Ectopic expression of C-type lectin Mincle renders mice susceptible to staphylococcal pneumonia.

Staphylococcus aureus is a prevalent pathogen in pneumonia and harbors glycolipids which may serve as molecular patterns in Mincle (Macrophage inducible C-type lectin) dependent pathogen recognition. We examined the role of Mincle in lung defense against S. aureus in WT, Mincle KO and Mincle transgenic (tg) mice. Two glycolipids, glucosyl-diacylglycerol (Glc-DAG) and diglucosyl-diacylglycerol (Glc2-DAG) were purified, of which only Glc-DAG triggered Mincle reporter cell activation and professional phagocyte responses. Proteomic profiling revealed that Glc2-DAG blocked Glc-DAG-induced cytokine responses, thereby acting as inhibitor of Glc-DAG/Mincle-signaling. WT mice responded to S. aureus with a similar lung pathology as Mincle KO mice, most likely due to Glc2-DAG-dependent inhibition of Glc-DAG/Mincle-signaling. In contrast, ectopic Mincle expression caused severe lung pathology in S. aureus-infected mice characterized by bacterial outgrowth and fatal pneumonia. Collectively, Glc2-DAG inhibits Glc-DAG/Mincle-dependent responses in WT mice, whereas sustained Mincle expression overrides Glc2-DAG-mediated inhibitory effects, conferring increased host susceptibility to S. aureus.

Frequency of positive anti-PF4/polyanion antibody tests after COVID-19 vaccination with ChAdOx1 nCoV-19 and BNT162b2.

Vaccination using the adenoviral vector COVID-19 vaccine ChAdOx1 nCoV-19 (AstraZeneca) has been associated with rare vaccine-induced immune thrombotic thrombocytopenia (VITT). Affected patients test strongly positive in platelet factor 4 (PF4)/polyanion enzyme immunoassays (EIAs), and serum-induced platelet activation is maximal in the presence of PF4. We determined the frequency of anti-PF4/polyanion antibodies in healthy vaccinees and assessed whether PF4/polyanion EIA+ sera exhibit platelet-activating properties after vaccination with ChAdOx1 nCoV-19 (n = 138) or BNT162b2 (BioNTech/Pfizer; n = 143). In total, 19 of 281 participants tested positive for anti-PF4/polyanion antibodies postvaccination (All: 6.8% [95% confidence interval (CI), 4.4-10.3]; BNT162b2: 5.6% [95% CI, 2.9-10.7]; ChAdOx1 nCoV-19: 8.0% [95% CI, 4.5% to 13.7%]). Optical densities were mostly low (between 0.5 and 1.0 units; reference range, <0.50), and none of the PF4/polyanion EIA+ samples induced platelet activation in the presence of PF4. We conclude that positive PF4/polyanion EIAs can occur after severe acute respiratory syndrome coronavirus 2 vaccination with both messenger RNA- and adenoviral vector-based vaccines, but many of these antibodies likely have minor (if any) clinical relevance. Accordingly, low-titer positive PF4/polyanion EIA results should be interpreted with caution when screening asymptomatic individuals after vaccination against COVID-19. Pathogenic platelet-activating antibodies that cause VITT do not occur commonly following vaccination.

Staphylococcus aureus nasal colonization among dental health care workers in Northern Germany (StaphDent study).

Methicillin-resistant Staphylococcus aureus (MRSA) can colonize dental patients and students, however, studies on the prevalence of MRSA and methicillin-susceptible S. aureus (MSSA) among dental health care workers (DHCW) including use of personal protective equipment (PPE) are scarce. We conducted an observational study (StaphDent study) to (I) determine the prevalence of MRSA and MSSA colonization in DHCW in the region of Mecklenburg Western-Pomerania, Germany, (II) resolve the S. aureus population structure to gain hints on possible transmission events between co-workers, and (III) clarify use of PPE. Nasal swabs were obtained from dentists (n = 149), dental assistants (n = 297) and other dental practice staff (n = 38). Clonal relatedness of MSSA isolates was investigated using spa typing and, in some cases, whole genome sequencing (WGS). PPE use was assessed by questionnaire. While 22.3% (108/485) of the participants were colonized with MSSA, MRSA was not detected. MSSA prevalence was not associated with size of dental practices, gender, age, or duration of employment. The identified 61 spa types grouped into 17 clonal complexes and four sequence types. Most spa types (n = 47) were identified only once. In ten dental practices one spa type occurred twice. WGS data analysis confirmed a close clonal relationship for 4/10 isolate pairs. PPE was regularly used by most dentists and assistants. To conclude, the failure to recover MRSA from DHCW reflects the low MRSA prevalence in this region. Widespread PPE use suggests adherence to routine hygiene protocols. Compared to other regional HCW MRSA rates the consequent usage of PPE seems to be protective.

