Publications by authors named "Teiichiro Shiino"

28 Publications

  • Page 1 of 1

Human Leukocyte Antigen-Associated HIV-1 CRF02_AG gag and vif Polymorphisms in Ghana.

Jpn J Infect Dis 2019 Nov 28;72(6):374-380. Epub 2019 Jun 28.

Joint Research Center for Human Retrovirus Infection, Kumamoto University.

In human immunodeficiency virus type-1 (HIV-1) infections, cytotoxic T-lymphocyte (CTL) responses targeting human leukocyte antigen (HLA)-restricted viral epitopes exert strong suppressive pressure on viral replication and frequently select for mutations resulting in viral escape from CTL recognition. Numerous data on these HLA-associated mutations in HIV-1 subtypes B and C have been amassed with few reports described in other subtypes. In the present study, we investigated the HLA-associated mutations in HIV-1 subtype CRF02_AG prevailing in Ghana, Western Africa. We determined viral gag sequences in 246 out of 324 HIV-1-infected Ghanaians. Phylogeny analysis revealed that 200 (81.3%) individuals were infected with HIV-1 CRF02_AG. Full gag and vif sequences were obtained from 199 and 138, respectively, out of the 200 individuals infected with CRF02_AG and subjected to determination of HLA-associated mutations. The analysis found HLA-associated HIV-1 CRF02_AG non-synonymous polymorphisms at 19 sites; 13 in gag and six in vif, including those that were newly determined. Generation of this data is an important contribution to our understanding of HIV-1 CRF02_AG and host T cell interaction.
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http://dx.doi.org/10.7883/yoken.JJID.2019.201DOI Listing
November 2019

Human leukocyte antigen-associated gag and nef polymorphisms in HIV-1 subtype A/E-infected individuals in Vietnam.

Microbes Infect 2019 03 30;21(2):113-118. Epub 2018 Oct 30.

Center of BioMedical Research, National Institute of Hygiene and Epidemiology, No.1 Yersin Street, Hanoi, Viet Nam. Electronic address:

Numbers of HLA-associated polymorphisms have been reported on HIV-1 subtypes B and C, but few on other subtypes. Here, we analyzed HLA-associated gag and nef polymorphisms in HIV-1 subtype A/E prevalent in Vietnam. We determined HLA-A, B and C genotypes in 179 HIV-1-infected Vietnamese by next generation sequencing and analyzed proviral genome sequences in 144 of them, showing that 142 of the 144 were subtype A/E. Analysis revealed HLA-associated subtype A/E gag and nef polymorphisms at nineteen residues including those newly determined. Accumulation of these data would contribute to our understanding of HIV-1 subtype A/E and host immune interaction.
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http://dx.doi.org/10.1016/j.micinf.2018.10.001DOI Listing
March 2019

In vivo virulence of MHC-adapted AIDS virus serially-passaged through MHC-mismatched hosts.

PLoS Pathog 2017 Sep 20;13(9):e1006638. Epub 2017 Sep 20.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan.

CD8+ T-cell responses exert strong suppressive pressure on HIV replication and select for viral escape mutations. Some of these major histocompatibility complex class I (MHC-I)-associated mutations result in reduction of in vitro viral replicative capacity. While these mutations can revert after viral transmission to MHC-I-disparate hosts, recent studies have suggested that these MHC-I-associated mutations accumulate in populations and make viruses less pathogenic in vitro. Here, we directly show an increase in the in vivo virulence of an MHC-I-adapted virus serially-passaged through MHC-I-mismatched hosts in a macaque AIDS model despite a reduction in in vitro viral fitness. The first passage simian immunodeficiency virus (1pSIV) obtained 1 year after SIVmac239 infection in a macaque possessing a protective MHC-I haplotype 90-120-Ia was transmitted into 90-120-Ia- macaques, whose plasma 1 year post-infection was transmitted into other 90-120-Ia- macaques to obtain the third passage SIV (3pSIV). Most of the 90-120-Ia-associated mutations selected in 1pSIV did not revert even in 3pSIV. 3pSIV showed lower in vitro viral fitness but induced persistent viremia in 90-120-Ia- macaques. Remarkably, 3pSIV infection in 90-120-Ia+ macaques resulted in significantly higher viral loads and reduced survival compared to wild-type SIVmac239. These results indicate that MHC-I-adapted SIVs serially-transmitted through MHC-I-mismatched hosts can have higher virulence in MHC-I-matched hosts despite their lower in vitro viral fitness. This study suggests that multiply-passaged HIVs could result in loss of HIV-specific CD8+ T cell responses in human populations and the in vivo pathogenic potential of these escaped viruses may be enhanced.
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http://dx.doi.org/10.1371/journal.ppat.1006638DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5624644PMC
September 2017

Simian Immunodeficiency Virus Targeting of CXCR3 CD4 T Cells in Secondary Lymphoid Organs Is Associated with Robust CXCL10 Expression in Monocyte/Macrophage Subsets.

