Shigellosis outbreak among persons experiencing homelessness-San Diego County, California, October-December 2021.

During October 2021, the County of San Diego Health and Human Services Agency identified five cases of shigellosis among persons experiencing homelessness (PEH). We conducted an outbreak investigation and developed interventions to respond to shigellosis outbreaks among PEH. Confirmed cases occurred among PEH with stool-cultured Shigella sonnei; probable cases were among PEH with Shigella-positive culture-independent diagnostic testing. Patients were interviewed to determine infectious sources and risk factors. Fifty-three patients were identified (47 confirmed, 6 probable); 34 (64%) were hospitalised. None died. No point source was identified. Patients reported inadequate access to clean water and sanitation facilities, including public restrooms closed because of the COVID-19 pandemic. After implementing interventions, including handwashing stations, more frequent public restroom cleaning, sanitation kit distribution, and isolation housing for ill persons, S. sonnei cases decreased to preoutbreak frequencies. Improving public sanitation access was associated with decreased cases and should be considered to prevent outbreaks among PEH.

The Many Hosts of Mycobacteria 9 (MHM9): A conference report.

The Many Hosts of Mycobacteria (MHM) meeting series brings together basic scientists, clinicians and veterinarians to promote robust discussion and dissemination of recent advances in our knowledge of numerous mycobacterial diseases, including human and bovine tuberculosis (TB), nontuberculous mycobacteria (NTM) infection, Hansen's disease (leprosy), Buruli ulcer and Johne's disease. The 9th MHM conference (MHM9) was held in July 2022 at The Ohio State University (OSU) and centered around the theme of "Confounders of Mycobacterial Disease." Confounders can and often do drive the transmission of mycobacterial diseases, as well as impact surveillance and treatment outcomes. Various confounders were presented and discussed at MHM9 including those that originate from the host (comorbidities and coinfections) as well as those arising from the environment (e.g., zoonotic exposures), economic inequality (e.g. healthcare disparities), stigma (a confounder of leprosy and TB for millennia), and historical neglect (a confounder in Native American Nations). This conference report summarizes select talks given at MHM9 highlighting recent research advances, as well as talks regarding the historic and ongoing impact of TB and other infectious diseases on Native American Nations, including those in Southwestern Alaska where the regional TB incidence rate is among the highest in the Western hemisphere.

Use of standard, contact, and droplet precautions with eye protection for the prevention of severe acute respiratory coronavirus virus 2 (SARS-CoV-2) transmission among home healthcare personnel in hospice and home healthcare settings-King and Snohomish counties, Washington, February-October 2020.

Among EvergreenHealth Home Care Service professionals, no coronavirus disease 2019 (COVID-19) cases were reported when they were instructed to use standard, contact, and droplet precautions with eye protection while providing home health care to patients diagnosed with laboratory-confirmed severe acute respiratory coronavirus virus 2 (SARS-CoV-2). These precautions might provide some level of protection against coronavirus disease 2019 (COVID-19) among home healthcare personnel.

Occupational exposures and mitigation strategies among homeless shelter workers at risk of COVID-19.

OBJECTIVE: To describe the work environment and COVID-19 mitigation measures for homeless shelter workers and assess occupational risk factors for COVID-19. METHODS: Between June 9-August 10, 2020, we conducted a self-administered survey among homeless shelter workers in Washington, Massachusetts, Utah, Maryland, and Georgia. We calculated frequencies for work environment, personal protective equipment use, and SARS-CoV-2 testing history. We used generalized linear models to produce unadjusted prevalence ratios (PR) to assess risk factors for SARS-CoV-2 infection. RESULTS: Of the 106 respondents, 43.4% reported frequent close contact with clients; 75% were worried about work-related SARS-CoV-2 infections; 15% reported testing positive. Close contact with clients was associated with testing positive for SARS-CoV-2 (PR 3.97, 95%CI 1.06, 14.93). CONCLUSIONS: Homeless shelter workers may be at risk of being exposed to individuals with COVID-19 during the course of their work. Frequent close contact with clients was associated with SARS-CoV-2 infection. Protecting these critical essential workers by implementing mitigation measures and prioritizing for COVID-19 vaccination is imperative during the pandemic.