Staphylococcus aureus Host Tropism and Its Implications for Murine Infection Models.

Staphylococcus aureus (S. aureus) is a pathobiont of humans as well as a multitude of animal species. The high prevalence of multi-resistant and more virulent strains of S. aureus necessitates the development of new prevention and treatment strategies for S. aureus infection. Major advances towards understanding the pathogenesis of S. aureus diseases have been made using conventional mouse models, i.e., by infecting naïve laboratory mice with human-adapted S.aureus strains. However, the failure to transfer certain results obtained in these murine systems to humans highlights the limitations of such models. Indeed, numerous S. aureus vaccine candidates showed promising results in conventional mouse models but failed to offer protection in human clinical trials. These limitations arise not only from the widely discussed physiological differences between mice and humans, but also from the lack of attention that is paid to the specific interactions of S. aureus with its respective host. For instance, animal-derived S. aureus lineages show a high degree of host tropism and carry a repertoire of host-specific virulence and immune evasion factors. Mouse-adapted S.aureus strains, humanized mice, and microbiome-optimized mice are promising approaches to overcome these limitations and could improve transferability of animal experiments to human trials in the future.

Staphylococcal enterotoxin-like X (SElX) is a unique superantigen with functional features of two major families of staphylococcal virulence factors.

Staphylococcus aureus is an opportunistic pathogen that produces many virulence factors. Two major families of which are the staphylococcal superantigens (SAgs) and the Staphylococcal Superantigen-Like (SSL) exoproteins. The former are immunomodulatory toxins that induce a Vß-specific activation of T cells, while the latter are immune evasion molecules that interfere with a wide range of innate immune defences. The superantigenic properties of Staphylococcal enterotoxin-like X (SElX) have recently been established. We now reveal that SElX also possesses functional characteristics of the SSLs. A region of SElX displays high homology to the sialyl-lactosamine (sLacNac)-specific binding site present in a sub-family of SSLs. By analysing the interaction of SElX with sLacNac-containing glycans we show that SElX has an equivalent specificity and host cell binding range to the SSLs. Mutation of key amino acids in this conserved region affects the ability of SElX to bind to cells of myeloid origin and significantly reduces its ability to protect S. aureus from destruction in a whole blood killing (WBK) assay. Like the SSLs, SElX is up-regulated early during infection and is under the control of the S. aureus exotoxin expression (Sae) two component gene regulatory system. Additionally, the structure of SElX in complex with the sLacNac-containing tetrasaccharide sialyl Lewis X (sLeX) reveals that SElX is a unique single-domain SAg. In summary, SElX is an 'SSL-like' SAg.

Bringing together what belongs together: Optimizing murine infection models by using mouse-adapted Staphylococcus aureus strains.

Staphylococcus (S.) aureus is a leading cause of bacterial infection world-wide, and currently no vaccine is available for humans. Vaccine development relies heavily on clinically relevant infection models. However, the suitability of mice for S. aureus infection models has often been questioned, because experimental infection of mice with human-adapted S. aureus requires very high infection doses. Moreover, mice were not considered to be natural hosts of S. aureus. The latter has been disproven by our recent findings, showing that both laboratory mice, as well as wild small mammals including mice, voles, and shrews, are naturally colonized with S. aureus. Here, we investigated whether mouse-and vole-derived S. aureus strains show an enhanced virulence in mice as compared to the human-adapted strain Newman. Using a step-wise approach based on the bacterial genotype and in vitro assays for host adaptation, we selected the most promising candidates for murine infection models out of a total of 254 S. aureus isolates from laboratory mice as well as wild rodents and shrews. Four strains representing the clonal complexes (CC) 8, 49, and 88 (n = 2) were selected and compared to the human-adapted S. aureus strain Newman (CC8) in murine pneumonia and bacteremia models. Notably, a bank vole-derived CC49 strain, named DIP, was highly virulent in BALB/c mice in pneumonia and bacteremia models, whereas the other murine and vole strains showed virulence similar to or lower than that of Newman. At one tenth of the standard infection dose DIP induced disease severity, bacterial load and host cytokine and chemokine responses in the murine bacteremia model similar to that of Newman. In the pneumonia model, DIP was also more virulent than Newman but the effect was less pronounced. Whole genome sequencing data analysis identified a pore-forming toxin gene, lukF-PV(P83)/lukM, in DIP but not in the other tested S. aureus isolates. To conclude, the mouse-adapted S. aureus strain DIP allows a significant reduction of the inoculation dose in mice and is hence a promising tool to develop clinically more relevant infection models.

