Comparison of the clinical characteristics of SARS-CoV-2 Delta (B.1.617.2) and Omicron (B.1.1.529) infected patients from a single hospitalist service.

BACKGROUND: While existing evidence suggests less severe clinical manifestations and lower mortality are associated with the Omicron variant as compared to the Delta variant. However, these studies fail to control for differences in health systems facilities and providers. By comparing patients hospitalized on a single medical service during the Delta and Omicron surges we were able to conduct a more accurate comparison of the two varaints' clinical manifestations and outcomes. METHODS: We conducted a prospective study of 364 Omicron (BA.1) infected patients on a single hospitalist service and compared these findings to a retrospective analysis of 241 Delta variant infected patients managed on the same service. We examined differences in symptoms, laboratory measures, and clinical severity between the two variants and assessed potential risk drivers for case mortality. FINDINGS: Patients infected with Omicron were older and had more underlying medical conditions increasing their risk of death. Although they were less severely ill and required less supplemental oxygen and dexamethasone, in-hospital mortality was similar to Delta cases, 7.14% vs. 4.98% for Delta (q-value = 0.38). Patients older than 60 years or with immunocompromised conditions had much higher risk of death during hospitalization, with estimated odds ratios of 17.46 (95% CI: 5.05, 110.51) and 2.80 (1.03, 7.08) respectively. Neither vaccine history nor variant type played a significant role in case fatality. The Rothman score, NEWS-2 score, level of neutrophils, level of care, age, and creatinine level at admission were highly predictive of in-hospital death. INTERPRETATION: In hospitalized patients, the Omicron variant is less virulent than the Delta variant but is associated with a comparable mortality. Clinical and laboratory features at admission are informative about the risk of death.

N-WASP deficiency reveals distinct pathways for cell surface projections and microbial actin-based motility.

The Wiskott-Aldrich syndrome protein (WASP) family of molecules integrates upstream signalling events with changes in the actin cytoskeleton. N-WASP has been implicated both in the formation of cell-surface projections (filopodia) required for cell movement and in the actin-based motility of intracellular pathogens. To examine N-WASP function we have used homologous recombination to inactivate the gene encoding murine N-WASP. Whereas N-WASP-deficient embryos survive beyond gastrulation and initiate organogenesis, they have marked developmental delay and die before embryonic day 12. N-WASP is not required for the actin-based movement of the intracellular pathogen Listeria but is absolutely required for the motility of Shigella and vaccinia virus. Despite these distinct defects in bacterial and viral motility, N-WASP-deficient fibroblasts spread by using lamellipodia and can protrude filopodia. These results imply a crucial and non-redundant role for N-WASP in murine embryogenesis and in the actin-based motility of certain pathogens but not in the general formation of actin-containing structures.

The actin released from profilin--actin complexes is insufficient to account for the increase in F-actin in chemoattractant-stimulated polymorphonuclear leukocytes.

Chemoattractant stimulation of polymorphonuclear leukocytes is associated with a nearly two-fold rise in actin filament content. We examined the role of the actin monomer sequestering protein, profilin, in the regulation of PMN actin filament assembly during chemoattractant stimulation using a Triton extraction method. Poly-L-proline-conjugated Sepharose beads were used to assess the relative concentration of actin bound to profilin with high enough affinity to withstand dilution (profilin-actin complex) and DNase I-conjugated beads to measure the relative concentration of actin in the Triton-soluble fraction not bound to profilin. Actin associated with the Triton-insoluble fraction (F-actin) was also measured. In unstimulated PMN, the relative concentration of actin bound to profilin was maximum. After FMLP stimulation, profilin released actin monomers within 10 s, with the profilin-actin complex concentration reaching a nadir by 40 s and remaining low as long as the cells were exposed to chemoattractant (up to 30 min). If FMLP was dissociated from PMN membrane receptors using t-BOC, actin reassociated with profilin within 20 s. Quantitative analysis of these reactions, however, revealed that profilin release of and rebinding to actin could account for only a small percentage of the total change in F-actin content. Determination of the total profilin and actin concentrations in PMN revealed that the molar ratio of profilin to actin was 1 to 5.2. When purified actin was polymerized in PMN Triton extract containing EGTA, removal of profilin from the extract minimally affected (12% reduction) the high apparent critical concentration at which actin began to assemble. Although profilin released actin at the appropriate time to stimulate actin assembly during exposure to chemoattractants, the concentration of profilin in PMN was insufficient to explain the high unpolymerized actin content in unstimulated PMN and the quantity of actin released from profilin too small to account for the large shifts from unpolymerized to polymerized actin associated with maximal chemoattractant stimulation.

