Emergence of methicillin resistance predates the clinical use of antibiotics.

The discovery of antibiotics more than 80 years ago has led to considerable improvements in human and animal health. Although antibiotic resistance in environmental bacteria is ancient, resistance in human pathogens is thought to be a modern phenomenon that is driven by the clinical use of antibiotics1. Here we show that particular lineages of methicillin-resistant Staphylococcus aureus-a notorious human pathogen-appeared in European hedgehogs in the pre-antibiotic era. Subsequently, these lineages spread within the local hedgehog populations and between hedgehogs and secondary hosts, including livestock and humans. We also demonstrate that the hedgehog dermatophyte Trichophyton erinacei produces two ß-lactam antibiotics that provide a natural selective environment in which methicillin-resistant S. aureus isolates have an advantage over susceptible isolates. Together, these results suggest that methicillin resistance emerged in the pre-antibiotic era as a co-evolutionary adaptation of S. aureus to the colonization of dermatophyte-infected hedgehogs. The evolution of clinically relevant antibiotic-resistance genes in wild animals and the connectivity of natural, agricultural and human ecosystems demonstrate that the use of a One Health approach is critical for our understanding and management of antibiotic resistance, which is one of the biggest threats to global health, food security and development.

Spiroplasma infection as a cause of severe congenital keratouveitis, cataract and glaucoma.

BACKGROUND: Only seven cases of ocular Spiroplasma infection have been reported to date, all presenting as congenital cataracts with concomitant intraocular inflammation. We describe the first case of Spiroplasma infection initially presenting as a corneal infiltrate. CASE PRESENTATION: A 1-month-old girl was referred for a corneal infiltrate in the left eye. She presented in our hospital with unilateral keratouveitis. Examination showed a stromal corneal infiltrate and dense white keratic precipitates in the left eye. Herpetic keratouveitis was suspected and intravenous acyclovir therapy was initiated. Two weeks later, the inflammation in the left eye persisted and was also noticed in the right eye. Acute angle-closure glaucoma and a cataract with dilated iris vessels extending onto the anterior lens capsule developed in the left eye. The inflammation resolved after treatment with azithromycin. Iridectomy, synechiolysis and lensectomy were performed. Bacterial metagenomic sequencing (16 S rRNA) and transmission electron microscopy revealed Spiroplasma ixodetis species in lens aspirates and biopsy. Consequently, a diagnosis of bilateral Spiroplasma uveitis was made. CONCLUSIONS: In cases of congenital cataract with concomitant intraocular inflammation, Spiroplasma infection should be considered. The purpose of this case report is to raise awareness of congenital Spiroplasma infection as a cause of severe keratouveitis, cataract and angle-closure glaucoma in newborns. Performing molecular testing on lens aspirates is essential to confirm diagnosis. Systemic macrolides are suggested as the mainstay of treatment.

Combined oropharyngeal/nasal swab is equivalent to nasopharyngeal sampling for SARS-CoV-2 diagnostic PCR.

BACKGROUND: Early 2020, a COVID-19 epidemic became a public health emergency of international concern. To address this pandemic broad testing with an easy, comfortable and reliable testing method is of utmost concern. Nasopharyngeal (NP) swab sampling is the reference method though hampered by international supply shortages. A new oropharyngeal/nasal (OP/N) sampling method was investigated using the more readily available throat swab. RESULTS: 35 patients were diagnosed with SARS-CoV-2 by means of either NP or OP/N sampling. The paired swabs were both positive in 31 patients. The one patient who tested negative on both NP and OP/N swab on admission, was ultimately diagnosed on bronchoalveolar lavage fluid. A strong correlation was found between the viral RNA loads of the paired swabs (r = 0.76; P < 0.05). The sensitivity of NP and OP/N analysis in hospitalized patients (n = 28) was 89.3% and 92.7% respectively. CONCLUSIONS: This study demonstrates equivalence of NP and OP/N sampling for detection of SARS-CoV-2 by means of rRT-PCR. Sensitivity of both NP and OP/N sampling is very high in hospitalized patients.

