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Genetic and serologic characterization of human herpesvirus type 8 (HHV -8) IN South Africa.Alagiozoglou, Pandeli (Lee) 06 March 2014 (has links)
Human herpes virus type 8 (HHV-8) is strongly implicated as the etiological agent of
Kaposi’s sarcoma (KS). The incidence of KS in South Africa is increasing in parallel with the HIV-1 epidemic. Molecular and serological prevalence of HHV-8 in HIV-1 infected individuals with and without KS was investigated. DNA fragments from ORF26 (capsid, 330BAM233) and ORF75 (tegument) regions were used to determine the prevalence of HHV-8 DNA in peripheral blood mononuclear cells (PBMC) from 429 HIV-1 infected individuals, 95 of whom had histologically confirmed KS. Of those without KS, 14 (4.2%) were PGR positive for HHV-8 DNA. In the individuals with KS, the proportion of HHV-8 DNA positive PBMC samples was 11 times higher (46/95,48%). Similarly, an immunofluorescence assay showed that 78% of KS patients had antibodies to HHV-8 compared to 16% of KS negative individuals. Among the KS group, 93% of PCRpositive samples were also HHV-8 antibody positive compared to only 66% of PCR negative samples indicating that viremia is associated with good antibody responses. Matched lymph node and PBMC samples were available for 8 patients. HHV-8 DNA was more
frequently detected in the lymph node (3/8) than in the blood (1/8), suggesting that the lymph nodes are a reser / o r for HHV-8. These data confirm the association between HHV-8 and KS and suggest that there is a high background prevalence of HHV-8 infection in HIV-1 infected individuals in South Africa.
The ORP 75 gene of 40 HHV-8 strains was sequenced and the phylogenetic relationships
between South African and already published sequences were investigated. The majority (n=29) of strains overlapped with the published A and B subgroups and were termed A/B variants.Three strains were classified as subgroup C while 8 sequences did not cluster with any of the previously classified subgroups and were termed novel (N) group. The DNA distance of this novel group differed from the A, B and C subgroups by 4.7%, 3.8% and 4,5% respectively although within the N group there was only 0.4% variation. The addition of this group significantly increased the number of subgroup-specific polymorphisms from 17 to 47 over a 804 bp region. There was sufficient inter-subgroup genetic diversity that single strand conformational polymorphisms (SSCP) could be used to rapidly identify them. Thus, based on the analysis of the ORF75 gene, a unique HHV-8 sub-group is present in South Africa which accounts for 20% of circulating strains. Further studies are required to determine the extent of evolutionary phytogeny, distribution and pathogenic potential of this novel group.
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Patobiologia do hespervírus associado ao Sarcoma de Kaposi/Hespervírus Humano Tipo-8: genotipagem de isolados virais em lesões de Sarcoma de Kaposi e imunoevasão dependente da inibição da síntese e modulação da degradação da proteína viral LANA-1Silva, Suzane Ramos da [UNESP] 29 April 2008 (has links) (PDF)
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Patobiologia do hespervírus associado ao Sarcoma de Kaposi/Hespervírus Humano Tipo-8 : genotipagem de isolados virais em lesões de Sarcoma de Kaposi e imunoevasão dependente da inibição da síntese e modulação da degradação da proteína viral LANA-1 /Silva, Suzane Ramos da. January 2008 (has links)
Orientador: Deilson Elgui de Oliveira / Banca: José Vassallo / Banca: Luisa Lina Villa / Banca: Sandra Cecília Botelho Costa / Banca: Claudio Sérgio Pannuti / Resumo: Não disponível. / Abstract: Not available. / Doutor
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Detecção do herpesvirus humano 8 (HHV-8) em pacientes com sarcoma de Kaposi, mieloma multiploo e em doadores de sangueCunha, Andrea Mendonça Gusmão 25 April 2001 (has links)
Orientador: Fernando Ferreira Costa / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-07-27T18:34:11Z (GMT). No. of bitstreams: 1
