RESUMO
Mice are normally unaffected by SARS coronavirus 2 (SARS-CoV-2) infection since the virus does not bind effectively to the murine version of the angiotensin-converting enzyme 2 (ACE2) receptor molecule. Here, we report that induced mild pulmonary morbidities rendered SARS-CoV-2-refractive CD-1 mice susceptible to this virus. Specifically, SARS-CoV-2 infection after application of low doses of the acute lung injury stimulants bleomycin or ricin caused severe disease in CD-1 mice, manifested by sustained body weight loss and mortality rates greater than 50%. Further studies revealed markedly higher levels of viral RNA in the lungs, heart, and serum of low-dose ricin-pretreated mice compared with non-pretreated mice. Furthermore, lung extracts prepared 2-3 days after viral infection contained subgenomic mRNA and virus particles capable of replication only when derived from the pretreated mice. The deleterious effects of SARS-CoV-2 infection were effectively alleviated by passive transfer of polyclonal or monoclonal antibodies generated against the SARS-CoV-2 receptor binding domain (RBD). Thus, viral cell entry in the sensitized mice seems to depend on viral RBD binding, albeit by a mechanism other than the canonical ACE2-mediated uptake route. This unique mode of viral entry, observed over a mildly injured tissue background, may contribute to the exacerbation of coronavirus disease 2019 (COVID-19) pathologies in patients with preexisting morbidities.
Assuntos
Bleomicina/toxicidade , COVID-19/patologia , Lesão Pulmonar , Ricina/toxicidade , Animais , Chlorocebus aethiops , Comorbidade , Modelos Animais de Doenças , Feminino , Lesão Pulmonar/induzido quimicamente , Lesão Pulmonar/virologia , Camundongos , Células Vero , Ligação Viral , Internalização do Vírus/efeitos dos fármacosRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of the ongoing coronavirus disease 2019 (COVID-19) pandemic1. To understand the pathogenicity and antigenic potential of SARS-CoV-2 and to develop therapeutic tools, it is essential to profile the full repertoire of its expressed proteins. The current map of SARS-CoV-2 coding capacity is based on computational predictions and relies on homology with other coronaviruses. As the protein complement varies among coronaviruses, especially in regard to the variety of accessory proteins, it is crucial to characterize the specific range of SARS-CoV-2 proteins in an unbiased and open-ended manner. Here, using a suite of ribosome-profiling techniques2-4, we present a high-resolution map of coding regions in the SARS-CoV-2 genome, which enables us to accurately quantify the expression of canonical viral open reading frames (ORFs) and to identify 23 unannotated viral ORFs. These ORFs include upstream ORFs that are likely to have a regulatory role, several in-frame internal ORFs within existing ORFs, resulting in N-terminally truncated products, as well as internal out-of-frame ORFs, which generate novel polypeptides. We further show that viral mRNAs are not translated more efficiently than host mRNAs; instead, virus translation dominates host translation because of the high levels of viral transcripts. Our work provides a resource that will form the basis of future functional studies.
Assuntos
Perfilação da Expressão Gênica , Genoma Viral/genética , Fases de Leitura Aberta/genética , Biossíntese de Proteínas , SARS-CoV-2/genética , Proteínas Virais/biossíntese , Proteínas Virais/genética , Animais , Linhagem Celular , Humanos , Anotação de Sequência Molecular , Peptídeos/genética , Peptídeos/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Viral/genética , RNA Viral/metabolismo , Ribossomos/metabolismo , SARS-CoV-2/imunologia , SARS-CoV-2/metabolismo , SARS-CoV-2/patogenicidade , Proteínas Virais/metabolismoRESUMO
A previously healthy young man presented with a chronic cavitary pulmonary infection that began while in Goa, India. Burkholderia pseudomallei was cultured from sputum samples. The infection fully resolved after prolonged antibiotic treatment. Other than traveling during the monsoon season, extensive use of well-water for water-pipe smoking of cannabis was identified as a possible risk factor for infection. This is one of the first reports of travel-associated melioidosis from India. Genomic and immunological characterization suggested that the B. pseudomallei isolate collected from the reported case exhibited limited similarity to other B. pseudomallei strains.