J Virol 2017 07 9;91(13). Epub 2017 Jun 9.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan

Glycosylation of Env defines pathogenic properties of simian immunodeficiency virus (SIV). We previously demonstrated that pathogenic SIVmac239 and a live-attenuated, quintuple deglycosylated Env mutant (Δ5G) virus target CD4 T cells residing in different tissues during acute infection. SIVmac239 and Δ5G preferentially infected distinct CD4 T cells in secondary lymphoid organs (SLOs) and within the lamina propria of the small intestine, respectively (C. Sugimoto et al., J Virol 86:9323-9336, 2012, https://doi.org/10.1128/JVI.00948-12). Here, we studied the host responses relevant to SIV targeting of CXCR3 CCR5 CD4 T cells in SLOs. Genome-wide transcriptome analyses revealed that Th1-polarized inflammatory responses, defined by expression of CXCR3 chemokines, were distinctly induced in the SIVmac239-infected animals. Consistent with robust expression of CXCL10, CXCR3 T cells were depleted from blood in the SIVmac239-infected animals. We also discovered that elevation of CXCL10 expression in blood and SLOs was secondary to the induction of CD14 CD16 monocytes and MAC387 macrophages, respectively. Since the significantly higher levels of SIV infection in SLOs occurred with a massive accumulation of infiltrated MAC387 macrophages, T cells, dendritic cells (DCs), and residential macrophages near high endothelial venules, the results highlight critical roles of innate/inflammatory responses in SIVmac239 infection. Restricted infection in SLOs by Δ5G also suggests that glycosylation of Env modulates innate/inflammatory responses elicited by cells of monocyte/macrophage/DC lineages. We previously demonstrated that a pathogenic SIVmac239 virus and a live-attenuated, deglycosylated mutant Δ5G virus infected distinct CD4 T cell subsets in SLOs and the small intestine, respectively (C. Sugimoto et al., J Virol 86:9323-9336, 2012, https://doi.org/10.1128/JVI.00948-12). Accordingly, infections with SIVmac239, but not with Δ5G, deplete CXCR3 CCR5 CD4 T (Th1) cells during the primary infection, thereby compromising the cellular immune response. Thus, we hypothesized that distinct host responses are elicited by the infections with two different viruses. We found that SIVmac239 induced distinctly higher levels of inflammatory Th1 responses than Δ5G. In particular, SIVmac239 infection elicited robust expression of CXCL10, a chemokine for CXCR3 cells, in CD14 CD16 monocytes and MAC387 macrophages recently infiltrated in SLOs. In contrast, Δ5G infection elicited only modest inflammatory responses. These results suggest that the glycosylation of Env modulates the inflammatory/Th1 responses through the monocyte/macrophage subsets and elicits marked differences in SIV infection and clinical outcomes.
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http://dx.doi.org/10.1128/JVI.00439-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5469254PMC
July 2017

Association of lymph-node antigens with lower Gag-specific central-memory and higher Env-specific effector-memory CD8(+) T-cell frequencies in a macaque AIDS model.

Sci Rep 2016 07 25;6:30153. Epub 2016 Jul 25.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo 162-8640, Japan.

Virus-specific CD8(+) T cells exert strong suppressive pressure on human/simian immunodeficiency virus (HIV/SIV) replication. These responses have been intensively examined in peripheral blood mononuclear cells (PBMCs) but not fully analyzed in lymph nodes (LNs), where interaction between CD8(+) T cells and HIV/SIV-infected cells occurs. Here, we investigated target antigen specificity of CD8(+) T cells in LNs in a macaque AIDS model. Analysis of virus antigen-specific CD8(+) T-cell responses in the inguinal LNs obtained from twenty rhesus macaques in the chronic phase of SIV infection showed an inverse correlation between viral loads and frequencies of CD8(+) T cells with CD28(+) CD95(+) central memory phenotype targeting the N-terminal half of SIV core antigen (Gag-N). In contrast, analysis of LNs but not PBMCs revealed a positive correlation between viral loads and frequencies of CD8(+) T cells with CD28(-)CD95(+) effector memory phenotype targeting the N-terminal half of SIV envelope (Env-N), soluble antigen. Indeed, LNs with detectable SIV capsid p27 antigen in the germinal center exhibited significantly lower Gag-N-specific CD28(+) CD95(+) CD8(+) T-cell and higher Env-N-specific CD28(-)CD95(+) CD8(+) T-cell responses than those without detectable p27. These results imply that core and envelope antigen-specific CD8(+) T cells show different patterns of interactions with HIV/SIV-infected cells.
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http://dx.doi.org/10.1038/srep30153DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4958968PMC
July 2016

