La déferlante COVID-19 : la PCR à la rescousse ! : Chronique d’une gestion de crise au Laboratoire de Microbiologie clinique du CHU de Liège

in Revue Médicale de Liège (2020), 75(S55-S61), 55-61

The SARS-CoV-2 outbreak in December 2019 in China and its expansion across the world and Europe have requested the participation of clinical laboratories as major players in the diagnosis of COVID-19, to ... [more ▼]

The SARS-CoV-2 outbreak in December 2019 in China and its expansion across the world and Europe have requested the participation of clinical laboratories as major players in the diagnosis of COVID-19, to perform PCR tests mainly on nasopharyngeal swabs. In Belgium, the first confirmed COVID-19 patient was diagnosed in early February, the first of many, especially travelers returning from winter sports. In order to meet the ever-increasing demands for testing, the Clinical Microbiology Laboratory of the CHU of Liege had to adapt to this situation: firstly, by developing manual PCR tests and then automated solutions, permitting to increase the number of analyzes by ensuring a short turnaround time of results. Then, a system for the communication of results on a large scale has been set up, and finally solutions to deal with the lack of sampling devices have been found. This first wave of the pandemic has also highlighted an unprecedented solidarity within the institution. In this article, we recount the chronology of the management of this unprecedented health crisis within the Clinical Microbiology Laboratory of the CHU of Liege. [less ▲]

Group B streptococcus neonatal invasive infections in Belgium 2010-2017, and characterization of isolated strains.

in INMIS, International Committee (Ed.) INMIS 2019 Abstract book (2019, September)

Introduction/Background & Aims: Where intrapartum antibiotic-prophylaxis (IAP) is given to pregnant women colonized with Group B Streptococcus (GBS), the incidence of neonatal early-onset disease (EOD ... [more ▼]

Introduction/Background & Aims: Where intrapartum antibiotic-prophylaxis (IAP) is given to pregnant women colonized with Group B Streptococcus (GBS), the incidence of neonatal early-onset disease (EOD) has been successfully reduced; nevertheless, GBS is still the leading cause of severe disease among newborns, notably because the incidence of GBS late-onset disease (LOD) is not affected by IAP. Another strategy such as maternal immunization for prevention of both EOD/LOD is highly desirable worldwide. Aiming to describe GBS epidemiology and characterization of relevant epidemiological markers for vaccine development, surveillance of isolates causing neonatal disease is needed. We provide here results from the Belgian surveillance organized by the National Reference Centre(NRC). Methods: A total of 292 strains of GBS isolated from blood culture/cerebro-spinal fluid of newborns with invasive disease (149 EOD; 143 LOD) were sent to NRC by laboratories of a surveillance network, through years 2010-2017. Capsular-polysaccharide (CPS)-typing and pili-typing were performed with multiplex PCR assays. Multilocus sequence-typing and assignment to the hypervirulent clonal-complex (CC)17 was determined. Results: CPS type III isolates were responsible for 38.9% (n=58) of EOD cases, followed mainly by types Ia, V and II (22.1%, 18.1%, 8.1%). LOD cases were mainly caused by type III isolates (n=107, 74.8%), followed by types Ia (12.6%), V, Ib, IV and II (4.2%, 3.5%,2.8%, 2.1%). These distributions did not vary during the study period. A pili type was assigned to all isolates: at least one pili gene, PI2a, PI2b, or a combination of genes PI1-PI2a and PI1-PI2b. In 2016-2017, the hypervirulent-clone CC17 accounted for 33.3% of EOD (70.4% of type III) and 67% of LOD (89% of type III). Conclusions: The Belgian CPS distributions of isolates from EOD/LOD were similar to European data. One or 2 of 3 pilus-genes were detected in all isolates. CPS type III was predominant in both EOD/LOD and was mainly represented by CC-17 strains. [less ▲]

Evaluation of the Abbott RealTime quantitative CMV and EBV assays using the maxCycle protocol in a laboratory automation context

in Journal of Virological Methods (2019), 270

Real-time PCR are often used for the diagnosis and monitoring of Cytomegalovirus (CMV)and Epstein-Barr virus (EBV)infections in susceptible populations. In this context, we evaluated the analytical ... [more ▼]