Recognition of two classes of oligoproline sequences in profilin-mediated acceleration of actin-based Shigella motility.

The gram negative rod Shigella flexneri uses it surface protein IcsA to induce host cell actin assembly and to achieve intracellular motility. Yet, the IcsA protein lacks the oligoproline sequences found in ActA, the surface protein required for locomotion of the gram positive rod Listeria monocytogenes. Microinjection of a peptide matching the second ActA oligoproline repeat (FEFPPPPTDE) stops Listeria locomotion (Southwick, F.S., and D.L. Purich. 1994a. Proc. Natl. Acad. Sci. USA. 91:5168-5172), and submicromolar concentrations (intracellular concentration 80-800 nM) similarly arrest Shigella rocket-tail assembly and intracellular motility. Coinjection of a binary solution containing profilin and the ActA analogue increased the observed rates of intracellular motility by a factor of three (mean velocity 0.90 +/- 0.07 mu m/s, SD n=16 before injection vs 0.3 +/- 0.1 mu m/s, n=33 postinjection, intracellular concentration = 80 nM profilin plus 80 nM ActA analogue). Recent evidence suggests the ActA analogue may act by displacing the profilin-binding protein VASP (Pistor, S.C., T. Chakaborty, V. Walter, and J. Wehland. 1995. Curr. Biol. 5:517-525). At considerably higher intracellular concentrations (10 muM), the VASP oligoproline sequence (GPPPPP)3 thought to represent the profilin-binding site (Reinhard, M., K. Giehl, K. Abel, C. Haffner, T. Jarchau, V. Hoppe, B.M. Jockusch, and U. Walter. 1995. EMBO (Eur. Mol. Biol. Organ.) J. 14:1583-1589) also inhibited Shigella movement. A binary mixture of the VASP analogue and profilin (each 10 muM intracellular concentration) led to a doubling of Shigella intracellular migration velocity (0.09 +/- 0.06 mu m/s, n = 25 preinjection vs 0.18 +/- 0.10 mu m/s, n = 61 postinjection). Thus, the two structurally divergent bacteria, Listeria and Shigella, have adopted convergent mechanisms involving profilin recognition of VASP oligoproline sequences and VASP recognition of oligoproline sequences in ActA or an ActA-like host protein to induce host cell actin assembly and to provide the force for intracellular locomotion and cell-cell spread.

Comparisons of CapG and gelsolin-null macrophages: demonstration of a unique role for CapG in receptor-mediated ruffling, phagocytosis, and vesicle rocketing.

Capping the barbed ends of actin filaments is a critical step for regulating actin-based motility in nonmuscle cells. The in vivo function of CapG, a calcium-sensitive barbed end capping protein and member of the gelsolin/villin family, has been assessed using a null Capg allele engineered into mice. Both CapG-null mice and CapG/gelsolin double-null mice appear normal and have no gross functional abnormalities. However, the loss of CapG in bone marrow macrophages profoundly inhibits macrophage colony stimulating factor-stimulated ruffling; reintroduction of CapG protein by microinjection fully restores this function. CapG-null macrophages also demonstrate approximately 50% impairment of immunoglobulin G, and complement-opsonized phagocytosis and lanthanum-induced vesicle rocketing. These motile functions are not impaired in gelsolin-null macrophages and no additive effects are observed in CapG/gelsolin double-null macrophages, establishing that CapG function is distinct from, and does not overlap with, gelsolin in macrophages. Our observations indicate that CapG is required for receptor-mediated ruffling, and that it is a major functional component of macrophage phagocytosis. These primary effects on macrophage motile function suggest that CapG may be a useful target for the regulation of macrophage-mediated inflammatory responses.

Polymorphonuclear leukocyte adherence induces actin polymerization by a transduction pathway which differs from that used by chemoattractants.

