Genotypic resistance determined by whole genome sequencing versus phenotypic resistance in 234 Escherichia coli isolates.

Whole genome sequencing (WGS) enables detailed characterization of bacteria at single nucleotide resolution. It provides data about acquired resistance genes and mutations leading to resistance. Although WGS is becoming an essential tool to predict resistance patterns accurately, comparing genotype to phenotype with WGS is still in its infancy. Additional data and validation are needed. In this retrospective study, we analysed 234 E. coli isolates from positive blood cultures using WGS as well as microdilution for 11 clinically relevant antibiotics, to compare the two techniques. We performed whole genome sequencing analyses on 234 blood culture isolates (genotype) to detect acquired antibiotic resistance. Minimal inhibitory concentrations (MIC) for E. coli were performed for amoxicillin, cefepime, cefotaxime, ceftazidime, meropenem, amoxicillin/clavulanic acid, piperacillin/tazobactam, amikacin, gentamicin, tobramycin, and ciprofloxacin, using the ISO 20776-1 standard broth microdilution method as recommended by EUCAST (phenotype). We then compared the two methods for statistical 'agreement'. A perfect (100%) categorical agreement between genotype and phenotype was observed for gentamicin and meropenem. However, no resistance to meropenem was observed. A high categorical agreement (> 95%) was observed for amoxicillin, cefepime, cefotaxime, ceftazidime, amikacin, and tobramycin. A categorical agreement lower than 95% was observed for amoxicillin/clavulanic acid, piperacillin/tazobactam, and ciprofloxacin. Most discrepancies occurred in isolates with MICs within ± 1 doubling dilution of the breakpoint and 22.73% of the major errors were samples that tested phenotypically susceptible at higher antibiotic exposure and were therefore considered as 'not resistant'. This study shows that WGS can be used as a valuable tool to predict phenotypic resistance against most of the clinically relevant antibiotics used for the treatment of E. coli bloodstream infections.

Molecular characterization of extraintestinal and diarrheagenic Escherichia coli blood isolates.

Pathogenic E. coli strains can be classified into two major groups, based on the presence of specific virulence factors: extraintestinal pathogenic E. coli (ExPEC) and diarrheagenic E. coli (DEC). Several case reports describe that DEC can cause bloodstream infections in some rare cases. This mainly concerns a few specific sequence types that express virulence factors from both ExPEC and DEC. In this study, we retrospectively analysed 234 E. coli blood isolates with whole genome sequencing (WGS). WGS was performed on an Illumina NovaSeq6000. Genotyping was performed using BioNumerics software. The presence of genes was determined with a minimum percentage sequence identity (ID) threshold of 95% and a minimum length for sequence coverage of 95%. Three of the 234 (1.28%) isolates were defined as DEC, 182 (77.78%) as ExPEC, and 49 (20.94%) did not carry pathotype-associated virulence genes. We identified 112 different virulence genes, 48 O-antigens, and 28 H-antigens 82 STs, among the 234 analyzed isolates. ST131 and ST88 were related to healthcare-associated infections. This study provides insight into the prevalence of virulence factors in a large set of E. coli blood isolates from the UZ Brussel. It illustrates high diversity in virulence profiles and highlights the potential of DEC to carry virulence factors associated with extraintestinal infections, making it possible for unusual pathotypes to invade and survive in the bloodstream causing bacteraemia. Diarrheagenic strains causing bacteremia are rare and presently underreported, but modern sequencing techniques will better underscore their importance.

Sink drains as reservoirs of VIM-2 metallo-ß-lactamase-producing Pseudomonas aeruginosa in a Belgian intensive care unit: relation to patients investigated by whole-genome sequencing.