Assuntos
Doenças Transmissíveis Importadas/diagnóstico , Melioidose/diagnóstico , Viagem , Adulto , Antibacterianos/uso terapêutico , Burkholderia pseudomallei/isolamento & purificação , Doenças Transmissíveis Importadas/microbiologia , Humanos , Índia , Israel , Masculino , Melioidose/tratamento farmacológico , Fatores de Risco , Escarro/microbiologiaRESUMO
We have previously shown that the cell morphogenesis NlpD lipoprotein is essential for virulence of the plague bacteria, Yersinia pestis. To elucidate the role of NlpD in Y. pestis pathogenicity, we conducted a whole-genome comparative transcriptome analysis of the wild-type Y. pestis strain and an nlpD mutant under conditions mimicking early stages of infection. The analysis suggested that NlpD is involved in three phenomena: (i) Envelope stability/integrity evidenced by compensatory up-regulation of the Cpx and Psp membrane stress-response systems in the mutant; (ii) iron acquisition, supported by modulation of iron metabolism genes and by limited growth in iron-deprived medium; (iii) activity of the twin-arginine (Tat) system, which translocates folded proteins across the cytoplasmic membrane. Virulence studies of Y. pestis strains mutated in individual Tat components clearly indicated that the Tat system is central in Y. pestis pathogenicity and substantiated the assumption that NlpD essentiality in iron utilization involves the activity of the Tat system. This study reveals a new role for NlpD in Tat system activity and iron assimilation suggesting a modality by which this lipoprotein is involved in Y. pestis pathogenesis.
Assuntos
Proteínas de Bactérias/metabolismo , Técnicas de Inativação de Genes , Ferro/metabolismo , Lipoproteínas/metabolismo , Sistema de Translocação de Argininas Geminadas/metabolismo , Fatores de Virulência/metabolismo , Yersinia pestis/enzimologia , Yersinia pestis/metabolismo , Animais , Proteínas de Bactérias/genética , Transporte Biológico , Feminino , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Lipoproteínas/genética , Camundongos , Fatores de Virulência/genética , Yersinia pestis/genéticaRESUMO
Ricin, a lethal toxin derived from castor oil beans, is a potential bio-threat due to its high availability and simplicity of preparation. Ricin is prepared according to simple recipes available on the internet, and was recently considered in terrorist, suicide, or homicide attempts involving the parenteral route of exposure. In-depth study of the morbidity developing from parenteral ricin poisoning is mandatory for tailoring appropriate therapeutic measures to mitigate ricin toxicity in such instances. The present study applies various biochemical, hematological, histopathological, molecular, and functional approaches to broadly investigate the systemic effects of parenteral intoxication by a lethal dose of ricin in a murine model. Along with prompt coagulopathy, multi-organ hemorrhages, and thrombocytopenia, ricin induced profound morpho-pathological and functional damage in the spleen, bone marrow, and cardiovascular system. In the heart, diffuse hemorrhages, myocyte necrosis, collagen deposition, and induction in fibrinogen were observed. Severe functional impairment was manifested by marked thickening of the left ventricular wall, decreased ventricular volume, and a significant reduction in stroke volume and cardiac output. Unexpectedly, the differential severity of the ricin-induced damage did not correlate with the respective ricin-dependent catalytic activity measured in the various organs. These findings emphasize the complexity of ricin toxicity and stress the importance of developing novel therapeutic strategies that will combine not only anti-ricin specific therapy, but also will target ricin-induced indirect disturbances.
Assuntos
Substâncias para a Guerra Química/toxicidade , Ricina/toxicidade , Animais , Medula Óssea/efeitos dos fármacos , Medula Óssea/patologia , Colágeno/metabolismo , Feminino , Fibrinogênio/metabolismo , Coração/efeitos dos fármacos , Coração/fisiologia , Injeções Intramusculares , Rim/efeitos dos fármacos , Rim/patologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Camundongos , Miocárdio/metabolismo , Miocárdio/patologia , Baço/efeitos dos fármacos , Baço/patologiaRESUMO
We report a case of monkeypox in a man who returned from Nigeria to Israel in 2018. Virus was detected in pustule swabs by transmission electron microscopy and PCR and confirmed by immunofluorescence assay, tissue culture, and ELISA. The West Africa monkeypox outbreak calls for increased awareness by public health authorities worldwide.