HIV-1 CRF01_AE and Subtype B Transmission Networks Crossover: A New AE/B Recombinant Identified in Japan.

AIDS Res Hum Retroviruses 2016 May 16;32(5):412-9. Epub 2015 Nov 16.

1 Clinical Research Center, National Hospital Organization Nagoya Medical Center , Nagoya, Japan .

The major circulating HIV-1 strains in Japan have been subtype B (B) followed by CRF01_AE (AE) in newly diagnosed HIV/AIDS cases. These two subtypes have distinct epidemiological characteristics; B predominates in men who have sex with men, while AE is observed mostly in heterosexuals engaging in high-risk sex. However, transmission networks of these two high-risk populations appear to be crossing over and diffusing. Here we report the emergence of previously unidentified HIV-1 AE/B recombinants in Japan. We initially identified 13 cases with discordant subtyping results with AE (gag MA)/B (pol PR-RT)/AE (env C2V3) by molecular phylogenetic analysis of 1,070 cases who visited Nagoya Medical Center from 1997 to 2012. Genetic characterization of full-length sequences demonstrated that they shared an identical recombinant structure, and was designated as CRF69_01B by the Los Alamos HIV National Laboratory. By reviewing gag, pol, and env sequences collected in the Japanese Drug Resistance HIV-1 Surveillance Network, we found five other CRF69_01B probable cases from different areas in Japan, suggesting that the strain is transmitted widely throughout the country. The time of the most recent common ancestor analyses estimated that CRF69_01B emerged between 1991 and 1995, soon after AE was introduced from neighboring countries in the mid-1990s. Understanding the current epidemic strains is important for the diagnosis and treatment of HIV/AIDS, as well as for the development of globally effective HIV vaccines.
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http://dx.doi.org/10.1089/aid.2015.0192DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4845645PMC
May 2016

Characteristics of Transmitted Drug-Resistant HIV-1 in Recently Infected Treatment-Naive Patients in Japan.

J Acquir Immune Defic Syndr 2016 Apr;71(4):367-73

*Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Japan; †Currently, HIV Dynamics and Replication Program, National Cancer Institute, National Institutes of Health, MD; ‡Infectious Disease Surveillance Center, National Institute of Infectious Diseases, Tokyo, Japan; §National Center for Global Health and Medicine, Tokyo, Japan; ‖Osaka Prefectural Institute of Public Health, Osaka, Japan; ¶National Hospital Organization Kyushu Medical Center, Fukuoka, Japan; #Saitama Institute of Public Health; **Tokyo Metropolitan Institute of Public Health, Tokyo, Japan; ††Kanagawa Prefectural Institute of Public Health, Chigasaki, Japan; ‡‡Division of Basic Medicine, Nagoya University Graduate School of Medicine, Nagoya, Japan; and §§Currently, GlaxoSmithKline KK, Tokyo, Japan.

Objectives: Progress in antiretroviral treatment has led to fewer virological failure cases, but 10%-20% of treatment-naive HIV/AIDS cases are reported to harbor drug-resistant strains, suggesting transmission of drug-resistant HIV. We aimed to determine the trend in prevalence of transmitted drug-resistant (TDR) HIV in Japan, particularly in recently infected patients.

Methods: Drug-resistance test was performed on 3904 HIV-1-infected cases newly diagnosed between 2007 and 2012. The number of cases infected within 6 months [recent seroconverters (RS)] was estimated by BED assay of 2700 plasma samples. Characteristics of RS cases were further analyzed.