Real-time PCR are often used for the diagnosis and monitoring of Cytomegalovirus (CMV)and Epstein-Barr virus (EBV)infections in susceptible populations. In this context, we evaluated the analytical performances of the Abbott RealTime CMV/EBV maxCycle protocol automated on the m2000 platform (Abbott). It was compared to our routinely-used procedure consisting of a NucleoMag® DNA extraction automated on a STARlet platform followed by manually processed CMV and EBV quantitative real-time PCR (Diagenode). In this study, we showed that both EBV assays exhibited a similar sensitivity but with a better precision for the EBV Abbott RealTime assay. For the CMV performances, the Abbott assay was more sensitive and more precise than our routine method. The use of WHO International Standards also indicated a slight underestimation of the viral loads (−0.25 log10 IU/mL and −0.21 log10 IU/mL for CMV and EBV assays respectively)while these were rather overestimated with the Starlet/Diagenode method (0.48 log10 IU/mL and 0.19 log10 IU/mL for CMV and EBV assays respectively). These trends were confirmed using relevant whole-blood clinical samples and external quality controls. The workflows were also compared and we highlighted a significant technician hands-on time reduction (−63%)using the Abbott CMV/EBV maxCycle automated protocol. © 2019 Elsevier B.V. [less ▲]

Retour de l'ECCMID

Scientific conference (2018, May 29)

VALIDATION, IMPLEMENTATION AND ROUTINE WORK WITH TOTAL LAB-AUTOMATION BD-KIESTRA : A TWO-YEAR EXPERIENCE AT THE UNIVERSITY HOSPITAL OF LIÈGE, BELGIUM

Poster (2018, April 21)

Background: Automation in Bacteriology is a major revolution in clinical microbiology laboratories. At the university hospital of Liège (Belgium), a BD-Kiestra Total Laboratory Automation (BD-K-TLA ... [more ▼]

Background: Automation in Bacteriology is a major revolution in clinical microbiology laboratories. At the university hospital of Liège (Belgium), a BD-Kiestra Total Laboratory Automation (BD-K-TLA) system has been installed in September 2015. Here, we describe the validation process achieved for the implementation of the BD-K-TLA system, and our two-year experience in the daily practice of the laboratory. Materials/methods: We present the validation plan (i.e. all the parameters that have to be checked before TLA-BD-K implementation) established by the Key User Group BD-Kiestra. In order to evaluate some of the added-value of this system compared to full-manual bacteriology, we measured two parameters for two similar period, before (February 2015) and after (February 2017) implementation of the BD-K-TLA: (1) the positivity rate (%), defined as the number of significant positive cultures among the total of samples analyzed and (2) the time-to-result (TTR), defined as the time (in hours) between reception of a specimen in the microbiology laboratory and the isolation of a significant pathogen in this sample. All specimens were included, except respiratory samples and blood cultures. Results: Validation plan included the checking process of each of the modules of BD-K-TLA: BarcodA, InoqulA, ProceedA, ReadA Compact and ReadA Browser. Furthermore, specimens were tested in parallel, on the BD-K-TLA and with full-manual bacteriological techniques: a minimum of 95% agreement was observed for qualitative and semi-quantitative assessment of Gram stain and cultures. The positivity rate went from 19,7% (pre-implementation – 1169 of 5941 samples analyzed), to 21,8% (post-implementation – 1391 of 6381 samples analyzed). Mean TTR for the specimens was of 26,95 h in February 2015, while it was 26,83 h in February 2017. Looking at these two measures, no significant time saving is currently observed. However, TLA-BD-K allows traceability through the whole process, quality of inoculation, fixed incubation times and better biological validation by clinical microbiologists thanks to remote pictures review. Conclusions: Working with BD-K-TLA allows a better quality and traceability of the bacteriological results, with a major benefit of remote pictures review. In order to observe time saving with BD-K-TLA, a well-considered configuration of the instrument and major reorganization of the workflow are mandatory. [less ▲]

Coulisse de la santé du patient : Dédale des prélèvements: 
de l’analyse microbiologique 
au diagnostic

Conference given outside the academic context (2018)

Parcours d'un échantillon destiné à des analyses microbiologiques: différentes étapes pour aboutir à un diagnostic. Diaporama complété par une video.

