SARS-CoV-2 infection predisposes patients to coinfection with Staphylococcus aureus.

Severe COVID-19 has been associated with coinfections with bacterial and fungal pathogens. Notably, patients with COVID-19 who develop Staphylococcus aureus bacteremia exhibit higher rates of mortality than those infected with either pathogen alone. To understand this clinical scenario, we collected and examined S. aureus blood and respiratory isolates from a hospital in New York City during the early phase of the pandemic from both SARS-CoV-2+ and SARS-CoV-2- patients. Whole genome sequencing of these S. aureus isolates revealed broad phylogenetic diversity in both patient groups, suggesting that SARS-CoV-2 coinfection was not associated with a particular S. aureus lineage. Phenotypic characterization of the contemporary collection of S. aureus isolates from SARS-CoV-2+ and SARS-CoV-2- patients revealed no notable differences in several virulence traits examined. However, we noted a trend toward overrepresentation of S. aureus bloodstream strains with low cytotoxicity in the SARS-CoV-2+ group. We observed that patients coinfected with SARS-CoV-2 and S. aureus were more likely to die during the acute phase of infection when the coinfecting S. aureus strain exhibited high or low cytotoxicity. To further investigate the relationship between SARS-CoV-2 and S. aureus infections, we developed a murine coinfection model. These studies revealed that infection with SARS-CoV-2 renders mice susceptible to subsequent superinfection with low cytotoxicity S. aureus. Thus, SARS-CoV-2 infection sensitizes the host to coinfections, including S. aureus isolates with low intrinsic virulence. IMPORTANCE: The COVID-19 pandemic has had an enormous impact on healthcare across the globe. Patients who were severely infected with SARS-CoV-2, the virus causing COVID-19, sometimes became infected with other pathogens, which is termed coinfection. If the coinfecting pathogen is the bacterium Staphylococcus aureus, there is an increased risk of patient death. We collected S. aureus strains that coinfected patients with SARS-CoV-2 to study the disease outcome caused by the interaction of these two important pathogens. We found that both in patients and in mice, coinfection with an S. aureus strain lacking toxicity resulted in more severe disease during the early phase of infection, compared with infection with either pathogen alone. Thus, SARS-CoV-2 infection can directly increase the severity of S. aureus infection.

Prophage-encoded methyltransferase drives adaptation of community-acquired methicillin-resistant Staphylococcus aureus.

We recently described the evolution of a community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) USA300 variant responsible for an outbreak of skin and soft tissue infections. Acquisition of a mosaic version of the Φ11 prophage (mΦ11) that increases skin abscess size was an early step in CA-MRSA adaptation that primed the successful spread of the clone. The present report shows how prophage mΦ11 exerts its effect on virulence for skin infection without encoding a known toxin or fitness genes. Abscess size and skin inflammation were associated with DNA methylase activity of an mΦ11-encoded adenine methyltransferase (designated pamA). pamA increased expression of fibronectin-binding protein A (fnbA; FnBPA), and inactivation of fnbA eliminated the effect of pamA on abscess virulence without affecting strains lacking pamA. Thus, fnbA is a pamA-specific virulence factor. Mechanistically, pamA was shown to promote biofilm formation in vivo in skin abscesses, a phenotype linked to FnBPA's role in biofilm formation. Collectively, these data reveal a novel mechanism-epigenetic regulation of staphylococcal gene expression-by which phage can regulate virulence to drive adaptive leaps by S. aureus.

Detection and Transmission of Proteus mirabilis in Immunodeficient Mice.

The exclusion of opportunistic pathogens is important for protecting animal health and ensuring desired research outcomes in highly immunodeficient mice. Proteus mirabilis has been associated with gastrointestinal tract lesions, septicemia, pyelonephritis, splenomegaly, and hepatitis and can influence select mouse models. To inform health-surveillance practices after we experienced difficulty in excluding P. mirabilis from our mouse colony, we aimed to determine the likelihood of detecting P. mirabilis-positive immunocompromised (SRG), immunovague (Fbn1+/-), and immunocompetent (CD1) colony mice through culture and PCR testing; to evaluate transmission via 2 sentinel-based approaches (direct contact and indirect dirty-bedding transfer); and to further characterize associated pathology. We hypothesized that immunocompromised mice would be better detectors and transmitters of P. mirabilis. Multiple logistic regression models were used for analysis and included PCR copy number, repeated testing, age, sex, and antibiotic-treated (trimethoprim-sulfamethoxazole) diet as covariates. Repeated testing over 10 wk showed that P. mirabilis -colonized immunocompromised colony mice were 95 times more likely than immunocompetent mice to test positive by culture and 30 times more likely by PCR assay. Sentinel mice were 15 times more likely to test positive by PCR assay for P. mirabilis when exposed by direct contact compared with dirty bedding and 18 times more likely to test positive when exposed to positive immunocompromised as compared with immunocompetent colony mice. After 10 wk of exposure, 3.8% of dirty-bedding sentinel PCR tests were positive, as compared with 30.7% of contact sentinels. Only immunocompromised mice on antibiotic diet (37.5%) developed lesions of the urogenital tract and abdominal cavity consistent with known pathology of P. mirabilis. Our findings suggest that PCR testing of dirty-bedding sentinels alone is not sufficient for the detection of P. mirabilis in mouse colonies. Direct-contact sentinels and testing of colony mice-especially if immunocompromised-with adjunct culture may facilitate successful bioexclusion.

Response of Canine Soft Tissue Sarcoma to Stereotactic Body Radiotherapy.

Canine soft tissue sarcoma (STS) has served as a preclinical model for radiation, hyperthermia, experimental therapeutics, and tumor microenvironmental research for decades. Stereotactic body radiotherapy (SBRT) demonstrates promising results for the control of various tumors in human and veterinary medicine; however, there is limited clinical data for the management of STS with SBRT. In this retrospective study, we aimed to define overall efficacy and toxicity of SBRT for the treatment of macroscopic canine STS to establish this preclinical model for comparative oncology research. Fifty-two canine patients met inclusion criteria. Total radiation dose prescribed ranged from 20-50 Gy delivered in 1-5 fractions. Median progression-free survival time (PFST) was 173 days and overall survival time (OST) 228 days. Best overall response was evaluable in 46 patients, with 30.4% responding to treatment (complete response n = 3; partial response n = 11). For responders, OST significantly increased to 475 days vs. 201 days (P = 0.009). Prognostic factors identified by multivariable Cox regressions included size of tumor and metastasis at presentation. Dogs were 3× more likely to progress (P = 0.009) or 3.5× more likely to experience death (P = 0.003) at all times of follow up if they presented with metastatic disease. Similarly, every 100-cc increase in tumor volume resulted in a 5% increase in the risk of progression (P = 0.002) and death (P = 0.001) at all times of follow up. Overall, 30.8% of patients developed acute toxicities, 7.7% grade 3; 28.8% of patients developed late toxicities, 11.5% grade 3. Increased dose administered to the skin significantly affected toxicity development. SBRT serves as a viable treatment option to provide local tumor control for canine macroscopic STS, particularly those with early-stage disease and smaller tumors. The results of this study will help to define patient inclusion criteria and to set dose limits for preclinical canine STS trials involving SBRT.