Results: The overall prevalence of TDR was 9.1%, ranging from 7.3% in 2008% to 12.5% in 2010. Among 1403 subtype B/E/D cases with >50 CD4 T cell counts and >1000 HIV copies per milliliter, 468 (33.4%) were estimated to be RS. The prevalence of RS was significantly higher among cases who were male, Japanese, and men who have sex with men. The prevalence of TDR did not differ significantly between recent and long-term seroconverters (8.5% vs. 9.2%, respectively, P = 0.68). Common mutations in both groups were M46I/L and T215 revertants. Furthermore, sequences with these mutations, K103N and D30N/N88D formed clusters on phylogenetic trees.

Conclusion: Our study clarified an increase in prevalence of TDR in Japan from 2007 to 2012. The phylogenetic clustering of cases with M46I/L or T215 revertants suggests that HIV with these mutations have become circulating strains. Furthermore, detailed analyses showed that Japanese men who have sex with men are more aware of their risk of HIV infection.
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http://dx.doi.org/10.1097/QAI.0000000000000861DOI Listing
April 2016

Longitudinal Detection and Persistence of Minority Drug-Resistant Populations and Their Effect on Salvage Therapy.

PLoS One 2015 11;10(9):e0135941. Epub 2015 Sep 11.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan; Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Japan; Department of AIDS Research, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Background: Drug-resistant HIV are more prevalent and persist longer than previously demonstrated by bulk sequencing due to the ability to detect low-frequency variants. To clarify a clinical benefit to monitoring minority-level drug resistance populations as a guide to select active drugs for salvage therapy, we retrospectively analyzed the dynamics of low-frequency drug-resistant population in antiretroviral (ARV)-exposed drug resistant individuals.

Materials And Methods: Six HIV-infected individuals treated with ARV for more than five years were analyzed. These individuals had difficulty in controlling viremia, and treatment regimens were switched multiple times guided by standard drug resistance testing using bulk sequencing. To detect minority variant populations with drug resistance, we used a highly sensitive allele-specific PCR (AS-PCR) with detection thresholds of 0.3-2%. According to ARV used in these individuals, we focused on the following seven reverse transcriptase inhibitor-resistant mutations: M41L, K65R, K70R, K103N, Y181C, M184V, and T215F/Y. Results of AS-PCR were compared with bulk sequencing data for concordance and presence of additional mutations. To clarify the genetic relationship between low-frequency and high-frequency populations, AS-PCR amplicon sequences were compared with bulk sequences in phylogenetic analysis.

Results: The use of AS-PCR enabled detection of the drug-resistant mutations, M41L, K103N, Y181C, M184V and T215Y, present as low-frequency populations in five of the six individuals. These drug resistant variants persisted for several years without ARV pressure. Phylogenetic analysis indicated that pre-existing K103N and T215I variants had close genetic relationships with high-frequency K103N and T215I observed during treatment.

Discussion And Conclusion: Our results demonstrate the long-term persistence of drug-resistant viruses in the absence of drug pressure. The rapid virologic failures with pre-existing mutant viruses detectable by AS-PCR highlight the clinical importance of low-frequency drug-resistant viruses. Thus, our results highlight the usefulness of AS-PCR and support its expanded evaluation in ART clinical management.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0135941PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4567277PMC
May 2016

High proportion of HIV serodiscordance among HIV-affected married couples in northern Vietnam.

PLoS One 2015 21;10(4):e0125299. Epub 2015 Apr 21.

AIDS Clinical Center, National Center of Global Health and Medicine, Tokyo, Japan.

Introduction: Little is known about the state of HIV transmission among married couples in Vietnam. This study aims to clarify HIV serostatus in this group and elucidate risk factors for intra-marital HIV transmission.

Methods: In 2012, we enrolled a group of HIV-positive married men registered at the HIV outpatient clinic of a referral hospital in northern Vietnam, along with their wives. Sociodemographic, behavioural and clinical data were collected from men and wives. HIV serodiscordant couples were followed until March 2014 to determine seroconversion rate. A phylogenetic analysis was performed based on env V3 sequence to detail cluster formation among men.

Results: Of the 163 HIV-positive men enrolled in the study, 101 (62.0%) had wives testing HIV-negative. Half of men reported injecting drug use (IDU) as a likely transmission route. Couples reported a high incidence of unprotected sexual intercourse prior to diagnosis; the median (inter quartile range) was 4 (4-8) times per month. Only 17 couples (10.4%) reported using condoms during at least half these instances. Multivariable analysis revealed IDU history among men was independently associated with HIV-negative wives (adjusted OR 0.31; 95% CI 0.10-0.95, p=0.041). Phylogenetic analysis of 80 samples indicated CRF01_AE. Of these, 69 (86.3%) clustered with IDU-associated viruses from Vietnam. No HIV seroconversion was identified during a follow-up of 61 serodiscordant couples, with 126.5 person-years of observation during which HIV-infected men were on antiretroviral drug therapy (ART).