Staphylococcus aureus colonization in hemodialysis patients: a prospective 25 months observational study.

BACKGROUND: Dialysis patients are frequently exposed to Staphylococcus aureus due to stays in dialysis centers, hospitals or rest homes. The hemodialysis vascular access is a potential entry site for S. aureus, in particular when using a central venous catheter (CVC) which increases the risk of sepsis compared to arteriovenous (AV) fistula. We prospectively followed a cohort of 86 hemodialysis patients from an outpatient dialysis center over 25 months analyzing S. aureus carrier status, S. aureus infection rates and mortality. METHODS: Demographic data and patients´ medical histories were collected and followed from all hemodialysis patients. Blood samples, nasal swabs and swabs from the hemodialysis vascular access site were taken every six months for a period of 25 months and tested for S. aureus. Strains were cultured and further characterized by spa PCR and microarray-based genotyping. Resulting data were compared with those from the general population. RESULTS: In cross-sectional analyses, an average of 40% of hemodialysis patients were S. aureus carriers compared to 27% in the general population. Longitudinally, a total of 65% were S. aureus carriers: 16% were persistent carriers, 43% were intermittently colonized. The most common S. aureus lineage in the dialysis patient cohort was the clonal complex (CC) 8 and the spa type t008, while in the general population, the clonal complex CC30 dominates. During the study period, we observed six S. aureus-associated blood stream infections with one S. aureus attributable death. S. aureus carriers with an AV fistula were more densely colonized in the nasal mucosa compared to patients with a CVC. Overall mortality was lower for hemodialysis patients with a positive S. aureus carrier status compared to non-carriers (hazard ratio of 0.19). CONCLUSIONS: Compared to the general population, hemodialysis patients were more frequently colonized with S. aureus and displayed both different S. aureus colonization densities as well as lineages, possibly explained by more frequent exposure to health care environments. The lower overall mortality in carriers compared to non-carriers is intriguing and will be investigated in detail in the future. TRIAL REGISTRATION: ISRCTN 14385893 , 2. October 2018, retrospectively registered.

Staphylococcal Protein A Contributes to Persistent Colonization of Mice with Staphylococcus aureus.

Staphylococcus aureus persistently colonizes the nasopharynx in humans, which increases the risk for invasive diseases, such as skin infection and bacteremia. Nasal colonization triggers IgG responses against staphylococcal surface antigens; however, these antibodies cannot prevent subsequent colonization or disease. Here, we describe S. aureus WU1, a multilocus sequence type 88 (ST88) isolate that persistently colonizes the nasopharynx in mice. We report that staphylococcal protein A (SpA) is required for persistence of S. aureus WU1 in the nasopharynx. Compared to animals colonized by wild-type S. aureus, mice colonized with the Δspa variant mount increased IgG responses against staphylococcal colonization determinants. Immunization of mice with a nontoxigenic SpA variant, which cannot cross-link B cell receptors and divert antibody responses, elicits protein A-neutralizing antibodies that promote IgG responses against colonizing S. aureus and diminish pathogen persistence.IMPORTANCEStaphylococcus aureus persistently colonizes the nasopharynx in about one-third of the human population, thereby promoting community- and hospital-acquired infections. Antibiotics are currently used for decolonization of individuals at increased risk of infection. However, the efficacy of antibiotics is limited by recolonization and selection for drug-resistant strains. Here, we propose a model of how staphylococcal protein A (SpA), a B cell superantigen, modifies host immune responses during colonization to support continued persistence of S. aureus in the nasopharynx. We show that this mechanism can be thwarted by vaccine-induced anti-SpA antibodies that promote IgG responses against staphylococcal antigens and diminish colonization.

