A ring trial to harmonize Toxoplasma gondii microsatellite typing: comparative analysis of results and recommendations for optimizationJoeres, M.; Cardron, G.; Passebosc-Faure, K.; Plault, N.; Fernández-Escobar, M.; Hamilton, C. M.; O’Brien-Anderson, L.; Calero-Bernal, R.; Galal, L.; Luttermann, C.; Maksimov, P.; Conraths, F. J.; Dardé, M. L.; Ortega-Mora, L. M.; Jokelainen, P.; Mercier, A.; Schares, G.
doi: 10.1007/s10096-023-04597-7pmid: 37093325
A ring trial among five European laboratories was organized to reach consistency in microsatellite (MS) typing of the zoonotic parasite Toxoplasma gondii. Three sample sets were circulated and analyzed by each laboratory following a previously published method that is based on fragment length polymorphism of 15 MS markers. The first sample set compared typing results in general and focused on effects of DNA concentration; the second sample set focused on the polymorphic fingerprinting markers that can differentiate T. gondii strains within the same archetypal lineage; and the third set focused on non-archetypal genotypes. Methodological variations between laboratories, including the software programs used to determine MS fragment length, were collated using a questionnaire. Overall, lineage-level typing results reached a high level of agreement, especially in samples with the highest DNA concentrations. However, laboratory-specific differences were observed for particular markers. Major median differences in fragment length, of up to 6 base pairs, were related to the fluorophore used to label fragment-specific primers. In addition, primer pairs with identical sequences obtained from different suppliers resulted in fragments of differing length. Furthermore, differences in the way the sequencing profiles were assessed and interpreted may have led to deviating results in fragment length determination. Harmonization of MS typing, for example, by using the same fluorophores or by numerical adjustments applied to the fragment-lengths determined, could improve the uniformity of the results across laboratories. This is the first interlaboratory comparison, providing guidelines (added as a supplement) for the optimization of this technique.
Incidence, seasonal pattern, and clinical manifestations of Streptococcus dysgalactiae subspecies equisimilis bacteremia; a population-based studyNevanlinna, Viivi; Huttunen, Reetta; Aittoniemi, Janne; Luukkaala, Tiina; Rantala, Sari
doi: 10.1007/s10096-023-04607-8pmid: 37119347
Streptococcus dysgalactiae subspecies equisimilis (SDSE) is a human pathogen causing severe invasive infections. Population-based studies on SDSE bacteremia are limited. The purpose of this study was to investigate the incidence, seasonal pattern, clinical manifestations, and recurrence of SDSE bacteraemia. Records regarding patients aged ≥ 18 years with SDSE bacteremia in the Pirkanmaa health district in August 2015 to July 2018 were retrospectively reviewed. A total of 230 SDSE bacteremia episodes were identified, with 217 episodes (involving 211 patients) available for analysis. The mean annual incidence rate of SDSE bacteremia was 16.9/100 000 inhabitants. Most episodes (33%) were detected in the summer (June to August) (p = 0.058). Episodes with bacteremic cellulitis were statistically significantly more common during the summer compared with other seasons (p = 0.008). Cellulitis was the most common presenting clinical manifestation of SDSE bacteremia (68% of all episodes). Risk factors of recurring bacteremia were chronic eczema and/or skin erosion (OR 3.96 [95% CI 1.11–14.1]), heart disease (OR 3.56 [95% CI 1.22–10.4]), diabetes (OR 3.77 [95% CI 1.35–10.5]) and a history of cellulitis. We found a remarkably high incidence of SDSE bacteraemia in the Pirkanmaa health district. Bacteraemic cellulitis, which was the predominant clinical manifestation is more often occurred in the summer. Risk factors of recurring SDSE bacteremia were a history of cellulitis, chronic eczema or skin erosion, diabetes, and heart disease.