Nitrobenzoxadiazole-phallacidin in combination with quantitative fluorescent microscopy have been used to measure F-actin concentrations in human polymorphonuclear leukocytes (PMN) as they adhere to a plastic surface. Like stimulation with chemoattractants, adherence is associated with a twofold rise in F-actin content. However unlike the rapid rise in F-actin induced by chemoattractants which peaks within 30 s, actin assembly induced by adherence is slower, maximum F-actin values not being observed until 10 min. Furthermore the rise in F-actin induced by adherence is persistent, remaining constant over 60 min while F-actin returns to near basal levels after 20 min exposure to chemoattractant. The combination of adherence (5 min) followed by chemoattractant (FMLP 5 x 10(-8) M for 40 s) resulted in an additive rise in F-actin content to greater than threefold over unstimulated values. Unlike chemoattractant induced actin assembly, adherence-associated PMN actin polymerization was not inhibited by pertussis toxin, but was markedly reduced by lowering extracellular Ca2+. Fluorescent micrographs of adherent PMN stained with nitrobenzoxadiazole-phallacidin revealed F-actin in the lamellipodia and in small foci on the adherent surface. These findings suggest that the transduction mechanisms by which adherence induces PMN actin polymerization differ from those used by chemoattractant receptors.

Vinculin proteolysis unmasks an ActA homolog for actin-based Shigella motility.

To generate the forces needed for motility, the plasma membranes of nonmuscle cells adopt an activated state that dynamically reorganizes the actin cytoskeleton. By usurping components from focal contacts and the actin cytoskeleton, the intracellular pathogens Shigella flexneri and Listeria monocytogenes use molecular mimicry to create their own actin-based motors. We raised an antibody (designated FS-1) against the FEFPPPPTDE sequence of Listeria ActA, and this antibody: (a) localized at the trailing end of motile intracellular Shigella, (b) inhibited intracellular locomotion upon microinjection of Shigella-infected cells, and (c) cross-reacted with the proteolytically derived 90-kD human vinculin head fragment that contains the Vinc-1 oligoproline sequence, PDFPPPPPDL. Antibody FS-1 reacted only weakly with full-length vinculin, suggesting that the Vinc-1 sequence in full-length vinculin may be masked by its tail region and that this sequence is unmasked by proteolysis. Immunofluoresence staining with a monoclonal antibody against the head region of vinculin (Vin 11-5) localized to the back of motile bacteria (an identical staining pattern observed with the anti-ActA FS-1 antibody), indicating that motile bacteria attract a form of vinculin containing an unmasked Vinc-1 oligoproline sequence. Microinjection of submicromolar concentrations of a synthetic Vinc-1 peptide arrested Shigella intracellular motility, underscoring the functional importance of this sequence. Western blots revealed that Shigella infection induces vinculin proteolysis in PtK2 cells and generates p90 head fragment over the same 1-3 h time frame when intracellular bacteria move within the host cell cytoplasm. We also discovered that microinjected p90, but not full-length vinculin, accelerates rates of pathogen motility by a factor of 3 +/- 0.4 in Shigella-infected PtK2 cells. These experiments suggest that vinculin p90 is a rate-limiting component in actin-based Shigella motility, and that supplementing cells with p90 stimulates rocket tail growth. Earlier findings demonstrated that vinculin p90 binds to IcsA (Suzuki, T.A., S. Saga, and C. Sasakawa. 1996. J. Biol. Chem. 271:21878-21885) and to vasodilator-stimulated phosphoprotein (VASP) (Brindle, N.P.J., M. R. Hold, J.E. Davies, C.J. Price, and D.R. Critchley. 1996. Biochem. J. 318:753-757). We now offer a working model in which proteolysis unmasks vinculin's ActA-like oligoproline sequence. Unmasking of this site serves as a molecular switch that initiates assembly of an actin-based motility complex containing VASP and profilin.

An analogue of the erythroid membrane skeletal protein 4.1 in nonerythroid cells.

