Biology subjects

Iredell, J. R.

Publications and source records attributed to Iredell, J. R..

9 recordsLinked to original sources

bioRxiv2026

Specific exclusion of conjugative plasmids from the gut microflora

The rise of dangerous antimicrobial resistance (AMR), especially to the newer carbapenem antibiotics, is largely driven by the spread of conjugative plasmids between bacteria. These plasmids rapidly disseminate among common gut organisms like Escherichia coli and Klebsiella pneumoniae, which together account for around half of lethal sepsis and septic shock. Current AMR control measures, such as surveillance and isolation, are often ineffective, and AMR is often detected for the first time when infection is well established. Natural plasmid entry-exclusion systems (EES) protect bacterial populations from repeated entry by plasmids that are already established, or indeed by any plasmid with a related cognate EES, and we demonstrate here the exploitation of this mechanism for therapeutic purposes. We combined exclusion genes from three major AMR plasmid types (IncM, IncL, and IncC) into an efficient conjugative plasmid backbone and showed that this engineered probiotic plasmid prevents invasive AMR plasmids from entering bacterial populations in vitro and in vivo, in the mouse gut. This offers a promising new strategy to control invasive AMR plasmids and prevent AMR acquisition in high-risk settings, such as in hospitals or AMR-endemic regions.

microbiology↗

bioRxiv2022

The paradoxical relationship between CRISPR-Cas and phage susceptibility in Pseudomonas aeruginosa

CRISPR-Cas systems are part of the pan-immune system of Pseudomonas aeruginosa and have been shown to limit horizontal gene transfers in that species. Indeed, isolates equipped with these systems tend to have smaller genomes and CRISPR spacers targeting integrative conjugative elements, phages, and plasmids. In this work, we investigate the genomic effects and phenotypic consequences of CRISPR-Cas systems in P. aeruginosa. First, we establish that the population structure is a confounding factor of the relation between genome sizes and the presence of CRISPR-Cas systems in P. aeruginosa as isolates from group II are, on average, 200 kbp larger than those from group I, and have a lower likelihood of possessing CRISPR-Cas systems. Second, we show that the impact on the genome size of CRISPR deactivation by anti-CRISPR proteins differs between the various CRISPR-Cas types found in this species (I-C, I-E, and I-F). Finally, we highlight a paradoxical, positive correlation between the presence of CRISPR-Cas systems and the chances of the host being infected by a set of distinct, strictly virulent Pseudomonas phages. We propose that this increased phage susceptibility in the presence of CRISPR-Cas is linked to a depletion of other accessory defense system genes in isolates with CRISPR-Cas systems. Note: This manuscript was published as a chapter in the doctoral dissertation of Cedric Lood [72]

microbiology↗

bioRxiv2021

L-form switching confers antibiotic, phage and stress tolerance in pathogenic Escherichia coli

The bacterial L-form is induced by exposure to cell wall targeting antibiotics or innate immune effectors such as lysozyme and is likely to be important in many human infections. Here, we demonstrate that the osmotically fragile L-form is a distinct physiological state in Escherichia coli that is highly tolerant of oxidative stress and resistant to powerful antibiotics and common therapeutic bacteriophages. L-forms quickly revert (<20h) to their cell-walled state after antibiotic withdrawal, with apparently normal physiology and with few or no changes in DNA sequence. T4-like phages that are obligately lytic in cell-walled E. coli preferentially pseudolysogenise their L-forms providing them with transient superinfection immunity. Our data indicate that L-form switching is a common response of pathogenic E. coli strains to cell wall-targeting antibiotics and that the most commonly used lytic bacteriophages are ineffective against them in this state.

microbiology↗

bioRxiv2020

Clinical Antibiotic Resistance Patterns Across 70 Countries

We sought global patterns of antibiotic resistant pathogenic bacteria within the AMR Research Initiative database, Atlas. This consists of 6.5M clinical minimal inhibitory concentrations (MICs) observed in 70 countries in 633k patients between 2004 and 2017. Stratifying MICs according to pathogens (P), antibiotics (A) and countries (C), we found that the frequency of resistance was higher in Atlas than other publicly available databases. We determined global MIC distributions and, after showing they are coherent between years, we predicted MIC changes for 43 pathogens and 827 pathogen-antibiotic (PAs) pairings that exhibit significant resistance dynamics, including MIC increases and even decreases. However, many MIC distributions are multi-modal and some PA pairs exhibit sudden changes in MIC. We therefore analysed Atlas after replacing the clinical classification of pathogens into susceptible, intermediate and resistant with an information-optimal, cluster-based classifier to determine subpopulations with differential resistance that we denote S and R. Accordingly, S and R clusters for different PA pairs exhibit signatures of stabilising, directional and disruptive selection because their respective MICs can have different dynamics. Finally, we discuss clinical applications of a (R, dR/dt) phase plane whereby the MIC of R is regressed against change in MIC (dR/dt), a methodology we use to detect PA pairs at risk of developing clinical resistance.