Wild rodents and shrews are natural hosts of Staphylococcus aureus.

Laboratory mice are the most commonly used animal model for Staphylococcus aureus infection studies. We have previously shown that laboratory mice from global vendors are frequently colonized with S. aureus. Laboratory mice originate from wild house mice. Hence, we investigated whether wild rodents, including house mice, as well as shrews are naturally colonized with S. aureus and whether S. aureus adapts to the wild animal host. 295 animals of ten different species were caught in different locations over four years (2012-2015) in Germany, France and the Czech Republic. 45 animals were positive for S. aureus (15.3%). Three animals were co-colonized with two different isolates, resulting in 48 S. aureus isolates in total. Positive animals were found in Germany and the Czech Republic in each studied year. The S. aureus isolates belonged to ten different spa types, which grouped into six lineages (clonal complex (CC) 49, CC88, CC130, CC1956, sequence type (ST) 890, ST3033). CC49 isolates were most abundant (17/48, 35.4%), followed by CC1956 (14/48, 29.2%) and ST890 (9/48, 18.8%). The wild animal isolates lacked certain properties that are common among human isolates, e.g., a phage-encoded immune evasion cluster, superantigen genes on mobile genetic elements and antibiotic resistance genes, which suggests long-term adaptation to the wild animal host. One CC130 isolate contained the mecC gene, implying wild rodents might be both reservoir and vector for methicillin-resistant . In conclusion, we demonstrated that wild rodents and shrews are naturally colonized with S. aureus, and that those S. aureus isolates show signs of host adaptation.

Global spread of mouse-adapted Staphylococcus aureus lineages CC1, CC15, and CC88 among mouse breeding facilities.

We previously reported that laboratory mice from all global vendors are frequently colonized with Staphylococcus aureus (S. aureus). Genotyping of a snap sample of murine S. aureus isolates from Charles River, US, showed that mice were predominantly colonized with methicillin-sensitive CC88 strains. Here, we expanded our view and investigated whether laboratory mice from other global animal facilities are colonized with similar strains or novel S. aureus lineages, and whether the murine S. aureus isolates show features of host adaptation. In total, we genotyped 230 S. aureus isolates from various vendor facilities of laboratory mice around the globe (Charles River facilities in the USA, Canada, France, and Germany; another US facility) and university- or company-associated breeding facilities in Germany, China and New Zealand. Spa typing was performed to analyse the clonal relationship of the isolates. Moreover, multiplex PCRs were performed for human-specific virulence factors, the immune-evasion cluster (IEC) and superantigen genes (SAg). We found a total of 58 different spa types that clustered into 15 clonal complexes (CCs). Three of these S. aureus lineages had spread globally among laboratory mice and accounted for three quarters of the isolates: CC1 (13.5%), CC15 (14.3%), and CC88 (47.0%). Compared to human colonizing isolates of the same lineages, the murine isolates frequently lacked IEC genes and SAg genes on mobile genetic elements, implying long-term adaptation to the murine host. In conclusion, laboratory mice from various vendors are colonized with host-adapted S. aureus-strains of a few lineages, predominantly the CC88 lineage. S. aureus researchers must be cautioned that S. aureus colonization might be a relevant confounder in infection and vaccination studies and are therefore advised to screen their mice before experimentation.

Staphylococcal enterotoxin A-activated regulatory T cells promote allergen-specific TH2 response to intratracheal allergen inoculation.