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Previous issue date: 2001 / Resumo: o Herpesvírus Humano 8 (HHV-8), ou Herpesvírus associado ao sarcoma de Kaposi (KSHV), foi identificado em 1994 por CHANG et aI em biópsia de pele de pacientes com sarcoma de Kaposi (SK) associado à Síndrome da Imunodeficiência Adquirida (SIDA / AIDS). O HHV-8 pertence à família Herpesviridae, sub-família Gamaherpesvirinae e gênero Rhadinovirus, o único do gênero a infectar humanos....Observação: O resumo, na integra, podera ser visualizado no texto completo da tese digital / Abstract: Human herpesvirus 8 (HHV-8), or Kaposi's sarcoma associated herpesvirus (KSHV), was identified in 1994 by Yuan Chang et ai in skin biopsies ftom patients with Acquired Immunodeficiency Syndrome (AIDS) related Kaposi's sarcoma. HHV-8 belongs to Herpesviridae family, Gamaherpesvirinae subfamily and Radinovirus genus, the only one ftom this genus which infects humans.... Note: The complete abstract is available with the full electronic digital thesis or dissertations / Mestrado / Mestre em Ciências Médicas
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A prospective randomized trial of two fractionation regimens of radiation therapy in the management of AIDS- associated Kaposi SarcomaSingh, Niveditha Bhavna 14 February 2007 (has links)
Student Number : 9201769X -
M Med research report -
School of Clinical Medicine -
Faculty of Health Sciences / A PROSPECTIVE RANDOMIZED TRIAL OF TWO FRACTIONATION
REGIMENS OF RADIATION THERAPY IN THE MANAGEMENT OF AIDSASSOCIATED
KAPOSI SARCOMA
OBJECTIVE:
To compare a standard fractionation scheme with a hypofractionated scheme in
the treatment of AIDS-associated Kaposi sarcoma with the aim of showing noninferiority
of the shorter schedule.
PATIENTS AND METHODS:
HIV positive patients with histologically proven Kaposi sarcoma presenting
consecutively to Radiation Oncology at Johannesburg Hospital were randomized
between January 2003 and May 2004 to receive a standard regimen of 24 Gy in
12 fractions (ARM A) or the study regimen of 20 Gy in 5 fractions (ARM B). The
radiation technique used was individualized for each site in accordance with
departmental practice. Follow-up assessment was done at monthly intervals.
Treatment response and toxicity were recorded at each follow-up visit.
RESULTS:
A total of 60 patients were recruited, of which 41 were male and 19 were female.
The median age was 36 years (range: 23 – 55 years). Thirteen patients died prior
to receiving treatment. The remaining 47 patients were treated to 65 sites, of
which 35 sites received 24 Gy in 12 fractions (ARM A) and 30 sites received 20
Gy in 5 fractions (ARM B). The main indications for treatment were pain (n=71),
oedema (n=44), functional impairment (n=35), cosmesis (n=14) and bleeding
(n=4). At the time of reporting 28 patients were alive and 32 patients have died.
The overall survival of the whole group was 37% at 1 year. A complete response
was recorded at 28 sites, a partial response at 19 sites and stable disease at 3
sites. The mean time to maximum objective response was 3 months (range: 1 –
14 months). The response rates were equal in the 2 treatment arms (p=0.73).
Local control was equal in the 2 treatment arms with a median local recurrence
free survival of 150 days for ARM A and 455 days for ARM B (p=0.11, log rank
test). Acute skin toxicity occurred at 27 sites. Moist desquamation developed at 7
sites while necrosis developed at 2 sites. Acute skin toxicity was equal in the 2
treatment arms (p=0.77). Acute mucosal toxicity occurred at 2 sites. Late skin
reactions developed at 21 sites, of which necrosis or ulceration occurred at 5
sites. Chronic skin reactions were equivalent in the 2 treatment arms (p=0.24).
Post radiation oedema developed at 5 sites.
CONCLUSION:
In our experience, 20 Gy in 5 fractions gave similar results to 24 Gy in 12
fractions in terms of treatment response, local recurrence free survival and
toxicity in this small group of patients.
