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Background and objectives: SARS-C0V-2 infections have varied manifestations among individuals ranging from asymptomatic or mild symptoms to severe disease and death. This study is done to look into various histopathological changes in lung, liver, and kidney tissues among Covid19 positive autopsies with cellular tropism and viral load among various organs by immunohistochemistry (IHC) for the SARS-C0V-2 viral marker. Method(s): A prospective descriptive study of core biopsies from covid19 positive autopsies from the lung, liver, and kidneys were taken from 20 cases. A routine histopathological examination of the tissues with IHC staining for SARS-CoV-2 cocktail antibodies was performed and assessed. Result(s): Histopathological changes in the lung, liver, and kidney tissues showed changes of varying severity. On IHC, in the lung, the tropism for SARS-CoV-2 was seen in pneumocytes, bronchial epithelial cells, endothelial cells, and macrophages. In the kidney, tropism was seen towards tubular epithelial cells and endothelial cells. In the liver, hepatocytes and bile duct epithelial cells were positive. Variable viral density was seen in different organs which varied from case to case. The density of the viral load was highest in the lung and lower in the kidney and least in the liver. Conclusion(s): In this study the various histopathological changes and cellular tropism of the SARS-CoV-2 among Lung, liver, and kidney tissues have been described and compared with various similar studies across the globe.Copyright © 2023 International Journal of Life Sciences Biotechnology and Pharma Research. All rights reserved.
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As hyperbaric oxygen (HBO) has been shown to mitigate the COVID-19 symptoms, we were interested in studying whether HBO exposure affects expression of viral entry genes and innate immune genes in the air-liquid interface (ALI)-cultured human bronchial epithelial cells (HBECs) derived from normal individuals (NHBECs) and age-matched COPD patients (DHBECs), which were cultured under normoxia or daily exposure of 40-min hyperbaric oxygen (HBO) with 100% O2 at 2.5 ATA for 28 days in total. We found for the first time that HBO exposure differentially regulated mucociliary differentiation of HBECs by respectively decreasing and increasing expression of CGRP, MUC5AC, FOXJ1, NOTCH3 and HEYL in NHBECs and DHBECs, and respectively decreased and increased FOXO1 expression whereas increased and decreased NFE2L2 and NFKB1 expression in NHBECs and DHBECs, in association with respectively decreased and increased expression the SARS-CoV-2 entry genes ACE2 and 2 TMPRSS2 and the tight junction protein genes TJP1 and TJP2, and in association with respectively increased and decreased expression of the cell growth and inflammatory transcription factors SRF, c-FOS and TP63, as well as the TLR pathway genes TLR3, AKT1, IL-1B, IL-5, IL-6, IL-33, IRAK4 and NFKBIA in NHBECs and DHBECs. (Figure Presented).
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Even after virus elimination, coronavirus disease 2019 (COVID-2019) leaves numerous sequelae. Growing evidence demonstrates that massive release of proinflammatory cytokines, which drives COVID-19 progression, severity, and mortality, remains elevated after acute phase of COVID-2019, playing a central role in the disease' sequelae. In this way, bronchial epithelial cells are the first cells hyperactivated by coronavirus-2 (SARS-Cov-2) leading to massive cytokine release, triggering leukocytes and other cells hyperactivation, mediating COVID-19 sequelae. So, proinflammatory cytokines are initiated by the host. Thus, this in vitro study tested the hypothesis that ImmuneRecovTM, a protein blend, could inhibit the hyperactivation of human bronchial epithelial cells (BEAS-2B) induced by SARS-Cov-2. BEAS-2B (5x104/mL/well) cells were co-cultivated with 1ml of blood of a SARS-Cov-2 infected patient for 4 hours and protein blend (1ug/mL) was added in the first minute of the co-culture. After 4 hours, the cells were recovered and used for analysis of cytotoxicity by MTT and for mRNA expression of IL-1beta, IL-6, IL-10. The supernatant was used to measure cytokines. SARS-Cov-2 incubation resulted in increased levels of IL-1beta and IL-6 by BEAS-2B cells (p<0.001). Treatment with the protein blend resulted in reduced levels of pro-inflammatory IL1beta and IL-6 (p<0.001), and increased the levels of anti-inflammatory IL-10 (p<0.001). Protein blend reduced SARS Cov-2-increased the mRNA expression of IL-1beta and IL-6, and increased the expression of IL-10 and IFN-gamma. In conclusion, protein blend presents important anti-inflammatory effects in the context of SARS-Cov-2 infection.
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Background: TGF-beta signaling plays an essential role in tissue fibrosis and mediates profibrotic programs after SARS-CoV-2 infection in the kidney and lung. SARS-CoV-2 also induces humoral immune responses controlled by cytokines, including TGF-beta. Studies have found that the incidence of SARS-CoV-2 infection and the severity of Covid-19 in cystic fibrosis (CF) patients is lower than the general population. We studied how SARS-CoV-2 regulates TGF-beta-mediated gene expression in the CF airway. Method(s): Small RNAseq was performed in human bronchial epithelial cells CFBE41o-from a patient homozygous for the F508del mutation in the CFTR gene on Illumina NextSeq 500's. Pathway analysis was done by Ingenuity Pathway Analysis (IPA) software (QIAGEN) and miRNet browser. IPA was used for analyzing coronavirus associated pathways affected by differentially regulated miRNAs. miRNAs predicted to target the coronavirus associated genes were collected from TargetScan Human release 7.2, miRmap, Diana-TarBase v.8, and miRBase bioinformatics tools. Anti-miRNA oligonucleotide miRCURY LNATM Power Inhibitors or control (Exiqon) were used. Cells expressing F508del or wild type CFTR were used to compare the results in CF and non-CF models. Findings were validated in primary human bronchial epithelial (HBE) cells. Result(s): Compared to vehicle control, TGF-beta1 dysregulated 48 miRNAs;38 and 19 pathways were uniquely affected by the upregulated and downregulated miRNAs, respectively. We found 43 miRNAs targeting 119 different mRNA of the proteins associated with coronavirus pathogenesis pathway and 21 miRNAs targeting 21 different mRNA of the proteins associated with coronavirus replication pathway. Two miRNAs upregulated by TGF-beta1 target the host receptor for SARS-CoV-2 invasion, angiotensin converting enzyme 2 (ACE2). We confirmed the results by qRT-PCR that TGF-beta1 increased expression of specific miRNAs targeting ACE2 mRNA. Upregulation of the miRNAs was followed by inhibition of ACE2 mRNA and protein levels and the effect was blocked by specific anti-miRNA oligonucleotides. The above results differed between the CF and non-CF cells. Conclusion(s): miRNAs may be important effectors of TGF-beta modulating SARSCoV-2 pathogenicity and replication in the CF airway. Ongoing studies focus on elucidating the mechanisms of SARS-CoV-2 invasion of kidney cells.
