Biology subjects

Kröger, C.

Publications and source records attributed to Kröger, C..

7 recordsLinked to original sources

bioRxiv2025

Differential roles of the type I and II secretion systems for the intracellular ABC141 Acinetobacter baumannii infection, which elicits an atypical hypoxia response in endothelial cells

Acinetobacter baumannii poses a substantial global health threat, causing severe multi-drug-resistant infections in hospitalized patients. Circulating clinical isolates present remarkable diversity, with a proportion capable of establishing a transient intracellular niche suitable for persistence, multiplication, and spread. Yet, it remains unknown which bacterial factors mediate the formation and maintenance of this niche, especially within non-phagocytic cells, nor what host responses are elicited. This work demonstrates that the invasive A. baumannii ABC141 strain does not secrete ammonia in endothelial cells as previously shown for other A. baumannii strains multiplying within macrophages but resides in an acidic vacuole devoid of active lysosomal degradative enzymes. This compartment mediates bacterial egress and infection of neighboring cells, promoting dissemination. Using a Dual-RNAseq approach, we mapped the host and bacterial gene expression during the replicative stage of the infection. An atypical hypoxia cell response was observed without significant induction of the HIF1 pathway, with no metabolic shift or disturbance of mitochondria. Surprisingly, ABC141 efficiently grew in hypoxic conditions in culture and within host cells. In addition, we found a bacterial signature reflective of an adaptation to a nutrient-deprived environment. Our work also highlights a differential role for ABC141 secretion systems, with the T1SS assisting intracellular multiplication and the T2SS required for host cell invasion, implicating for the first time the T2SS in the intracellular lifecycle of invasive ABC141 in endothelial cells. Author summaryAcinetobacter baumannii is a major burden for healthcare facilities worldwide, causing severe infections in hospitalized and intensive care patients that are challenging to treat because of extremely high levels of resistance to most antimicrobials. Traditionally seen as extracellular, recent studies have highlighted that a proportion of clinical isolates currently circulating in clinics can invade and multiply inside epithelial and endothelial cells. Yet, how these strains establish an intracellular protected niche remains unknown. Here, we undertook a detailed characterization of the intracellular compartment enclosing multiplying bacteria, the induced host response, and the bacterial genes that contribute to the adaptation to this intracellular environment. We found that the A. baumannii ABC141 clinical strain is well equipped to resist acidic, low oxygen, and low nutrient environments. Furthermore, we discovered it uses its type II secretion system to invade human endothelial cells, the first step of this intracellular lifecycle. This study opens new avenues of research to help develop new antimicrobial strategies to combat this bacteria when shielded within intracellular compartments in the host.

microbiology↗

bioRxiv2023

Global analysis of the RNA-RNA interactome in Acinetobacter baumannii AB5075 uncovers a small regulatory RNA repressing the virulence-related outer membrane protein CarO

Acinetobacter baumannii is an opportunistic Gram-negative pathogen that infects critically ill patients. The emergence of antimicrobial resistant A. baumannii has exacerbated the need to functionally characterise environmental adaptation, antibiotic resistance and pathogenicity of this organism and their genetic regulators to inform intervention strategies. Critical to rapid adaptation to changing environments in bacteria are small regulatory RNAs (sRNAs), however, the role that sRNAs play in the biology of A. baumannii is poorly understood. To assess the regulatory function of sRNAs and to uncover their RNA interaction partners in A. baumannii, we employed an RNA proximity ligation and sequencing method (Hi-GRIL-seq) in three different environmental conditions. We found that 40 sRNA candidates were ligated to sRNA-RNA chimeric sequencing reads, suggesting that sRNA-mediated gene regulation is pervasive in A. baumannii and that sRNAs act as direct regulators of mRNA molecules through antisense base-pairing. In-depth characterisation uncovered the sRNA Aar to be a post-transcriptional regulator of four mRNA targets including that of the outer membrane protein CarO and the siderophore receptor BfnH. We show that Aar initiates base-pairing with these mRNA molecules using a conserved seed region of nine nucleotides, sequestering the ribosome binding sites and inhibiting translation. Aar is differentially expressed in response to multiple stress stimuli suggesting a role in fine-tuning translation of the Aar-target molecules in A. baumannii under hostile conditions. Together, our study provides mechanistic insights into sRNA-mediated gene expression control in A. baumannii and represents a valuable resource for future RNA-centric research endeavours in this ESKAPE pathogen.