Pitfalls in the detection of myositis specific antibodies by lineblot in clinically suspected idiopathic inflammatory myopathy.

OBJECTIVES: Today, the contribution of myositis-specific autoantibodies (MSA) in the diagnostic workup of idiopathic inflammatory myopathies (IIM) is on the rise. The aim of this study was to document MSA frequency as detected by lineblot in a set of consecutive MSA requests and to correlate the results with clinical diagnosis, IIM subtype and indirect immunofluorescence (IIF) findings. Additionally, a comparison between two lineblots was performed. METHODS: A total of 118 consecutive samples of patients with suspicion of IIM were analysed on IIF and two lineblots. A total of 107 patients with autoimmune rheumatic diseases served as controls. RESULTS: MSA were detected in 55% of IIM patients (n=31) and 7.9% (n=12) of patients without clinical diagnosis of IIM or myositis overlap syndrome. All the IIM patients had a MSA-compatible clinical subtype. There was no to fair agreement between both lineblots for the individual antibodies, with most discrepancies observed for anti-TIF1γ (κ=-0.021), anti-SRP (κ=-0.006) and anti-SAE (κ=0.395). Differences between both assays were mostly observed in the non-IIM patients, also showing signi cantly lower blot signal intensities compared to IIM patients (p=0.0013). MSA in the non-IIM patients frequently showed an incompatible IIF pattern. CONCLUSIONS: Lineblot seems to be an interesting tool for MSA detection in a clinical context, allowing the identification of clinical subtypes. However, considerable caution must be exercised in interpreting the results in case of low positive MSA signal intensity, discordant lineblot results and/or an incompatible IIF pattern.

National surveillance of Staphylococcus epidermidis recovered from bloodstream infections in Belgian hospitals.

OBJECTIVES: The objectives of this study were: (i) to determine the species diversity of CoNS isolated from bloodstream infections collected during a national surveillance study; and (ii) to examine the antimicrobial resistance and genomic diversity among Staphylococcus epidermidis isolates. METHODS: Eighty CoNS were identified by MALDI-TOF. Antimicrobial resistance determination, molecular characterization of resistance and virulence genes, and molecular typing were performed for S. epidermidis isolates. RESULTS: The majority (76%) of CoNS were identified as S. epidermidis. Among these S. epidermidis, 77% were resistant to methicillin [methicillin-resistant S. epidermidis (MRSE)] and showed multiresistance to other antimicrobials. Genes implicated in resistance were erm(C), erm(A) and msr(A) for erythromycin, aacA-aphD and aadC for aminoglycosides, tet(K) for tetracycline and mupA for high-level resistance to mupirocin. Molecular typing showed that 34/40 MRSE isolates (85%) belonged to clonal complex (CC) 2 that could be subdivided into CC2-I (ST2) and CC2-II (ST5, ST59 and ST88). In contrast, methicillin-susceptible S. epidermidis displayed high genomic diversity. The majority (70%) of S. epidermidis isolates contained an icaA or arcA gene. The icaA gene was found in the CC2-I subgroup, whereas arcA was more common in methicillin-susceptible S. epidermidis. CONCLUSIONS: S. epidermidis was frequently recovered among CoNS isolated from bloodstream infections with a high proportion of MRSE being multiresistant. A large number of S. epidermidis belonged to CC2, a clone that is disseminated worldwide. More studies are needed to understand its clonal evolutionary success.

Evaluation of the automated Vitek 2 system for detection of various mechanisms of macrolide and lincosamide resistance in Staphylococcus aureus.

We evaluated the performance of the automated Vitek 2 system against disk diffusion for susceptibility testing of Staphylococcus aureus strains showing various resistance mechanisms to macrolides and lincosamides (ML). The Vitek 2 system showed 100% concordance with the D-zone test in detection of the most common resistance mechanisms to ML, including methylase and efflux systems.

