Biology subjects

Jaafar, T.

Publications and source records attributed to Jaafar, T..

4 recordsLinked to original sources

bioRxiv2026

Clinical Campylobacter jejuni isolates: genomes and genetic tools

Campylobacter jejuni is a major cause of food-borne gastroenteritis and is responsible for substantial mortality and economic losses in meat and dairy production. Detecting C. jejuni in contaminated food samples remains difficult because current assays are culture-based, slow, and can yield false positives. As a result, contamination may not be identified for several days, limiting detection at the point of production. Developing improved assays has also been challenging because Campylobacter genetics and the biology of clinical isolates remain poorly understood. Here, we expand the C. jejuni genetic toolbox by sequencing two strains, HC1 and RM1164, derived from patient and food samples. We identified two cryptic plasmids in HC1, one potentially capable of conjugation and another conferring tetracycline resistance. We also engineered a mobilizable plasmid carrying an OriT sequence that can be transferred from Escherichia coli donor strains to C. jejuni RM1164 by conjugation. Together, these clinical isolates and the plasmid system expand the genetic tools available for C. jejuni.

synthetic biology↗

bioRxiv2025

Conjugation-mediated DNA delivery to the filamentous fungus Ustilago maydis

Phytopathogenic fungi are ubiquitous throughout the environment and threaten global food security. This issue is further amplified by the increasing resistance of pathogens to antimicrobials. Current chemical-based antifungals target cells by inhibiting growth or metabolic function, making them ideal for fungal gain of resistance mutations. Biofungicides are a rising class of antifungals that have low potential for negative environmental impact and provide the fungi almost no potential for gaining resistance. Conjugative plasmids which play a role in the natural mechanism of horizontal gene transfer in bacteria, have been repurposed to deliver toxic genetic cargo to recipient cells, showing promise as next-generation antimicrobial agents. In this work, we have demonstrated the first protocol for delivering DNA from Escherichia coli to the filamentous phytopathogen, Ustilago maydis through conjugation. DNA delivery was confirmed using PCR screening of DNA isolated from the re-streaked transconjugants. Although challenges such as reduced conjugation efficiency and extrachromosomal replication persist, this work establishes the first step towards creating a conjugation-based biofungicide.

synthetic biology↗

bioRxiv2025

Resolving replication incompatibility between chloroplast and conjugative plasmids in E. coli

Chloroplast genomes present a promising chassis for engineering photosynthetic eukaryotes, but efficient delivery of large DNA constructs back into the organelle remains a major technical barrier. Conventional transformation methods rely on purified DNA and physical force to drive uptake into the chloroplast, often resulting in DNA shearing and thus low transfer efficiency for large constructs. Bacterial conjugation offers an attractive alternative as this is an entirely in vivo process, enabling DNA transfer without any physical manipulation. To assess the feasibility of this approach, we attempted to generate an Escherichia coli donor strain carrying both the broad-host-range conjugative plasmid pTA-Mob (52.7 kb) and the cloned Phaeodactylum tricornutum chloroplast genome (pPt_Cp, 132.9 kb). Unexpectedly, pTA-Mob and pPt_Cp proved incompatible: co-maintenance could not be achieved via electroporation, and conjugation with a self-transmissible pTA-Mob variant resulted in a [~]10{square}-fold decrease in transfer efficiency. Systematic testing of pTA-Mob and pPt_Cp plasmid variants as well as sequence analysis of evolved transconjugants revealed that the incompatibility arose from the pBBR1 replicon present in pTA-Mob. Guided by these insights, we identified an alternative conjugative plasmid, pRL443, that was compatible with pPt_Cp. Together, these findings provide a framework for dissecting plasmid incompatibility when working with large constructs and establish a functional conjugative system capable of mobilizing the P. tricornutum chloroplast genome.

synthetic biology↗

bioRxiv2025

Engineering Conjugative Plasmids for Inducible Horizontal DNA Transfer

Rapidly developing microbial resistance to existing antimicrobials poses a growing threat to public health and global food security. Current chemical-based treatments target cells by inhibiting growth or metabolic function, but their effectiveness is diminishing. To address the growing antimicrobial resistance crisis, there is an urgent need for innovative therapies. Conjugative plasmids, a natural mechanism of horizontal gene transfer in bacteria, have been repurposed to deliver toxic genetic cargo to recipient cells, showing promise as next-generation antimicrobial agents. However, the ecological risks posed by unintended gene transfer require robust biocontainment strategies. In this study, we developed inducible conjugative plasmids to solve these challenges. Utilizing an arabinose-inducible promoter, we evaluated 13 plasmids with single essential gene deletions, identifying trbC and trbF as strong candidates for stringent regulation. These plasmids demonstrated inducibility in both cis and trans configurations, with induction resulting in up to a 5-log increase in conjugation efficiency compared to uninduced conditions. Although challenges such as reduced conjugation efficiency and promoter leakiness persist, this work establishes a foundation for the controlled transfer of plasmids, paving the way for safer and more effective antimicrobial technologies.

synthetic biology↗