Shelter Characteristics, Infection Prevention Practices, and Universal Testing for SARS-CoV-2 at Homeless Shelters in 7 US Urban Areas.

Objectives. To examine shelter characteristics and infection prevention practices in relation to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection point prevalence during universal testing at homeless shelters in the United States.Methods. SARS-CoV-2 testing was offered to clients and staff at homeless shelters, irrespective of symptoms. Site assessments were conducted from March 30 to June 1, 2020, to collect information on shelter characteristics and infection prevention practices. We assessed the association between SARS-CoV-2 infection prevalence and shelter characteristics, including 20 infection prevention practices by using crude risk ratios (RRs) and exact unconditional 95% confidence intervals (CIs).Results. Site assessments and SARS-CoV-2 testing results were reported for 63 homeless shelters in 7 US urban areas. Median infection prevalence was 2.9% (range = 0%-71.4%). Shelters implementing head-to-toe sleeping and excluding symptomatic staff from working were less likely to have high infection prevalence (RR = 0.5; 95% CI = 0.3, 0.8; and RR = 0.5; 95% CI = 0.4, 0.6; respectively); shelters with medical services available were less likely to have very high infection prevalence (RR = 0.5; 95% CI = 0.2, 1.0).Conclusions. Sleeping arrangements and staffing policies are modifiable factors that might be associated with SARS-CoV-2 infection prevalence in homeless shelters. Shelters should follow recommended practices to reduce the risk of SARS-CoV-2 transmission.

MG53 suppresses interferon-ß and inflammation via regulation of ryanodine receptor-mediated intracellular calcium signaling.

TRIM family proteins play integral roles in the innate immune response to virus infection. MG53 (TRIM72) is essential for cell membrane repair and is believed to be a muscle-specific TRIM protein. Here we show human macrophages express MG53, and MG53 protein expression is reduced following virus infection. Knockdown of MG53 in macrophages leads to increases in type I interferon (IFN) upon infection. MG53 knockout mice infected with influenza virus show comparable influenza virus titres to wild type mice, but display increased morbidity accompanied by more accumulation of CD45+ cells and elevation of IFNß in the lung. We find that MG53 knockdown results in activation of NFκB signalling, which is linked to an increase in intracellular calcium oscillation mediated by ryanodine receptor (RyR). MG53 inhibits IFNß induction in an RyR-dependent manner. This study establishes MG53 as a new target for control of virus-induced morbidity and tissue injury.

COVID-19 Outbreak Among Three Affiliated Homeless Service Sites - King County, Washington, 2020.

On March 30, 2020, Public Health - Seattle and King County (PHSKC) was notified of a confirmed case of coronavirus disease 2019 (COVID-19) in a resident of a homeless shelter and day center (shelter A). Residents from two other homeless shelters (B and C) used shelter A's day center services. Testing for SARS-CoV-2, the virus that causes COVID-19, was offered to available residents and staff members at the three shelters during March 30-April 1, 2020. Among the 181 persons tested, 19 (10.5%) had positive test results (15 residents and four staff members). On April 1, PHSKC and CDC collaborated to conduct site assessments and symptom screening, isolate ill residents and staff members, reinforce infection prevention and control practices, provide face masks, and advise on sheltering-in-place. Repeat testing was offered April 7-8 to all residents and staff members who were not tested initially or who had negative test results. Among the 118 persons tested in the second round of testing, 18 (15.3%) had positive test results (16 residents and two staff members). In addition to the 31 residents and six staff members identified through testing at the shelters, two additional cases in residents were identified during separate symptom screening events, and four were identified after two residents and two staff members independently sought health care. In total, COVID-19 was diagnosed in 35 of 195 (18%) residents and eight of 38 (21%) staff members who received testing at the shelter or were evaluated elsewhere. COVID-19 can spread quickly in homeless shelters; rapid interventions including testing and isolation to identify cases and minimize transmission are necessary. CDC recommends that homeless service providers implement appropriate infection control practices, apply physical distancing measures including ensuring resident's heads are at least 6 feet (2 meters) apart while sleeping, and promote use of cloth face coverings among all residents (1).