Conclusion: High HIV serodiscordance was observed among HIV-affected married couples in northern Vietnam. A large number of at-risk wives therefore remain HIV-negative and can be protected with measures including proper use of ART if couples are made aware of the serodiscordance through screening.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0125299PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4405585PMC
March 2016

Development and customization of a color-coded microbeads-based assay for drug resistance in HIV-1 reverse transcriptase.

PLoS One 2014 14;9(10):e109823. Epub 2014 Oct 14.

Division of Infectious Diseases, Advanced Clinical Research Center, the Institute of Medical Science, the University of Tokyo, Tokyo, Japan; Department of Infectious Disease Control, International Research Center for Infectious Diseases, the Institute of Medical Science, the University of Tokyo, Tokyo, Japan.

Background: Drug resistance (DR) of HIV-1 can be examined genotypically or phenotypically. Although sequencing is the gold standard of the genotypic resistance testing (GRT), high-throughput GRT targeted to the codons responsible for DR may be more appropriate for epidemiological studies and public health research.

Methods: We used a Japanese database to design and synthesize sequence-specific oligonucleotide probes (SSOP) for the detection of wild-type sequences and 6 DR mutations in the clade B HIV-1 reverse transcriptase region. We coupled SSOP to microbeads of the Luminex 100 xMAP system and developed a GRT based on the polymerase chain reaction (PCR)-SSOP-Luminex method.

Results: Sixteen oligoprobes for discriminating DR mutations from wild-type sequences at 6 loci were designed and synthesized, and their sensitivity and specificity were confirmed using isogenic plasmids. The PCR-SSOP-Luminex DR assay was then compared to direct sequencing using 74 plasma specimens from treatment-naïve patients or those on failing treatment. In the majority of specimens, the results of the PCR-SSOP-Luminex DR assay were concordant with sequencing results: 62/74 (83.8%) for M41, 43/74 (58.1%) for K65, 70/74 (94.6%) for K70, 55/73 (75.3%) for K103, 63/73 (86.3%) for M184 and 68/73 (93.2%) for T215. There were a number of specimens without any positive signals, especially for K65. The nucleotide position of A2723G, A2747G and C2750T were frequent polymorphisms for the wild-type amino acids K65, K66 and D67, respectively, and 14 specimens had the D67N mutation encoded by G2748A. We synthesized 14 additional oligoprobes for K65, and the sensitivity for K65 loci improved from 43/74 (58.1%) to 68/74 (91.9%).

Conclusions: We developed a rapid high-throughput assay for clade B HIV-1 DR mutations, which could be customized by synthesizing oligoprobes suitable for the circulating viruses. The assay could be a useful tool especially for public health research in both resource-rich and resource-limited settings.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0109823PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4196989PMC
June 2015

Vaccine-induced CD107a+ CD4+ T cells are resistant to depletion following AIDS virus infection.

J Virol 2014 Dec 1;88(24):14232-40. Epub 2014 Oct 1.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan The Institute of Medical Science, The University of Tokyo, Tokyo, Japan

Unlabelled: CD4(+) T-cell responses are crucial for effective antibody and CD8(+) T-cell induction following virus infection. However, virus-specific CD4(+) T cells can be preferential targets for human immunodeficiency virus (HIV) infection. HIV-specific CD4(+) T-cell induction by vaccination may thus result in enhancement of virus replication following infection. In the present study, we show that vaccine-elicited CD4(+) T cells expressing CD107a are relatively resistant to depletion in a macaque AIDS model. Comparison of virus-specific CD107a, macrophage inflammatory protein-1β, gamma interferon, tumor necrosis factor alpha, and interleukin-2 responses in CD4(+) T cells of vaccinated macaques prechallenge and 1 week postchallenge showed a significant reduction in the CD107a(-) but not the CD107a(+) subset after virus exposure. Those vaccinees that failed to control viremia showed a more marked reduction and exhibited significantly higher viral loads at week 1 than unvaccinated animals. Our results indicate that vaccine-induced CD107a(-) CD4(+) T cells are depleted following virus infection, suggesting a rationale for avoiding virus-specific CD107a(-) CD4(+) T-cell induction in HIV vaccine design.