BACKGROUND: Hospital-acquired infections caused by VIM-encoded metallo-ß-lactamase-positive Pseudomonas aeruginosa are a major problem in intensive care units (ICUs) worldwide. A previous study conducted in the UZ Brussel hospital revealed that sink drains of the ICU were a possible source of various multidrug-resistant pathogenic bacteria. AIM: To investigate the presence and persistence of VIM P. aeruginosa in the sink drains of the four adult ICUs and their role in nosocomial infections, emphasizing sink-to-patient transmission. METHODS: Thirty-six sinks located in the ICUs of the UZ Brussel were sampled and screened for the presence of VIM P. aeruginosa in August and October 2019. Whole-genome sequencing (WGS) was performed on all positive sink drain isolates together with 61 isolates from patients who were retrospectively selected (ICU patients 2019-2020, N = 46; non-ICU patients 2019, N = 6). FINDINGS: Twenty sinks were found positive for P. aeruginosa at both sampling time-points. WGS revealed that the predominating environmental cluster belonged to sequence type ST111. Ten additional STs were identified. VIM-2 was detected among all ST17 (N = 2) and ST111 (N = 14) sink drain isolates. Based on whole-genome multi-locus sequence typing analysis of all genomes, 15 clusters of highly related isolates were identified, of which seven included both sink drain and clinical isolates. CONCLUSION: Our findings confirm that sink drains are a possible source of VIM-2 P. aeruginosa, probably after being contaminated with clinical waste from patients. Patients could be exposed to VIM-2 P. aeruginosa dispersed in their environment because of colonized sink drains.

Performance evaluation of the Diasorin LIAISON® XL Zika capture IgM CLIA test.

Because Zika virus (ZIKV) can cause serious birth defects and is involved in cases of Guillain-Barré syndrome, the ZIKV outbreak in the American continent in 2015 resulted in an enormous need for ZIKV diagnostic tools. We evaluated the LIAISON® XL Zika Capture IgM test on 106 samples from patients, mainly travelers, with a confirmed or probable ZIKV infection. Sensitivity between 0 and 84 days after onset of symptoms was 92.5%. Specificity was evaluated on a panel of 56 samples known to cause possible cross-reactions. Cross-reaction with DENV antibodies was limited (10.5%) but false-positive results occurred in samples from patients with malaria, CMV and EBV infections.

Validation of a New Web Application for Identification of Fungi by Use of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.

Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry has emerged as a reliable technique to identify molds involved in human diseases, including dermatophytes, provided that exhaustive reference databases are available. This study assessed an online identification application based on original algorithms and an extensive in-house reference database comprising 11,851 spectra (938 fungal species and 246 fungal genera). Validation criteria were established using an initial panel of 422 molds, including dermatophytes, previously identified via DNA sequencing (126 species). The application was further assessed using a separate panel of 501 cultured clinical isolates (88 mold taxa including dermatophytes) derived from five hospital laboratories. A total of 438 (87.35%) isolates were correctly identified at the species level, while 26 (5.22%) were assigned to the correct genus but the wrong species and 37 (7.43%) were not identified, since the defined threshold of 20 was not reached. The use of the Bruker Daltonics database included in the MALDI Biotyper software resulted in a much higher rate of unidentified isolates (39.76 and 74.30% using the score thresholds 1.7 and 2.0, respectively). Moreover, the identification delay of the online application remained compatible with real-time online queries (0.15 s per spectrum), and the application was faster than identifications using the MALDI Biotyper software. This is the first study to assess an online identification system based on MALDI-TOF spectrum analysis. We have successfully applied this approach to identify molds, including dermatophytes, for which diversity is insufficiently represented in commercial databases. This free-access application is available to medical mycologists to improve fungal identification.

Neuroprotective efficacy of subcutaneous insulin-like growth factor-I administration in normotensive and hypertensive rats with an ischemic stroke.

The aim of this study was to test the insulin-like growth factor-I (IGF-I) as a neuroprotective agent in a rat model for ischemic stroke and to compare its neuroprotective effects in conscious normotensive and spontaneously hypertensive rats. The effects of subcutaneous IGF-I injection were investigated in both rat strains using the endothelin-1 rat model for ischemic stroke. Motor-sensory functions were measured using the Neurological Deficit Score. Infarct size was assessed by Cresyl Violet staining. Subcutaneous administration of IGF-I resulted in significantly reduced infarct volumes and an increase in motor-sensory functions in normotensive rats. In these rats, IGF-I did not modulate blood flow in the striatum and had no effect on the activation of astrocytes as assessed by GFAP staining. In hypertensive rats, the protective effects of IGF-I were smaller and not always significant. Furthermore, IGF-I significantly reduced microglial activation in the cortex of hypertensive rats, but not in normotensive rats. More detailed studies are required to find out whether the reduction by IGF-I of microglial activation contributes to an impairment IGF-I treatment efficacy. Indeed, we have shown before that microglia in hypertensive rats have different properties compared to those in control rats, as they exhibit a reduced responsiveness to ischemic stroke and lipopolysaccharide.