2015 Surveillance of Group B Streptococcus strains isolated from invasive diseases among adults in Belgium : bacteriological and clinical characteristics.

in Abstract book of 1st ISSAD 2018 (2018)

Background Even if Group B streptococcus (GBS) disease risk is highest during the first 3 months of life, GBS also causes significant morbidity and mortality among adults, especially in the elderly and ... [more ▼]

Background Even if Group B streptococcus (GBS) disease risk is highest during the first 3 months of life, GBS also causes significant morbidity and mortality among adults, especially in the elderly and immunocompromised or with chronic illnesses. We here provide an overview of bacteriological and clinical characteristics of GBS causing invasive diseases in non-pregnant adults in Belgium. Methods Overall 143 GBS strains isolated from invasive diseases among non-pregnant adults sent, on a voluntary-base, to the National Reference Centre (NRC) during the year 2015 by any laboratory located in Belgium were characterized: capsular polysaccharide (CPS)-typing by agglutination and/or with PCR, pili-typing with PCR, antimicrobial susceptibility testing, and detection of resistance genes with PCR. Results Among adult invasive isolates, CPS-type Ia was predominant (25.1%) followed by V, III, II, IV, Ib (23%, 19.6%, 9.8%, 9.8%, 7%) and VI, VII, VIII (1.4% each), IX (0.7%). One strain remained non typeable even with PCR. All strains were susceptible to penicillin. Rate of resistance to macrolides/lincosamides was 35.7%. ErmB, ErmTr and MefA genes were detected respectively within 45.2%, 27.4% and 19.5% among the resistant strains. One strain presenting the L profile (isolated resistance to clindamycin) harboured the LsaC gene. About pili-typing, the combined PI1, PI2a genes were predominant with 51.2% of the cases, followed by PI2a alone, the combined PI1, PI2b and PI2b alone (33.3%, 14.7%, 1.4%). The vast majority of strains were isolated from blood: 43% of bacteremia without reported focus, 19% originating from skin-and-soft tissue infections, 4.9% urosepsis, 4.2% bone-and-join infections, 2.8% endocarditis and 1.4% meningitis. Conclusion Among invasive adult diseases, GBS bacteriological characteristics were consistent with reported data among European countries. Macrolides/lincosamides resistance rate has slightly increased. Bacteremia without identified focus and skin-soft tissue infections were highly predominant; meningitis and endocarditis were less common but associated with serious morbidity and mortality. [less ▲]

A propos de diphtérie

Scientific conference (2017, December 05)

Cette présentation relate un cas clinique de diphtérie respiratoire à Corynebacterium ulcerans, et aborde la pathogénèse, les manifestations cliniques, l'épidémiologie, le diagnostic microbiologique ainsi ... [more ▼]

Cette présentation relate un cas clinique de diphtérie respiratoire à Corynebacterium ulcerans, et aborde la pathogénèse, les manifestations cliniques, l'épidémiologie, le diagnostic microbiologique ainsi que la prise en charge des close contacts en lien avec cette bactérie. [less ▲]

Evolution of rate and genotypes of resistance to macrolide/lincosamide among invasive Group B Streptococcus (GBS): Development of a multiplex PCR tool for simultaneous detection of ErmB, ErmTr, MefA and LsaC resistance genes.

Poster (2017, October)

Methods: A multiplex-PCR, using a set of specifically designed or already described (Kataja, 1999; Malbruny, 2011) primers was developed and used to detect, in GBS, three genes for erythromyc resistance ... [more ▼]