Assessment of SARS-CoV-2 Infection Prevalence in Homeless Shelters - Four U.S. Cities, March 27-April 15, 2020.

In the United States, approximately 1.4 million persons access emergency shelter or transitional housing each year (1). These settings can pose risks for communicable disease spread. In late March and early April 2020, public health teams responded to clusters (two or more cases in the preceding 2 weeks) of coronavirus disease 2019 (COVID-19) in residents and staff members from five homeless shelters in Boston, Massachusetts (one shelter); San Francisco, California (one); and Seattle, Washington (three). The investigations were performed in coordination with academic partners, health care providers, and homeless service providers. Investigations included reverse transcription-polymerase chain reaction testing at commercial and public health laboratories for SARS-CoV-2, the virus that causes COVID-19, over approximately 1-2 weeks for residents and staff members at the five shelters. During the same period, the team in Seattle, Washington, also tested residents and staff members at 12 shelters where a single case in each had been identified. In Atlanta, Georgia, a team proactively tested residents and staff members at two shelters with no known COVID-19 cases in the preceding 2 weeks. In each city, the objective was to test all shelter residents and staff members at each assessed facility, irrespective of symptoms. Persons who tested positive were transported to hospitals or predesignated community isolation areas.

COVID-19 in a Long-Term Care Facility - King County, Washington, February 27-March 9, 2020.

On February 28, 2020, a case of coronavirus disease (COVID-19) was identified in a woman resident of a long-term care skilled nursing facility (facility A) in King County, Washington.* Epidemiologic investigation of facility A identified 129 cases of COVID-19 associated with facility A, including 81 of the residents, 34 staff members, and 14 visitors; 23 persons died. Limitations in effective infection control and prevention and staff members working in multiple facilities contributed to intra- and interfacility spread. COVID-19 can spread rapidly in long-term residential care facilities, and persons with chronic underlying medical conditions are at greater risk for COVID-19-associated severe disease and death. Long-term care facilities should take proactive steps to protect the health of residents and preserve the health care workforce by identifying and excluding potentially infected staff members and visitors, ensuring early recognition of potentially infected patients, and implementing appropriate infection control measures.

Epidemiology of Covid-19 in a Long-Term Care Facility in King County, Washington.

BACKGROUND: Long-term care facilities are high-risk settings for severe outcomes from outbreaks of Covid-19, owing to both the advanced age and frequent chronic underlying health conditions of the residents and the movement of health care personnel among facilities in a region. METHODS: After identification on February 28, 2020, of a confirmed case of Covid-19 in a skilled nursing facility in King County, Washington, Public Health-Seattle and King County, aided by the Centers for Disease Control and Prevention, launched a case investigation, contact tracing, quarantine of exposed persons, isolation of confirmed and suspected cases, and on-site enhancement of infection prevention and control. RESULTS: As of March 18, a total of 167 confirmed cases of Covid-19 affecting 101 residents, 50 health care personnel, and 16 visitors were found to be epidemiologically linked to the facility. Most cases among residents included respiratory illness consistent with Covid-19; however, in 7 residents no symptoms were documented. Hospitalization rates for facility residents, visitors, and staff were 54.5%, 50.0%, and 6.0%, respectively. The case fatality rate for residents was 33.7% (34 of 101). As of March 18, a total of 30 long-term care facilities with at least one confirmed case of Covid-19 had been identified in King County. CONCLUSIONS: In the context of rapidly escalating Covid-19 outbreaks, proactive steps by long-term care facilities to identify and exclude potentially infected staff and visitors, actively monitor for potentially infected patients, and implement appropriate infection prevention and control measures are needed to prevent the introduction of Covid-19.

Interferon-induced transmembrane proteins inhibit cell fusion mediated by trophoblast syncytins.