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Aids and endemic kaposi's sarcoma development : comparison by histopathology, virology (HHV-8/KSHV) and cytogenetics /Pyakurel, Pawan, January 2005 (has links)
Diss. (sammanfattning) Stockholm : Karol. inst., 2005. / Härtill 5 uppsatser.
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HHV-8/KSHV association with tumor cells during development of Kaposi sarcoma /Pak, Fatemeh. January 2006 (has links)
Lic.-avh. (sammanfattning) Stockholm : Karolinska institutet, 2006. / Härtill 3 uppsatser.
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Produção de vetores recombinantes para análise das propriedades biológicas e cancerígenas da proteína K1 do herpesvírus associado ao sarcoma de Kaposi (KSHV/HHV-8)Squiavinato, Annie Cristhine Moraes Sousa [UNESP] 25 June 2014 (has links) (PDF)
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000793411.pdf: 2869881 bytes, checksum: dde8b87e326aaccf874b6b62350ca99d (MD5) / O herpesvírus associado ao sarcoma de Kaposi (KSHV) é um gammaherpesvírus associado ao sarcoma de Kaposi (SK). A proteína K1 do KSHV é conhecida por aumentar a sobrevivência e proliferação celular em células infectadas. A ORF-K1 viral apresenta elevada variabilidade, de modo a discriminar diferentes genótipos do KSHV. Até o momento não se sabe se diferentes genótipos virais apresentam características biológicas próprias. Assim, vetores recombinantes contendo a ORF-K1 de genótipos virais A e B foram gerados. A ORF-K1 foi obtida a partir do DNA de linhagens de linfoma de efusão primária (PEL) constitutivamente infectadas pelo KSHV. O amplicon da ORF-K1 e vetor comercial foram digeridos, ligados e clonados em bactéria E. coli DH5α. Clones selecionados foram então submetidos à sequenciamento automatizado de DNA. As sequências geradas dos vetores recombinantes foram alinhadas e comparadas com sequências-protótipo de ORF-K1 do KSHV. Sequência de aminoácidos dos vetores recombinantes foram geradas e analisadas quanto a região codificadora do ITAM de K1. Um clone validado de cada vetor recombinante foi transfectado estavelmente em células HEK293 e a expressão de K1 foi avaliada por Western blot (WB) O sequenciamento de DNA demonstrou que a ORF-K1 dos vetores recombinantes de genótipo A, B e C corresponde a de sequências-protótipo depositadas de linhagens de PEL. A presença de K1 no lisado de células HEK293 transfectadas foi demonstrada por WB. A análise da sequência de aminoácidos de K1 codificada nos vetores recombinantes revelou que o domínio ITAM de K1 do genótipo B apresenta aminoácidos distintos em relação ao ITAM de K1 de genótipos A e C. Portanto, vetores recombinantes de ORF-K1 foram produzidos e validados, e serão úteis para se estabelecer modelo experimental para análises das propriedades biológicas da proteína K1 do KSHV / Kaposi's sarcoma-associated herpesvirus (KSHV) is a gammaherpesvirus associated with the development of Kaposi's sarcoma. The K1 protein of KSHV has been shown to induce increases the survival and proliferation of infected cells. The viral ORF-K1 shows high variability, so it is possible to distinguish different KSHV genotypes (genotypes A, B, C, D). So far, it is unclear whether different viral genotypes have their own biological characteristics. To this intent, recombinant vectors were generated containing the ORFK1 genotypes A and B. ORF-K1 was obtained from the DNA of primary effusion lymphoma (PEL) cell lines. The amplicon generated and the commercial vector were digested, bonded and cloned in E.coli DH5α. Selected clones underwent automated DNA sequencing. The generated sequences were compared with prototype sequences of ORF-K1. The amino acid sequences from vectors were generated and analyzed in the K1 ITAM-coding region. Validated clones were stably transfected into HEK293 cells and K1 expression was evaluated using Western blot (WB). DNA sequencing showed that the ORF-K1 recombinant vectors corresponds to the prototype sequences deposited from PEL cell lines. WB demonstrated the presence of K1 in the lysate of HEK293 transfected cells. Analysis of the K1 amino acid sequence encoded in the vectors revealed that ITAM domain of K1 (genotype B) has distinct amino acids from the ones in the ITAM domain of K1 (genotypes A and C). Therefore ORF-K1 recombinant vectors were produced and validated, and will be useful to establish an experimental model for analysis of the biological properties of the K1 protein