Genetic diversity among methicillin-resistant Staphylococcus aureus isolates carrying the mecC gene in Belgium.

OBJECTIVES: A mecA homologue gene, named mecC, has been reported in methicillin-resistant Staphylococcus aureus (MRSA) isolates from humans and from diverse animal species. We investigated the proportion, and the phenotypic and genotypic characteristics, of mecC-carrying MRSA recovered from humans in Belgium. METHODS: A total of 4869 S. aureus isolates, collected by the National Reference Centre from 2003 to 2012, were retrospectively analysed for the presence of mecC. The mecC-carrying MRSA isolates were tested for phenotypic resistance and the presence of toxin genes. Genotyping was performed using spa typing and multilocus sequence typing. RESULTS: Nine S. aureus isolates, mecA negative but cefoxitin resistant (MIC 16-64 mg/L), were found to carry the mecC gene. Among these, eight showed resistance to oxacillin (MIC 4-64 mg/L). These isolates remained fully susceptible to all non-ß-lactam antimicrobials. Although the proportion of mecC-carrying MRSA in Belgium was low (<1% per year), mecC-MRSA were assigned to three distinct genetic lineages corresponding to clonal complex (CC) 130, CC49 and CC1943. CONCLUSIONS: This first Belgian nationwide analysis showed a low occurrence of mecC-MRSA. Further studies should be conducted to better understand the reservoirs and risk factors for mecC-MRSA acquisition.

High genetic diversity of methicillin-susceptible Staphylococcus aureus (MSSA) from humans and animals on livestock farms and presence of SCCmec remnant DNA in MSSA CC398.

OBJECTIVES: To investigate the genetic diversity of methicillin-susceptible Staphylococcus aureus (MSSA) carriage isolates from animals and humans on pig, veal, dairy, beef and broiler farms. METHODS: S. aureus isolates were genotyped using spa typing and multilocus sequence typing (MLST). Antimicrobial susceptibility phenotypes and genotypes were determined. The presence of staphylococcal cassette chromosome mec (SCCmec)-associated DNA was characterized by PCR and sequencing among isolates of clonal complex (CC) 398. RESULTS: Overall, 41 MSSA isolates in humans and 141 in animals were found, originating from all farm types. These MSSA were mainly assigned to CC398, CC1, CC5, CC9, CC30, CC97, CC133 and CC705/151. MSSA CC398 showed resistance to tetracycline, trimethoprim, macrolides and/or lincosamides, aminoglycosides, ciprofloxacin, spectinomycin and quinupristin/dalfopristin, whereas non-CC398 MSSA showed considerably less resistance. Three porcine MSSA CC398-t011 isolates harboured remnant DNA of a composite SCCmec V(5C2&5)c element that lacked the mec gene complex. This resulted from an MRSA-to-MSSA conversion due to recombination between the ccrC genes flanking the mec gene complex. The SCC remnant still contained an intact J1 region harbouring czrC and tet(K), encoding zinc and tetracycline resistance, respectively, thereby illustrating the capacity of S. aureus CC398 to adapt to different antibiotic selection pressures in the farming environment. Processes such as mec gene complex deletion probably contribute to the enormous diversity of SCC(mec) elements observed in staphylococci. CONCLUSIONS: MSSA CC398 precursors from which MRSA CC398 might (re)emerge were present on pig, veal and broiler farms, all of which are livestock sectors commonly known to be affected by MRSA CC398. The multiresistance phenotype of S. aureus CC398 appears to be independent of methicillin resistance.

The use of information technology to improve interdisciplinary communication during infectious diseases ward rounds on the paediatric intensive care unit.

