title: Analysis of Dominant HIV Quasispecies Suggests Independent Viral Evolution Within Spinal Granulomas Coinfected with Mycobacterium tuberculosis and HIV-1 Subtype C.
abstract: Extrapulmonary tuberculosis (TB) is a significant public health challenge in South Africa and worldwide, largely fuelled by the HIV epidemic. In spinal TB, Mycobacteria infect the spinal column without dissemination to the spinal cord. The immune microenvironment, target cell characteristics, and other evolutionary forces within granulomas during HIV/TB coinfection are poorly characterized. We investigated whether spinal TB granulomas represent a sequestered anatomical site where independent HIV evolution occurs, and assessed the role of macrophages as a target cell for both HIV and mycobacteria. RNA was extracted from plasma and granulomatous tissue from six antiretroviral-naive HIV-1/spinal TB-coinfected patients, RT-PCR amplified, and the C2-V5 env segment was cloned and sequenced. Analysis of genetic diversity, phylogeny and coalescence patterns was performed on clonal sequences. To investigate their role in HIV sequestration, macrophages and the HIV-1 p24 protein were immune localized and ultrastructural features were studied. Intercompartment diversity measurements and phylogenetic reconstruction revealed anatomically distinct monophyletic HIV-1 clusters in four of six patients. Genotypic CCR5-tropic variants were predominant (98.9%) with conservation of putative N-linked glycosylation sites in both compartments. CD68(+) reactivity was associated with higher tissue viral load (r = 1.0; p < 0.01) but not greater intrapatient diversity (r = 0.60; p > 0.05). Ultrastructural imaging revealed the presence of bacterial and virus-like particles within membrane-bound intracellular compartments of macrophages. Spinal tuberculosis granulomas may form anatomically discreet sites of divergent viral evolution. Macrophages in these granulomas harbored both pathogens, suggesting that they may facilitate the process of viral sequestration within this compartment.
authors.affiliation: 1 Africa Centre for Health and Population Studies, Nelson R. Mandela School of Medicine, University of KwaZulu-Natal (UKZN) , Durban, South Africa .
authors.forename: Sivapragashini
authors.initials: S
authors.lastname: Danaviah
authors.affiliation: 1 Africa Centre for Health and Population Studies, Nelson R. Mandela School of Medicine, University of KwaZulu-Natal (UKZN) , Durban, South Africa .
authors.forename: Tulio
authors.initials: T
authors.lastname: de Oliveira
authors.affiliation: 2 Department of Virology, Nelson R. Mandela School of Medicine , UKZN, Durban, South Africa .|3 KwaZulu-Natal Research Institute for Tuberculosis and HIV (K-RITH) , UKZN, Durban, South Africa .
authors.forename: Michelle
authors.initials: M
authors.lastname: Gordon
authors.affiliation: 4 Department of Orthopedics, Nelson R. Mandela School of Medicine , UKZN, Durban, South Africa .
authors.forename: Shunmugam
authors.initials: S
authors.lastname: Govender
authors.affiliation: 5 School of Public Health, Sefako Makgatho Health Sciences University , Medunsa, South Africa .
authors.forename: Paul
authors.initials: P
authors.lastname: Chelule
authors.affiliation: 1 Africa Centre for Health and Population Studies, Nelson R. Mandela School of Medicine, University of KwaZulu-Natal (UKZN) , Durban, South Africa .
authors.forename: Sureshnee
authors.initials: S
authors.lastname: Pillay
authors.affiliation: 6 Optics and Imaging Centre, Nelson R. Mandela School of Medicine , UKZN, Durban, South Africa .
authors.forename: Thajasvarie
authors.initials: T
authors.lastname: Naicker
authors.affiliation: 7 MRC Inflammation and Immunity Unit, Department of Immunology, University of Pretoria , Pretoria, South Africa .
authors.forename: Sharon
authors.initials: S
authors.lastname: Cassol
authors.affiliation: 3 KwaZulu-Natal Research Institute for Tuberculosis and HIV (K-RITH) , UKZN, Durban, South Africa .|8 HIV Pathogenesis Programme, Doris Duke Medical Research Institute , Nelson R. Mandela School of Medicine, UKZN, Durban, South Africa .|9 Max Planck Institute for Infection Biology , Chariteplatz, Berlin, Germany .|10 The Ragon Institute of MGH, MIT and Harvard University , Cambridge, Massachusetts.
authors.forename: Thumbi
authors.initials: T
authors.lastname: Ndung'u
mesh_terms: D000328:Adult; D002675:Child, Preschool; D003001:Cloning, Molecular; D060085:Coinfection / Q000821:virology; D005260:Female; D014644:Genetic Variation; D005838:Genotype; D006099:Granuloma / Q000821:virology*; D015658:HIV Infections / Q000150:complications* / Q000821:virology*; D015497:HIV-1 / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D008297:Male; D010802:Phylogeny; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D013019:South Africa; D014399:Tuberculosis, Spinal / Q000150:complications*; D054299:env Gene Products, Human Immunodeficiency Virus / Q000235:genetics
chemical_list: D012367:RNA, Viral; D054299:env Gene Products, Human Immunodeficiency Virus