Spaced training programs employ short, frequent CPR training sessions to improve provider skills. The optimum training frequency for CPR skill acquisition and retention has not been determined. We ...aimed to determine the training interval associated with the highest quality CPR performance at one year.
Participants were randomized to 1-month, 3-month, 6-month, and 12-month CPR training intervals over the course of a 12-month study period. Practice sessions included repeated two-minute CPR practice sessions with visual feedback and verbal coaching until Excellent CPR was achieved, to a maximum of three attempts. Excellent CPR was defined as a two-minute CPR session with ≥90% of compressions with a depth of 50–60 millimeters, a rate of 100–120 per minute, and with complete chest recoil. CPR performance was assessed in all groups at 12 months. The primary outcome was the proportion of participants able to perform Excellent CPR in each group.
A total of 167 participants were included in the analysis. Baseline assessment showed no difference in CPR performance (p = 0.38). Participants who were trained monthly had a significantly higher proportion of Excellent CPR performance (58%) than those in all other groups (26% in the 3-month group, p = 0.008; 21% in the 6-month group, p = 0.002; and 15% in the 12-month group, p < 0.001).
Short-duration, distributed CPR training on a manikin with real-time visual feedback is effective in improving CPR performance, with monthly training more effective than training every 3, 6, or 12 months.
One vital aspect of emergency medicine management is communication after episodes of care to improve future performance through group reflection on the shared experience. This reflective activity in ...teams is known as debriefing, and despite supportive evidence highlighting its benefits, many practitioners experience barriers to implementing debriefing in the clinical setting. The aim of this article is to review the current evidence supporting postevent debriefing and discuss practical approaches to implementing debriefing in the emergency department. We will address the who, what, when, where, why, and how of debriefing and provide a practical guide for the clinician to facilitate debriefing in the clinical environment.
HIV-1 must overcome multiple innate antiviral mechanisms to replicate in CD4
T lymphocytes and macrophages. Previous studies have demonstrated that the apolipoprotein B mRNA editing enzyme ...polypeptide-like 3 (APOBEC3, A3) family of proteins (at least A3D, A3F, A3G, and stable A3H haplotypes) contribute to HIV-1 restriction in CD4
T lymphocytes. Virus-encoded virion infectivity factor (Vif) counteracts this antiviral activity by degrading A3 enzymes allowing HIV-1 replication in infected cells. In addition to A3 proteins, Vif also targets other cellular proteins in CD4
T lymphocytes, including PPP2R5 proteins. However, whether Vif primarily degrades only A3 proteins during viral replication is currently unknown. Herein, we describe the development and characterization of
-,
-, and
-to-
-null THP-1 cells. In comparison to Vif-proficient HIV-1, Vif-deficient viruses have substantially reduced infectivity in parental and
-null THP-1 cells, and a more modest decrease in infectivity in
-null cells. Remarkably, disruption of A3A-A3G protein expression completely restores the infectivity of Vif-deficient viruses in THP-1 cells. These results indicate that the primary function of Vif during infectious HIV-1 production from THP-1 cells is the targeting and degradation of A3 enzymes. IMPORTANCE HIV-1 Vif neutralizes the HIV-1 restriction activity of A3 proteins. However, it is currently unclear whether Vif has additional essential cellular targets. To address this question, we disrupted
3
to
3
genes in the THP-1 myeloid cell line using CRISPR and compared the infectivity of wild-type HIV-1 and Vif mutants with the selective A3 neutralization activities. Our results demonstrate that the infectivity of Vif-deficient HIV-1 and the other Vif mutants is fully restored by ablating the expression of cellular A3A to A3G proteins. These results indicate that A3 proteins are the only essential target of Vif that is required for fully infectious HIV-1 production from THP-1 cells.
Bats have attracted attention in recent years as important reservoirs of viruses deadly to humans and other mammals. These infections are typically nonpathogenic in bats raising questions about ...innate immune differences that might exist between bats and other mammals. The APOBEC3 gene family encodes antiviral DNA cytosine deaminases with important roles in the suppression of diverse viruses and genomic parasites. Here, we characterize pteropid APOBEC3 genes and show that species within the genus Pteropus possess the largest and most diverse array of APOBEC3 genes identified in any mammal reported to date. Several bat APOBEC3 proteins are antiviral as demonstrated by restriction of retroviral infectivity using HIV-1 as a model, and recombinant A3Z1 subtypes possess strong DNA deaminase activity. These genes represent the first group of antiviral restriction factors identified in bats with extensive diversification relative to homologues in other mammals.
