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Peptidoglycan synthesis drives a single population of septal cell wall synthases during division in Bacillus subtilis

Kevin D. Whitley, James Grimshaw, David M. Roberts, Eleni Karinou, Phillip J. Stansfeld & Seamus Holden

Bacterial cell division requires septal peptidoglycan (sPG) synthesis by the divisome complex. Treadmilling of the essential tubulin homologue FtsZ has been implicated in septal constriction, though its precise role remains unclear. Here we used live-cell single-molecule imaging of the divisome transpeptidase PBP2B to investigate sPG synthesis dynamics in Bacillus subtilis. In contrast to previous models, we observed a single population of processively moving PBP2B molecules whose motion is driven by peptidoglycan synthesis and is not associated with FtsZ treadmilling. However, despite the asynchronous motions of PBP2B and FtsZ, a partial dependence of PBP2B processivity on FtsZ treadmilling was observed. Additionally, through single-molecule counting experiments we provide evidence that the divisome synthesis complex is multimeric. Our results support a model for B. subtilis division where a multimeric synthesis complex follows a single track dependent on sPG synthesis whose activity and dynamics are asynchronous with FtsZ treadmilling.

Nature Microbiology. March 2024


The contribution of an X chromosome QTL to non-Mendelian inheritance and unequal chromosomal segregation in A. freiburgense

Talal Al-Yazeedi, Sally Adams, Sophie Tandonnet, Anisa Turner, Jun Kim, Junho Lee, Andre Pires-daSilva

Auanema freiburgense is a nematode with males, females, and selfing hermaphrodites. When XO males mate with XX females, they typically produce a low proportion of XO offspring because they eliminate nullo-X spermatids. This occurs because of an unequal distribution of essential cellular organelles during sperm formation. Intriguingly, the interbreeding of two A. freiburgense strains results in hybrid males capable of producing viable nullo-X sperm. Consequently, when these hybrid males mate with females, they yield a high percentage of male offspring. To uncover the genetic basis of nullo-spermatid elimination and X- chromosome drive, we generated a genome assembly for A. freiburgense, and genotyped the intercrossed lines. This analysis identified a Quantitative Trait Locus spanning several X chromosome genes linked to the non-Mendelian inheritance patterns observed in A. freiburgense. This finding provides valuable clues to the underlying factors involved in asymmetric organelle partitioning during male meiotic division and thus non-Mendelian transmission of the X chromosome and sex ratios.

Genetics. March 2024

Thu 21 Mar 2024, 08:21 | Tags: Cells & Development Environment & Ecology

Detection of a historic reservoir of bedaquiline/clofazimine resistance-associated variants in Mycobacterium tuberculosis

Camus Nimmo, Arturo Torres Ortiz, Cedric C. S. Tan, Juanita Pang, Mislav Acman, James Millard, Nesri Padayatchi, Alison D. Grant, Max O’Donnell, Alex Pym, Ola B. Brynildsrud, Vegard Eldholm, Louis Grandjean, Xavier Didelot, François Balloux & Lucy van Dorp

Drug resistance in tuberculosis (TB) poses a major ongoing challenge to public health. The recent inclusion of bedaquiline into TB drug regimens has improved treatment outcomes, but this advance is threatened by the emergence of strains of Mycobacterium tuberculosis (Mtb) resistant to bedaquiline. Clinical bedaquiline resistance is most frequently conferred by off-target resistance-associated variants (RAVs) in the mmpR5 gene (Rv0678), the regulator of an efflux pump, which can also confer cross-resistance to clofazimine, another TB drug. We compiled a dataset of 3682 Mtb genomes, including 180 carrying variants in mmpR5, and its immediate background (i.e. mmpR5 promoter and adjacent mmpL5 gene), that have been associated to borderline (henceforth intermediate) or confirmed resistance to bedaquiline. We characterised the occurrence of all nonsynonymous mutations in mmpR5 in this dataset and estimated, using time-resolved phylogenetic methods, the age of their emergence.

