotics and Prebiotics and the Implications for Early Career Researchers. Front Microbiol 2020; 11:1400. [PMID: 32714306 PMCID: PMC7344207 DOI: 10.3389/fmicb.2020.01400] [Citation(s) in RCA: 22] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2020] [Accepted: 05/29/2020] [Indexed: 12/24/2022] Open
Abstract
The opportunities in the fields of probiotics and prebiotics to a great degree stem from what we can learn about how they influence the microbiota and interact with the host. We discuss recent insights, cutting-edge technologies and controversial results from the perspective of early career researchers innovating in these areas. This perspective emerged from the 2019 meeting of the International Scientific Association for Probiotics and Prebiotics - Student and Fellows Association (ISAPP-SFA). Probiotic and prebiotic research is being driven by genetic characterization and modification of strains, state-of-the-art in vitro, in vivo, and in silico techniques designed to uncover the effects of probiotics and prebiotics on their targets, and metabolomic tools to identify key molecules that mediate benefits on the host. These research tools offer unprecedented insights into the functionality of probiotics and prebiotics in the host ecosystem. Young scientists need to acquire these diverse toolsets, or form inter-connected teams to perform comprehensive experiments and systematic analysis of data. This will be critical to identify microbial structure and co-dependencies at body sites and determine how administered probiotic strains and prebiotic substances influence the host. This and other strategies proposed in this review will pave the way for translating the health benefits observed during research into real-life outcomes. Probiotic strains and prebiotic products can contribute greatly to the amelioration of global issues threatening society. The intent of this article is to provide an early career researcher's perspective on where the biggest opportunities lie to advance science and impact human health.
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Affiliation(s)
- Irina Spacova
- Laboratory of Applied Microbiology and Biotechnology, Department of Bioscience Engineering, University of Antwerp, Antwerp, Belgium
| | - Hemraj B. Dodiya
- Department of Neurobiology, The University of Chicago, Chicago, IL, United States
| | - Anna-Ursula Happel
- Division of Immunology, Department of Pathology, Institute of Infectious Disease & Molecular Medicine, University of Cape Town, Cape Town, South Africa
| | - Conall Strain
- APC Microbiome Ireland, University College Cork, Cork, Ireland
- Teagasc Food Research Centre, Fermoy, Ireland
| | - Dieter Vandenheuvel
- Laboratory of Applied Microbiology and Biotechnology, Department of Bioscience Engineering, University of Antwerp, Antwerp, Belgium
- Institute for Biomedical Sciences, Shinshu University, Nagano, Japan
| | - Xuedan Wang
- Department of Zoology, University of Oxford, Oxford, United Kingdom
- Department of Biochemistry, University of Oxford, Oxford, United Kingdom
| | - Gregor Reid
- Centre for Human Microbiome and Probiotic Research, Lawson Health Research Institute, London, ON, Canada
- Department of Microbiology and Immunology, The University of Western Ontario, London, ON, Canada
- Department of Surgery, The University of Western Ontario, London, ON, Canada
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