The APOBEC3 family of DNA cytosine deaminases comprises an important arm of the innate antiviral defense system. The gamma-herpesviruses Epstein-Barr virus and Kaposi's sarcoma-associated herpesvirus ...and the alpha-herpesviruses herpes simplex virus (HSV)-1 and HSV-2 have evolved an efficient mechanism to avoid APOBEC3 restriction by directly binding to APOBEC3B and facilitating its exclusion from the nuclear compartment. The only viral protein required for APOBEC3B relocalization is the large subunit of the ribonucleotide reductase (RNR). Here, we ask whether this APOBEC3B relocalization mechanism is conserved with the beta-herpesvirus human cytomegalovirus (HCMV). Although HCMV infection causes APOBEC3B relocalization from the nucleus to the cytoplasm in multiple cell types, the viral RNR (UL45) is not required. APOBEC3B relocalization occurs rapidly following infection suggesting the involvement of an immediate early or early (IE/E) viral protein. In support of this possibility, genetic (IE1 mutant) and pharmacologic (cycloheximide) strategies that prevent the expression of IE/E viral proteins also block APOBEC3B relocalization. In comparison, the treatment of infected cells with phosphonoacetic acid, which interferes with viral late protein expression, still permits A3B relocalization. These results combine to indicate that the beta-herpesvirus HCMV uses an RNR-independent, yet phenotypically similar, molecular mechanism to antagonize APOBEC3B. IMPORTANCE Human cytomegalovirus (HCMV) infections can range from asymptomatic to severe, particularly in neonates and immunocompromised patients. HCMV has evolved strategies to overcome host-encoded antiviral defenses to achieve lytic viral DNA replication and dissemination and, under some conditions, latency and long-term persistence. Here, we show that HCMV infection causes the antiviral factor, APOBEC3B, to relocalize from the nuclear compartment to the cytoplasm. This overall strategy resembles that used by related herpesviruses. However, the HCMV relocalization mechanism utilizes a different viral factor(s) and available evidence suggests the involvement of at least one protein expressed at the early stages of infection. This knowledge is important because a greater understanding of this mechanism could lead to novel antiviral strategies that enable APOBEC3B to naturally restrict HCMV infection.
An integral part of the antiviral innate immune response is the APOBEC3 family of single-stranded DNA cytosine deaminases, which inhibits virus replication through deamination-dependent and ...-independent activities. Viruses have evolved mechanisms to counteract these enzymes, such as HIV-1 Vif-mediated formation of a ubiquitin ligase to degrade virus-restrictive APOBEC3 enzymes. A new example is Epstein-Barr virus (EBV) ribonucleotide reductase (RNR)-mediated inhibition of cellular APOBEC3B (A3B). The large subunit of the viral RNR, BORF2, causes A3B relocalization from the nucleus to cytoplasmic bodies and thereby protects viral DNA during lytic replication. Here, we use coimmunoprecipitation and immunofluorescence microscopy approaches to ask whether this mechanism is shared with the closely related gammaherpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV) and the more distantly related alphaherpesvirus herpes simplex virus 1 (HSV-1). The large RNR subunit of KSHV, open reading frame 61 (ORF61), coprecipitated multiple APOBEC3s, including A3B and APOBEC3A (A3A). KSHV ORF61 also caused relocalization of these two enzymes to perinuclear bodies (A3B) and to oblong cytoplasmic structures (A3A). The large RNR subunit of HSV-1, ICP6, also coprecipitated A3B and A3A and was sufficient to promote the relocalization of these enzymes from nuclear to cytoplasmic compartments. HSV-1 infection caused similar relocalization phenotypes that required ICP6. However, unlike the infectivity defects previously reported for BORF2-null EBV, ICP6 mutant HSV-1 showed normal growth rates and plaque phenotypes. Combined, these results indicate that both gamma- and alphaherpesviruses use a conserved RNR-dependent mechanism to relocalize A3B and A3A and furthermore suggest that HSV-1 possesses at least one additional mechanism to neutralize these antiviral enzymes.
The APOBEC3 family of DNA cytosine deaminases constitutes a vital innate immune defense against a range of different viruses. A novel counterrestriction mechanism has recently been uncovered for the gammaherpesvirus EBV, in which a subunit of the viral protein known to produce DNA building blocks (ribonucleotide reductase) causes A3B to relocalize from the nucleus to the cytosol. Here, we extend these observations with A3B to include a closely related gammaherpesvirus, KSHV, and a more distantly related alphaherpesvirus, HSV-1. These different viral ribonucleotide reductases also caused relocalization of A3A, which is 92% identical to A3B. These studies are important because they suggest a conserved mechanism of APOBEC3 evasion by large double-stranded DNA herpesviruses. Strategies to block this host-pathogen interaction may be effective for treating infections caused by these herpesviruses.