microbiology↗

bioRxiv2020

Specialised functions of two common plasmid mediated toxin-antitoxin systems, ccdAB and pemIK, in Enterobacteriaceae

Toxin-antitoxin systems (TAS) are commonly found on bacterial plasmids and are involved in plasmid maintenance. Even though the same TAS are present in a variety of plasmid types and bacterial species, differences in their sequences, expression and functions are not well defined. Here, we aimed to identify commonly occurring plasmid TAS in Escherichia coli and Klebsiella pneumoniae and compare the sequence, expression and plasmid stability function of their variants. 27 putative type II TAS were identified from 1063 plasmids of Klebsiella pneumoniae in GenBank. Among these, ccdAB and pemIK were found to be most common, also occurring in plasmids of E. coli. Comparisons of ccdAB variants, taken from E. coli and K. pneumoniae, revealed sequence differences, while pemIK variants from IncF and IncL/M plasmids were almost identical. Similarly, the expression and plasmid stability functions of ccdAB variants varied according to the host strain and species, whereas the expression and functions of pemIK variants were consistent among host strains. The specialised functions of some TAS may determine the host specificity and epidemiology of major antibiotic resistance plasmids.

microbiology↗

bioRxiv2019

CCMetagen: comprehensive and accurate identification of eukaryotes and prokaryotes in metagenomic data

High-throughput sequencing of DNA and RNA from environmental and host-associated samples (metagenomics and metatranscriptomics) is a powerful tool to assess which organisms are present in a sample. Taxonomic identification software usually align individual short sequence reads to a reference database, sometimes containing taxa with complete genomes only. This is a challenging task given that different species can share identical sequence regions and complete genome sequences are only available for a fraction of organisms. A recently developed approach to map sequence reads to reference databases involves weighing all high scoring read-mappings to the data base as a whole to produce better-informed alignments. We used this novel concept in read mapping to develop a highly accurate metagenomic classification pipeline named CCMetagen. Using simulated fungal and bacterial metagenomes, we demonstrate that CCMetagen substantially outperforms other commonly used metagenome classifiers, attaining a 3 - 1580 fold increase in precision and a 2 - 922 fold increase in F1 scores for species-level classifications when compared to Kraken2, Centrifuge and KrakenUniq. CCMetagen is sufficiently fast and memory efficient to use the entire NCBI nucleotide collection (nt) as reference, enabling the assessment of species with incomplete genome sequence data from all biological kingdoms. Our pipeline efficiently produced a comprehensive overview of the microbiome of two biological data sets, including both eukaryotes and prokaryotes. CCMetagen is user-friendly and the results can be easily integrated into microbial community analysis software for streamlined and automated microbiome studies.

bioinformatics↗

bioRxiv2019

Safety and Tolerability of Bacteriophage Therapy in Severe Staphylococcus aureus Infection

ImportanceThe effect of IV administration of a bacteriophage cocktail produced under GMP conditions on patients with severe S. aureus infection, including complicated bacteraemia, endocarditis and septic shock, is unknown.\n\nObjectiveTo assess safety and tolerability of adjunctive bacteriophage therapy in patients with severe S. aureus infections.\n\nDesign, Setting, ParticipantsObservational, open-label clinical trial of thirteen critically-ill patients admitted to a tertiary-referral hospital with S. aureus bacteraemia (including infective endocarditis, n=6) were assessed by the treating clinician and two consulting infectious diseases physicians to independently verify that routine medical and surgical therapy was optimal and that a poor outcome remained likely. Compassionate access to therapy was approved by both US and Australian regulators and by the Westmead Hospital Human Research Ethics Committee.\n\nInterventionA GMP-quality preparation of three combined Myoviridae bacteriophages with specific activity against S. aureus (AB-SA01), was administered intravenously in conjunction with optimal antibiotic therapy.\n\nMain Outcome and MeasurementsPhysiological, haematological and biochemical markers of infection, bacterial and bacteriophage kinetics in blood, development of resistance to bacteriophages, and mortality at 28 (D28) and 90 (D90) days were measured. Main outcomes were safety and tolerability.\n\nResultsBacteriophage therapy was initiated 4-10 days after antibiotic commencement, at 109 plaque-forming units (PFU) twice daily. Infecting staphylococci were typical of common local subtypes. Initial input ratio of phages to bacteria in the bloodstream (MOIinput) was >100. Five of the thirteen patients died by D28 and a sixth patient suffered sudden cardiac death on D90. Bacteriophage therapy coincided with a marked reduction in staphylococcal bacterial DNA in the blood and in sepsis-associated inflammatory responses in almost all cases. No bacteriophage-attributable adverse events were identified. Development of bacteriophage resistance was not observed. Population analysis revealed no significant effect of bacteriophage therapy on the gut microflora.\n\nConclusions and RelevanceAdjunctive bacteriophage therapy appears to be safe and well-tolerated in critically ill patients with severe S. aureus infection. Two weeks of twice daily intravenous administration may be a suitable protocol. Controlled trials are needed.\n\nTrial RegistrationWestmead Hospital Human Research Ethics Committee approval July 11, 2017; ClinicalTrials.gov Identifier: NCT03395769, AB-SA01-EAP01 (January 10, 2018); Clinical Trials Notification (Australian Therapeutic Goods Association): CT-2018-CTN-02372-1 (July 23, 2018).\n\nKey Points\n\nQuestionIs intravenous (IV) administration of investigational bacteriophage (phage) therapy safe and well-tolerated in patients with severe Staphylococcus aureus infection?\n\nFindingsThirteen patients with severe S. aureus infections received AB-SA01, a bacteriophage product prepared according to Good Manufacturing Practices (GMP), as adjunctive therapy to antibiotics. AB-SA01 was well-tolerated with no adverse events identified. Bacterial burden and inflammatory responses were reduced and no phage-resistant staphylococci were isolated during or after therapy.\n\nMeaningOur results will inform future randomised controlled trials assessing the antibacterial and anti-inflammatory potential of bacteriophages in the treatment of severe S. aureus infection.