Genome Medicine. February 2024

Mon 18 Mar 2024, 07:58 | Tags: Microbiology & Infectious Disease

Early Detection of Fusarium Basal Rot Infection in Onions and Shallots Based on VOC Profiles Analysis

Malgorzata Wesoly, Emma Daulton, Sascha Jenkins, Sarah van Amsterdam, John Clarkson and James A. Covington

In this work, we employed Gas chromatography ion-mobility spectrometry (GC-IMS) technology for early detection of Fusarium basal rot in brown onion, red onion, and shallot bulbs and for tracking disease progression during storage. The volatile profiles of the infected and healthy control bulbs were characterized using GC-IMS and gas chromatography-time-of-flight mass spectrometry (GC-TOF-MS). GC-IMS data combined with principal component analysis and supervised methods provided discrimination between infected and healthy control bulbs as early as 1 day after incubation with the pathogen, classification regarding the proportion of infected to healthy bulbs in a sample, and prediction of the infection’s duration with an average R2 = 0.92. Furthermore, GC-TOF-MS revealed several compounds, mostly sulfides and disulfides, that could be uniquely related to Fusarium basal rot infection.

Agricultural and Food Chemistry. February 2024

Thu 14 Mar 2024, 08:12 | Tags: Plant & Agricultural Bioscience

The long and short of it: benchmarking viromics using Illumina, Nanopore and PacBio sequencing technologies

Ryan Cook​, Nathan Brown​, Branko Rihtman​, Slawomir Michniewski​, Tamsin Redgwell, Martha Clokie​, Dov J. Stekel, Yin Chen​, David J. Scanlan​, Jon L. Hobman, Andrew Nelson​, Michael A. Jones​, Darren Smith​ and Andrew Millard

ecology. Long-read sequencing and hybrid assembly approaches that combine long- and short-read technologies are now being widely implemented in bacterial genomics and metagenomics. However, the use of long-read sequencing to investigate viral communities is still in its infancy. While Nanopore and PacBio technologies have been applied to viral metagenomics, it is not known to what extent different technologies will impact the reconstruction of the viral community. Thus, we constructed a mock bacteriophage community of previously sequenced phage genomes and sequenced them using Illumina, Nanopore and PacBio sequencing technologies and tested a number of different assembly approaches. Overall the best approach was assembly by a combination of Illumina and Nanopore reads, which reduced error rates to levels comparable with short-read-only assemblies. When using a single technology, Illumina only was the best approach. These findings will provide a starting point for others in the choice of reads and assembly algorithms for the analysis of viromes.

Microbial Genomics. February 2024


Genome assembly and transcriptomic analysis to elucidate the ability of Nasonovia ribisnigri to break host plant resistance

Dion Garrett, Graham Teakle, Rosemary Collier, James R. Bell, Sergio Cerezo-Medina, Ramiro Morales-Hojas

The currant-lettuce aphid (Nasonovia ribisnigri (Hemiptera: Aphididae) (Mosley)) is a cosmopolitan pest of outdoor lettuce. Until recently, the use of resistant cultivars was an effective method for managing N. ribisnigri. A resistant cultivar containing a single gene (Nr-locus), introduced in the 1980s, conferred complete resistance to feeding. Overreliance of this Nr-locus in lettuce resulted in N. ribisnigri's ability to break resistance mechanism. Our work attempts to understand which candidate gene(s) are associated with this resistance-breaking mechanism. We present two de novo draft assembles for N. ribisnigri genomes, corresponding to both avirulent (Nr-locus susceptible) and virulent (Nr-locus resistant) biotypes.. Out of the 18,872 differentially expressed genes, a single gene/locus was identified in N. ribisnigri that was shared between two resistant-breaking biotypes. This locus was further explored and validated in Real-Time Quantitative Reverse Transcription PCR (qRT-PCR) experiments and has predicted localisations in both the cytoplasm and nucleus. This is the first study to provide evidence that a single gene/locus is likely responsible for the ability of N. ribisnigri to overcome the Nr-locus resistance in the lettuce host.

Insect Molecular Biology. February 2024


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