Prospective audit with feedback during infectious diseases ward rounds (IDWR) is a common antimicrobial stewardship (AMS) practice on the Paediatric Intensive Care Unit (PICU). These interdisciplinary meetings rely on the quality of handover, with high risk of omission of information. We developed an electronic platform integrating infection-related patient data (COSARAPed). In the mixed PICU of a Belgian tertiary hospital we conducted an observational prospective cohort study comparing patient handovers during IDWRs using the COSARAPed-platform to those with access only to conventional resources. The quality of handover was investigated directly by assessment if the narrative was in accordance with Situation-Background-Assessment-Recommendation principles and if adequate demonstration of diagnostic information occurred, and also indirectly by registration if this was only achieved after intervention by the non-presenting AMS team members. We also recorded all AMS-recommendations. During a 6-month study period, 24 IDWRs and 82 patient presentations were assessed. We could only find a statistically significant advantage in favor of COSARAPed by indirect evaluation. We registered 92 AMS-recommendations, mainly resulting in reduced antibiotic pressure. We concluded that the IDWR is an appropriate platform for AMS on the PICU and that the utilisation of COSARAPed may enhance the quality of patient handover.

Prevalence, risk factors and genetic diversity of methicillin-resistant Staphylococcus aureus carried by humans and animals across livestock production sectors.

OBJECTIVES: This study aimed to assess the prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in animals and humans on veal, dairy, beef and broiler farms and to compare the risk for human MRSA carriage with that of strictly horticulture farmers. The genetic background, resistance phenotypes and genotypes and toxin gene content of the isolated MRSA strains were compared with MRSA collected on MRSA clonal complex (CC)398-positive pig farms. METHODS: MRSA carriage isolates were genotyped (spa, SCCmec and multilocus sequence typing), resistance to 16 antimicrobials was determined and resistance and toxin genes were detected. RESULTS: MRSA carriage rates were higher (P<0.01) on veal farms (calves, 64%; farmers, 72%) compared with on dairy (cows, 1%), beef (cows, 5%; farmers, 11%), broiler (pooled broths, 5%; farmers, 3%) and horticulture (farmers, 3%) farms. The intensity of animal contact was identified as a risk factor for human MRSA carriage. The vast majority of MRSA (n=344), including those from pigs, were CC398 (98%). SCCmec V(5C2), V(5C2&5)c, IV(2B) and IV(2B&5) predominated. MRSA CC130 and CC599 carrying mecC were detected in beef and dairy cattle. MRSA from veal calves were significantly more resistant than MRSA from pigs (P<0.01). A few isolates, including mecC-carrying MRSA, harboured pyrogenic superantigen toxins. Human- and animal-derived MRSA from individual farms showed similar characteristics. CONCLUSIONS: This systematic cross-sector survey revealed a high prevalence of multiresistant livestock-associated MRSA on Belgian veal calf farms as compared with other farm types. MRSA harbouring mecC was detected at a low frequency in beef and dairy cows, but not in humans.

Characterization of methicillin-resistant non-Staphylococcus aureus staphylococci carriage isolates from different bovine populations.

OBJECTIVES: This study aimed at investigating bovine non-Staphylococcus aureus staphylococci for their role as a potential reservoir for methicillin resistance. METHODS: Nasal swab samples were collected from 150 veal calves on 15 veal farms, 100 dairy cows on 10 dairy farms and 100 beef cows on 10 beef farms. Suspected staphylococcal isolates were investigated by PCR for the presence of the classic mecA and mecA(LGA251). Methicillin-resistant non-S. aureus staphylococci (MRNAS) were genotypically identified and were characterized by broth microdilution antimicrobial susceptibility testing and staphylococcal cassette chromosome mec (SCCmec) typing. RESULTS: The MRNAS (n = 101) carriage rate was estimated as 30.29% (95% CI 6.14%-74.28%) in veal calves, 13.1% (95% CI 1.28%-63.72%) in dairy cows and 24.8% (95% CI 11.97%-44.42%) in beef cows. Carriage rates were not significantly different between the three populations (P > 0.05). mecA(LGA251) was not detected. Most (n = 80) MRNAS were identified as Staphylococcus sciuri, Staphylococcus lentus or Staphylococcus fleurettii. Resistance to aminoglycosides, macrolide-lincosamide-streptogramin antimicrobials, tetracycline and ciprofloxacin was frequently detected. Two linezolid-resistant MRNAS from veal calves carried the multidrug-resistance gene cfr. SCCmec cassettes of type III predominated (n = 46); another 40 SCCmec cassettes harboured a class A mec complex without identifiable ccr complex; type IVa, type V and several other non-typeable cassettes were detected in low frequencies, especially in methicillin-resistant Staphylococcus epidermidis. CONCLUSIONS: The SCCmec types predominating in bovine MRNAS differ from those mostly detected in livestock-associated methicillin-resistant S. aureus strains. Yet, the detection of cfr and the high level of other antimicrobial resistances suggest a potentially important role of bovine MRNAS as a reservoir for resistance determinants other than SCCmec.