clinical trials↗

bioRxiv2018

High levels of antibiotic resistance gene expression among birds living in a wastewater treatment plant

Antibiotic resistance is rendering common bacterial infections untreatable. Wildlife can incorporate and disperse antibiotic resistant bacteria in the environment, such as water systems, which in turn serve as reservoirs of resistance genes for human pathogens. We used bulk RNA-sequencing (meta-transcriptomics) to assess the diversity and expression levels of functionally active resistance genes in the microbiome of birds with aquatic behavior. We sampled birds across a range of habitats, from penguins in Antarctica to ducks in a wastewater treatment plant in Australia. This revealed 81 antibiotic resistance genes in birds from all localities, including {beta}-lactam, tetracycline and chloramphenicol resistance in Antarctica, and genes typically associated with multidrug resistance plasmids in areas with high human impact. Notably, birds feeding at a wastewater treatment plant carried the greatest resistance gene burden, suggesting that human waste, even if it undergoes treatment, contributes to the spread of antibiotic resistance genes to the wild. Differences in resistance gene burden also reflected the birds ecology, taxonomic group and microbial functioning. Ducks, which feed by dabbling, carried a higher abundance and diversity of resistance genes than turnstones, avocets and penguins, that usually prey on more pristine waters. In sum, this study helps to reveal the complex factors explaining the distribution of resistance genes and their exchange routes between humans and wildlife.

ecology↗

bioRxiv2018

Host adaptation and convergent evolution increases antibiotic resistance without loss of virulence in a major human pathogen

As human population density and antibiotic exposure increases, specialised bacterial subtypes have begun to emerge. Arising among species that are common commensals and infrequent pathogens, antibiotic-resistant high-risk clones have evolved to better survive in the modern human. Here, we show that the major matrix porin (OmpK35) of Klebsiella pneumoniae is not required in the mammalian host for colonisation, pathogenesis, nor for antibiotic resistance, and that it is commonly absent in pathogenic isolates. This is found in association with, but apparently independent of, a highly specific change in the co-regulated partner porin, the osmoporin (OmpK36), which provides enhanced antibiotic resistance without significant loss of fitness in the mammalian host. These features are common in well-described high-risk clones of K. pneumoniae, as well as in unrelated members of this species and similar adaptations are found in other members of the Enterobacteriaceae that share this lifestyle. Available sequence data indicates evolutionary convergence, with implications for the spread of lethal antibiotic-resistant pathogens in humans.\n\nAuthor summaryKlebsiella pneumoniae is a Gram-negative enterobacteria and a significant cause of human disease. It is a frequent agent of pneumonia, and systemic infections can have high mortality rates (60%). OmpK35 and OmpK36 are the major co-regulated outer membrane porins of K. pneumoniae. OmpK36 absence has been related to antibiotic resistance but decreased bacterial fitness and diminished virulence. A mutation that constricts the porin channel (Gly134Asp135 duplication in loop 3 of the porin, OmpK36GD) has been previously observed and suggested as a solution to the fitness cost imposed by loss of OmpK36.\n\nIn the present study we constructed isogenic mutants to verify this and test the impact of these porin changes on antimicrobial resistance, fitness and virulence. Our results show that loss of OmpK35 has no significant cost in bacterial survival in nutrient-rich environments nor in the mammalian host, consistent with a predicted role outside that niche. When directly compared with the complete loss of the partner osmoporin OmpK36, we found that isogenic OmpK36GD strains maintain high levels of antibiotic resistance and that the GD duplication significantly reduces neither gut colonisation nor pathogenicity in a pneumonia mouse model. These changes are widespread in unrelated genomes. Our data provide clear evidences that specific variations in the loop 3 of OmpK36 and the absence of OmpK35 in K. pneumoniae clinical isolates are examples of successful adaptation to human colonization/infection and antibiotic pressure, and are features of a fundamental evolutionary shift in this important human pathogen.

microbiology↗