BACKGROUND: TH2 responses are implicated in asthma pathobiology. Epidemiologic studies have found a positive association between asthma and exposure to staphylococcal enterotoxins. OBJECTIVE: We used a mouse model of asthma to determine whether staphylococcal enterotoxins promote TH2 differentiation of allergen-specific CD4 conventional T (Tcon) cells and asthma by activating allergen-nonspecific regulatory T (Treg) cells to create a TH2-polarizing cytokine milieu. METHODS: Ovalbumin (OVA)-specific, staphylococcal enterotoxin A (SEA)-nonreactive naive CD4 Tcon cells were cocultured with SEA-reactive allergen-nonspecific Treg or CD4 Tcon cells in the presence of OVA and SEA. The OVA-specific CD4 T cells were then analyzed for IL-13 and IFN-γ expression. SEA-activated Treg cells were analyzed for the expression of the TH2-polarizing cytokine IL-4 and the T-cell activation markers CD69 and CD62L. For asthma induction, mice were intratracheally sensitized with OVA or cat dander extract (CDE) alone or together with SEA and then challenged with OVA or CDE. Mice were also subject to transient Treg cell depletion before sensitization with OVA plus SEA. Asthma features and TH2 differentiation in these mice were analyzed. RESULTS: SEA-activated Treg cells induced IL-13 but suppressed IFN-γ expression in OVA-specific CD4 Tcon cells. SEA-activated Treg cells expressed IL-4, upregulated CD69, and downregulated CD62L. Sensitization with OVA plus SEA but not OVA alone induced asthma, and SEA exacerbated asthma induced by CDE. Depletion of Treg cells abolished these effects of SEA and IL-13 expression in OVA-specific T cells. CONCLUSION: SEA promoted TH2 responses of allergen-specific T cells and asthma pathogenesis by activating Treg cells.

Molecular Epidemiology of Staphylococcus aureus in the General Population in Northeast Germany: Results of the Study of Health in Pomerania (SHIP-TREND-0).

Population-based studies on Staphylococcus aureus nasal colonization are scarce. We examined the prevalence, resistance, and molecular diversity of S. aureus in the general population in Northeast Germany. Nasal swabs were obtained from 3,891 adults in the large-scale population-based Study of Health in Pomerania (SHIP-TREND). Isolates were characterized using spa genotyping, as well as antibiotic resistance and virulence gene profiling. We observed an S. aureus prevalence of 27.2%. Nasal S. aureus carriage was associated with male sex and inversely correlated with age. Methicillin-resistant S. aureus (MRSA) accounted for 0.95% of the colonizing S. aureus strains. MRSA carriage was associated with frequent visits to hospitals, nursing homes, or retirement homes within the previous 24 months. All MRSA strains were resistant to multiple antibiotics. Most MRSA isolates belonged to the pandemic European hospital-acquired MRSA sequence type 22 (HA-MRSA-ST22) lineage. We also detected one livestock-associated MRSA ST398 (LA-MRSA-ST398) isolate, as well as six livestock-associated methicillin-susceptible S. aureus (LA-MSSA) isolates (clonal complex 1 [CC1], CC97, and CC398). spa typing revealed a diverse but also highly clonal S. aureus population structure. We identified a total of 357 spa types, which were grouped into 30 CCs or sequence types. The major seven CCs (CC30, CC45, CC15, CC8, CC7, CC22, and CC25) included 75% of all isolates. Virulence gene patterns were strongly linked to the clonal background. In conclusion, MSSA and MRSA prevalences and the molecular diversity of S. aureus in Northeast Germany are consistent with those of other European countries. The detection of HA-MRSA and LA-MRSA within the general population indicates possible transmission from hospitals and livestock, respectively, and should be closely monitored.

Antibacterial Activity of Cold Atmospheric Pressure Argon Plasma against 78 Genetically Different (mecA, luk-P, agr or Capsular Polysaccharide Type) Staphylococcus aureus Strains.

Previous studies on the antimicrobial activity of cold atmospheric pressure argon plasma showed varying effects against mecA+ or mecA-Staphylococcus aureus strains. This observation may have important clinical and epidemiological implications. Here, the antibacterial activity of argon plasma was investigated against 78 genetically different S. aureus strains, stratified by mecA, luk-P, agr1-4, or the cell wall capsule polysaccharide types 5 and 8. kINPen09® served as the plasma source for all experiments. On agar plates, mecA+luk-P-S. aureus strains showed a decreased susceptibility against plasma compared to other S. aureus strains. This study underlines the high complexity of microbial defence against antimicrobial treatment and confirms a previously reported strain-dependent susceptibility of S. aureus to plasma treatment.

Immune control of Staphylococcus aureus - regulation and counter-regulation of the adaptive immune response.

Successful vaccination relies on immune memory, a core competence of the adaptive immune system. This review summarizes the current knowledge about the adaptive immune response to Staphylococcus aureus as well as the bacterial escape mechanisms. The Janus-faced bacteria, both life-threatening pathogens and peaceful cohabitants of their human host, have so far frustrated all attempts at vaccine development. This begs the question of whether the adaptive immune system is at all able to protect against S. aureus infection. In search of an answer the main functions of the adaptive immune system are probed for potential mechanisms of protection against S. aureus, which may be put to the test in future research.