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Background: People with cystic fibrosis (PwCF) have chronic, pronounced respiratory damage and have been considered among those at highest risk for serious harm from SARS-CoV-2. Numerous clinical studies have reported that individuals with CF in North America and Europe, although highly susceptible to COVID-19, do not have mortality levels that exceed those of the general population. Method(s): To understand features that might influence lethality of COVID- 19 in PwCF, we tested potential relationships between CFTR and viral pathogenesis. As one approach to evaluate impact of CF transmembrane conductance regulator (CFTR) on COVID-19 severity, independent sets of blood samples fromvirally infected individualswere genotyped. Bloodwas obtained from 424 U.S. patients hospitalized with severe COVID-19 and a much larger European cohort of 7147 healthy individuals and 2587 individuals with severe COVID-19. Deoxyribonucleic acid in both studies was probed for the F508del variant. In other experiments, we investigated the possibility that lack of CFTR might alter viral binding and propagation. We used human bronchial epithelial cell (HBEC) monolayers from individuals without functional CFTR for this purpose. Finally, we examined effects of CF airway secretions and features such as viscosity, pH, and protease/anti-protease imbalance during SARS-CoV-2 infection. Result(s): We found no evidence of a relationship between deficient CFTR function (based on carrier status for the severe F508del defect) and clinical outcomes from COVID-19. In addition, viral propagation studies using airway epithelial monolayers (a model that reproduces many aspects of in vivo tissue biology) were not influenced by homozygous absence of CFTR. We show that levels of angiotensin converting enzyme-2 receptor messenger ribonucleic acid (mRNA) appear normal in CF primary epithelium, whereas transmembrane serine protease 2 mRNA is variable but lower ( p < 0.001) in a manner that correlates with viral infectivity (R2 = 0.76). Dependence of viral proliferation on features of CF mucosal fluid-including pH (viral replication optimum at pH 7-7.5), viscosity (diminished propagation in highly viscous apical media), and protease/ anti-protease imbalancewere identified as likely contributors to efficiency of SARS-CoV-2 replication and pathogenesis. Conclusion(s): These findings using patient data, CF and non-CF primary airway epithelia, and CF airway secretions fail to demonstrate a causal relationship between loss of CFTR and susceptibility to severe COVID-19. Notwithstanding the caveat that addition of virus in small buffer volumes disrupts airway surface liquid depth and composition, our findings also argue against a role for CFTR during acute infection of airway cells in vitro. On the other hand, chronic disruption of periciliary liquid, diminished pH, altered protease/anti-protease homeostasis, and increased fluid viscosity (sequelae that occur in CF lungs) were implicated as contributors to impaired SARS-CoV-2 propagation. Such studies provide a basis for future work to test relationships between CFTR and severity of COVID-19. Copyright © 2022, European Cystic Fibrosis Society. All rights reserved
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Purpose/Objectives: <Most used lower respiratory tract models consist of cell monolayers which lack of tissue and organ level response and of in-vivo phenotype. Ex-vivo lung tissues have short viability and limited availability. Lung organoids, which recapitulates better the 3D cellular complex structures, architecture, and in-vivo function, fail to reach maturity even after 85 -185 days of culture. Therefore, these models have a limited use to study fetal lung diseases. Other lung models, consist of only one structure of the lower track, such as bronchial tubes or alveoli, but fail to recapitulate the whole organ structure. In this work, cell microenvironment was used to promote the self-organization of epithelial and mesenchymal cells into macro-structures, aiming to mimic the whole and adult lower respiratory tract model> Methodology: <Different parts of the microenvironment were considered to create a compliant matrix. Alginate-Gelatin hydrogels were used for 3D encapsulation of mesenchymal origin cells. This hydrogel provided a stiffness like the one on the lung. Base membrane zone proteins were used to induce the attachment and guidance of epithelial cells into 3D structures. The interactions between both cell types, further guided them into lung fate. The morphology of resulting organoids was analyzed using immunostaining and confocal microscopy, LSM710, with the purpose of evaluate polarization, protein markers, and different cell populations. Quantitative PCR was performed to evaluate and compare the expression of lung fate genes with traditional cell monocultures.> Results: <The engineered microenvironment and protocol development done in this work resulted in macro-scale structures, in which branching morphogenesis occurred at day 21. Different structures were identified in the organoid including bronchial tube, bronchioles, and alveoli. Polarization of the organoids was confirmed by visualization of E-cadherin, and ZO-1. Expression of Surfactant Protein B and C into the organoids confirmed the presence of alveolar type II cells, which are only present in the later development stage. Surfactant Protein B, Transmembrane protease, serine 2, TMPRSS-2, and Angiotensin-converting enzyme 2, ACE2 were found to be significantly higher expressed into the organoids in comparison with traditional epithelial cells monolayers.> Conclusion/Significance: <Growth factors are normally used to induce the fate of stem cells into lung organoids;however, these fail to reach maturity. Here, we developed a new methodology to induce the formation of the organoids based on the cell microenvironment. The resulting organoids require less time for development. The initial stage of adult cells can be modulated through culture conditions induce a 3D structure like the adult lung. As such, these organoids have the potential to be used for modeling adult diseases and to develop specific models from patient cells, which is one step forward to personalized medicine. SFTPB is one of the main proteins which facilitates the breathing process. Its high expression into our model may indicate that breathing occurs into our lung organoids. The higher expression of TMPRSS-2 and ACE2 into the organoids has a major significance in the field of virology since both proteins are the mainly entrance of SARS-CoV-2, and influenza H1N1.>.
