bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.64898/2026.10.09.758105

A tetracycline-inducible promoter for gene expression in Gram-negative bacteria including Acinetobacter baumannii

Stella, N. A.·Ginde, R. K.·Calvario, R. C.·Doi, Y.·Shanks, R. M. Q.
View original source ↗Export metadata ↓

Abstract

Carbohydrate inducible promoters on expression plasmids are widely used for biotechnology and basic science research. However, using carbohydrates for inducers has drawbacks including influencing bacterial metabolism, the inducer being used as a carbon source by the bacteria, high background expression, and only a subset of cells experiencing promoter activation. Furthermore, some bacteria including the pathogen Acinetobacter baumannii currently lack good carbohydrate-inducible promoters. To address this issue, a tetracycline-inducible promoter, PtetB, isolated from A. baumannii was cloned and evaluated in a number of important bacteria: Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Pseudomonas fluorescens, and Serratia marcescens. A highly sensitive luxCDABE reporter was used to evaluate promoter activity. S. marcescens strains with elevated and reduced cyclic-AMP levels were used to evaluate the impact of cAMP on PtetB function. The PtetB promoter was induced over 800-fold in A. baumannii and 10-750-fold inducible in the other tested species. Background expression was lower from PtetB in the pseudomonads than previously described carbohydrate inducible promoters. Data indicated that the PtetB promoter system may be a useful tool for future basic science research with the tested bacteria.

Explore related subjects

microbiologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Stella, N. A., Ginde, R. K., Calvario, R. C., Doi, Y., Shanks, R. M. Q.. 2026-10-10. A tetracycline-inducible promoter for gene expression in Gram-negative bacteria including Acinetobacter baumannii. https://doi.org/10.64898/2026.10.09.758105

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Effects of Alternating Polarity Frequency on the Assembly of Electro-Methanogenic Microbial Communities

Electro-methanogenesis can enhance methane production and has attracted extensive interest, but the current strategies for building electro-methanogenic microbial communities suffer from several limitations. Alternating polarity has been shown as a new strategy to build electro-methanogenic communities, but the impact of alternation frequency on community assembly remains to be understood. In this study, the impact was evaluated using bioelectrochemical systems amended with granular activated carbon, and the working-electrode potential was alternated between +0.4 and -0.4 V vs. SHE at six frequencies ranging from 0.5 to 12 days. Among the tested conditions, 4-d alternation led to the best overall performance, producing approximately 250 L/L/kg cumulative biogas, 96% methane content, and a 45% methane yield, while preserving a stable and reproducible community structure. The community contained two Methanobacterium populations, two putatively electroactive Geobacter populations, and one putatively electroactive Pseudomonas population, together with diverse fermentative and syntrophic bacteria. Transcript-based analyses revealed clear frequency- and potential-dependent shifts in the activity of these functional groups. Redundancy analysis associated the Methanobacterium formicicum-related population with cathodic current and methane production, whereas the Geobacter sulfurreducens-related population was linked to anodic current and methane production. Bayesian and local-similarity networks further connected methanogen dynamics with specific electroactive, fermentative, and syntrophic partners. Collectively, these results establish alternating polarity frequency as a critical parameter for building robust communities capable of high-rate, high-quality methane production.

microbiology↗
bioRxiv2026

Benchmarking Microbiome Methods for Improved Metagenomic Sequencing of Lower Respiratory Tract Samples

Bronchoalveolar lavage is a common diagnostic strategy used to sample immune cells and microbiota of the lower respiratory tract. Metagenomic sequencing of raw bronchoalveolar lavage fluid reveals a low biomass microbial community often obscured by abundant host DNA. Here, we set out to determine an optimal strategy to robustly purify microbial DNA in excess of human DNA using a combinatorial lysis, host-depletion, DNA extraction approach. We identify strategies that improve metagenomic sequencing success that are commensurate with purification challenges that reduce signal from respiratory microbiota. Finally, we present a general framework to further improve microbial sequencing of lower respiratory tract samples in a robust and reproducible manner.

microbiology↗
bioRxiv2026

Shared genomic features of oral bacterial habitat specialization

The oral microbiome exhibits spatial organization, with related bacteria occupying different surfaces despite continual dispersal between habitats. Whether this ecological partitioning is accompanied by genomic functions that recur across evolutionarily divergent lineages remains unclear. Here we compared plaque- and mucosa-associated bacteria representing 47 human microbial taxa, 13 genera and six phyla, using 1,121 reference genome assemblies and habitat-affinity estimates assembled from six oral microbiome abundance summaries. Across 13 genera, a genus-balanced screen of 13,917 nonredundant genomic profiles prioritised 14 exploratory annotations with nominal associations in the same direction before and after adjustment for genome composition with none passing genome-wide family-wise error correction. Proteins carrying recurrent domains belonged to multiple sequence clusters. Closely spaced, divergently oriented HxlR-SnoaL_2 gene pairs occurred in 66 genomes associated with plaque biofilms across four genera, although overall HxlR-SnoaL_2 linkage was not significantly associated with plaque preference. Using source-family-balanced habitat scores, nested cross-validation with entire genera held out supported prediction of habitat preference (macro-average within-genus AUC, 0.821; empirical P = 0.010). Predictive performance was lower after adjustment for genome size, GC content and coding-sequence number (AUC, 0.776; empirical P = 0.059). These findings reveal partial recurrence of habitat-associated genomic annotations amid lineage-dependent protein repertoires and genetic arrangements. They provide a comparative framework for distinguishing broadly shared ecological genomic signatures from alternative, potentially lineage-specific routes to oral habitat specialization.

microbiology↗