Assuntos
Doenças Transmissíveis Importadas/diagnóstico , Doenças Transmissíveis Importadas/epidemiologia , Surtos de Doenças , Monkeypox virus , Mpox/diagnóstico , Mpox/epidemiologia , Animais , Biópsia , Chlorocebus aethiops , Doenças Transmissíveis Importadas/história , Doenças Transmissíveis Importadas/virologia , História do Século XXI , Humanos , Israel/epidemiologia , Mpox/história , Mpox/virologia , Pele/patologia , Pele/virologia , Células VeroRESUMO
Vaccinia virus protein A33 (A33VACV) plays an important role in protection against orthopoxviruses, and hence is included in experimental multi-subunit smallpox vaccines. In this study we show that single-dose vaccination with recombinant Sindbis virus expressing A33VACV, is sufficient to protect mice against lethal challenge with vaccinia virus WR (VACV-WR) and ectromelia virus (ECTV) but not against cowpox virus (CPXV), a closely related orthopoxvirus. Moreover, a subunit vaccine based on the cowpox virus A33 ortholog (A33CPXV) failed to protect against cowpox and only partially protected mice against VACV-WR challenge. We mapped regions of sequence variation between A33VACV and A33CPXVand analyzed the role of such variations in protection. We identified a single protective region located between residues 104-120 that harbors a putative H-2Kd T cell epitope as well as a B cell epitope - a target for the neutralizing antibody MAb-1G10 that blocks spreading of extracellular virions. Both epitopes in A33CPXV are mutated and predicted to be non-functional. Whereas vaccination with A33VACV did not induce in-vivo CTL activity to the predicted epitope, inhibition of virus spread in-vitro, and protection from lethal VACV challenge pointed to the B cell epitope highlighting the critical role of residue L118 and of adjacent compensatory residues in protection. This epitope's critical role in protection, as well as its modifications within the orthopoxvirus genus should be taken in context with the failure of A33 to protect against CPXV as demonstrated here. These findings should be considered when developing new subunit vaccines and monoclonal antibody based therapeutics against orthopoxviruses, especially variola virus, the etiologic agent of smallpox.
Assuntos
Vírus da Varíola Bovina/imunologia , Vírus da Ectromelia/imunologia , Ectromelia Infecciosa/prevenção & controle , Glicoproteínas de Membrana/imunologia , Vaccinia virus/imunologia , Vacínia/prevenção & controle , Proteínas do Envelope Viral/imunologia , Vacinas Virais/imunologia , Imunidade Adaptativa , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Modelos Animais de Doenças , Portadores de Fármacos , Epitopos de Linfócito B/genética , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Feminino , Variação Genética , Vetores Genéticos , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos BALB C , Sindbis virus/genética , Proteínas do Envelope Viral/genética , Vacinas Virais/administração & dosagemRESUMO
To assess the putative correlation between comparative genomic hybridization (CGH)-detectable genetic alterations in epithelial ovarian cancer and disease recurrence, conventional CGH was performed on 45 epithelial ovarian cancers: 26 tumors from sporadic, BRCA mutation noncarriers and 11 and 8 tumors from BRCA1 and BRCA2 mutation carriers, respectively. Relevant clinical data, including histology, grade, stage, size of residual tumor, recurrence, and survival, were obtained from outpatient and inpatient charts. Among the 45 cases, the most common regions involving gain of DNA copy number were 3q (n = 23; 51%), 8q (n = 21; 47%), and 1q (n = 14; 31%), and the most common regions with loss were 19 and 22 at 9 cases (20%) each, followed by 5q (n = 6; 13%). In multivariate analysis, the total number of genetic alterations was not associated with risk of recurrence, but gain in 5p was associated with a higher risk of recurrence (hazard ratio HR = 6.06, P = 0.0399), and gain in 1p as well as loss in 5q were associated with a significant decrease in recurrence (HR = 0.08, P = 0.0079, and HR = 0.10, P = 0.0143, respectively). Recurrence rate in patients with epithelial ovarian cancer is seemingly associated with specific genetic alterations detected by CGH, but the specific genes involved and the implications of these findings await further studies.