Herpesvirus lytic infection causes cells to arrest at the G1/S phase of the cell cycle by poorly defined mechanisms. In a prior study using fluorescent ubiquitination-based cell cycle indicator ...(FUCCI) cells that express fluorescently tagged proteins marking different stages of the cell cycle, we showed that the Epstein-Barr virus (EBV) protein BORF2 induces the accumulation of G1/S cells, and that BORF2 affects p53 levels without affecting the p53 target protein p21. We also found that BORF2 specifically interacted with APOBEC3B (A3B) and forms perinuclear bodies with A3B that prevent A3B from mutating replicating EBV genomes. We now show that BORF2 also interacts with p53 and that A3B interferes with the BORF2-p53 interaction, although A3B and p53 engage distinct surfaces on BORF2. Cell cycle analysis showed that G1/S induction by BORF2 is abrogated when either p53 or A3B is silenced or when an A3B-binding mutant of BORF2 is used. Furthermore, silencing A3B in EBV lytic infection increased cell proliferation, supporting a role for A3B in G1/S arrest. These data suggest that the p53 induced by BORF2 is inactive when it binds BORF2, but is released and induces G1/S arrest when A3B is present and sequesters BORF2 in perinuclear bodies. Interestingly, this mechanism is conserved in the BORF2 homologue in HSV-1, which also re-localizes A3B, induces and binds p53, and induces G1/S dependent on A3B and p53. In summary, we have identified a new mechanism by which G1/S arrest can be induced in herpesvirus lytic infection. IMPORTANCE In lytic infection, herpesviruses cause cells to arrest at the G1/S phase of the cell cycle in order to provide an optimal environment for viral replication; however, the mechanisms involved are not well understood. We have shown that the Epstein-Barr virus BORF2 protein and its homologue in herpes simplex virus 1 both induce G1/S, and do this by similar mechanisms which involve binding p53 and APOBEC3B and induction of p53. Our study identifies a new mechanism by which G1/S arrest can be induced in herpesvirus lytic infection and a new role of APOBEC3B in herpesvirus lytic infection.
Herpesvirus lytic infection causes cells to arrest at the G
/S phase of the cell cycle by poorly defined mechanisms. In a prior study using fluorescent ubiquitination-based cell cycle indicator ...(FUCCI) cells that express fluorescently tagged proteins marking different stages of the cell cycle, we showed that the Epstein-Barr virus (EBV) protein BORF2 induces the accumulation of G
/S cells, and that BORF2 affects p53 levels without affecting the p53 target protein p21. We also found that BORF2 specifically interacted with APOBEC3B (A3B) and forms perinuclear bodies with A3B that prevent A3B from mutating replicating EBV genomes. We now show that BORF2 also interacts with p53 and that A3B interferes with the BORF2-p53 interaction, although A3B and p53 engage distinct surfaces on BORF2. Cell cycle analysis showed that G
/S induction by BORF2 is abrogated when either p53 or A3B is silenced or when an A3B-binding mutant of BORF2 is used. Furthermore, silencing A3B in EBV lytic infection increased cell proliferation, supporting a role for A3B in G
/S arrest. These data suggest that the p53 induced by BORF2 is inactive when it binds BORF2, but is released and induces G
/S arrest when A3B is present and sequesters BORF2 in perinuclear bodies. Interestingly, this mechanism is conserved in the BORF2 homologue in HSV-1, which also re-localizes A3B, induces and binds p53, and induces G
/S dependent on A3B and p53. In summary, we have identified a new mechanism by which G
/S arrest can be induced in herpesvirus lytic infection.
In lytic infection, herpesviruses cause cells to arrest at the G
/S phase of the cell cycle in order to provide an optimal environment for viral replication; however, the mechanisms involved are not well understood. We have shown that the Epstein-Barr virus BORF2 protein and its homologue in herpes simplex virus 1 both induce G
/S, and do this by similar mechanisms which involve binding p53 and APOBEC3B and induction of p53. Our study identifies a new mechanism by which G
/S arrest can be induced in herpesvirus lytic infection and a new role of APOBEC3B in herpesvirus lytic infection.
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