Importance: Induction of effective antibody and/or CD8(+) T-cell responses is a principal vaccine strategy against human immunodeficiency virus (HIV) infection. CD4(+) T-cell responses are crucial for effective antibody and CD8(+) T-cell induction. However, virus-specific CD4(+) T cells can be preferential targets for HIV infection. Here, we show that vaccine-induced virus-specific CD107a(-) CD4(+) T cells are largely depleted following infection in a macaque AIDS model. While CD4(+) T-cell responses are important in viral control, our results indicate that virus-specific CD107a(-) CD4(+) T-cell induction by vaccination may not lead to efficient CD4(+) T-cell responses following infection but rather be detrimental and accelerate viral replication in the acute phase. This suggests that HIV vaccine design should avoid virus-specific CD107a(-) CD4(+) T-cell induction. Conversely, this study found that vaccine-induced CD107a(+) CD4(+) T cells are relatively resistant to depletion following virus challenge, implying that induction of these cells may be an alternative approach toward HIV control.
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http://dx.doi.org/10.1128/JVI.02032-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4249136PMC
December 2014

Phylodynamic analysis reveals CRF01_AE dissemination between Japan and neighboring Asian countries and the role of intravenous drug use in transmission.

PLoS One 2014 15;9(7):e102633. Epub 2014 Jul 15.

Department of Infectious Diseases and Immunology, Clinical Research Center, Nagoya Medical Center, Nagoya, Japan; Department of AIDS Research, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Background: One major circulating HIV-1 subtype in Southeast Asian countries is CRF01_AE, but little is known about its epidemiology in Japan. We conducted a molecular phylodynamic study of patients newly diagnosed with CRF01_AE from 2003 to 2010.

Methods: Plasma samples from patients registered in Japanese Drug Resistance HIV-1 Surveillance Network were analyzed for protease-reverse transcriptase sequences; all sequences undergo subtyping and phylogenetic analysis using distance-matrix-based, maximum likelihood and Bayesian coalescent Markov Chain Monte Carlo (MCMC) phylogenetic inferences. Transmission clusters were identified using interior branch test and depth-first searches for sub-tree partitions. Times of most recent common ancestor (tMRCAs) of significant clusters were estimated using Bayesian MCMC analysis.

Results: Among 3618 patient registered in our network, 243 were infected with CRF01_AE. The majority of individuals with CRF01_AE were Japanese, predominantly male, and reported heterosexual contact as their risk factor. We found 5 large clusters with ≥5 members and 25 small clusters consisting of pairs of individuals with highly related CRF01_AE strains. The earliest cluster showed a tMRCA of 1996, and consisted of individuals with their known risk as heterosexual contacts. The other four large clusters showed later tMRCAs between 2000 and 2002 with members including intravenous drug users (IVDU) and non-Japanese, but not men who have sex with men (MSM). In contrast, small clusters included a high frequency of individuals reporting MSM risk factors. Phylogenetic analysis also showed that some individuals infected with HIV strains spread in East and South-eastern Asian countries.

Conclusions: Introduction of CRF01_AE viruses into Japan is estimated to have occurred in the 1990s. CFR01_AE spread via heterosexual behavior, then among persons connected with non-Japanese, IVDU, and MSM. Phylogenetic analysis demonstrated that some viral variants are largely restricted to Japan, while others have a broad geographic distribution.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0102633PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4099140PMC
October 2015

Highly-sensitive allele-specific PCR testing identifies a greater prevalence of transmitted HIV drug resistance in Japan.

PLoS One 2013 16;8(12):e83150. Epub 2013 Dec 16.

AIDS Research Center, National Institute of Infectious Diseases (NIID), Tokyo, Japan ; Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Japan ; Department of AIDS Research, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Background: The transmission of drug-resistant HIV in newly identified infected populations has become an underlying epidemic which can be better assessed with sensitive resistance testing. Since minority drug resistant variants cannot be detected by bulk sequencing, methods with improved sensitivity are required. Thus, the goal of this study was to evaluate if transmitted drug resistance mutations at minority levels in Japanese patients could be identified using highly sensitive allele-specific PCR (AS-PCR).