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Produção de vetores recombinantes para análise das propriedades biológicas e cancerígenas da proteína K1 do herpesvírus associado ao sarcoma de Kaposi (KSHV/HHV-8) /Squiavinato, Annie Cristhine Moraes Sousa. January 2014 (has links)
Orientador: Deilson Elgui de Oliveira / Banca: Claudia Esther Alicia Rocio Hassan / Banca: João Pessoa Araújo Junior / Resumo: O herpesvírus associado ao sarcoma de Kaposi (KSHV) é um gammaherpesvírus associado ao sarcoma de Kaposi (SK). A proteína K1 do KSHV é conhecida por aumentar a sobrevivência e proliferação celular em células infectadas. A ORF-K1 viral apresenta elevada variabilidade, de modo a discriminar diferentes genótipos do KSHV. Até o momento não se sabe se diferentes genótipos virais apresentam características biológicas próprias. Assim, vetores recombinantes contendo a ORF-K1 de genótipos virais A e B foram gerados. A ORF-K1 foi obtida a partir do DNA de linhagens de linfoma de efusão primária (PEL) constitutivamente infectadas pelo KSHV. O amplicon da ORF-K1 e vetor comercial foram digeridos, ligados e clonados em bactéria E. coli DH5α. Clones selecionados foram então submetidos à sequenciamento automatizado de DNA. As sequências geradas dos vetores recombinantes foram alinhadas e comparadas com sequências-protótipo de ORF-K1 do KSHV. Sequência de aminoácidos dos vetores recombinantes foram geradas e analisadas quanto a região codificadora do ITAM de K1. Um clone validado de cada vetor recombinante foi transfectado estavelmente em células HEK293 e a expressão de K1 foi avaliada por Western blot (WB) O sequenciamento de DNA demonstrou que a ORF-K1 dos vetores recombinantes de genótipo A, B e C corresponde a de sequências-protótipo depositadas de linhagens de PEL. A presença de K1 no lisado de células HEK293 transfectadas foi demonstrada por WB. A análise da sequência de aminoácidos de K1 codificada nos vetores recombinantes revelou que o domínio ITAM de K1 do genótipo B apresenta aminoácidos distintos em relação ao ITAM de K1 de genótipos A e C. Portanto, vetores recombinantes de ORF-K1 foram produzidos e validados, e serão úteis para se estabelecer modelo experimental para análises das propriedades biológicas da proteína K1 do KSHV / Abstract: Kaposi's sarcoma-associated herpesvirus (KSHV) is a gammaherpesvirus associated with the development of Kaposi's sarcoma. The K1 protein of KSHV has been shown to induce increases the survival and proliferation of infected cells. The viral ORF-K1 shows high variability, so it is possible to distinguish different KSHV genotypes (genotypes A, B, C, D). So far, it is unclear whether different viral genotypes have their own biological characteristics. To this intent, recombinant vectors were generated containing the ORFK1 genotypes A and B. ORF-K1 was obtained from the DNA of primary effusion lymphoma (PEL) cell lines. The amplicon generated and the commercial vector were digested, bonded and cloned in E.coli DH5α. Selected clones underwent automated DNA sequencing. The generated sequences were compared with prototype sequences of ORF-K1. The amino acid sequences from vectors were generated and analyzed in the K1 ITAM-coding region. Validated clones were stably transfected into HEK293 cells and K1 expression was evaluated using Western blot (WB). DNA sequencing showed that the ORF-K1 recombinant vectors corresponds to the prototype sequences deposited from PEL cell lines. WB demonstrated the presence of K1 in the lysate of HEK293 transfected cells. Analysis of the K1 amino acid sequence encoded in the vectors revealed that ITAM domain of K1 (genotype B) has distinct amino acids from the ones in the ITAM domain of K1 (genotypes A and C). Therefore ORF-K1 recombinant vectors were produced and validated, and will be useful to establish an experimental model for analysis of the biological properties of the K1 protein / Mestre