EUCAST rapid antimicrobial susceptibility testing (RAST) compared to conventional susceptibility testing: implementation and potential added value in a tertiary hospital in Belgium.

OBJECTIVES: EUCAST breakpoints for short incubation disk diffusion allow rapid antimicrobial susceptibility testing (RAST) directly from positive blood cultures. We evaluate the RAST methodology and assess its potential added value in a setting of low prevalence of multidrug-resistant (MDR) organisms. METHODS: In our two-part study, we performed RAST on 127 clinical blood cultures at 6 and 8 h and determined categorical agreement with direct susceptibility testing. We also measure the impact of susceptibility results on antimicrobial therapy compared to empirical treatment. RESULTS: Categorical agreement was 96.2% at 6 h (575/598 isolate-drug combinations) and 96.6% at 8 h (568/588 combinations). Major errors involved piperacillin/tazobactam in 16 of 31 cases. The second part of our study shows that AST reporting proved essential in correcting ineffective empirical therapy in 6.3% of the patients (8/126). CONCLUSION: EUCAST RAST is an inexpensive and reliable method of susceptibility testing, although care must be taken with reporting piperacillin/tazobactam. In support of RAST implementation, we show that AST remains of great importance in providing effective therapy, even in a setting of low MDR prevalence and elaborate antibiotic guidelines.

Sanitary installations and wastewater plumbing as reservoir for the long-term circulation and transmission of carbapenemase producing Citrobacter freundii clones in a hospital setting.

BACKGROUND: Accumulating evidence shows a role of the hospital wastewater system in the spread of multidrug-resistant organisms, such as carbapenemase producing Enterobacterales (CPE). Several sequential outbreaks of CPE on the geriatric ward of the Ghent University hospital have led to an outbreak investigation. Focusing on OXA-48 producing Citrobacter freundii, the most prevalent species, we aimed to track clonal relatedness using whole genome sequencing (WGS). By exploring transmission routes we wanted to improve understanding and (re)introduce targeted preventive measures. METHODS: Environmental screening (toilet water, sink and shower drains) was performed between 2017 and 2021. A retrospective selection was made of 53 Citrobacter freundii screening isolates (30 patients and 23 environmental samples). DNA from frozen bacterial isolates was extracted and prepped for shotgun WGS. Core genome multilocus sequence typing was performed with an in-house developed scheme using 3,004 loci. RESULTS: The CPE positivity rate of environmental screening samples was 19.0% (73/385). Highest percentages were found in the shower drain samples (38.2%) and the toilet water samples (25.0%). Sink drain samples showed least CPE positivity (3.3%). The WGS data revealed long-term co-existence of three patient sample derived C. freundii clusters. The biggest cluster (ST22) connects 12 patients and 8 environmental isolates taken between 2018 and 2021 spread across the ward. In an overlapping period, another cluster (ST170) links eight patients and four toilet water isolates connected to the same room. The third C. freundii cluster (ST421) connects two patients hospitalised in the same room but over a period of one and a half year. Additional sampling in 2022 revealed clonal isolates linked to the two largest clusters (ST22, ST170) in the wastewater collection pipes connecting the rooms. CONCLUSIONS: Our findings suggest long-term circulation and transmission of carbapenemase producing C. freundii clones in hospital sanitary installations despite surveillance, daily cleaning and intermittent disinfection protocols. We propose a role for the wastewater drainage system in the spread within and between rooms and for the sanitary installations in the indirect transmission via bioaerosol plumes. To tackle this problem, a multidisciplinary approach is necessary including careful design and maintenance of the plumbing system.