microbiology↗

bioRxiv2019

A window into lysogeny: Revealing temperate phage biology with transcriptomics

Integrated phage elements, known as prophages, are a pervasive feature of bacterial genomes. Prophages can enhance the fitness of their bacterial hosts by conferring beneficial functions, such as virulence, stress tolerance or phage resistance, which are encoded by accessory loci. Whilst the majority of phage-encoded genes are repressed during lysogeny, accessory loci are often highly expressed. However, novel prophage accessory loci are challenging to identify based on DNA sequence data alone. Here, we use bacterial RNA-seq data to examine the transcriptional landscapes of five Salmonella prophages. We show that transcriptomic data can be used to heuristically enrich for prophage features that are highly expressed within bacterial cells and often represent functionally-important accessory loci. Using this approach we identify a novel anti-sense RNA species in prophage BTP1, STnc6030, which mediates superinfection exclusion of phage BTP1 and immunity to closely-related phages. Bacterial transcriptomic datasets are a powerful tool to explore the molecular biology of temperate phages.

microbiology↗

bioRxiv2019

The diversity, evolution and ecology of Salmonella in venomous snakes

BackgroundReptile-associated Salmonella are a major, but often neglected cause of both gastrointestinal and bloodstream infection globally. The diversity of Salmonella enterica has not yet been determined in venomous snakes, however other cold-blooded animals have been reported to carry a broad range of Salmonella bacteria. We investigated the prevalence and assortment of Salmonella in a collection of venomous snakes in comparison with non-venomous reptiles.\n\nMethodology/Principle FindingsWe used a combination of selective enrichment techniques and whole-genome sequencing. We established a unique dataset of reptilian isolates to study Salmonella enterica species-level evolution and ecology and investigated differences between phylogenetic groups. We observed that 91% of venomous snakes carried Salmonella, and found substantial diversity between the serovars (n=58) carried by reptiles. The Salmonella serovars belonged to four of the six Salmonella enterica subspecies: diarizonae, enterica, houtanae and salamae. Subspecies enterica isolates were distributed among two distinct phylogenetic clusters, previously described as clade A (52%) and clade B (48%). We identified metabolic differences between S. diarizonae, S. enterica clade A and clade B involving growth on lactose, tartaric acid, dulcitol, myo-inositol and allantoin.\n\nSignificanceWe present the first whole genome-based comparative study of the Salmonella bacteria that colonise venomous and non-venomous reptiles and shed new light on Salmonella evolution. The findings raise the possibility that venomous snakes are a reservoir for human Salmonellosis in Africa. The proximity of venomous snakes to human dwellings in rural Africa may result in contaminated faecal matter being shed on surfaces and in water sources used for human homes and to irrigate salad crops. Because most of the venomous snakes had been captured in Africa, we conclude that the high level of Salmonella diversity reflects the African environmental niches where the snakes have inhabited.\n\nAuthor SummarySalmonella enterica is a remarkable bacterial species that causes Neglected Tropical Diseases globally. The burden of disease is greatest in some of the most poverty-afflicted regions of Africa, where salmonellosis frequently causes bloodstream infection with fatal consequences. The bacteria have the ability to colonise the gastrointestinal tract of a wide range of animals including reptiles. Direct or indirect contact between reptiles and humans can cause Salmonellosis. In this study, we determined the prevalence and diversity of Salmonella in a collection of African venomous snakes for the first time. Using the power of genomics, we showed that the majority of venomous snakes (91%) carry Salmonella, two thirds of which belonged to a subspecies of S. enterica called enterica, which is associated with most cases of human salmonellosis. Within the S. enterica subspecies we identified two evolutionary groups which display distinct growth patterns on infection relevant carbon sources. Our findings could have particular significance in Africa where venomous snakes wander freely around human dwellings and potentially shed contaminated faecal matter in water sources and on surfaces in rural homes.