Assuntos
Carcinoma/genética , Hibridização Genômica Comparativa , Recidiva Local de Neoplasia/genética , Neoplasias Ovarianas/genética , Idoso , Carcinoma/etiologia , Carcinoma/metabolismo , Feminino , Perfilação da Expressão Gênica/métodos , Humanos , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/etiologia , Recidiva Local de Neoplasia/metabolismo , Neoplasias Ovarianas/metabolismoRESUMO
Ovarian cancer, the leading cause of death due to gynecological malignancy, is diagnosed in most cases at an advanced stage. Combined with the paucity of symptoms of early-stage disease, the need to develop novel effective markers for the detection of potentially curable, early-stage disease is self-evident. Comprehensive analyses of somatic gene expression patterns in ovarian cancer were reported previously (n=17) and yielded substantial information on somatically altered genes, information that can potentially be useful in developing early detection markers. To further substantiate the role that these genes play in ovarian cancer tumorogenesis, we surveyed these reports and arranged the significantly altered genes from all reported studies by their chromosomal location (in silico chromosomal clustering). Subsequent comparison of this clustering to known genomic somatic alterations at the DNA level from data obtained using comparative genomic hybridization (CGH) was carried out. The major chromosomal regions that displayed overexpressed genes were correlated with the major CGH-detectable DNA amplification areas at 20q (harboring HE4, SLPI, MYBL2, UBE2C, and SDC4) and 1q (harboring MUC1). These genes may provide insights into ovarian cancer pathogenesis and may also prove to be useful as early detection tools.
Assuntos
Mapeamento Cromossômico , Perfilação da Expressão Gênica , Neoplasias Ovarianas/genética , Feminino , Dosagem de Genes , Humanos , Hibridização de Ácido Nucleico , Análise de Sequência com Séries de OligonucleotídeosRESUMO
Epithelial ovarian cancer is the most lethal gynecologic malignancy in the western world. In 75% of patients, peritoneal metastases are found at the time of primary surgery. However, the genetic events leading to the development of ovarian tumors and to the genetic progression toward metastasis remain unclear. To gain insight into this issue, the types and patterns of DNA copy number changes were compared between primary ovarian tumors and their respective metastases by using comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH). The genetic alterations (deletions and amplifications) detected by CGH were similar in the primary tumors and in their respective metastases. Moreover, the FISH results show a similar pattern of chromosomal abnormalities. Our results imply that the major gross genetic changes in ovarian cancer take place in the primary tumor, and the additional genetic changes that may occur in the metastases are not detectable by CGH.
Assuntos
Neoplasias Ovarianas/genética , Deleção Cromossômica , Feminino , Amplificação de Genes , Humanos , Hibridização in Situ Fluorescente , Pessoa de Meia-Idade , Hibridização de Ácido Nucleico/métodos , Neoplasias Peritoneais/genética , Neoplasias Peritoneais/secundárioRESUMO
OBJECTIVE: Our purpose was to get an overview of the genetic events leading to the development of familial and sporadic ovarian tumors and to identify chromosomal regions that may contain genes important in tumor progression. METHODS: The comparative genomic hybridization (CGH) technique was employed in a total of 46 epithelial ovarian or peritoneal tumors: 27 sporadic tumors, 11 tumors disected from BRCA1 mutation (185delAG) carriers, and eight from BRCA2 mutation (6174delT) carriers (familial tumors). RESULTS: The average number of genetic alterations (deletions and amplifications) was significantly (alpha=0.0069) higher in familial tumors (9.17 +/- 4.25 alterations per tumor in the BRCA1 mutation carriers and 7.25 +/- 6.06 in the BRCA2 mutation carriers) compared to the sporadic group (4.26 +/- 3.61 alterations per tumor). The pattern of the chromosome amplifications resembled in the three groups and the most common amplifications detected were at chromosomes 8q, 3q, and 2q. The pattern of the chromosomal deletions varied between the groups. Among the BRCA1 group, the most common deletions were in chromosomes 9 and 19. The BRCA2 group showed a lower frequency of deletions. Deletion of chromosome 16 and 22 were the most frequent ones. No specific chromosomal deletion was significantly indicated in the sporadic group. CONCLUSIONS: Familial ovarian tumors exhibit a significantly higher number of chromosomal aberrations and genomic imbalances and nonrandom genetic changes were characterized in the BRCA1 and BRCA2 groups.