Type I interferon (IFN) induced by virus infections during pregnancy can cause placental damage, but the mechanisms and identities of IFN-stimulated genes that are involved in this damage remain under investigation. The IFN-induced transmembrane proteins (IFITMs) inhibit virus infections by preventing virus membrane fusion with cells and by inhibiting fusion of infected cells (syncytialization). Fusion of placental trophoblasts via expression of endogenous retroviral fusogens known as syncytins forms the syncytiotrophoblast, a multinucleated cell structure essential for fetal development. We found here that IFN blocks fusion of BeWo human placental trophoblasts. Stably expressed IFITM1, -2, and -3 also blocked fusion of these trophoblasts while making them more resistant to virus infections. Conversely, stable IFITM knockdowns in BeWo trophoblasts increased their spontaneous fusion and allowed fusion in the presence of IFN while also making the cells more susceptible to virus infection. We additionally found that exogenous expression of IFITMs in HEK293T cells blocked fusion with cells expressing syncytin-1 or syncytin-2, confirming the ability of IFITMs to block individual syncytin-mediated fusion. Overall, our data indicate that IFITMs inhibit trophoblast fusion and suggest that there may be a critical balance between these antifusogenic effects and the beneficial antiviral effects of IFITMs in virus infections during pregnancy.

IFITM3 protects the heart during influenza virus infection.

Influenza virus can disseminate from the lungs to the heart in severe infections and can induce cardiac pathology, but this has been difficult to study due to a lack of small animal models. In humans, polymorphisms in the gene encoding the antiviral restriction factor IFN-induced transmembrane protein 3 (IFITM3) are associated with susceptibility to severe influenza, but whether IFITM3 deficiencies contribute to cardiac dysfunction during infection is unclear. We show that IFITM3 deficiency in a new knockout (KO) mouse model increases weight loss and mortality following influenza virus infections. We investigated this enhanced pathogenesis with the A/PR/8/34 (H1N1) (PR8) influenza virus strain, which is lethal in KO mice even at low doses, and observed increased replication of virus in the lungs, spleens, and hearts of KO mice compared with wild-type (WT) mice. Infected IFITM3 KO mice developed aberrant cardiac electrical activity, including decreased heart rate and irregular, arrhythmic RR (interbeat) intervals, whereas WT mice exhibited a mild decrease in heart rate without irregular RR intervals. Cardiac electrical dysfunction in PR8-infected KO mice was accompanied by increased activation of fibrotic pathways and fibrotic lesions in the heart. Infection with a sublethal dose of a less virulent influenza virus strain (A/WSN/33 [H1N1]) resulted in a milder cardiac electrical dysfunction in KO mice that subsided as the mice recovered. Our findings reveal an essential role for IFITM3 in limiting influenza virus replication and pathogenesis in heart tissue and establish IFITM3 KO mice as a powerful model for studying mild and severe influenza virus-induced cardiac dysfunction.

From APOBEC to ZAP: Diverse mechanisms used by cellular restriction factors to inhibit virus infections.

Antiviral restriction factors are cellular proteins that inhibit the entry, replication, or spread of viruses. These proteins are critical components of the innate immune system and function to limit the severity and host range of virus infections. Here we review the current knowledge on the mechanisms of action of several restriction factors that affect multiple viruses at distinct stages of their life cycles. For example, APOBEC3G deaminates cytosines to hypermutate reverse transcribed viral DNA; IFITM3 alters membranes to inhibit virus membrane fusion; MXA/B oligomerize on viral protein complexes to inhibit virus replication; SAMHD1 decreases dNTP intracellular concentrations to prevent reverse transcription of retrovirus genomes; tetherin prevents release of budding virions from cells; Viperin catalyzes formation of a nucleoside analogue that inhibits viral RNA polymerases; and ZAP binds virus RNAs to target them for degradation. We also discuss countermeasures employed by specific viruses against these restriction factors, and mention secondary functions of several of these factors in modulating immune responses. These important examples highlight the diverse strategies cells have evolved to combat virus infections.

IFITM3 Restricts Human Metapneumovirus Infection.

