7th joint conference "Joint Microbiology & Infection Conference 2024" of the DGHM and VAAM

Chair

Healthcare-associated infections and pathogens: Prevention, surveillance, outbreaks und antibiotic stewardship 2

PD. Dr.

Prof. Dr.

Ralstonia pickettii bloodstream infections due to contaminated saline solution – a global problem?

Healthcare-associated infections and pathogens: Prevention, surveillance, outbreaks und antibiotic stewardship

QuestionRalstonia pickettii is a gram-negative rod which may cause invasive infections due to the use of contaminated liquid medical products.MethodsAfter R. pickettii was detected in blood cultures and a stem cell product from three patients in a tertiary-care hospital in Germany, environmental sampling was performed and subsequently whole genome sequencing (WGS) of the three patient isolates and two water isolates from the environment.ResultsCore Genome Multi Locus Sequence Typing (cgMLST) analysis showed that the three patient isolates were closely related and there was a large number of allele differences to the environmental isolates. The genomic comparison of the patients" isolates with a R. pickettii strain from an outbreak in Australia suspected to be caused by contaminated saline produced in India revealed only few differences.ConclusionsOur data and information point towards an ongoing risk by medical products contaminated with R. pickettii potentially distributed worldwide. Identification of medical products applied to patients with R. pickettii infections and timely whole genome sequencing of all available R. pickettii isolates may help to identify the source of this potentially global outbreak.

Oral Presentation

Influence of antibiotics on microbial communities in hospital drain biofilms and their resistance profile

Introduction &amp; AimSince biofilms are known to contain resistant bacterial species and many hospital-acquired infections are believed to be biofilm-associated, their occurrence in hospitals is critical. As drains get in contact with antimicrobial residues like antibiotics, these may act as resistance drivers. Class 1 integrons as a multi-resistance marker allow for monitoring changes in resistances, checking for integrase gene intI1.Our research aimed to investigate the impact of antibiotics on the composition of hospital drain biofilms and their genetic and phenotypic resistance profile based on consumption data of different wards and experimental approaches. Methods20 bathroom sink biofilms were sampled from 4 wards of a hospital. The cell count of viable cells, Enterobacteriaceae and yeasts/moulds was determined. Additionally, relevant pathogens were isolated, identified and analyzed for resistances with VITEK®2 compact system and by Epsilometer-testing. intI1-prevalence was measured by qPCR. Further testing took place by adding different antibiotics to the media during biofilm growth. After DNA extraction, changes of DNA in a high resolution melting analysis (HRMA) and the intI1 prevalence were evaluated.ResultsOur data show that cell counts of the wards were not significantly different. MIC-values for antibiotics obtained by VITEK®2 and Epsilometer-testing showed no correlation to the administration of recommended daily doses (RDD)/100 bed days. In contrast, the median intI prevalence correlated with the RDD. While adding sub-inhibitory concentrations of antibiotics during growth, differences in HRMA were observed depending on the biofilm and antibiotic (Fig.1). Moreover, shifts in HRMA data correlated with changes in intI1 prevalences.SummaryOur approach gave an insight into correlations between resistance values and the RDD of hospital wards. Expected pathogens were found. Genetic resistance markers correlated with the amount of prescribed antibiotics, although phenotypes didn´t. Thus, a selective pressure by sub-inhibitory antibiotic concentrations were found to cause community shifts accompanied by changes in the intI1 prevalence.

Microbiome translocation and within-patient evolution promotes opportunistic, nosocomial infections

Alarmingly, many commensal bacteria that peacefully reside in the human microbiome are also able to cause acute opportunistic infections. Thought to be triggered randomly in patients with impaired immunity, intriguing evidence suggests that within-host evolution can promote infection. The major barrier to studying the progression of opportunistic pathogens from carriage to acute infections is the challenge of sampling human patients prospectively. Here we test whether adaptive de novo mutations in asymptomatically colonizing opportunistic pathogen lineages facilitate translocation within the patient's body and promote acute infections. Therefore, we have enrolled 14 critically ill patients who are longitudinally sampled at four microbiome niches before the onset of healthcare-associated infection (HAI). Among four patients, we observe twelve HAIs caused by ten different opportunistic pathogens. Using culture-based sequencing we address within-patient evolution by generating over 600 whole genomes of the pathogen populations to characterize acquisition timing, spatial translocation, and whether adaptive de novo mutations promote acute infection. We find that in 70% of HAI cases the pathogen lineages can be identified within the microbiome prior to or at the time of disease onset. Moreover, using a molecular clock we infer that the pathogen lineages are regularly acquired already before hospitalization of the patient. In a single patient we observe a short-lived mutation in a fimbriae gene associated with HAI. Phenotypic tests reveal the mutation upregulates fimbriae production increasing cell adhesion and leads to higher virulence in a Drosophila infection model. In summary, our prospective approach allowed us to explore the hidden evolutionary paths leading up to HAI and underlines the importance to investigate microbiome dynamics at a population-wide whole genome level during disease.