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SESSION TITLE: Drug-Induced and Associated Critical Care Cases Posters 1 SESSION TYPE: Case Report Posters PRESENTED ON: 10/19/2022 12:45 pm - 01:45 pm INTRODUCTION: Interstitial pneumonitis (ILD) is inflammation of lung interstitium leading to scarring and pulmonary fibrosis. Various etiologies include idiopathic, connective tissue disorders, sarcoidosis and drug induced1. Many chemotherapy agents have been implicated in drug related ILD such as bleomycin, taxanes. However, newer chemotherapeutic drugs such as molecular agents such as anti-VEGF, anti-EGFR (panitumumab) could be causative of drug induced ILD. CASE PRESENTATION: A 75-year-old female with stage IV sigmoid colon cancer treated with surgery, adjuvant FOLFOX chemotherapy and Panitumumab. She presented to the emergency department with shortness of breath and hypoxia after known COVID-19 exposure. Initial imaging with chest radiography showed bilateral ground glass opacities. A chest CT pulmonary embolism protocol was negative for pulmonary embolism but showed bilateral ground glass opacities (GGOs) and some interstitial thickening (L>R) not typical of COVID-19 infection. She was treated with remdesivir and dexamethasone, however her oxygen requirements continued to rapidly escalate. A repeat CT chest without contrast showed bilateral asymmetric interstitial thickening and GGOs. Given persistence of CT chest abnormalities, workup for interstitial lung disease was initiated. The results include ANA titer 1:80, otherwise negative ANCA profile, rheumatoid factor, anti-CCP, Scl-70, Sjogren antibodies. Given clinical history and imaging findings, diagnosis of ILD was suspected, and she was started on solumedrol 1 mg/kg. Her oxygen requirements decreased significantly over the next 2 days, and she was discharged home on oral steroid taper and pneumocystis pneumonia prophylaxis. DISCUSSION: Panitumumab is a fully humanized monoclonal antibody against EGFR. Approved by the US Food and Drug Administration in 2006 for advanced or recurrent colorectal cancer exhibiting wild-type KRAS mutation.2 ILD is rarely reported with panitumumab monotherapy, but higher incidence when used as a combination treatment such as with FOLFOX or FOLFIRI. A Japanese post-marketing surveillance study from 2010-2015 showed an ILD incidence of 1.3% but mortality rates of 51.3%.2 EGFR is expressed on basal cells and non-cilia cells of the bronchioles and type II cells of the alveolus. EGFR mediated mechanisms are important in tissue repair.3 Therefore inhibition of this pathway has been postulated to play a role in development of ILD. Another mechanism was decreased surfactant production by type II cells in pre-clinical study.4,5 ILD secondary to Panitumumab can occur at any point during therapy and up to 1 year after administration of drug.6 The role of infectious processes, in this case, COVID-19 pneumonia, could synergistically worsen ILD presentation. CONCLUSIONS: Although the incidence of ILD is low, the mortality rate is high, therefore early recognition and treatment is associated with improved clinical outcomes. Reference #1: Mudawi D, Heyes K, Hastings R, Rivera-Ortega P, Chaudhuri N. An update on interstitial lung disease. Br J Hosp Med (Lond). Jul 2 2021;82(7):1-14. Reference #2: Osawa M, Kudoh S, Sakai F, et al. Clinical features and risk factors of panitumumab-induced interstitial lung disease: a postmarketing all-case surveillance study. Int J Clin Oncol. Dec 2015;20(6):1063-1071. Reference #3: The FASEB Journal - 2000 - Puddicombe - Involvement of the epidermal growth factor receptor in epithelial repair in asthma.pdf. DISCLOSURES: No relevant relationships by Navitha Ramesh No relevant relationships by Uba Udeh
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SESSION TITLE: Pulmonary Manifestations of Systemic Disease Case Posters SESSION TYPE: Case Report Posters PRESENTED ON: 10/19/2022 12:45 pm - 01:45 pm INTRODUCTION: Organizing pneumonia (OP) is a form of interstitial lung disease with a distinct histopathological pattern where bronchioles and alveoli become inflamed. It is associated with many clinical conditions including infections and connective tissue disease. OP has also been seen in patients with hematologic malignancies, however, primary pulmonary presentation of chronic lymphocytic leukemia (CLL) is uncommon. We present a rare case of OP as an initial presentation of CLL. CASE PRESENTATION: A 62 year-old male with a sixty pack year smoking history and COVID-19 infection one month ago presents with complaints of worsening dyspnea, headaches, productive cough, and congestion of 10 days duration. Patient is unvaccinated and did not require hospitalization for his COVID-19 infection. His vital signs on admission were significant for tachypnea with respiratory rate of 35 and hypoxia with oxygen saturation of 84% on room air. He initially required oxygen supplementation via non-rebreather mask to maintain oxygen saturation >88%. A chest tomography (CT) scan was completed and revealed bilateral dense consolidations with ground-glass opacities and air bronchograms consistent with OP. The scan was also significant for bulky mediastinal lymphadenopathy. The patient denied any personal or family history of autoimmune disease, occupational exposures, and recent travel. Evaluation for infection and for underlying connective tissue disease was unremarkable. He was started on broad spectrum antibiotics and high dose steroids. Due to fluctuating lymphocytosis, bulky lymphadenopathy, and negative infectious workup despite clinical improvement, he underwent a bronchoscopy with endobronchial ultrasound guided transbronchial needle aspiration of lymph nodes. Immunohistochemical (IHC) stains of these samples were compatible with CLL. Additionally, peripheral blood flow cytometry was also diagnostic of CLL. Oncology was consulted for further evaluation and treatment of CLL. The patient's respiratory symptoms improved and oxygen requirements decreased with steroid treatment and he was discharged home. DISCUSSION: OP occurring in patients with hematologic malignancies has multiple etiologies. Most case reports describe patients with previous exposure to