Protein 4.1 is a crucial component of the erythrocyte membrane skeleton. Responsible for the amplification of the spectrin-actin interaction, its presence is required for the maintenance of erythrocyte integrity. We have demonstrated a 4.1-like protein in nonerythroid cells. An antibody was raised to erythrocyte protein 4.1 purified by KCl extraction (Tyler, J. M., W. R. Hargreaves, and D. Branton, 1979, Proc. Natl. Acad. Sci. USA, 76:5192-5196), and used to identify a serologically cross-reactive protein in polymorphonuclear leukocytes, platelets, and lymphoid cells. The cross-reactive protein(s) were localized to various regions of the cells by immunofluorescence microscopy. Quantitative adsorption studies indicated that at least 30-60% of the anti-4.1 antibodies reacted with this protein, demonstrating significant homology between the erythroid and nonerythroid species. A homologous peptide doublet was observed on immunopeptide maps, although there was not complete identity between the two proteins. When compared with erythrocyte protein 4.1, the nonerythroid protein(s) displayed a lower molecular weight--68,000 as compared with 78,000-and did not bind spectrin or the nonerythroid actin-binding protein filamin. There was no detectable cross-reactivity between human acumentin or human tropomyosin-binding protein, which are similarly sized actin-associated proteins, and erythrocyte protein 4.1. The possible origin and significance of 4.1-related protein(s) in nonerythroid cells are discussed.

Neutrophil actin dysfunction is a genetic disorder associated with partial impairment of neutrophil actin assembly in three family members.

A male infant with a severe neutrophil motility disorder and poorly polymerizable actin in PMN extracts was reported over a decade ago to have neutrophil actin dysfunction (NAD) (1974. N. Engl. J. Med. 291:1093-1099). Polymerized actin (F-actin) content of fixed and permeabilized intact neutrophils from the father, mother, and sister of the NAD index case have been measured using nitrobenzoxadiazole-phallacidin, a fluorescent compound which binds specifically to actin filaments. F-actin content of unstimulated PMN from all three family members was significantly lower than unstimulated control PMN (mean 23.6 +/- 0.4 SEM fluorescent units vs. 32.6 +/- 0.6 for controls). After stimulation with the chemotactic peptide FMLP, maximal F-actin content of NAD family member PMN was below that of controls (52.7 +/- 1.3 vs. 72.6 +/- 1.8). F-actin content of detergent insoluble cytoskeletons after stimulation with FMLP was also significantly lower in PMN from NAD family members as compared with controls (21 +/- 6% vs. 73 +/- 8%). PMN extracts from the father and mother, when treated with 0.6 M KCl, polymerized half as much actin as controls. Whereas diisopropylfluorophosphate treatment of normal PMN decreased actin polymerizability in cell extracts, this treatment increased the assembly of actin in parental PMN extract. Addition of purified actin to NAD extracts failed to reveal an abnormal actin polymerization inhibitory activity, and no obvious structural defect in actin purified from the father's PMNs was noted by HPLC and two dimensional thin layer chromatography of tryptic digests. The present studies of actin assembly in intact PMNs confirm that NAD is associated with a true defect in PMN actin assembly and is a genetic disorder that is recessively inherited.

Septic thrombosis of the dural venous sinuses.

From 1940 to 1984, 19 cases of septic dural-sinus thrombosis have been diagnosed at the Massachusetts General Hospital, and some 136 cases have been reported from other institutions. Septic thrombosis most frequently involves the cavernous sinuses (96 cases). Facial or sphenoid air sinus infection often precede cavernous-sinus disease. In addition to the classical signs of proptosis, chemosis, and oculomotor paralysis, isolated sixth-nerve palsy and hypo- or hyperesthesia of the fifth nerve may be found. The major pathogens associated with cavernous-sinus infection include Staphylococcus aureus, other gram-positive organisms, and anaerobes. Septic lateral-sinus thrombosis (64 cases) is almost exclusively a complication of otitis media and/or mastoid infection. Organisms causing this infection include Proteus species, Escherichia coli, S. aureus, and anaerobes. Septic thrombosis of the superior sagittal sinus (23 cases) most frequently accompanies bacterial meningitis or air sinus infection. Causative organisms include Streptococcus pneumoniae, S. aureus, other streptococci, and Klebsiella species. Because septic dural-sinus thrombosis is rare, this disease is frequently misdiagnosed. Evaluation should include lumbar puncture, air sinus films, and computed tomographic scan with contrast. Other helpful diagnostic tests may include carotid angiography, and dynamic brain scan. Orbital venography is the most definitive study in cases of chronic cavernous-sinus thrombosis. Therapy should include intravenous antibiotics and early surgical drainage of purulent exudate in the air sinuses or mastoid regions. Retrospective analysis suggests that treatment with heparin may reduce mortality in carefully selected cases of septic cavernous-sinus thrombosis. Anticoagulation is not recommended in other forms of septic dural-sinus thrombosis. Mortality in the antibiotic-era remains high, particularly in patients with septic thrombosis of the cavernous (30%) and superior sagittal (78%) sinuses.

