Detection of Mpox Virus Using Microbial Cell-Free DNA: The Potential of Pathogen-Agnostic Sequencing for Rapid Identification of Emerging Pathogens.

Background: The 2022 global outbreak of Monkeypox virus (MPXV) highlighted challenges with polymerase chain reaction detection as divergent strains emerged and atypical presentations limited the applicability of swab sampling. Recommended testing in the United States requires a swab of lesions, which arise late in infection and may be unrecognized. We present MPXV detections using plasma microbial cell-free DNA (mcfDNA) sequencing.

Reductions in inpatient fluoroquinolone use and postdischarge infection (CDI) from a systemwide antimicrobial stewardship intervention.

Objective: To determine the impact of an inpatient stewardship intervention targeting fluoroquinolone use on inpatient and postdischarge infection (CDI).

Design: We used an interrupted time series study design to evaluate the rate of hospital-onset CDI (HO-CDI), postdischarge CDI (PD-CDI) within 12 weeks, and inpatient fluoroquinolone use from 2 years prior to 1 year after a stewardship intervention.

Setting: An academic healthcare system with 4 hospitals.

Decreased T Follicular Regulatory Cell/T Follicular Helper Cell (TFH) in Simian Immunodeficiency Virus-Infected Rhesus Macaques May Contribute to Accumulation of TFH in Chronic Infection.

T follicular helper cells (TFH) are critical for the development and maintenance of germinal center (GC) and humoral immune responses. During chronic HIV/SIV infection, TFH accumulate, possibly as a result of Ag persistence. The HIV/SIV-associated TFH expansion may also reflect lack of regulation by suppressive follicular regulatory CD4(+) T cells (TFR).

Induction of human plasmablasts during infection with antibiotic-resistant nosocomial bacteria.

Objectives: Nosocomial pathogens such as Acinetobacter baumannii are a growing public health threat, due in part to their increasing resistance to antibiotics. Since some strains are resistant to all available antibiotics, novel therapies are urgently needed. Plasmablasts are short-lived B cells found in the blood that can be collected and harnessed to produce therapeutic antibodies.

Discovery of a diaminoquinoxaline benzenesulfonamide antagonist of HIV-1 Nef function using a yeast-based phenotypic screen.

Background: HIV-1 Nef is a viral accessory protein critical for AIDS progression. Nef lacks intrinsic catalytic activity and binds multiple host cell signaling proteins, including Hck and other Src-family tyrosine kinases. Nef binding induces constitutive Hck activation that may contribute to HIV pathogenesis by promoting viral infectivity, replication and downregulation of cell-surface MHC-I molecules.

Allosteric loss-of-function mutations in HIV-1 Nef from a long-term non-progressor.

Activation of Src family kinases by human immunodeficiency virus type 1 (HIV-1) Nef may play an important role in the pathogenesis of HIV/AIDS. Here we investigated whether diverse Nef sequences universally activate Hck, a Src family member expressed in macrophages and other HIV-1 target cells. In general, we observed that Hck activation is a highly conserved Nef function.

Functional characterization and conformational analysis of the Herpesvirus saimiri Tip-C484 protein.

Tyrosine kinase interacting protein (Tip) of Herpesvirus saimiri (HVS) activates the lymphoid-specific member of the Src family kinase Lck. The Tip:Lck interaction is essential for transformation and oncogenesis in HVS-infected cells. As there are no structural data for Tip, hydrogen-exchange mass spectrometry was used to investigate the conformation of a nearly full-length form (residues 1-187) of Tip from HVS strain C484.

HIV-1 Nef selectively activates Src family kinases Hck, Lyn, and c-Src through direct SH3 domain interaction.

Nef is an HIV-1 virulence factor that promotes viral pathogenicity by altering host cell signaling pathways. Nef binds several members of the Src kinase family, and these interactions have been implicated in the pathogenesis of HIV/AIDS. However, the direct effect of Nef interaction on Src family kinase (SFK) regulation and activity has not been systematically addressed.

Activation of the Src family kinase Hck without SH3-linker release.

Src family protein-tyrosine kinases are regulated by intramolecular binding of the SH2 domain to the C-terminal tail and association of the SH3 domain with the SH2 kinase-linker. The presence of two regulatory interactions raises the question of whether disruption of both is required for kinase activation. To address this question, we engineered a high affinity linker (HAL) mutant of the Src family member Hck in which an optimal SH3 ligand was substituted for the natural linker.