chemotherapy, radiotherapy, or bone marrow transplant. However, our patient had no such exposure history and no prior diagnosis of a hematologic malignancy. Infectious and autoimmune etiology were considered, but serologic evaluation was unremarkable. Flow cytometric analysis of lymph node tissue along with lymphocytic bronchoalveolar lavage was consistent with initial diagnosis of CLL. CONCLUSIONS: Despite the low incidence, hematologic malignancy should be considered as a differential diagnosis in all patients who present with organizing pneumonia. Prednisone therapy for 6-12 month duration has been shown to reduce respiratory symptoms and may improve survival. Reference #1: Craig E. Daniels, Jeffrey L. Myers, James P. Utz, Svetomir N. Markovic, Jay H. Ryu. Organizing pneumonia in patients with hematologic malignancies: A steroid-responsive lesion. Respiratory Medicine, 101 (1) (2007), pp. 162-168. Reference #2: M. Mokhtari, P.B. Bach, P.A. Tietjen, D.E. Stover. Bronchiolitis obliterans organizing pneumonia in cancer: a case series. Respiratory Medicine, 96 (4) (2002), pp. 280-286. DISCLOSURES: no disclosure on file for Guillermo Garrido;No relevant relationships by Anita Gopalakrishnan No relevant relationships by Rameez Rao No relevant relationships by Mohammad Salimian
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SESSION TITLE: Critical Diffuse Lung Disease Cases 2 SESSION TYPE: Case Report Posters PRESENTED ON: 10/19/2022 12:45 pm - 01:45 pm INTRODUCTION: Recurrent episodes of community acquired pneumonia (CAP) have been shown to be common in elderly patients. Cryptogenic organizing pneumonia (COP) is an interstitial lung disease that is often mistaken for pneumonia, especially in the older population. Here, we present a 100-year-old woman diagnosed with COP after multiple failed courses of antibiotics for CAP. CASE PRESENTATION: A 100-year-old female with a history of cardiomyopathy, pulmonary hypertension, and autoimmune hemolytic anemia previously on prednisone, who presented with shortness of breath and non-productive cough. CT of the chest showed dense left upper and lower lobe consolidations. She was admitted 2 months prior with similar symptoms and found to have extensive right sided consolidations with concerns of CAP. She was treated with antibiotics without resolution of her symptoms. CXR from two years prior revealed right upper and right lower lobe consolidations. This admission, she was started on antibiotics with no improvement and required supplemental oxygen. She had no leukocytosis. COVID-19 testing was negative and she was unable to produce any sputum for culture. The patient declined bronchoscopy. She was seen by speech and swallow with no concern for aspiration. Prednisone was started empirically for COP and the patient experienced rapid improvement in symptoms and oxygenation. Ultimately, she was discharged on 20 mg of prednisone daily as well as Bactrim for PCP prophylaxis. She continued a slow taper as an outpatient with overall improvement in her clinical symptoms. Serial CT scans demonstrate complete resolution of the infiltrates with no recurrence or new infiltrates. DISCUSSION: Cryptogenic organizing pneumonia is a rare interstitial lung disease known to affect bronchioles and alveoli. Its etiology is unclear and symptoms often mimic other types of infectious pneumonia leading to frequent mis-diagnosis. The average age of onset is typically 50-60. Establishing this diagnosis can be difficult due to the non-specific symptomatology of dry cough and dyspnea. Bronchoscopy with lavage and transbronchial biopsies can be performed to rule out infectious and non-infectious etiologies but is not necessary for diagnosis. The most common radiographic pattern is multifocal asymmetrical parenchymal consolidations with air bronchograms that tend to migrate and appear in different sites over time. Less common presentations include ground glass opacities, nodular densities, and progressive fibrotic patterns. Steroids with a slow taper as outpatient are mainstay of therapy and the majority of patients respond with symptom and radiographic improvement. CONCLUSIONS: While elderly patients are particularly susceptible to recurrent CAP, the diagnosis of COP should be considered part of the differential diagnosis in those with recurrent unexplained consolidations on chest radiography without an infectious etiology. Reference #1: Hedlund J, Kalin M, Ortqvist A. Recurrence of pneumonia in middle-aged and elderly adults after hospital-treated pneumonia: aetiology and predisposing conditions. Scand J Infect Dis. 1997;29(4):387-92. doi: 10.3109/00365549709011836. PMID: 9360255. Reference #2: Tiralongo F, Palermo M, Distefano G, et al. Cryptogenic Organizing Pneumonia: Evolution of Morphological Patterns Assessed by HRCT. Diagnostics (Basel). 2020;10(5):262. Published 2020 Apr 29. doi:10.3390/diagnostics10050262 Reference #3: Lee JW, Lee KS, Lee HY, Chung MP, Yi CA, Kim TS, Chung MJ. Cryptogenic organizing pneumonia: serial high-resolution CT findings in 22 patients. AJR Am J Roentgenol. 2010 Oct;195(4):916-22. doi: 10.2214/AJR.09.3940. PMID: 20858818. DISCLOSURES: No relevant relationships by Vincent Chan No relevant relationships by Mackenzie Kramer No relevant relationships by John Madara No relevant relationships by Stephanie Tzarnas No relevant relationships by Laura Walters