Methods: A multiplex-PCR, using a set of specifically designed or already described (Kataja, 1999; Malbruny, 2011) primers was developed and used to detect, in GBS, three genes for erythromyc resistance, ermB, ermTR, mefA and one gene for clindamycin-resistance lsaC. AdhP gene amplification was used as control for GBS identification. All(219) GBS isolates from invasive infections in newborns and adults received by the Belgian National Reference Center for GBS in 2015, and control strains were tested for erythromycin/clindamycin susceptibility (disk-diffusion/broth- microdilution) and for detection of resistance genes. Results: PCR products demonstrated the expected respective sizes. The method has been validated successfully according to ISO15189 analytical requirements. Of the 219 isolates, 67(30,67%) w resistant to erythromycin and/or clindamycin: 44/67(65,78%) showed a constitutive-MLS phenotype and 10/67(14,9%) the inducible-MLS phenotype. Among the constitutive-MLS strains, 73% harboured ErmB gene, 13% ErmTR, 7% ErmB+mefA and 7% ermB together with LsaC gene. The inducible-MLS strains harboured mostly ErmTr gene (89%) and the others the ErmB gene. Among the 10/67(14,9%) GBS strains with an M-phenotype (isolated resistance to erythromycin), the MefA gene was exclusively detected. Among the 3(4,48%) strains showing an isolated resistance to clindamycin (L-phenotype), the LsaC gene was detected. Conclusion: The developed multiplex PCR is able to detect simultaneously four genes involved in MLS resistance in GBS. In 2015, 30,6% of the invasive GBS strains isolated in Belgium were resist to macrolides and/or lincosamides. The emergence of the L-phenotype in GBS described since 2010, justifies the relevance to also detect LsaC gene together with ErmB, ErmTr and MefA. [less ▲]

A 5-year survey of dermatophytes strains circulating in Belgium

Poster (2017, October)

Objectives Dermatophytosis refers to superficial fungal infections of keratinized tissues caused by keratinophilic dermatophytes. They are the most common cause of superficial fungal infections worldwide ... [more ▼]

Objectives Dermatophytosis refers to superficial fungal infections of keratinized tissues caused by keratinophilic dermatophytes. They are the most common cause of superficial fungal infections worldwide. Epidemiological studies regarding dermatophytes infections have been conducted in several countries and differences in the incidence and in etiological agents have been reported. That is why national surveillance of circulating strains causing dermatophytosis is critical. The National Reference Center (NRC) for mycoses conducted a survey on dermatophytes strains circulating in Belgium from 2012 to 2016. The present study was performed to assess the profile of dermatophytosis and to identify the species involved. Methods The Belgian NRC for mycoses (Leuven and Liège) collected 14227 strains between January 2012 and December 2016. The strains were obtained from clinically suspected fungal infections of skin, hair and nails. Strains were collected from Belgian laboratories in order to confirm the fungal identification which was performed by microscopy and in case of doubt by ITS sequencing. Results Among the 14227 samples collected, 6248 were identified as dermatophytes (44%). Trichophyton rubrum was the most prevalent species accounting for 61,3% (n=3820) of the strains collected from all sources, followed by T. mentagrophytes complex (19,2%, n=1199) according to the ancient classification (including T. interdigitale, T. benhamiae and T. mentagrophytes). Other less prevalent species were also recorded: M. audouinii (n=507, 8,1%), M. canis (n=210, 3,3%), T. tonsurans (n= 140, 2,2%), T. violaceum (n=133, 2,1%), T. soudanense (n=125, 2%), M. praecox (n=60, 0,96%) and E. floccosum (n=19, 0,3%) for the main ones. Our data show the predominance of anthropophilic species causing tinea capitis especially M. audouinii responsible for 43,4% (n=303/716) of hair/scalp infection with an increasing number from 2014 to 2016. Trichophyton soudanense, rarely observed in Belgium in the past, is an emerging agent of tinea capitis particularly since 2013, accounting for 11,3% (n=81) of the cases during the 5-year study period. Zoophilic strains such as M. canis which were well represented in the past epidemiology of tinea capitis are decreasing accounting for only 8,8% (n=63) of hair/scalp infection. Finally, our data confirm the high prevalence of T. rubrum as the main etiologic agent of onychomycosis (78,1%, n=3094/3968) followed by T. mentagrophytes complex (18,8%, n=743/3968). Both latter strains were also responsible for the majority of skin infections as they were isolated respectively in 46,2% (n=693/1612) and 21,7% (n=348/1612) of skin samples. Conclusions The present epidemiological survey provides recent data on the prevalence of all dermatophytes circulating in Belgium. Analyzing such data is critical for the establishment of measures for prevention and control of dermatophytes infections. Our study confirms the predominance of T. rubrum followed by species from the ancient T. mentagrophytes complex (T. interdigitale + T. benhamiae) in the Belgian population. This survey highlights also the persistent predominance of M. audouinii and the emergence of T. soudanense as causative agents of tinea capitis. [less ▲]