Human metapneumovirus (hMPV) utilizes a bifurcated cellular entry strategy, fusing either with the plasma membrane or, after endocytosis, with the endosome membrane. Whether cellular factors restrict or enhance either entry pathway is largely unknown. We found that the interferon-induced transmembrane protein 3 (IFITM3) inhibits hMPV infection to an extent similar to endocytosis-inhibiting drugs, and an IFITM3 variant that accumulates at the plasma membrane in addition to its endosome localization provided increased virus restriction. Mechanistically, IFITM3 blocks hMPV F protein-mediated membrane fusion, and inhibition of infection was reversed by the membrane destabilizing drug amphotericin B. Conversely, we found that infection by some hMPV strains is enhanced by the endosomal protein toll-like receptor 7 (TLR7), and that IFITM3 retains the ability to restrict hMPV infection even in cells expressing TLR7. Overall, our results identify IFITM3 as an endosomal restriction factor that limits hMPV infection of cells.

The palmitoyltransferase ZDHHC20 enhances interferon-induced transmembrane protein 3 (IFITM3) palmitoylation and antiviral activity.

Interferon-induced transmembrane protein 3 (IFITM3) is a cellular endosome- and lysosome-localized protein that restricts numerous virus infections. IFITM3 is activated by palmitoylation, a lipid posttranslational modification. Palmitoylation of proteins is primarily mediated by zinc finger DHHC domain-containing palmitoyltransferases (ZDHHCs), but which members of this enzyme family can modify IFITM3 is not known. Here, we screened a library of human cell lines individually lacking ZDHHCs 1-24 and found that IFITM3 palmitoylation and its inhibition of influenza virus infection remained strong in the absence of any single ZDHHC, suggesting functional redundancy of these enzymes in the IFITM3-mediated antiviral response. In an overexpression screen with 23 mammalian ZDHHCs, we unexpectedly observed that more than half of the ZDHHCs were capable of increasing IFITM3 palmitoylation with ZDHHCs 3, 7, 15, and 20 having the greatest effect. Among these four enzymes, ZDHHC20 uniquely increased IFITM3 antiviral activity when both proteins were overexpressed. ZDHHC20 colocalized extensively with IFITM3 at lysosomes unlike ZDHHCs 3, 7, and 15, which showed a defined perinuclear localization pattern, suggesting that the location at which IFITM3 is palmitoylated may influence its activity. Unlike knock-out of individual ZDHHCs, siRNA-mediated knockdown of both ZDHHC3 and ZDHHC7 in ZDHHC20 knock-out cells decreased endogenous IFITM3 palmitoylation. Overall, our results demonstrate that multiple ZDHHCs can palmitoylate IFITM3 to ensure a robust antiviral response and that ZDHHC20 may serve as a particularly useful tool for understanding and enhancing IFITM3 activity.

Human Genetic Determinants of Viral Diseases.

Much progress has been made in the identification of specific human gene variants that contribute to enhanced susceptibility or resistance to viral diseases. Herein we review multiple discoveries made with genome-wide or candidate gene approaches that have revealed significant insights into virus-host interactions. Genetic factors that have been identified include genes encoding virus receptors, receptor-modifying enzymes, and a wide variety of innate and adaptive immunity-related proteins. We discuss a range of pathogenic viruses, including influenza virus, respiratory syncytial virus, human immunodeficiency virus, human T cell leukemia virus, human papilloma virus, hepatitis B and C viruses, herpes simplex virus, norovirus, rotavirus, parvovirus, and Epstein-Barr virus. Understanding the genetic underpinnings that affect infectious disease outcomes should allow tailored treatment and prevention approaches in the future.

IFITM3 requires an amphipathic helix for antiviral activity.

Interferon-induced transmembrane protein 3 (IFITM3) is a cellular factor that blocks virus fusion with cell membranes. IFITM3 has been suggested to alter membrane curvature and fluidity, though its exact mechanism of action is unclear. Using a bioinformatic approach, we predict IFITM3 secondary structures and identify a highly conserved, short amphipathic helix within a hydrophobic region of IFITM3 previously thought to be a transmembrane domain. Consistent with the known ability of amphipathic helices to alter membrane properties, we show that this helix and its amphipathicity are required for the IFITM3-dependent inhibition of influenza virus, Zika virus, vesicular stomatitis virus, Ebola virus, and human immunodeficiency virus infections. The homologous amphipathic helix within IFITM1 is also required for the inhibition of infection, indicating that IFITM proteins possess a conserved mechanism of antiviral action. We further demonstrate that the amphipathic helix of IFITM3 is required to block influenza virus hemagglutinin-mediated membrane fusion. Overall, our results provide evidence that IFITM proteins utilize an amphipathic helix for inhibiting virus fusion.