Materials And Methods: Samples were taken from newly diagnosed HIV/AIDS cases at the National Nagoya Hospital from January 2008 to December 2009. All samples were bulk sequenced for HIV protease and reverse transcriptase. To detect minority populations with drug resistance, we used AS-PCR with mutation-specific primers designed for seven reverse transcriptase inhibitor resistance mutations, M41L, K65R, K70R, K103N, Y181C, M184V, and T215F/Y, and for three protease inhibitor resistance mutations, M46I/L and L90M.

Results: We studied 149 newly identified HIV cases. Bulk sequencing detected 8 cases with NRTI resistance mutations (one with A62V, one D67E, one T215D, one T215E, two with T215L and two T215S) and 15 with PI resistance mutations (one with N88D and 14 with M46I). Results obtained by AS-PCR and bulk sequencing demonstrated good concordance but the AS-PCR enabled the detection of seven additional drug-resistant cases (one M41L, two with K65R, two with K70R, and one M184V) in the RT region. Additionally, AS-PCR assays identified 15 additional cases with M46I, five with M46L and four cases with L90M in the protease region.

Conclusions: Using AS-PCR substantially increased the detection of transmitted drug resistance in this population from 15.4% to 26.8%, further supporting the benefit of sensitive testing among drug-naïve populations. Since the clinical impact of minority drug-resistant populations is not fully comprehended for all mutations, follow-up studies are needed to understand their significance for treatment.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0083150PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3865156PMC
October 2014

A novel protective MHC-I haplotype not associated with dominant Gag-specific CD8+ T-cell responses in SIVmac239 infection of Burmese rhesus macaques.

PLoS One 2013 14;8(1):e54300. Epub 2013 Jan 14.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan.

Several major histocompatibility complex class I (MHC-I) alleles are associated with lower viral loads and slower disease progression in human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) infections. Immune-correlates analyses in these MHC-I-related HIV/SIV controllers would lead to elucidation of the mechanism for viral control. Viral control associated with some protective MHC-I alleles is attributed to CD8+ T-cell responses targeting Gag epitopes. We have been trying to know the mechanism of SIV control in multiple groups of Burmese rhesus macaques sharing MHC-I genotypes at the haplotype level. Here, we found a protective MHC-I haplotype, 90-010-Id (D), which is not associated with dominant Gag-specific CD8+ T-cell responses. Viral loads in five D+ animals became significantly lower than those in our previous cohorts after 6 months. Most D+ animals showed predominant Nef-specific but not Gag-specific CD8+ T-cell responses after SIV challenge. Further analyses suggested two Nef-epitope-specific CD8+ T-cell responses exerting strong suppressive pressure on SIV replication. Another set of five D+ animals that received a prophylactic vaccine using a Gag-expressing Sendai virus vector showed significantly reduced viral loads compared to unvaccinated D+ animals at 3 months, suggesting rapid SIV control by Gag-specific CD8+ T-cell responses in addition to Nef-specific ones. These results present a pattern of SIV control with involvement of non-Gag antigen-specific CD8+ T-cell responses.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0054300PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3544795PMC
August 2013

Phylodynamic analysis of a viral infection network.

Authors:
Teiichiro Shiino

Front Microbiol 2012 31;3:278. Epub 2012 Jul 31.

Infectious Diseases Surveillance Center, National Institute of Infectious Diseases Tokyo, Japan.

Viral infections by sexual and droplet transmission routes typically spread through a complex host-to-host contact network. Clarifying the transmission network and epidemiological parameters affecting the variations and dynamics of a specific pathogen is a major issue in the control of infectious diseases. However, conventional methods such as interview and/or classical phylogenetic analysis of viral gene sequences have inherent limitations and often fail to detect infectious clusters and transmission connections. Recent improvements in computational environments now permit the analysis of large datasets. In addition, novel analytical methods have been developed that serve to infer the evolutionary dynamics of virus genetic diversity using sample date information and sequence data. This type of framework, termed "phylodynamics," helps connect some of the missing links on viral transmission networks, which are often hard to detect by conventional methods of epidemiology. With sufficient number of sequences available, one can use this new inference method to estimate theoretical epidemiological parameters such as temporal distributions of the primary infection, fluctuation of the pathogen population size, basic reproductive number, and the mean time span of disease infectiousness. Transmission networks estimated by this framework often have the properties of a scale-free network, which are characteristic of infectious and social communication processes. Network analysis based on phylodynamics has alluded to various suggestions concerning the infection dynamics associated with a given community and/or risk behavior. In this review, I will summarize the current methods available for identifying the transmission network using phylogeny, and present an argument on the possibilities of applying the scale-free properties to these existing frameworks.
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http://dx.doi.org/10.3389/fmicb.2012.00278DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3441063PMC
October 2012

Association of major histocompatibility complex class I haplotypes with disease progression after simian immunodeficiency virus challenge in burmese rhesus macaques.