Non-cross-reactive epitopes dominate the humoral immune response to COVID-19 vaccination - kinetics of plasma antibodies, plasmablasts and memory B cells.

Introduction: COVID-19 vaccines are highly effective in inducing protective immunity. While the serum antibody response to COVID-19 vaccination has been studied in depth, our knowledge of the underlying plasmablast and memory B cell (Bmem) responses is still incomplete. Here, we determined the antibody and B cell response to COVID-19 vaccination in a naïve population and contrasted it with the response to a single influenza vaccination in a primed cohort. In addition, we analyzed the antibody and B cell responses against the four endemic human coronaviruses (HCoVs). Methods: Measurement of specific plasma IgG antibodies was combined with functional analyses of antibody-secreting plasmablasts and Bmems. SARS-CoV-2- and HCoV-specific IgG antibodies were quantified with an in-house bead-based multiplexed immunoassay. Results: The antibody and B cell responses to COVID-19 vaccination reflected the kinetics of a prime-boost immunization, characterized by a slow and moderate primary response and a faster and stronger secondary response. In contrast, the influenza vaccinees possessed robust immune memory for the vaccine antigens prior to vaccination, and the recall vaccination moderately boosted antibody production and Bmem responses. Antibody levels and Bmem responses waned several months after the 2nd COVID-19 vaccination, but were restored upon the 3rd vaccination. The COVID-19 vaccine-induced antibodies mainly targeted novel, non-cross-reactive S1 epitopes of the viral spike protein, while cross-reactive S2 epitopes were less immunogenic. Booster vaccination not only strongly enhanced neutralizing antibodies against an original SARS-CoV-2 strain, but also induced neutralizing antibodies against the Omicron BA.2 variant. We observed a 100% plasma antibody prevalence against the S1 subunits of HCoVs, which was not affected by vaccination. Discussion: Overall, by complementing classical serology with a functional evaluation of plasmablasts and memory B cells we provide new insights into the specificity of COVID-19 vaccine-induced antibody and B cell responses.

TLR4 sensing of IsdB of Staphylococcus aureus induces a proinflammatory cytokine response via the NLRP3-caspase-1 inflammasome cascade.

IMPORTANCE: The prevalence of multidrug-resistant Staphylococcus aureus is of global concern, and vaccines are urgently needed. The iron-regulated surface determinant protein B (IsdB) of S. aureus was investigated as a vaccine candidate because of its essential role in bacterial iron acquisition but failed in clinical trials despite strong immunogenicity. Here, we reveal an unexpected second function for IsdB in pathogen-host interaction: the bacterial fitness factor IsdB triggers a strong inflammatory response in innate immune cells via Toll-like receptor 4 and the inflammasome, thus acting as a novel pathogen-associated molecular pattern of S. aureus. Our discovery contributes to a better understanding of how S. aureus modulates the immune response, which is necessary for vaccine development against the sophisticated pathogen.

Toxin exposure and HLA alleles determine serum antibody binding to toxic shock syndrome toxin 1 (TSST-1) of Staphylococcus aureus.

Life-threatening toxic shock syndrome is often caused by the superantigen toxic shock syndrome toxin-1 (TSST-1) produced by Staphylococcus aureus. A well-known risk factor is the lack of neutralizing antibodies. To identify determinants of the anti-TSST-1 antibody response, we examined 976 participants of the German population-based epidemiological Study of Health in Pomerania (SHIP-TREND-0). We measured anti-TSST-1 antibody levels, analyzed the colonization with TSST-1-encoding S. aureus strains, and performed a genome-wide association analysis of genetic risk factors. TSST-1-specific serum IgG levels varied over a range of 4.2 logs and were elevated by a factor of 12.3 upon nasal colonization with TSST-1-encoding S. aureus. Moreover, the anti-TSST-1 antibody levels were strongly associated with HLA class II gene loci. HLA-DRB1*03:01 and HLA-DQB1*02:01 were positively, and HLA-DRB1*01:01 as well as HLA-DQB1*05:01 negatively associated with the anti-TSST-1 antibody levels. Thus, both toxin exposure and HLA alleles affect the human antibody response to TSST-1.