Professor

Dr. med.

WGS-based characterization of putative hypervirulent Klebsiella pneumoniae identified in a tertiary care hospital in Germany

Introduction: Hypervirulent Klebsiella pneumoniae strains (hvKp), in contrast to classical K. pneumoniae strains, can cause invasive community-acquired infections in healthy patients of all ages. In this study, K. pneumoniae isolates from routine microbiological diagnostics were tested via string-test for hypermucoviscous phenotype and PCR for virulence genes to screen for hvKp, which were further analyzed by whole-genome sequencing (WGS).Goals: The prevalence of &quot;hidden&quot; hypervirulent hvKp, including non-invasive strains and their population structure should be analyzed in a tertiary-care hospital in Southern Germany.Materials/methods: For the study period 1.5 years, all K. pneumoniae isolates were string-tested. A multiplex PCR for hvKp genes was applied to all string-test positive isolates. PCR-positive isolates were subjected to WGS to determine genotyping and phylogeny.Results: A total of 10.9% (331/3044) isolates with hmKp phenotype were detected by string-test. The patients&quot; age rangend from 0 to 95 years, with a mean of 69 years. In total, 13.3% (44/331) isolates were tested positive by PCR for genes associated with hvKp. cgMLST revealed that 41.5% of sequenced isolates belonged to international hvKp clonal lineages ST23/K1 with high virulence scores and close phylogenetic relationships. In contrast, 22.6% of isolates belonged to the ST86/K2 with lower virulence scores. Isolates of liver abscesses (7.5%) belonged to ST23, ST25 and ST268, but without phylogenetic relatedness to other isolates.Summary: We identified hypervirulent K. pneumoniae within the study period with an overall prevalence of 1.4%. No transmissions were identified. Isolates were assigned to the international hvKp lineage ST23/K1 and lineage ST86, bearing the potential to spread further in the community. The known association of multidrug-resistance ST86, was not observed in this study. Presented cases of liver abscesses seem to represent individual occurrences, so far. In conclusion, the screening of hmKp phenotypes in routine diagnostics seems to be a suitable surveillance method. Further, the population size and structure of &quot;hidden&quot; putative hvKp were more complex than expected.

Genetic analysis of carbapenem resistant Acinetobacter baumannii isolates associated with the war in Ukraine

QuestionThe worldwide spread and increase of multidrug-resistant Gramnegative bacteria represents a serious threat to public health. However, the prevalence of resistance mechanisms may differ between different regions or countries. In case of Ukraine, high AMR proportions in Gram-negative bacterial isolates have been reported in 2021, including carbapenem resistant Acinetobacter spp. (73%) and carbapenem resistant Klebsiella pneumoniae (64%)1. Recently, an increase in NDM-1 and NDM-1/OXA-48-producing K. pneumoniae in Germany was observed that derived at least partially from patients with prior exposure in Ukraine2.Here, we report genetic surveillance of carbapenem resistant A. baumannii isolates in Germany from patients with prior exposure in Ukraine.MethodsCarbapenemase detection was performed at the German National Refence Laboratory for multidrug-resistant Gram-negative bacteria (NRC) by modified Hodge test, combined disk test with EDTA and PCR for carbapenemase genes. Whole-cell DNA was subjected to WGS on an Illumina MiSeq platform with 2 x 300 bp paired end reads and subsequent genome assembly and typing was conducted using Ridom SeqSphere.ResultsIn total, 58 A. baumannii isolates from patients with prior exposure in Ukraine were sequenced. Most of the isolates produced the carbapenemases OXA-23 or OXA-72. The most prevalent sequence types (STs) were ST2 for OXA-23 positive isolates and ST78 for OXA-72 positive isolates. Comparison via core genome MLST (cgMLST) revealed several clusters distributed all over Germany.ConclusionsWe report genetic analysis of carbapenemase producing A. baumannii isolates from patients with prior exposure in Ukraine. The distribution of closely related isolates all over Germany suggests that transmission may have occurred during hospitalization in Ukraine or transfer to Germany.

Oral Session
Pathogenicity and Reservoirs of Nosocomial Pathogens

Termin

Chair

Themen

Beschreibung

Programm

Oral SessionPathogenicity and Reservoirs of Nosocomial Pathogens

Termin

Chair

Themen

Beschreibung

Programm

Oral Session
Pathogenicity and Reservoirs of Nosocomial Pathogens