Staphylococcus aureus CC398 clade associated with human-to-human transmission.

Staphylococcus aureus clonal complex 398 (CC398) isolates colonize livestock and can spread to human contacts. Genetic analysis of isolates epidemiologically associated with human-to-human, but not livestock, transmission in multiple countries and continents identified a common clade that was negative for tet(M) and positive for bacteriophage 3. Another group of human-to-human-transmitted isolates belonged to the common livestock-associated clade but had acquired a unique 7 bacteriophage.

Different long-term avidity maturation for IgG anti-spike and anti-nucleocapsid SARS-CoV-2 in hospitalized COVID-19 patients.

INTRODUCTION: The high variability of SARS-CoV-2 serological response after COVID-19 infection hampers its use as indicator of the timing of infection. A potential alternative method is the determination of affinity maturation of SARS-CoV-2 IgG, expressed as the SARS-CoV-2 IgG avidity. METHODS: SARS-CoV-2 IgG concentration and avidity were measured in sera of hospitalized COVID-19 patients sampled at two weeks and ≥12 weeks post symptom onset using an in-house developed protocol based on EUROIMMUN (anti-spike) and EDI™ (anti-nucleocapsid) SARS-CoV-2 IgG ELISA protocols. RESULTS: We included 68 confirmed COVID-19 patients that tested positive for SARS-CoV-2 IgG in both the initial and follow-up specimen sampled at a median of 14 (range 10-18) days and 120 (range 84-189) days, respectively, post symptom onset. The median anti-spike and anti-nucleocapsid SARS-CoV-2 IgG avidity response was 40% (range 9-93%) and 72% (range 27-104%), respectively, for the first sample, and 66% (range 28-90%) and 57% (range 25-94%), respectively, for the second sample. The proportion of SARS-CoV-2 IgG avidity results ≥60% was significantly lower for anti-spike compared to anti-nucleocapsid IgG for initial samples (p< 0.01) and vice versa for follow-up samples (p< 0.01). CONCLUSION: Anti-nucleocapsid SARS-CoV-2 IgG maturation occurs faster and avidity decreases faster than anti-spike IgG, indicating different kinetics of anti-spike and anti-nucleocapsid IgG. Further, affinity maturation after SARS-CoV-2 infection is frequently incomplete.

Effects of cascade reporting of susceptibility profiles for Enterobacterales on broad-spectrum antibiotics use and resistance.