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Soroprevalencia e epidemiologia molecular do herpesvirus humano 8 (HHV-8) em populações brasileirasCunha, Andrea Mendonça Gusmão 24 February 2005 (has links)
Orientador: Fernando Ferreira Costa, Sandra Cecilia Botelho Costa / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-04T04:21:19Z (GMT). No. of bitstreams: 1
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Previous issue date: 2005 / Resumo: O Herpesvírus Humano 8 (HHV-8) foi identificado em 1994 por CHANG et al em biópsia de pele de pacientes com sarcoma de Kaposi associado à Síndrome da Imunodeficiência Adquirida (SIDA). O HHV-8 é um oncovírus membro da família Herpesviridae, sub-família Gamaherpesvirinae e gênero Rhadinovirus, o único do gênero a infectar humano. Possui ultraestrutura semelhante à de outros herpesvírus, apresentando regiões de DNA homólogas a dois gama-herpesvírus: Epstein-Barr Vírus (EBV) e o Herpes Vírus Saimiri (HVS), ambos com potencial oncogênico. Estudos revelaram a associação entre o HHV-8 e todas as formas de SK: clássica, endêmica, relacionadas à AIDS e associado a transplantes, além de outras lesões proliferativas das linhagens linfóides, relacionadas ou não à AIDS, como o linfoma primário de serosas (PEL) ou linfoma de cavidades do corpo (BCBL) e a doença de Castleman multicêntrica (DCM). No presente estudo, com a utilização dos ensaios sorológicos de imunofluorescência indireta foi possível determinar que o HHV-8 é endêmico em duas tribos indígenas (Tiriyó e Waiampi), localizadas na região Amazônica. Anticorpos anti-HHV-8 foram detectados em 56,8% dos índios (558/982), em todas as faixas etárias (0-81 anos) e em ambos os sexos. Nessas populações, a elevada prevalência em crianças menores de 2 anos (44,4%) e crianças de 2-9 anos (35.0%) sugerem a existência de vias de transmissão não-sexual do HHV-8, através da transmissão vertical ou pelo contato com secreções contaminadas. A soroprevalência do HHV-8 em doadores de sangue da cidade de Campinas foi baixa (2,8%), sendo todos os casos positivos pertencentes ao sexo masculino (9/319) e faixa etária de 31 a 50 anos. Curiosamente, todos os casos de pacientes com SK acompanhados no Hospital de Clínicas da Unicamp também pertenceram ao sexo masculino, sendo detectados anticorpos anti-HHV-8 em todos os casos de SK avaliados. A genotipagem do HHV-8 através do sequenciamento de regiões hipervariáveis do genoma viral, como a ORF-K1, possibilitou a classificação dos principais subtipos virais (A, B, C, D e E). Para realizar a genotipagem do HHV-8 nós padronizamos técnicas moleculares para amplificação dos fragmentos VR1 e VR2 da região hipervariável ORF-K1 do HHV-8, possibilitando a construção de árvores filogenéticas e a determinação dos subtipos virais. Em pacientes com SK da cidade de Campinas foram detectados os subtipos A, B e C do HHV-8, com maior percentual do subtipo C. Em índios da região Amazônica foram detectados os subtipos A e E do HHV-8. Nosso estudo foi o primeiro a realizar genotipagem em amostras de indivíduos com sorologia positiva para o Vírus da Imunodeficiência Humana (HIV) da cidade de Salvador, sendo detectados os subtipos B e subtipo indeterminado do HHV-8. Com isso, foi possível determinar que os subtipos A, B, C e E do HHV-8 estão presentes em populações brasileiras. Todas as seqüências nucleotídicas dos fragmentos VR1 e VR2 do HHV-8 obtidas no presente estudo serão depositadas no banco de dados NCBI/Nucleotide Sequence Database (GenBank) informando a comunidade científica mundial dados relevantes referentes às cepas do HHV-8 circulantes no Brasil. Em