Biology subjects

Romling, U.

Publications and source records attributed to Romling, U..

7 recordsLinked to original sources

bioRxiv2026

Functional diversification of two Lon homologs enhances stress adaptation in Pseudomonas aeruginosa

The Lon protease is a highly conserved ATP-dependent protease that contributes to protein quality control and regulatory processes across all domains of life. The opportunistic pathogen Pseudomonas aeruginosa, along with other members of the Pseudomonadales, encodes AsrA (aminoglycoside-induced stress response ATP-dependent protease), a second LonA-type protease in addition to canonical Lon. Although AsrA is upregulated by aminoglycoside stress, its biochemical activity, substrate spectrum, and cellular functions have remained elusive. Here, we demonstrate that AsrA is a temperature- and antimicrobial stress-induced protease with specialized functions and partial redundancy with Lon. Quantitative proteomics revealed a distinct AsrA substrate profile, while comparative biochemical analyses showed that AsrA and Lon share a substantial number of substrates in vitro but differ in their degradation kinetics, indicating divergent substrate preferences. Among the proteins preferentially degraded by AsrA is the quorum-sensing anti-activator QslA, and we show that AsrA-dependent QslA degradation under tobramycin stress induces quorum-sensing gene expression. Together, our findings demonstrate how duplication of a conserved protease can generate specialized regulatory functions through differential expression and substrate preference, expanding the proteolytic network that enables bacterial adaptation to stress.

microbiology↗

bioRxiv2026

Regulatory plasticity of the rdar biofilm morphotype in clinical uropathogenic Escherichia coli and its modulation by ciprofloxacin

Red, dry and rough (rdar) biofilm formation in Escherichia coli is characterized by the coordinated production of extracellular cellulose and amyloid curli fimbriae. Although rdar biofilm formation has been extensively characterized in laboratory strains, its clinical relevance and implications for antimicrobial treatment remain poorly understood. Here, we systematically investigate rdar biofilm formation of 150 consecutively isolated E. coli strains recovered from patients with urinary tract infections and correlate it to antimicrobial resistance. Genetic analysis of rdar regulation revealed distinct nucleotide signatures within the csgD promoter region that discriminate semi-constitutive rdar expression from temperature-dependent phenotypes, highlighting regulatory plasticity among clinical isolates. Whole-genome sequencing-based phylogenetic analysis further demonstrated that rdar-positive isolates are distributed across diverse E. coli phylogroups and sequence types, indicating that rdar biofilm formation is not restricted to specific clonal lineages. Strikingly, phenotypic assays revealed that the fluoroquinolone antibiotic ciprofloxacin suppresses rdar biofilm formation and associated extracellular matrix architecture in ciprofloxacin-resistant isolates at subinhibitory concentrations, suggesting that ciprofloxacin modulates biofilm-associated pathways beyond its canonical bactericidal targets. Together, our findings establish the rdar morphotype as a clinically relevant biofilm phenotype in uropathogenic E. coli and reveal an antibiofilm activity of ciprofloxacin that is uncoupled from antibiotic resistance. These results underscore the importance of considering antibiotic-mediated modulation of biofilm behavior when interpreting treatment responses and designing strategies to combat persistent urinary tract infections.

microbiology↗

bioRxiv2025

Diverse horizontally transferred cellulose biosynthesis gene clusters in Escherichia coli strains

The phosphoethanolamine modified exopolysaccharide cellulose is a major extracellular matrix component of Escherichia coli and Salmonella typhimurium. Upon enhanced acute virulence, however, cellulose production can be diminished or entirely abolished. Here, we report that homologs of the bcsABC core genes of the core genome bcs cellulose biosynthesis operon and even entire bcs operons can be mobilized on plasmids and are occasionally manifested on E. coli chromosomes at alternative locations. While BcsA2 and BcsA3 cellulose synthases are restricted to the genus Escherichia, the BcsA4 cellulose synthase and the entire associated bcs gene cluster are highly similar to one of the two chromosomal bcs gene clusters harboured by Klebsiella pneumoniae. Cyclic di-GMP turnover proteins known to post-translationally regulate cellulose biosynthesis are frequently localized on bcs operon bearing plasmids. In thermotolerant meat derived E. coli 740v1 harbouring a type 4 bcs operon on a plasmid, chemical and genetic evidence indicates that cellulose is produced in M9 minimal medium and upon activation by the second messenger cyclic di-GMP, an allosteric activator of the cellulose synthase, respectively. With gene duplication and horizontal gene transfer to contribute to the multiplication of bcs operons in a number of different bacterial species, the ecological role(s) of multiplication, transfer and replacement of cellulose biosynthesis operons also in species other than E. coli still need to be unraveled.

genomics↗

bioRxiv2024

Previously uncharacterized aliphatic amino acid positions modulate the apparent catalytic activity of the EAL domain of ZMO_1055 and other cyclic di-GMP specific EAL phosphodiesterases