OBJECTIVE: To reduce the inappropriate use of broad-spectrum antibiotics in a 1000+ bed acute tertiary care hospital by the introduction of cascade antimicrobial susceptibility reporting for Enterobacterales. METHODS: Over a 1-year period, we selectively suppressed reporting of susceptibility to the broad-spectrum antibiotics piperacillin-tazobactam (TZP) and meropenem (MEM) for Enterobacterales strains susceptible to amoxicillin-clavulanic acid (AMC) and negative for extended-spectrum ß-lactamase (ESBL). We measured the effects on hospital-wide antibiotic consumption (defined daily doses/1000 admissions) and resistance of Escherichia coli and Klebsiella pneumoniae on two levels. First, we compared resistance and antibiotic use for the antibiotics impacted by the intervention (AMC, TZP and MEM) with control antibiotics that were consistently reported (fluoroquinolones, trimethoprim-sulfamethoxazole and third-generation cephalosporins). Second, we compared the resistance for TZP and MEM with a control pathogen (Pseudomonas aeruginosa) and studied the impact on rate of Clostridioides difficile-associated diarrhoea in our hospital. RESULTS: We observed an overall increased use of AMC relative to overall antibiotic consumption (20.0%, p<0.0001) together with a decreased use of TZP (-11.9%, p=0.049) and unchanged use of MEM (p=0.68) relative to overall antibiotic consumption. As for resistance, the number of ESBL-positive K. pneumoniae strains diminished by 5.9% (p<0.0001). When focusing on intensive care units, the carbapenemase-producing Enterobacterales (CPE) rate also decreased by 4.5% (p=0.0091). For E. coli, no significant difference in ESBL (p=0.33) and CPE (p=0.48) rates were observed. No significant difference in the rate of C. difficile infections was observed (p=0.40). CONCLUSIONS: Restricted susceptibility reporting of TZP and MEM was associated with a significant increased use of AMC and decreased use of TZP relative to overall antibiotic consumption and significant reduction in ESBL- and CPE-positive K. pneumoniae strains.

Lower persistence of anti-nucleocapsid compared to anti-spike antibodies up to one year after SARS-CoV-2 infection.

We retrospectively compared the long-term evolution of IgG anti-spike (S) and anti-nucleocapsid (N) levels (Abbott immunoassays) in 116 non-severe and 115 severe SARS-CoV-2 infected patients from 2 university hospitals up to 365 days post positive RT-PCR. IgG anti-S and anti-N antibody levels decayed exponentially up to 365 days after a peak 0 to 59 days after positive RT-PCR. Peak antibody level/cut-off ratio 0 to 59 days after positive RT-PCR was more than 70 for anti-S compared to less than 6 for anti-N (P < 0.01). Anti-S and anti-N were significantly higher in severe compared to non-severe patients up to 180 to 239 days and 300 to 365 days, respectively (P < 0.05). Despite similar half-lives, the estimated time to 50% seronegativity was more than 2 years for anti-S compared to less than 1 year for anti-N in non-severe and severe COVID-19 patients, due to the significantly higher peak antibody level/cut-off ratio for anti-S compared to anti-N.

Sensitivity and specificity of 14 SARS-CoV-2 serological assays and their diagnostic potential in RT-PCR negative COVID-19 infections.

BACKGROUND: Molecular detection of SARS-CoV-2 in respiratory samples is the gold standard for COVID-19 diagnosis but it has a long turnaround time and struggles to detect low viral loads. Serology could help to diagnose suspected cases which lack molecular confirmation. Two case reports are presented as illustration. OBJECTIVES: The aim of this study was to evaluate the performance of several commercial assays for COVID-19 serology. We illustrated the added value of COVID-19 serology testing in suspect COVID-19 cases with negative molecular test. STUDY DESIGN: Twenty-three sera from 7 patients with a confirmed molecular diagnosis of SARS-CoV-2 were tested using 14 commercial assays. Additionally, 10 pre-pandemic sera and 9 potentially cross-reactive sera were selected. We calculated sensitivity and specificity. Furthermore, we discuss the diagnostic relevance of COVID-19 serology in a retrospective cohort of 145 COVID-19 cases in which repetitive molecular and serological SARS-CoV-2 tests were applied. RESULTS: The interpretation of the pooled sensitivity of IgM/A and IgG resulted in the highest values (range 14-71% on day 2-7; 88-94% on day 8-18). Overall, the specificity of the assays was high (range 79-100%). Among 145 retrospective cases, 3 cases (2%) remained negative after sequential molecular testing but positive on final SARS-CoV-2 serology. CONCLUSION: Sensitivity of COVID-19 serological diagnosis was variable but consistently increased at >7 days after symptom onset. Specificity was high. Our data suggest that serology can complement molecular testing for diagnosis of COVID-19, especially for patients presenting the 2nd week after symptom onset or later.