Antenatal pyelonephritis: a three-year retrospective cohort study of two Irish maternity centresBarry, Rachel; Houlihan, Elaine; Knowles, Susan J.; Eogan, Maeve; Drew, Richard J.
doi: 10.1007/s10096-023-04609-6pmid: 37126130
Pyelonephritis affects 1–2% of pregnant women, and is associated with significant maternal and fetal morbidity. Antenatal pyelonephritis has been associated with PPROM (preterm premature rupture of membranes), preterm labour, low birth weight (LBW) and prematurity. A three-year retrospective dual-centre cohort study of antenatal pyelonephritis cases was conducted in two neighbouring Irish maternity hospitals – the Rotunda Hospital (RH) and the National Maternity Hospital (NMH). Patient demographics, clinical presentation, investigations, management and maternal/neonatal outcomes were recorded. A total of 47,676 deliveries (24,768 RH; 22,908 NMH) were assessed. 158 cases of antenatal pyelonephritis were identified (n = 88 RH, n = 70 NMH), with an incidence of 0.33%. The median age was 28 years. The median gestation was 27 + 6 weeks, with 51% presenting before 28 weeks’ gestation. Risk factors included; obesity (18.4%), diabetes mellitus (13.3%) and self-reported clinical history of recurrent urinary tract infection (28.5%). Rate of relapse with UTI in the same pregnancy was 8.2%. Renal ultrasound was performed in 30.4%. Predominant uropathogens were Escherichia coli (60%), Klebsiella pneumoniae (11%) and Proteus mirabilis (5%). 7.5% of cases had a concurrent bloodstream infection, 13.3% of cases were complicated by sepsis and 1.9% with septic shock. Complications including PPROM (6.3%), preterm delivery < 37 weeks’ gestation (11%), LBW < 2,500 g (8.2%) were comparable between sites. Delivery within 72 hours of diagnosis was noted in 7% (n = 11) of patients, of which three were preterm and one had LBW. Appropriate and prompt investigation and management of antenatal pyelonephritis is essential given the associated maternal and neonatal morbidity.
An assessment of the downstream implications of blood culture collection and transitDavies, Peter J. B.; Jones, Timothy P. W.; Macleod, Mairi
doi: 10.1007/s10096-023-04610-zpmid: 37131082
The implications of the variables within the pre-analytical phase of blood culture processing are poorly understood. This study aims to explore the effect of transit times (TT) and culture volume, on time to microbiological diagnosis and patient outcomes. Blood cultures received between 1st March and 31st July 2020/21 were identified. TT, time in incubator (TII), and for positive samples, request to positivity times (RPT) were calculated. Demographic details were recorded for all samples, and culture volume, length of stay (LoS), and 30-day mortality for patients with positive samples. Statistical analysis examined how culture volume and TT effected culture positivity and outcome; in the context of the 4-h national TT target. Totally, 14,375 blood culture bottles were received from 7367 patients; 988 (13.4%) were positive for organisms. There was no significant difference between TT of negative and positive samples. The RPT was significantly lower for samples with TT < 4 h (p < 0.001). Culture bottle volume did not affect RPT (p = 0.482) or TII (p = 0.367). A prolonged TT was associated with a longer length-of-stay in those with a bacteraemia with a significant organism (p = 0.001). We found shorter blood culture transportation time was associated with a significantly faster time of positive culture reporting, while optimal blood culture volume did not make a significant impact. Delays in reporting for significant organisms correspond to a prolonged LoS. Laboratory centralisation makes achieving the 4-h target a logistical challenge; however, this data suggests such targets have significant microbiological and clinical impacts.
Molecular detection of Nocardia: development and application of a real-time PCR assay in sputum and bronchoalveolar lavage fluid samplesWang, Shuai; Wang, Peng; Liu, Jun; Yang, Chunxia; Li, Tianmeng; Yang, Jingxian; Gu, Li; Wei, Ming
doi: 10.1007/s10096-023-04619-4pmid: 37156981
The diagnosis of pulmonary nocardiosis remains challenging. Rapid detection of Nocardia is of primary importance for early diagnosis and precise treatment of nocardiosis. In this study, our objective was to develop and validate a new TaqMan real-time PCR (qPCR) assay for rapidly detecting Nocardia spp. in respiratory samples. Based on published sequence data, primers in a conserved region of the 16S rRNA gene and a probe within that region that was specific for Nocardia were designed. The distinction effect of the qPCR assay was assessed between Nocardia and other respiratory-associated bacteria. Furthermore, the specificity and sensitivity of the assay were evaluated in respiratory clinical samples (n = 205), compared to the results of 16S rRNA gene amplicon sequencing and clinical diagnosis. The qPCR assay exhibited high specificity, sensitivity, repeatability, and reproducibility. The limit of detection of standard plasmid DNA was 3 × 102 copies/mL. Additionally, the qPCR assay was applied to the direct detection of 205 clinical respiratory samples. The specificity and sensitivity of the qPCR were all 100% compared to 16S rRNA gene amplicon sequencing, as well as 98.4% and 100% compared to clinical diagnosis respectively. The qPCR yielded results within 3 h of sample processing, compared to several days for culture, significantly reducing turnaround time. The results suggest that the new qPCR assay developed in this study provides reliable and rapid detection of Nocardia spp. in the respiratory tracts and is expected to reduce the time required for diagnosing and treating nocardiosis.