The ubiquitous second messenger cyclic di-GMP signaling system decides bacterial life style transition between sessility and motility. GGDEF diguanylate cyclase and EAL phosphodiesterase domains conventionally direct the turnover of this signaling molecule being subject to micro- and macroevolution. While the highly conserved signature amino acids involved in divalent ion binding and catalysis have readily been identified, recognition of amino acid substitutions that modulate the catalytic activity is rare. Associated with development towards cellulose-mediated self-flocculation in Zymomonas mobilis ZM401, the A526V substitution, previously not been recognized to affect the functionality of the EAL domain, downregulates the apparent catalytic activity of the PAS-GGDEF-EAL ZMO1055 phosphodiesterase compared to parental Z. mobilis ZM4 and equally in homologous protein domains independently of the genetic background. Substitution of A526, which is conserved among homologs, with amino acids with longer aliphatic side chains than valine have an even more pronounced effect. Thus single amino acid substitutions that lead to alterations in the catalytic activity of cyclic di-GMP turnover domains amplify the signaling output and thus significantly contribute to the flexibility and adaptability of the cyclic di-GMP signaling network. In this context, ZMO1055 seems to be a current evolutionary target.

microbiology↗

bioRxiv2022

Evolution towards small colony variants of pandemic multidrug resistant ST131 Escherichia coli isolates from a 10-year bone infection

Chronic wounds are difficult to treat because underlying medical conditions can impair the mechanical and physiological first-line innate immune defenses, leading to persistent microbial infections. We report here the isolation, molecular and phenotypic characterization of seven E. coli strains that were isolated concomitantly with Enterococcus faecalis after an open foot fracture caused by the 2004 tsunami resulting in a 10-year chronic bone and joint infection. Initially present antimicrobial resistant E. coli ST405 and ST940 isolates were followed by host adapted isolates of ubiquitous ST131 clone presumably acquired from the environment already upon initial foot fracture. The E. coli ST131 clade C1 strains showed genomic alterations associated with virulence and persistence including large chromosomal inversions and, subsequently, a large deletion to cause small colony variants and higher susceptibility to formaldehyde and other stress provoking In this context deletion of hemB catalyzing an early step in the pathway for heme biosynthesis was the major, but presumably not the only cause of small colony variant emergence. Surprisingly, ST131 isolates did not display pronounced biofilm formation in conventional biofilm assays suggesting unconventional modes of persistence. In summary, the genomes of ST131 clone members are highly plastic which enables their persistence in novel ecological niches. In individuals with underlying metabolic diseases such as diabetes wound infection can prepare for colonization with ST131 E. coli isolates. FundingThis work was partially funded by ALF, the Petrus and Augusta Hedlunds Foundation and the Karolinska Institutet.

microbiology↗

bioRxiv2020

Morphological and physiological effects of a single amino acid substitution in the patatin-like phospholipase CapV in Escherichia coli

In rod-shaped bacteria morphological plasticity occurs in response to stress, which blocks cell division to promote filamentation. We demonstrate here that overexpression of the patatin-like phospholipase variant CapVQ329R but not CapV causes pronounced sulA-independent pyridoxine-inhibited cell filamentation and restriction of swimming and flagella production of Escherichia coli K-12 derivative MG1655. Mutational analyses of CapVQ329R indicated conserved amino acids in canonical patatin-like phospholipase A motifs, but not the nucleophilic serine to be required for the observed phenotypes. Furthermore, CapVQ329R alters rdar biofilm formation including expression of the biofilm activator CsgD. Moreover, commensal and pathogenic E. coli strains and Salmonella typhimurium also responded with cell filamentation and alteration in biofilm formation. In conclusion, this work identifies the CapV variant CapVQ329R as a pleiotropic regulator, emphasizes a scaffold function for patatin-like phospholipases and highlights the role of a single amino acid change for the evolution of protein functionality.

microbiology↗

bioRxiv2020

Clarithromycin Exerts an Antibiofilm Effect against Salmonella typhimurium rdar Biofilm Formation, and Transforms the Physiology towards an Apparent Oxygen-depleted Energy and Carbon Metabolism

Upon biofilm formation, production of extracellular matrix components and alteration in physiology and metabolism allows bacteria to build up multicellular communities which can facilitate nutrient acquisition during unfavorable conditions and provide protection towards various forms of environmental stresses to individual cells. Thus, bacterial cells become tolerant against antimicrobials and the immune system within biofilms. In the current study, we evaluated the antibiofilm activity of the macrolides clarithromycin and azithromycin. Clarithromycin showed antibiofilm activity against rdar (red, dry and rough) biofilm formation of the gastrointestinal pathogen Salmonella typhimurium ATCC14028 Nalr at 1.56 M subinhibitory concentration in standing culture and dissolved cell aggregates at 15 M in a microaerophilic environment suggesting that the oxygen level affects the activity of the drug. Treatment with clarithromycin significantly decreased transcription and production of the rdar biofilm activator CsgD, with biofilm genes such as csgB and adrA to be consistently downregulated. While fliA and other flagellar regulon genes were upregulated, apparent motility was downregulated. RNA sequencing showed a holistic cell response upon clarithromycin exposure, whereby not only genes involved in the biofilm-related regulatory pathways, but also genes that likely contribute to intrinsic antimicrobial resistance, and the heat shock stress response were differentially regulated. Most significantly, clarithromycin exposure shifts the cells towards an apparent oxygen- and energy-depleted status, whereby the metabolism that channels into oxidative phosphorylation is downregulated, and energy gain by degradation of propane 1,2-diol, ethanolamine and L-arginine catabolism is upregulated. This initial analysis will allow the subsequent identification of novel intrinsic antimicrobial resistance determinants.

microbiology↗