nosso estudo, a análise de amostras provenientes de doadores de sangue e pacientes com sarcoma de Kaposi da cidade de Campinas, pacientes HIV positivos de Salvador e de índios da região Amazônica resultou na obtenção de dados de soroprevalência e de epidemiologia molecular do HHV-8 em populações brasileiras. O conhecimento de técnicas sorológicas e moleculares aplicado no presente estudo poderá ser utilizado pelos Laboratórios de Biologia Molecular da Unicamp e do LASP/CPqGM/Fiocruz, possibilitando a implantação de técnicas para o diagnóstico e genotipagem do HHV-8 nos referidos centros / Abstract: CHANG et al first identified human Herpesvirus Type 8 (HHV-8) in 1994 from skin biopsies of patients with Acquired Immunodeficiency Syndrome (AIDS) associated Kaposi¿s Sarcoma (KS). HHV-8 is an oncovirus from the Herpesviridae family, Gamaherpesvirinae subfamily and Rhadinovirus genus. It is the only one of its genus to infect humans. Its ultra structure reminds other herpesvirus, presenting DNA regions similar to two gama-herpesvirus: Epstein-Barr Virus (EBV) and Saimiri Herpesvirus (SHV), both with oncogenic potential. Studies have revealed the association between HHV-8 and all forms of KS: classic, endemic, related to AIDS and associated to transplants, besides other lymphoid proliferative diseases related or not to AIDS, such as primary effusion lymphoma (PEL), body cavity lymphoma (BCBL) and Castleman's disease (CD). In this study, using indirect immunofluorescence serologic assays, it was possible to determine that HHV-8 is endemic in two Amerindian tribes (Tiriyo and Waiampi), from the Amazon region. Anti-HHV-8 antibodies were detected in 56.8% of the Amerindians (558/982), in all ages (0-81 years old) and both sexes. In these populations, high prevalence in children younger than 2 years old (44.4%) and children from 2 to 9 years old (35.0%) suggests non-sexual routs of transmission of HHV-8, through vertical transmission or contact to contaminated secretions. HHV-8 seroprevalence in blood donors from Campinas was low (2.8%). All positive cases were male (9/319) with 31 to 50 years of age. Curiously, all KS patients assessed in our study were male and anti-HHV-8 antibodies were detected in all cases. Genotyping of HHV-8 through sequencing of hypervariable regions of the viral genome, such as ORF-K1, made possible formulate a classification of the main viral subtype (A, B, C, D and E) and its variants. In order to perform HHV-8 genotyping, we standardized molecular techniques for amplification and sequencing of two fragments (VR1 and VR2) from the hypervariable ORF-K1 region of HHV-8, building up phylogenetic trees and determining viral subtypes. Patients with SK from the city of Campinas had subtypes A, B and C detected, with greater frequency of subtype C. Subtypes A and E were detected in Amazonic Amerindians. This study was the first to perform genotyping in samples of patients with Human Immunodeficiency Virus (HIV) positive serology from the city of Salvador, detecting subtype B and an undetermined subtype. Thus, it was possible to determine that HHV-8 subtypes A, B, C and E are present in Brazilian populations. All nucleotide sequences of HHV-8 fragments VR1 and VR2 found during this study will be deposit at the NCBI/Nucleotide Sequence Database (GenBank), informing the scientific community with important data of HHV-8 strains in Brazil. Analysis of samples from blood donors and Kaposi¿s Sarcoma in Campinas, HIV positive patients in Salvador and Amerindians from the Amazon region made up a databank of seroprevalence and molecular epidemiology of HHV-8 in Brazilian populations. The knowledge of serological and molecular techniques developed in this study may be used by Molecular Biology Laboratories at Unicamp and LASP/Fiocruz, allowing the implantation of HHV-8 diagnostic and genotyping techniques in these centers / Doutorado / Ciencias Biomedicas / Doutor em Ciências Médicas
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