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SESSION TITLE: Post-COVID-19 Infection Complications SESSION TYPE: Case Report Posters PRESENTED ON: 10/17/2022 12:15 pm - 01:15 pm INTRODUCTION: COVID-19 has affected over 200 million people worldwide. Clinicians continue to observe unusual manifestations of this disease. In an attempt to improve our understanding of COVID-19 pneumonia, we present the details of one patient who developed large bilateral pulmonary cysts. CASE PRESENTATION: A 40-year-old woman with no known medical problems presented with the chief complaint of fever, nausea, vomiting, generalized weakness followed by difficulty breathing that developed over a few days. Her vital signs on admission included temperature 98.4° F, heart rate 104 beats/minute, respiratory rate 48 breaths/minute, O2 saturation 88 percent on 15 liters of oxygen through a non-rebreather mask, and Body Mass Index 42 kg/m2. The patient tested positive for COVID-19. Computed tomography (CT) of the chest to rule out a pulmonary embolism showed bilateral extensive ground-glass opacities and reticular and nodular opacities. She was intubated for acute hypoxic respiratory failure. Twenty days into the hospital admission, she was noted to have a bulla in the right lower lobe. A repeat CT chest on day 45 revealed an increase in the number and size of cysts bilaterally. Patient was discharged to rehab and later readmitted for worsening respiratory status. This time she tested positive for human metapneumovirus. A CT chest showed increase in the size of the right sided lung cysts;the left sided lung cysts had resolved. DISCUSSION: The first COVID-19 related pulmonary cystic lesions were reported in May 2020(1). Since then, several reports have now established a relationship between an infection and cyst formation. The most common distribution is peripheral in the lower lobes. The pathogenesis remains uncertain, but several mechanisms have been proposed. Microthrombi in the pulmonary circulation could lead to ischemia and subsequent remodeling of interstitial matrix and bronchial obstruction with distal hyperinflation due to check valve mechanism. (1,2). Hamad et al. propose that pneumatoceles are formed by air leaked in to the interstitium which causes stripping and separation of a thin layer of lung parenchyma with further injury to the small blood vessels and bronchioles. The rate of barotrauma in non-COVID-19 related ARDS is 0.5%;the rate in COVID-19 ARDS is 15% (3). This suggests a close relation between COVID-19 pneumonia and subsequent development of pulmonary cysts. Our patient had no preexisting pulmonary disease and was noted to have pulmonary cysts after being on mechanical ventilation for almost 2 weeks. The patient later contracted the human metapneumovirus infection and CT showed that the right-sided lung cysts had become bigger in size. However, the left-sided cysts which had a maximum diameter of 4.8 cm had resolved. CONCLUSIONS: We need to follow patients with COVID 19 induced lung cysts clinically and radiologically to understand the clinical course and best management strategies. Reference #1: Kefu Liu et al. COVID 19 with cystic features on Computed tomography;Medicine (Baltimore) 2020May;99(18): e20175. PMCID: PMC7486878 Reference #2: Galindo J, Jimenez L, Lutz J et al. Spontaneous pneumothorax with or without pulmonary cysts, in patients with COVID 19 Pneumonia. Journal of infections in developing countries 2021;15(10);1404-1407 Reference #3: McGuinness G, Zhan C, Rosenberg N, Azour L, Wickstrom M, Mason DM, Thomas KM, Moore WH. Increased incidence of barotrauma in patients with COVID-19 on invasive mechanical ventilation. Radiology. 2020;297(2): E252–E262. doi: 10.1148/radiol.2020202352 DISCLOSURES: No relevant relationships by Arunee Motes No relevant relationships by Kenneth Nugent No relevant relationships by Tushi Singh No relevant relationships by Myrian Vinan Vega
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Background: Acute respiratory infection (ARI) is the leading infectious cause of pediatric death worldwide, comprising 15% of all deaths in children under 5 years old. Human metapneumovirus (HMPV) is a primary cause of ARI, and accounts for a major portion of ARI-related hospitalizations in infants and young children. Although nearly every person is infected with HMPV during early childhood, re-infections occur often, highlighting the difficulty in building long-term immunity. There are no approved vaccines or antiviral therapies. Early host responses to HMPV are poorly characterized, and further understanding could identify important antiviral pathways and potential therapeutic targets. Type I (IFN-α/β) and III interferons (IFN-λ) display antiviral activity against numerous respiratory viruses and are currently being investigated for therapeutic use in several respiratory infections including SARS-CoV-2. However, their roles in HMPV infection remain largely unknown. Our laboratory has previously shown that type I IFN is critical for HMPV pathogenesis, as loss of IFN-α/β signaling reduces lung inflammation and lessens HMPV disease severity in mice. Here, we describe distinct antiviral roles for type I and III IFNs during HMPV infection using an established mouse model. Methods: In vivo studies were conducted using mice lacking either the IFN-α/ β receptor (IFNAR-/-) or IFN-λ receptor (IFNLR-/-). Early immune responses to HMPV strains TN/94-49 and C2-202 were assessed by clinical disease scoring, plaque assay, Luminex immunoassay, and spectral cytometry of mouse lung samples. In vitro studies were performed using CMT 64-61 mouse bronchial epithelial cells. Responses to TN/94-49 and C2-202 were measured by qPCR, plaque assay, and Luminex immunoassay of cell lysates and supernatants. Results: IFNAR-/- mice exhibited lower clinical disease scores, reduced lung levels of inflammatory cytokines IL6, MIP-1α, and MCP-1, and decreased numbers of lung interstitial macrophages during HMPV infection, highlighting their critical role in HMPV immune-mediated pathogenesis. IFNLR-/- mice with intact IFNAR showed moderate clinical disease, higher lung levels of inflammatory cytokines IL-6, MCP-1, and IFN-γ, and increased lung interstitial macrophage recruitment. A reduction in HMPV disease was also recapitulated by IFNAR-neutralizing antibody treatment of IFNLR-/- mice. Interestingly, IFNLR-/- showed higher HMPV viral titers, while IFNAR-/- mice showed no differences or slightly lower viral titers, compared to wild-type mice. Moreover, IFN-λ pre-treatment of infected CMT 64-61 cells reduced HMPV viral titers and decreased supernatant levels of inflammatory cytokines IL-6, IL-1β, TNFα, and MCP-1. Conclusion: These findings suggest that type I IFN is necessary for HMPV pathogenesis, while type III IFN is critical for limiting HMPV replication in the lungs but does not contribute to HMPV inflammatory disease. This work uncovers key functional differences between type I and III IFNs during HMPV infection, an important feature of innate immune responses to HMPV that may be utilized to inform treatment.