Cardiac risk factors and complications in non-cardiac surgery.

In an attempt to assess cardiac risk in non-cardiac surgery, 1001 patients over 40 years of age who underwent major operative procedures were examined preoperatively, observed through surgery, studied with at least one postoperative electrocardiogram, and followed until hospital discharge or death. Documented postoperative myocardial infarction occurred in only 18 patients; though most of these patients had some pre-existing heart disease, there were few preoperative factors which were statistically correlated with postoperative infarction. Postoperative pulmonary edema was strongly correlated with preoperative heart failure, but 21 of the 36 patients who developed pulmonary edema did not have any prior history of heart failure. Nearly all of these 21 patients were elderly, had abnormal preoperative electrocardiograms, and had intraabdominal or intrathoracic surgery. In the absence of an acute infarction, bifascicular conduction defects, with or without PR interval prolongation, never progressed to complete heart block. Spinal anesthesia protected against postoperative heart failure but not against other cardiac complication. By multivariate regression analysis, postoperative cardiac death was significantly correlated with (a) myocardial infarction in the previous 6 months; (b) third heart sound or jugular venous distention immediately preoperatively; (c) more than five premature ventricular contractions per minute documented at any time preoperatively; (d) rhythm other than sinus, or premature atrial contractions on preoperative electrocardiogram; (e) age over 70 years; (f) significant valvular aortic stenosis; (g) emergency operation; (h) a 33% or greater fall in systolic blood pressure for more than 10 minutes intraoperatively. Notably unimportant factors included smoking, glucose intolerance, hyperlipidemia, hypertension, peripheral atherosclerotic vascular disease, angina, and distant myocardial infarction.

Hemophilus influenzae in hospitalized adults: current perspectives.

In an eight year period 16 cases of serious extrapulmonary Hemophilus influenzae infection in adults were identified, including cases of meningitis, pericarditis, epiglottitis, empyema, cellulitis, osteomyelitis, endometritis, urinary tract infection, orbital cellulitis, primary peritonitis, mesenteric lymphadenitis and aortic graft infection. An 18 month prospective study of H. influenzae infection in hospitalized adults identified 10 cases of bronchitis, 25 of pneumonia and 65 of respiratory tract colonization, but there were no extrapulmonary infections. In 29 percent of the respiratory tract infections, H. influenzae appeared to be a nosocomial pathogen; in 71 percent, the infection was mixed. Finally, 110 clinical isolates of H. influenzae were studied for antimicrobial susceptibility. Eight percent were ampicillin resistant, two strains were resistant to tetracycline and one to chloramphenicol, but all were susceptible to trimethoprim-sulfamethoxazole and cefamandole.

Chemotherapy of experimental streptococcal endocarditis. 3. Failure of a bacteriostatic agent (tetracycline) in prophylaxis.

Bacteriostatic agents are frequently recommended as alternatives to penicillin for prophylaxis of bacterial endocarditis. To test the efficacy of this group of antimicrobials, prophylaxis of experimental streptococcal endocarditis was attempted with tetracycline. The number of streptococci colonizing the aortic valves of rabbits was not affected by inhibitory levels of tetracycline, but multiplication was checked. Streptococcis urvived in vegetations for seven days despite the continuous presence of tetracycline, and multiplied when the drug was withdrawn. It is therefore suggested that bacteriostatic agents may be valueless for prophylaxis of bacterial endocarditis.

Who was caring for Mary? 1993.

An academic physician gives a personal account of his wife's progressive deterioration during her hospitalization at a university medical center. He encounters physicians who are too distracted by other academic pursuits to care adequately for his wife. Her hospital course is complicated by bilateral pulmonary emboli which occur during inadequate heparin therapy and are followed by myocardial infarction, shock, and the adult respiratory distress syndrome. His experience exemplifies the importance of closely supervised care. He calls on the leaders of academic medical centers to make excellence in patient care a top priority and recommends that clinical as well as research skills be rewarded.

Sphenoid sinusitis.

We review our clinical experience with infectious sphenoid sinusitis in an effort to characterize the clinical presentation, bacteriology and associated severe complications of this frequently misdiagnosed infection.