The Greifswald Post COVID Rehabilitation Study and Research (PoCoRe)-Study Design, Characteristics and Evaluation Tools.

(1) Background: COVID-19 is often associated with significant long-term symptoms and disability, i.e., the long/post-COVID syndrome (PCS). Even after presumably mild COVID-19 infections, an increasing number of patients seek medical help for these long-term sequelae, which can affect various organ systems. The pathogenesis of PCS is not yet understood. Therapy has so far been limited to symptomatic treatment. The Greifswald Post COVID Rehabilitation Study (PoCoRe) aims to follow and deeply phenotype outpatients with PCS in the long term, taking a holistic and comprehensive approach to the analysis of their symptoms, signs and biomarkers. (2) Methods: Post-COVID outpatients are screened for symptoms in different organ systems with a standardized medical history, clinical examination, various questionnaires as well as physical and cardiopulmonary function tests. In addition, biomaterials are collected for the analysis of immunomodulators, cytokines, chemokines, proteome patterns as well as specific (auto)antibodies. Patients are treated according to their individual needs, adhering to the current standard of care. PoCoRe's overall aim is to optimize diagnostics and therapy in PCS patients.

Anti-neuronal antibodies against brainstem antigens are associated with COVID-19.

BACKGROUND: Understanding how SARS-CoV-2 affects respiratory centres in the brainstem may help to preclude assisted ventilation for patients in intensive care setting. Viral invasion appears unlikely, although autoimmunity has been implicated, the responsible antigens remain unknown. We previously predicted the involvement of three epitopes within distinct brainstem proteins: disabled homolog 1 (DAB1), apoptosis-inducing-factor-1 (AIFM1), and surfeit-locus-protein-1 (SURF1). METHODS: Here, we used microarrays to screen serum from COVID-19 patients admitted to intensive care and compared those with controls who experienced mild course of the disease. FINDINGS: The results confirm the occurrence of IgG and IgM antibodies against the hypothesised epitopes in COVID-19 patients. Importantly, while IgM levels were similar in both groups, IgG levels were significantly elevated in severely ill patients compared to controls, suggesting a pathogenic role of IgG. INTERPRETATION: The newly discovered anti-neuronal antibodies might be promising markers of severe disease and the targeted peptide epitopes might be used for targeted immunomodulation. Further work is needed to determine whether these antibodies may play a role in long-COVID. FUNDING: AF, CF and PR received support from the German Research Foundation (grants FL 379/22-1, 327654276-SFB 1315, FR 4479/1-1, PR 1274/8-1). SH, DR, and DB received support from the Ministry of Economy, State of Mecklenburg Western Pomerania, Germany (grant COVIDPROTECT: "Optimisation of diagnostic and therapeutic pathways for COVID-19 patients in MV"). SH received support from the Research Group Molecular Medicine University of Greifswald (FVMM, seed funding FOVB-2021-01). AV received support from the Else Kröner Fresenius Foundation and the Alzheimer Research Initiative.

Genomic and proteomic characterization of Staphylococcus aureus mastitis isolates of bovine origin.

Staphylococcus aureus colonizes and infects humans as well as animals. In the present study, 17 S. aureus strains isolated from cows suffering from mastitis were characterized. The well-established multilocus sequence typing (MLST) technique and a diagnostic microarray covering 185 S. aureus virulence and resistance genes were used for genetic and epidemiological analyses. Virulence gene expression studies were performed by analyzing the extracellular protein pattern of each isolate on 2-D gels. By this way, a pronounced heterogeneity of the extracellular proteome between the bovine isolates has been observed which was attributed to genome plasticity and variation of gene expression. Merely 12 proteins were expressed in at least 80% of the isolates, i.e. Atl, Aur, GlpQ, Hla, LtaS, Nuc, PdhB, SAB0846, SAB2176, SAB0566, SspA, and SspB forming the core exoproteome. Fifteen extracellular proteins were highly variably expressed and only present in less than 20% of the isolates. This includes the serine proteases SplB, C, and F, and the superantigens SEC-bov, SEL and TSST-1. Compared to human isolates we identified at least six proteins with significantly different expression frequencies. While SAB0846 was expressed more frequently in bovine isolates, LytM, EbpS, Spa, Geh, and LukL1 were seen less frequently in these isolates.