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RATIONALE: Treatments for the coronavirus disease of 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), are urgently needed but remain limited. SARS-CoV-2 infects cells through the interactions of its spike (S) protein with ACE2 and TMPRSS2 on host cells. Multiple cells and organs are targeted, particularly airway epithelial cells. OM-85, a standardized lysate of human airway bacteria with strong immunomodulating properties and an impeccable safety profile, is widely used to prevent recurrent respiratory infections. Our finding that the airway administration of OM-85 inhibits Ace2 and Tmprss2 transcription in mouse lungs prompted us to investigate whether and how OM-85 may protect non-human primate and human epithelial cells against SARS-CoV-2 infection. METHODS: ACE2 and TMPRSS2 mRNA and protein expression, cell binding of SARS-CoV-2 S1 protein, cell entry of SARS-CoV-2 S protein-pseudotyped lentiviral particles, and SARS-CoV-2 cell infection were measured in kidney, lung and intestinal epithelial cell lines, primary human bronchial epithelial cells, and ACE2- transfected HEK293T cells treated with OM-85 in vitro. RESULTS: OM-85 significantly downregulated ACE2 and TMPRSS2 mRNA in epithelial cell lines and primary bronchial epithelial cells, and strongly inhibited SARS-CoV-2 S protein binding to, SARS-CoV-2 S proteinpseudotyped lentivirus entry into, and SARS-CoV-2 infection of epithelial cells. These effects of OM-85 appeared to depend on the downregulation of SARS-CoV-2 receptor expression. CONCLUSIONS: OM-85 inhibits SARS-CoV-2 epithelial cell infection in vitro by downregulating SARS-CoV-2 receptor expression. Further studies are warranted to assess whether OM-85 may prevent and/or reduce the severity of COVID-19.
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Rationale: Cell-penetrating peptides are able to cross membranes and deliver cargoes in a functional form. Our prior work identified a 12-amino acid, cardiac targeting peptide (APWHLSSQYSRT). Studies into its mechanism of transduction led to the identification of two lung targeting peptides (LTPs), S7A and R11A. Here we report on a) the comparative lung uptake of S7A versus R11A, b) complete biodistribution of R11A, c) show that cyclic versions are -100-fold more efficient than linear counterparts, d) uptake is via a non-endocytic pathway, and e) cyclic R11A's (cR11A) ability to deliver siRNA targeting structural proteins of SARS-CoV-2 and act as an anti-viral. Methods: Linear LTPs were synthesized with N-terminal labeled with Cyanine 5.5 (Cy5.5). Cyclic versions were synthesized with lysine added to the N-terminus, cyclized through a peptide bond, with a side NH-group labeled with Cy5.5. cR11A was conjugated to siRNA duplexes via a DTME linker. Wild-type, CD1 mice, were injected with S7A or R11A at 10, 5, and 1mg/Kg, peptides allowed to circulate for 15mins, mice euthanized, lung along with multiple other organs dissected and imaged using In Vivo Imaging Systems (IVIS, Perkin-Elmer) followed by confocal microscopy. CD1 mice were injected with R11A, 5mg/Kg, and euthanized at different time intervals for biodistribution studies. Endocytosis studies were done using serum-starved human bronchial epithelial cells (HBEC) incubated with fluorescently labeled transferrin and LTP-S7A or LTP- R11A. Lastly, anti-viral activity was tested in HBECs pre-treated with cR11A-siRNA followed by viral infection. Results: Mice injected with LTP-S7A or LTP-R11A showed robust uptake of the peptides by lung tissue, with R11A showing an increasingly favorable lung:liver ratio with decreasing dose. Lung uptake of R11A peaked at 120mins with complete dissipation of fluorescence by 24 hours. In Vitro studies in HBECs showed no co-localization of transferrin with LTPs, ruling out endocytosis as a mechanism of uptake. Comparison of linear versus cyclic peptides using FACS showed cyclic peptides had -100-fold increased transduction efficiency over their linear counterparts. cR11A conjugated to ant-spike, and anti-envelop proteins showed an anti-viral effect with EC90 of 0.6uM and 1.0μM, respectively. Conclusions: We have identified two novel lung-targeting peptides capable of acting as delivery vectors. Peak uptake of R11A occurred at 120mins. Furthermore, this uptake was not via endocytosis, and cyclic versions were -100-fold more efficiently taken up. Lastly, as proof of concept, we show cR11A acts as a vector and delivers siRNA to HBECs in a functional form, and act as anti-virals.
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Background: A novel coronavirus (SARS-CoV-2) has led to the worldwide spread of pandemic proportions and currently no effective therapy is available. The minor pulmonary surfactant lipids, palmitoyl-oleoyl-phosphatidylglycerol (POPG), and phosphatidylinositol (PI), are potent regulators of inflammatory processes, and are effective as anti-viral agents for multiple respiratory viruses including Respiratory syncytial virus (RSV), Influenza A virus (IAV) and Rhinoviruses (RVs). Objective: The primary objectives of this study are to determine whether POPG or PI are potent against SARS-CoV-2 in vitro, using human airway epithelial cells, and examine the potency of PI against SARS-CoV-2 in vivo, in a hamster model. Methods: We examined efficacies of POPG or PI against SARS-CoV-2 (USA WA/2020) in human bronchial epithelial cells, and nasal epithelial cells from healthy control subjects differentiated by ALI cultures. We quantified SARS-CoV-2 replication by quantitative plaque assays and qRT-PCR. We determined the potency of PI against SARS-CoV- 2 in golden Syrian hamster as in vivo model for SARS-CoV-2 infection. Results: We examined the efficacies of POPG and PI using primary human tracheal and nasal epithelial cells, differentiated in ALI culture. Cells were treated with POPG (10mg/ml) and PI (4mg/ml) added to apical media alone for 16hrs. Subsequently, cells were infected with SARS-CoV-2 at m.o.i = 0.02, for 48hrs, harvested for RNA extraction and qRT-PCR. SARS-CoV-2 replicated in tracheal cells with a 106-fold increase in mRNA. POPG and PI reduced viral mRNA expression by 70% and 85%, respectively (subject numbers n=3). In nasal epithelia, SARS-CoV-2 mRNA expression increased 105 -fold compared to sham infected cultures. Both POPG and PI attenuated the increase in viral mRNA expression by 70% - 82% (subject numbers n=6). We determined the PI effect in an in vivo study in hamsters. Hamsters were challenged with 103 pfu of SARS-CoV-2, either with, or without PI (2mg/hamster) administered intranasally. Hamsters were harvested at Day 3, and lungs were processed for histopathology. Pharyngeal swabs were used to examine viral burden by plaque assays. PI reduced plaque numbers compared to viral infection alone groups at day1 (Virus alone: 2.4±2.7(X104pfu/ml), Virus+PI: 0.9±2.1(X106pfu/ml), p<0.05). PI reduced lung histopathology score at day 3 (Virus alone: 28.0±15.6, Virus+PI: 6.7±7.0, p<0.05). Conclusions: POPG and PI significantly reduced SARS-CoV2 replication in human differentiated airway epithelial cells. PI inhibited SARS-CoV-2 infection in vivo in hamsters. These findings suggest that inhalation of POPG, or PI might be effective as novel anti-viral compounds for treating and preventing SARSCoV- 2 infection.