The conserved process of TCR/CD3 complex down-modulation by SIV Nef is mediated by the central core, not endocytic motifs.

The Nef protein of Simian immunodeficiency virus (SIV) associates with multiple T lymphocyte signaling proteins, including the T cell receptor (TCR) zeta chain. We demonstrate here that these interactions are conserved and highly specific. Nefs derived from genetically diverse strains of SIV (SIV(mac)239, SIV(smm)PBj, and SIV(smm)DeltaB670) all interacted with TCR zeta on two separate domains, referred to as SIV Nef interaction domains (SNIDs), as examined in both yeast two-hybrid and glutathione-S-transferase (GST) fusion protein pull-down assays.

Down-modulation of the costimulatory molecule, CD28, is a conserved activity of multiple SIV Nefs and is dependent on histidine 196 of Nef.

In this study Nef proteins derived from simian immunodeficiency virus (SIV) and human immunodeficiency virus (HIV) were compared to assess their abilities to down-modulate the cell surface levels of the T-cell costimulatory molecule CD28. We demonstrate that in addition to Nef derived from the prototypic SIVmac239, Nef proteins encoded by the pathogenic SIVsmmPBj molecular clone and the SIVsmmB670 isolate also down-modulate cell surface CD28. In contrast, Nef proteins derived from HIV failed to down-modulate CD28.

Dynamic actin polymerization drives T cell receptor-induced spreading: a role for the signal transduction adaptor LAT.

T cell activation induces functional changes in cell shape and cytoskeletal architecture. To facilitate the collection of dynamic, high-resolution images of activated T cells, we plated T cells on coverslips coated with antibodies to the T cell receptor (TCR). Using these images, we were able to quantitate the morphological responses of individual cells over time.

Association of Grb2, Gads, and phospholipase C-gamma 1 with phosphorylated LAT tyrosine residues. Effect of LAT tyrosine mutations on T cell angigen receptor-mediated signaling.

The linker for activation of T cells (LAT) is a critical adaptor molecule required for T cell antigen receptor (TCR)-mediated signaling and thymocyte development. Upon T cell activation, LAT becomes highly phosphorylated on tyrosine residues, and Grb2, Gads, and phospholipase C (PLC)-gamma1 bind LAT via Src homology-2 domains. In LAT-deficient mutant Jurkat cells, TCR engagement fails to induce ERK activation, Ca(2+) flux, and activation of AP-1 and NF-AT.

Functional analysis of LAT in TCR-mediated signaling pathways using a LAT-deficient Jurkat cell line.

The adaptor molecule LAT (linker for activation of T cells) is a palmitoylated integral membrane protein that localizes to the glycolipid-enriched microdomains in the plasma membrane. Upon TCR engagement, LAT becomes phosphorylated on multiple tyrosine residues and then binds several critical signaling molecules. Here, we describe the generation and characterization of a LAT-deficient cell line.

Essential role of LAT in T cell development.

The linker molecule LAT is a substrate of the tyrosine kinases activated following TCR engagement. Phosphorylated LAT binds many critical signaling molecules. The central role of this molecule in TCR-mediated signaling has been demonstrated by experiments in a LAT-deficient cell line.

LAT palmitoylation: its essential role in membrane microdomain targeting and tyrosine phosphorylation during T cell activation.

The linker molecule LAT is a critical substrate of the tyrosine kinases activated upon TCR engagement. Phosphorylated LAT binds Grb2, PLC-gamma1, and other signaling molecules. We demonstrate that human LAT is palmitoylated and that palmitoylated LAT predominantly localizes into glycolipid-enriched microdomains (GEMs).

LAT: the ZAP-70 tyrosine kinase substrate that links T cell receptor to cellular activation.

Despite extensive study, several of the major components involved in T cell receptor-mediated signaling remain unidentified. Here we report the cloning of the cDNA for a highly tyrosine-phosphorylated 36-38 kDa protein, previously characterized by its association with Grb2, phospholipase C-gamma1, and the p85 subunit of phosphoinositide 3-kinase. Deduced amino acid sequence identifies a novel integral membrane protein containing multiple potential tyrosine phosphorylation sites.