microbiology↗

bioRxiv2018

Adding function to the genome of African Salmonella ST313

Salmonella Typhimurium ST313 causes invasive nontyphoidal Salmonella (iNTS) disease in sub-Saharan Africa, targeting susceptible HIV+, malarial or malnourished individuals. An in-depth genomic comparison between the ST313 isolate D23580, and the well-characterized ST19 isolate 4/74 that causes gastroenteritis across the globe, revealed extensive synteny. To understand how the 856 nucleotide variations generated phenotypic differences, we devised a large-scale experimental approach that involved the global gene expression analysis of strains D23580 and 4/74 grown in sixteen infection-relevant growth conditions. Comparison of transcriptional patterns identified virulence and metabolic genes that were differentially expressed between D23580 versus 4/74, many of which were validated by proteomics. We also uncovered the S. Typhimurium D23580 and 4/74 genes that showed expression differences during infection of murine macrophages. Our comparative transcriptomic data are presented in a new enhanced version of the Salmonella expression compendium SalComD23580: bioinf.gen.tcd.ie/cgi-bin/salcom_v2.pl. We discovered that the ablation of melibiose utilization was caused by 3 independent SNP mutations in D23580 that are shared across ST313 lineage 2, suggesting that the ability to catabolise this carbon source has been negatively selected during ST313 evolution. The data revealed a novel plasmid maintenance system involving a plasmid-encoded CysS cysteinyl-tRNA synthetase, highlighting the power of large-scale comparative multi-condition analyses to pinpoint key phenotypic differences between bacterial pathovariants.

microbiology↗

bioRxiv2017

The primary transcriptome, small RNAs, and regulation of antimicrobial resistance in Acinetobacter baumannii

AO_SCPCAPBSTRACTC_SCPCAPWe present the first high-resolution determination of transcriptome architecture in the priority pathogen Acinetobacter baumannii. Pooled RNA from 16 laboratory conditions was used for differential RNA-seq (dRNA-seq) to identify 3731 transcriptional start sites (TSS) and 110 small RNAs, including the first identification in A. baumannii of sRNAs encoded at the 3 end of coding genes. Most sRNAs were conserved among sequenced A. baumannii genomes, but were only weakly conserved or absent in other Acinetobacter species. Single nucleotide mapping of TSS enabled prediction of -10 and -35 RNA polymerase binding sites and revealed an unprecedented base preference at position +2 that hints at an unrecognized transcriptional regulatory mechanism. To apply functional genomics to the problem of antimicrobial resistance, we dissected the transcriptional regulation of the drug efflux pump responsible for chloramphenicol resistance, craA. The two craA promoters were both down-regulated >1000-fold when cells were shifted to nutrient limited medium. This conditional down-regulation of craA expression renders cells sensitive to chloramphenicol, a highly effective antibiotic for the treatment of multidrug resistant infections. An online interface that facilitates open data access and visualization is provided as \"AcinetoCom\" (http://bioinf.gen.tcd.ie/acinetocom/).

microbiology↗

bioRxiv2017

The role of a single non-coding nucleotide in the evolution of an epidemic African clade of Salmonella

Introductory ParagraphSalmonella enterica serovar Typhimurium ST313 is a relatively newly emerged sequence type that is causing a devastating epidemic of bloodstream infections across sub-Saharan Africa. Analysis of hundreds of Salmonella genomes has revealed that ST313 is closely-related to the ST19 group of S. Typhimurium that cause gastroenteritis across the world. The core genomes of ST313 and ST19 vary by just 1000 single-nucleotide polymorphisms (SNPs). We hypothesised that the phenotypic differences that distinguish African Salmonella from ST19 are caused by certain SNPs that directly modulate the transcription of virulence genes.\n\nHere we identified 3,597 transcriptional start sites (TSS) of the ST313 strain D23580, and searched for a gene expression signature linked to pathogenesis of Salmonella. We identified a SNP in the promoter of the pgtE gene that caused high expression of the PgtE virulence factor in African S. Typhimurium, increased the degradation of the factor B component of human complement, contributed to serum resistance and modulated virulence in the chicken infection model. The PgtE protease is known to mediate systemic infection in animal models. We propose that high levels of expression PgtE of by African S. Typhimurium ST313 promotes bacterial survival and bacterial dissemination during human infection.\n\nOur finding of a functional role for an extra-genic SNP shows that approaches used to deduce the evolution of virulence in bacterial pathogens should include a focus on non-coding regions of the genome.

molecular biology↗