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Background: Humoral innate immunity consists of a limited, but diverse, set of humoral fluid phase pattern recognition molecules (PRMs) that represent a first line of resistance against microbial invaders by promoting pathogen disposal by phagocytosis, complement activation and inflammation. These factors encompass complement, ficolin, collectin and pentraxin family of proteins. Methods: We have analyzed the activity of PRMs for their potential capacity of inhibiting SARS-CoV-2 entry and replication into epithelial cells by a microneutralization assay based on a lentiviral particles pseudotyped with the SARS-CoV-2 spike protein in HEK293T cells overexpressing the angiotensin converting enzyme 2 (ACE2). Either SARS-CoV-2 or target cells were incubated with Mannose Binding Lectin (MBL, concentration range: 1-50 μ g/ml) to further characterize its anti-viral activity for 1 h prior to infection in both human Calu-3 cells and air-liquid interface cultures of human bronchial epithelial cells (HBEC). Binding experiments were carried out with SARS-CoV-2 Spike protein and recombinant MBL to further investigate its antiviral action. Results: Among 12 PRMs tested, only MBL inhibited viral entry in the pseudotyped neutralization assay. Furthermore, MBL protein inhibited SARS-CoV-2 viral replication in Calu-3 and HBEC by ca. one log10 at the top concentration (10 μ g/ml and 50 μ g/ml, respectively). MBL antiviral activity was confirmed also against alpha, beta and gamma SARS-CoV-2 variants of concern. Binding experiments showed that MBL specifically interacts with the trimeric form of SARS-CoV-2 spike. Conclusion: MBL binds to the Spike protein in its active trimeric conformation leading to the inhibition of SARS-CoV-2 infection and replication in vitro. These results suggest that MBL possesses an antiviral activity against SARS-CoV-2 that could bear therapeutic potential.
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Introduction: Hypercalcemia is a common clinical diagnosis. Hyperparathyroidism is one of the most common etiologies. Rarely hypercalcemia is associated with intense inflammation secondary to IL-6 production. Herein we present an interesting case of hypercalcemia associated with COVID-19. Case Description: 36-year-old woman with history of Cirrhosis secondary to hepatitis C and alcohol abuse initially admitted for COVID/ARDS and cryptococcemia without CNS involvement. She was initially treated with amphotericin B and continued on fluconazole. Patient was re-admitted after 2 weeks with abdominal pain, constipation and hypercalcemia (Ca 14.2 mg/dl (normal range 8.7-10.1mg/dl). Her ionized calcium was 1.78 mmol/L (normal range 1.12-1.32 mmol/L). Serum phosphorus was 2.5 mg/dl (normal range 2.5-4.5mg/dl). Intact PTH level was 6.7 pg/ml (normal range 6-48pg/ml). Vitamin D 25-OH level was 26.8 ng/ml (sufficient range 32-100 ng/ml);Vitamin D 1,25-OH level was 7.4 pg/ml (normal 19.9-79.3pg/ml). PTH-rp was unmeasurable (< 2pmol/l). CRP was elevated at 31.7mg/L (normal range 0.2-8 mg/L). She was not on calcium, Vitamin D supplementation or thiazide diuretics. Her renal functions were normal. She was given Intravenous fluids & Intravenous pamidronate. Steroids were not used due to an ongoing fungal infection. It was proposed that the patient had Interleukin-induced hypercalcemia secondary to COVID-19 infection. Her serum calcium normalized with improvement in clinical status Discussion: Recent literature suggests COVID-19 is associated with inflammatory response with cytokines & interleukins production. IL-6 production is significantly upregulated especially in severe cases of COVID-19 known as “Cytokine storm”. IL-6 is produced by bronchial epithelial cells. High levels of IL-6 are associated with worse outcomes and much more severe disease. IL-6 in turn causes osteoclast activation, bone resorption & hypercalcemia. In our patient other potential causes of hypercalcemia were ruled out. The proposed mechanism of her hypercalcemia is an intense inflammatory response associated with COVID-19 infection. Conclusion: We present a rare sequelae of COVID-19 infection which presents a teaching point for clinicians to consider while managing such novel disease
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Introduction: After the aftermath of covid 19 we are left to learn and understand the multiple respiratory manifestations of post covid 19 sequele The presence of bullous lung disease in post covid patients is one such a rare entity ,has been infrequently reported, studied Eventhough the exact mechanism of formation of bullae in post covid 19 are unknown, an emerging association has been observed.A bulla is an air containing space within the lung parenchyma that arises from destruction dilatation and confluence of airspaces distal to terminal bronchioles and is larger than 1 cm in diameter .Its wall are composed of attenuated and compressed parenchyma. Here in this case series we describe this unique presentation of bullous lung diseases in post covid 19 patients. Case Series: Here wepresent a case series of 7 patients without any known comorbidities who were diagnosed with post covid bullous lung disease Diagnosis was made with the help of contrast enchanced computed tomography aided by other serological and microbilogical workup. Patient was managed conservatively antibiotics, analgesics and other supportive measures. Learning Points: We are well aware of lung fibrosis post covid, our intention was to throw light into the new entity of bullous lung disease in post covid period. Bullous lung disease (with or without pneumothorax) should be part of differential diagnosis in a patient returning with chest pain and dyspnoea after SARSCov- 2 infection. CT imaging essential to differentiate radiographically presumed complex pneumothoraces from large bullae to prevent erroneous chest drain insertion into a bulla.
