Emergence of novel ST1299 vanA lineages as possible cause for the striking rise of vancomycin resistance among invasive strains of Enterococcus faecium at a German university hospital.

IMPORTANCE: The proportion of VREfm among all Enterococcus faecium isolated from blood cultures in German hospitals has increased in the period 2015-2020 from 11.9% to 22.3% with a country-wide spread of the clonal lineage ST117/CT71 vanB. In this study, we provided useful information about the genetic diversity of invasive strains of E. faecium. Moreover, our findings confirm the nosocomial spread of novel ST1299 vanA lineages, which recently had a rapid expansion in Austria and the south-eastern part of Germany.

Reactogenicity Correlates Only Weakly with Humoral Immunogenicity after COVID-19 Vaccination with BNT162b2 mRNA (Comirnaty®).

mRNA vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), such as BNT162b2 (Comirnaty®), have proven to be highly immunogenic and efficient but also show marked reactogenicity, leading to adverse effects (AEs). Here, we analyzed whether the severity of AEs predicts the antibody response against the SARS-CoV-2 spike protein. Healthcare workers without prior SARS-CoV-2 infection, who received a prime-boost vaccination with BNT162b2, completed a standardized electronic questionnaire on the duration and severity of AEs. Serum specimens were collected two to four weeks after the boost vaccination and tested with the COVID-19 ELISA IgG (Vircell-IgG), the LIAISON® SARS-CoV-2 S1/S2 IgG CLIA (DiaSorin-IgG) and the iFlash-2019-nCoV NAb surrogate neutralization assay (Yhlo-NAb). A penalized linear regression model fitted by machine learning was used to correlate AEs with antibody levels. Eighty subjects were enrolled in the study. Systemic, but not local, AEs occurred more frequently after the boost vaccination. Elevated SARS-CoV-2 IgG antibody levels were measured in 92.5% of subjects with Vircell-IgG and in all subjects with DiaSorin-IgG and Yhlo-NAb. Gender, age and BMI showed no association with the antibody levels or with the AEs. The linear regression model identified headache, malaise and nausea as AEs with the greatest variable importance for higher antibody levels (Vircell-IgG and DiaSorin-IgG). However, the model performance for predicting antibody levels from AEs was very low for Vircell-IgG (squared correlation coefficient r2 = 0.04) and DiaSorin-IgG (r2 = 0.06). AEs did not predict the surrogate neutralization (Yhlo-NAb) results. In conclusion, AEs correlate only weakly with the SARS-CoV-2 spike protein antibody levels after COVID-19 vaccination with BNT162b2 mRNA.

Induction of proliferation and pro-inflammatory cytokine production in rheumatoid arthritis peripheral blood mononuclear cells by a 65 KDa chondrocyte membrane-specific, constitutive target autoantigen (CH65).

OBJECTIVE: To analyze proliferation and pro-inflammatory cytokine production of peripheral blood mononuclear cells (PBMC) from rheumatoid arthritis (RA) patients following stimulation with a purified chondrocyte membrane-associated autoantigen (CH65). METHODS: CH65 was highly purified from bovine chondrocyte membranes by solubilization and ion exchange chromatography. PBMC of RA patients (n = 37; 28 seropositive, nine seronegative) and non-arthritic donors (n = 20) were isolated by ficoll centrifugation and used in cell proliferation assays. The levels of interleukin (IL)-1, tumo necrosis factor (TNF) and IL-6 produced after stimulation with CH65 were determined by enzyme-linked immunosorbent assay (ELISA). Statistical analysis was performed using Mann-Whitney U-test and Spearman rank test and the software SPSS 13.0TM (SPSS Inc.; Chicago, IL, USA). RESULTS: Peripheral blood mononuclear cells exhibited a strong proliferative response to purified CH65 in approximately 50% of the RA patients (seropositive > seronegative), with a maximum reactivity at 0.15 or 0.30 µg/mL culture medium. In contrast, PBMC from normal donors did not show a proliferative response to CH65 at any dose. The proliferative response in RA patients peaked at days 7-9 and returned to control levels at day 13, indicating an antigen-driven process. CH65-stimulated RA PBMC produced moderate to high amounts of IL-1, TNF and IL-6. This was comparable to the response after exposure to isolated whole chondrocyte membranes or purified collagen type II. CONCLUSION: These results demonstrate a significant cellular immune response to CH65 protein in RA patients. Given the high similarity between bovine and human CH65, the results suggest a pathogenetic involvement of this molecule as a cartilage-specific potential target autoantigen in RA.

Pro-inflammatory IL-1beta and/or TNF-alpha up-regulate matrix metalloproteases-1 and -3 mRNA in chondrocyte subpopulations potentially pathogenic in osteoarthritis: in situ hybridization studies on a single cell level.

AIM: In osteoarthritis chondrocytes, matrix metalloproteases (MMPs) and their inhibitors are induced by interleukin (IL)-1beta or tumor necrosis factor (TNF)-alpha and balanced by inhibitors, but their messenger RNA (mRNA) expression has not been studied in individual cells. METHODS: Normal articular chondrocytes (10 donors; age 50 ± 6 years, mean ± SEM) were stimulated in a monolayer for 24 h with IL-1beta, TNF-alpha, or transforming growth factor (TGF)-beta1 (10 ng/mL each), alone or in combination. mRNA expression for MMP-1, MMP-3 and tissue inhibitor of metalloproteinase (TIMP)-1 was studied by in situ hybridization ((35) S-cRNA) and quantitative reverse transcription polymerase chain reaction (RT-PCR) (n ≥ 3 each). RESULTS: Whereas < 5% chondrocytes constitutively expressed MMP-1, a higher percentage expressed MMP-3 and TIMP-1 (31.1 ± 1.8%; 36.7 ± 2.8%, respectively). Upon stimulation with IL-1beta, TNF-alpha or IL-1beta/TNF-alpha, the percentage of cells positive for MMP-1, MMP-3 and TIMP-1 rose significantly (IL-1beta: 31.5%, 54.5% and 60.2%, respectively; TNF-alpha: 35.4%, 56.6%, 50.9%; IL-1beta/TNF-alpha: 38.8%, 45.2%, 52.1%). In bulk population (RT-PCR), mRNA for MMP-1 and MMP-3 was also induced by IL-1beta (11.9-fold, 1.2-fold, respectively), TNF-alpha (4.8-fold, 1.0-fold) or IL-1beta/TNF-alpha (14.7-fold, 1.4-fold), an effect attenuated by TGF-beta1. TIMP-1 mRNA, in contrast, was down-regulated by IL-1beta, TNF-alpha or IL-1beta/TNF-alpha, an effect again partially reverted by TGF-beta1. Finally, collagen type II mRNA was down-regulated by IL-1beta, TNF-alpha or IL-1beta/TNF-alpha (by 90%, 50% and 98%, respectively) and that of collagen type I was up-regulated (5.7-fold, 3.0-fold, 3.7-fold). CONCLUSIONS: Up-regulation of MMP-1/MMP-3 by IL-1beta and/or TNF-alpha in a fraction of chondrocytes in vitro suggests that a subpopulation of catabolic cells may also exist in osteoarthritis. These cells may undergo considerable dedifferentiation, as indicated by a decreased collagen-II/collagen-I ratio.