E3 Ubiquitin Ligase NEDD4 Promotes Influenza Virus Infection by Decreasing Levels of the Antiviral Protein IFITM3.

Interferon (IFN)-induced transmembrane protein 3 (IFITM3) is a cell-intrinsic factor that limits influenza virus infections. We previously showed that IFITM3 degradation is increased by its ubiquitination, though the ubiquitin ligase responsible for this modification remained elusive. Here, we demonstrate that the E3 ubiquitin ligase NEDD4 ubiquitinates IFITM3 in cells and in vitro. This IFITM3 ubiquitination is dependent upon the presence of a PPxY motif within IFITM3 and the WW domain-containing region of NEDD4. In NEDD4 knockout mouse embryonic fibroblasts, we observed defective IFITM3 ubiquitination and accumulation of high levels of basal IFITM3 as compared to wild type cells. Heightened IFITM3 levels significantly protected NEDD4 knockout cells from infection by influenza A and B viruses. Similarly, knockdown of NEDD4 in human lung cells resulted in an increase in steady state IFITM3 and a decrease in influenza virus infection, demonstrating a conservation of this NEDD4-dependent IFITM3 regulatory mechanism in mouse and human cells. Consistent with the known association of NEDD4 with lysosomes, we demonstrate for the first time that steady state turnover of IFITM3 occurs through the lysosomal degradation pathway. Overall, this work identifies the enzyme NEDD4 as a new therapeutic target for the prevention of influenza virus infections, and introduces a new paradigm for up-regulating cellular levels of IFITM3 independently of IFN or infection.

IFITMs from Mycobacteria Confer Resistance to Influenza Virus When Expressed in Human Cells.

Interferon induced transmembrane proteins (IFITMs) found in vertebrates restrict infections by specific viruses. IFITM3 is known to be essential for restriction of influenza virus infections in both mice and humans. Vertebrate IFITMs are hypothesized to have derived from a horizontal gene transfer from bacteria to a primitive unicellular eukaryote. Since bacterial IFITMs share minimal amino acid identity with human IFITM3, we hypothesized that examination of bacterial IFITMs in human cells would provide insight into the essential characteristics necessary for antiviral activity of IFITMs. We examined IFITMs from Mycobacterium avium and Mycobacterium abscessus for potential antiviral activity. Both of these IFITMs conferred a moderate level of resistance to influenza virus in human cells, identifying them as functional homologues of IFITM3. Analysis of sequence elements shared by bacterial IFITMs and IFITM3 identified two hydrophobic domains, putative S-palmitoylation sites, and conserved phenylalanine residues associated with IFITM3 interactions, which are all necessary for IFITM3 antiviral activity. We observed that, like IFITM3, bacterial IFITMs were S-palmitoylated, albeit to a lesser degree. We also demonstrated the ability of a bacterial IFITM to co-immunoprecipitate with IFITM3 suggesting formation of a complex, and also visualized strong co-localization of bacterial IFITMs with IFITM3. However, the mycobacterial IFITMs lack the endocytic-targeting motif conserved in vertebrate IFITM3. As such, these bacterial proteins, when expressed alone, had diminished colocalization with cathepsin B-positive endolysosomal compartments that are the primary site of IFITM3-dependent influenza virus restriction. Though the precise evolutionary origin of vertebrate IFITMs is not known, our results support a model whereby transfer of a bacterial IFITM gene to eukaryotic cells may have provided a selective advantage against viral infection that was refined through the course of vertebrate evolution to include more robust signals for S-palmitoylation and localization to sites of endocytic virus trafficking.