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Objective: To see the effects of Raj Nirwan Bati (RNB) on the hematobiochemical parameters, coagulation tests, and histopathological changes in the lungs, liver, kidneys and spleen and also to evaluate the immunomodulatory activity of RNBin Wistar rats. Methods: A total of 24 adult albino Wistar rats (of bodyweight 200-250 g) of either sex were divided into 3 groups. In the normal control group (n=8), no drug was administered and in the rest of the groups (A and B), RNB@ 26 mg/kg body weight./day and 260 mg/kg body weight/day respectively were administered orally for a period of 14 d. The blood samples were collected from the jugular vein at zero d (before drug administration) and after the 14th d of drug administration in both groups (A and B). The organ samples (lungs, liver, kidneys, and spleen) were collected after euthanizing the rats using Ketamine anesthesia overdose intraperitoneally (IP) after the 14th d of drug administration. White Blood Cells (WBC), Red Blood Cells (RBC), Hemoglobin (Hb), Hematocrit (HCT), Mean Corpuscular Volume (MCV), Mean Corpuscular Hemoglobin(MCH), Mean Corpuscular Hemoglobin Concentration(MCHC), number of platelets, Differential Leucocyte Count(DLC) i.e. the percentage of neutrophils, lymphocytes, eosinophils, monocytes and basophils, neutrophil adhesion percentage, Prothrombin test (PT), Activated Partial Thromboplastin Time (APTT), fibrinogen, D-dimer, Lactate Dehydrogenase (LDH), urea, creatinine, Aspartate Amino Transferase (AST), Alanine amino Transferase (ALT), Alkaline Phosphatase (ALP), C-Reactive Protein (CRP) were evaluated and histological examination of organs were done. Results: After statistical analysis, it was found that the decrease in TLC, RBC, Hb, HCT, and LDH in Wistar rats after RNB intervention in Group A as compared to that of before RNB intervention, was found to be statistically significant (P=0.001, P=0.002, P=0.001, P=0.039, and P=0.008). On the other hand, an increase was observed in MCV, Urea, Creatinine and ALT values in the Wistar rats after RNB intervention in Group ‘A’ as compared to that of before RNB intervention and this increase in values was statistically significant (P=0.007, P=0.001, P<0.001 and P=0.038). After RNB intervention in Group B, the increase in MCH, fibrinogen concentration, and monocytes percentage, was found to be statistically significant (P=0.004, P=0.033, and P=0.001) as well as the decrease in PT and APTT was statistically significant (P=0.007and P=0.002). After comparing the Mean Hematobiochemical and coagulation test parameters in the rats of Group A and Group B, after RNB intervention, it was observed that the concentration of Urea, Creatinine, APTT, and D-dimer were less in Group B as compared to that of Group A and this difference was statistically significant(P<0.001, P<0.001, P<0.001 and P=0.022). Histologically the findings in the lungs of group B were more distortion of lung architecture, most of the alveoli become collapse and make emphysematous changes, more diffuse inflammatory infiltrate within interalveolar septa and around bronchioles as compared to Group A. In the liver of group B rats, the histological findings were mild to moderate distortion of lobular architecture, healthy hepatocytes with more activation of kupffer cells as well as larger and more aggregates of inflammatory cells as compared to group A. Histological findings of kidneys in group A and group B rats were similar to that of control group rats. Conclusion: The results suggest that the RNB is having an immunomodulatory effect. It might be helpful in the restoration of coagulation factors and can help treat the COVID patients. No harmful effects on the lungs, liver, kidney, and spleen were seen. These findings may act as baseline data for planning further clinical trials in human study subjects to evaluate the effects on various comorbidities.
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Background: Severe acute respiratory coronavirus 2 (SARS-CoV-2) is the infectious agent of the current COVID-19 pandemic and is responsible for more than 2 million deaths worldwide. The virus utilizes the surface receptor angiotensin-converting enzyme 2 (ACE2) to infiltrate its target cells. It has been postulated that ACE2 is a human interferon-stimulated gene (ISG) and is upregulated by interferon (IFN) and virus stimulation. However, previous studies showed that not ACE2, moreover a novel short isoform of the enzyme is mainly expressed after IFN stimulation. Method: In this study, we aimed to investigate the impact of IFN stimulation on ACE2 expression in human bronchial epithelial cells. We measured the expression levels of two ACE2 isoforms, namely the full-length isoform and the IFN-induced truncated isoform proposed by Onabajo et al. (truncACE2) at the transcription and protein level following stimulation of Calu-3 and primary human bronchial epithelial cells (NHBE) with type-I,-II and-III IFNs. Specific primers along the ACE2 coding sequence were used for quantitative and semi-quantitative analysis of the transcript isoforms. Induced ACE2 protein isoforms were additionally analyzed using immunoblotting. Results: In both, the Calu3 cell line and the NHBE, truncACE2 was dose-dependently upregulated at the transcription level after 6h of IFNγ stimulation, whereas the full-length transcript levels did not change. IFNs of type-I and type-III stimulation induced a dosedependent upregulation of both ACE2 versions at the mRNA level, although the truncACE2 showed a higher expression level. In the immunoblotting analysis, neither the full-length nor the truncated ACE2 version showed a difference after IFNγ stimulation of NHBE. Conclusion: This study is in line with previous findings that only a truncated ACE2 isoform acts as an ISG. However, our data additionally show that not only type-II, but also type-I and type-III interferons induce this truncated version of ACE2. This is especially important, as type-I and type-III IFNs are secreted during the initial epithelial host response to the virus. It remains to be uncovered whether SARS-CoV-2 can utilize also truncACE2 to enter cells thereby using the initial host response to increase truncACE2 and facilitate viral spread.
ABSTRACT
The COVID-19 pandemic has devastated individuals, families, and institutions throughout the world. Despite the breakneck speed of vaccine development, the human population remains at risk of further devastation. The decision to not become vaccinated, the protracted rollout of available vaccine, vaccine failure, mutational forms of the SARS virus, which may exhibit mounting resistance to our molecular strike at only one form of the viral family, and the rapid ability of the virus(es) to hitch a ride on our global transportation systems, means that we are will likely continue to confront an invisible, yet devastating foe. The enemy targets one of our human physiology's most important and vulnerable life-preserving body tissues, our broncho-alveolar gas exchange apparatus. Notwithstanding the fear and the fury of this microbe's potential to raise existential questions across the entire spectrum of human endeavor, the application of an early treatment intervention initiative may represent a crucial tool in our defensive strategy. This strategy is driven by evidence-based medical practice principles, those not likely to become antiquated, given the molecular diversity and mutational evolution of this very clever "world traveler".