Epidemiology and antimicrobial resistance profile of Neisseria gonorrhoeae in Catalonia, Spain, 2016–2019Herrero, Mercè; Broner, Sonia; Cruells, Adrià; Esteve, Silvia; Ferré, Lourdes; Mendioroz, Jacobo; Jané, Mireia; Ciruela, Pilar; ,
doi: 10.1007/s10096-023-04601-0pmid: 37162616
Antimicrobial resistance data for Neisseria gonorrhoeae is globally sparse and resistant strains are emerging in Catalonia. We aim to describe epidemiological and antimicrobial resistance in all patients infected with N. gonorrhoeae during the period from 2016 to 2019, using available antimicrobial susceptibility data. We retrospectively analysed confirmed N. gonorrhoeae cases notified to Catalonia’s microbiological reporting system. Antibiotic susceptibility testing (azithromycin, cefixime, ceftriaxone, ciprofloxacin, penicillin, spectinomycin, and tetracycline) was assessed using clinical breakpoints published by the European Committee on Antimicrobial Susceptibility Testing. Incidence rates were calculated and proportions were compared using the χ2 test or Fisher’s exact test, and analysed using the Statistical Package for Social Sciences (SPSS 18.0). A total of 14,251 confirmed cases of N. gonorrhoeae were notified. Incidence increased from 30.7 cases/100,000 person-years (p < 0.001) in 2016 to 64.7 in 2019. Culture was available in 6,292 isolates (44.2%), of which 5,377 (85.5%) were resistant to at least one of the antibiotics tested. Azithromycin resistance rose from 6.1% in 2016 to 16% in 2019 (p < 0.001). Only 1.0% (45 cases) were resistant to ceftriaxone. Multidrug-resistant N. gonorrhoeae increased from 0.25% in 2016 to 0.42% in 2019 (p = 0.521). One case presented extensively drug-resistant N. gonorrhoeae. In Catalonia, 10% of the N. gonorrhoeae isolates were resistant to azithromycin in the 2016–2019 period. According to World Health Organization guidelines, resistance above 5% indicates an alert to review treatment guidelines. Antimicrobial susceptibility testing in clinical practice followed by surveillance and interventions are essential to monitor trends and prevent the spread of antimicrobial resistance.
Assessing the quality of the anaerobic environment — a method developed to support EUCAST disk diffusion of anaerobic bacteriaJustesen, Ulrik Stenz; Åhman, Jenny; Matuschek, Erika; Kahlmeter, Gunnar
doi: 10.1007/s10096-023-04622-9pmid: 37171541
The purpose of this study is to establish a method for assessing the anaerobic environment for EUCAST disk diffusion antimicrobial susceptibility testing (AST) of anaerobic bacteria on fastidious anaerobe agar with 5% mechanically defibrinated horse blood (FAA-HB). The method utilizes the association between a decrease in the metronidazole disk zone diameter and increasing oxygen levels with an aerotolerant Clostridium perfringens strain DSM 25589 (CCUG 75076 and NCTC 14679). The C. perfringens strain was tested on FAA-HB with a McFarland 1 inoculum and a metronidazole 5 μg disk. FAA-HB was incubated for 16–20 h at 35–37°C. The association between oxygen levels (0, 0.16, 1, 2, and 4% oxygen) and the metronidazole zone diameter was determined. Reproducibility at 0% oxygen was investigated as part of a European multi-centre study of disk diffusion of anaerobic bacteria. The median zone diameters (n=12) at each oxygen level were 29 mm (0%), 21 mm (0.16%), 16 mm (1%), 15 mm (2%), and 15 mm (4%). The metronidazole zone diameters at 0% oxygen from the multi-centre reproducibility-study had a median of 29 mm and a 95%-percentile range of 25–33 mm (n=236). Only one reading was below 25 mm. Based on our results, a zone diameter of ≥25 mm using a metronidazole 5 μg disk and the C. perfringens strain, tested with EUCAST recommendations, can be used to indicate that the anaerobic environment is of sufficient quality for culture and disk diffusion AST. EUCAST has included the method as part of the quality control for AST of anaerobic bacteria.