Methicillin-susceptible Staphylococcus aureus ST398-t571 harbouring the macrolide-lincosamide-streptogramin B resistance gene erm(T) in Belgian hospitals.

OBJECTIVES: Methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible S. aureus (MSSA), collected from 109 Belgian acute-care hospitals during a national survey in 2008, were investigated for macrolide-lincosamide (ML) resistance with particular emphasis on the analysis of erm(T)-carrying isolates. METHODS: In total, 314 MRSA and 212 MSSA were collected and characterized by spa typing. The SCCmec type of MRSA was determined. Resistance to ML antibiotics was detected by agar dilution and resistant strains were screened by PCR for erm(A), erm(C) and msr(A). Five ML-resistant MSSA isolates, negative by PCR for the aforementioned genes, were further characterized. RESULTS: Half of all MRSA isolates (n = 157; 50.0%) were resistant to erythromycin and harboured the gene erm(A) (n = 112), erm(C) (n = 41), erm(A) + erm(C) (n = 3) or msr(A) (n = 1). The erm(A) gene was mainly present in MRSA spa-CC002-ST5-SCCmec II and spa-CC008-ST8-SCCmec IV (where CC stands for clonal complex and ST stands for sequence type); the distribution of erm(C) was more diverse. Thirty-five of the 40 erythromycin-resistant MSSA (18.9%) carried the gene erm(A) (n = 17), erm(C) (n = 9) or msr(A) (n = 9). The remaining five MSSA were ST398-t571 isolates, which exhibited closely related ApaI PFGE patterns, harboured the gene erm(T) in the chromosomal DNA and did not exhibit additional resistances. These isolates were from severe infections in patients, of whom four had no contact and one had only indirect contact with livestock via a family member working in animal husbandry. CONCLUSIONS: The ML-streptogramin B ('MLS(B)') resistance genes erm(A) or erm(C) were detected in the majority of ML-resistant MRSA and MSSA isolates. The erm(T) gene was identified in MSSA ST398 isolates from five independent patients who lacked direct contact with livestock.

Longitudinal follow-up of IgG anti-nucleocapsid antibodies in SARS-CoV-2 infected patients up to eight months after infection.

BACKGROUND: Most SARS-CoV-2 infected patients develop IgG antibodies within 2-3 weeks after symptom onset. Antibody levels have been shown to gradually decrease in the first months after infection, but few data are available at six months or later. METHODS: A retrospective multi-center study was performed using 652 samples of 236 PCR-confirmed SARS-CoV-2 infected patients from 2 Belgian University hospitals. Patients were included if at least two samples were available (range 2-7 samples); including at least one sample collected 30 days or later after first positive PCR (range 0-240 days). Of those 236 patients, 19.1 % were classified as mild/asymptomatic (mild) and 80.9 % as moderate to critical (severe). IgG anti-nucleocapsid antibodies (anti-N) were measured using the Abbott Architect immunoassay. RESULTS: 22.2 % of mild and 2.6 % of severe COVID-19 cases never seroconverted (p < 0.001). Of the mild patients who seroconverted 0-59 days after PCR; 18.8 %, 40.0 % and 61.1 % were seronegative in the windows 60-119 days, 120-179 days and 180-240 days after PCR, respectively. In severe patients, these numbers were 1.9 %, 10.8 % and 29.4 % respectively (p < 0.05 each). Antibody levels were significantly higher in severe patients compared to mild patients in each 60 day window (p < 0.001 each). CONCLUSIONS: SARS-CoV-2 anti-N IgG antibody levels steadily decreased after 2 months up to 8 months post PCR. Of severe COVID-19 patients, 70.6 % remained positive up to eight months after infection. Antibody levels were significantly lower in mild SARS-CoV-2 infected patients and 61.1 % became seronegative within 6 months after the first positive PCR.