J Virol 2012 Jun 4;86(12):6481-90. Epub 2012 Apr 4.

AIDS Research Center, National Institute of Infectious Diseases, Toyama, Shinjuku-ku, Tokyo, Japan.

Nonhuman primate AIDS models are essential for the analysis of AIDS pathogenesis and the evaluation of vaccine efficacy. Multiple studies on human immunodeficiency virus and simian immunodeficiency virus (SIV) infection have indicated the association of major histocompatibility complex class I (MHC-I) genotypes with rapid or slow AIDS progression. The accumulation of macaque groups that share not only a single MHC-I allele but also an MHC-I haplotype consisting of multiple polymorphic MHC-I loci would greatly contribute to the progress of AIDS research. Here, we investigated SIVmac239 infections in four groups of Burmese rhesus macaques sharing individual MHC-I haplotypes, referred to as A, E, B, and J. Out of 20 macaques belonging to A(+) (n = 6), E(+) (n = 6), B(+) (n = 4), and J(+) (n = 4) groups, 18 showed persistent viremia. Fifteen of them developed AIDS in 0.5 to 4 years, with the remaining three at 1 or 2 years under observation. A(+) animals, including two controllers, showed slower disease progression, whereas J(+) animals exhibited rapid progression. E(+) and B(+) animals showed intermediate plasma viral loads and survival periods. Gag-specific CD8(+) T-cell responses were efficiently induced in A(+) animals, while Nef-specific CD8(+) T-cell responses were in A(+), E(+), and B(+) animals. Multiple comparisons among these groups revealed significant differences in survival periods, peripheral CD4(+) T-cell decline, and SIV-specific CD4(+) T-cell polyfunctionality in the chronic phase. This study indicates the association of MHC-I haplotypes with AIDS progression and presents an AIDS model facilitating the analysis of virus-host immune interaction.
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http://dx.doi.org/10.1128/JVI.07077-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3393527PMC
June 2012

Impact of vaccination on cytotoxic T lymphocyte immunodominance and cooperation against simian immunodeficiency virus replication in rhesus macaques.

J Virol 2012 Jan 9;86(2):738-45. Epub 2011 Nov 9.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan.

Cytotoxic T lymphocyte (CTL) responses play a central role in viral suppression in human immunodeficiency virus (HIV) infections. Prophylactic vaccination resulting in effective CTL responses after viral exposure would contribute to HIV control. It is important to know how CTL memory induction by vaccination affects postexposure CTL responses. We previously showed vaccine-based control of a simian immunodeficiency virus (SIV) challenge in a group of Burmese rhesus macaques sharing a major histocompatibility complex class I haplotype. Gag(206-216) and Gag(241-249) epitope-specific CTL responses were responsible for this control. In the present study, we show the impact of individual epitope-specific CTL induction by prophylactic vaccination on postexposure CTL responses. In the acute phase after SIV challenge, dominant Gag(206-216)-specific CTL responses with delayed, naive-derived Gag(241-249)-specific CTL induction were observed in Gag(206-216) epitope-vaccinated animals with prophylactic induction of single Gag(206-216) epitope-specific CTL memory, and vice versa in Gag(241-249) epitope-vaccinated animals with single Gag(241-249) epitope-specific CTL induction. Animals with Gag(206-216)-specific CTL induction by vaccination selected for a Gag(206-216)-specific CTL escape mutation by week 5 and showed significantly less decline of plasma viral loads from week 3 to week 5 than in Gag(241-249) epitope-vaccinated animals without escape mutations. Our results present evidence indicating significant influence of prophylactic vaccination on postexposure CTL immunodominance and cooperation of vaccine antigen-specific and non-vaccine antigen-specific CTL responses, which affects virus control. These findings provide great insights into antigen design for CTL-inducing AIDS vaccines.
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http://dx.doi.org/10.1128/JVI.06226-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3255834PMC
January 2012

Identification of a current hot spot of HIV type 1 transmission in Mongolia by molecular epidemiological analysis.

AIDS Res Hum Retroviruses 2011 Oct 25;27(10):1073-80. Epub 2011 May 25.

AIDS Clinical Center, National Center for Global Health and Medicine, Tokyo, Japan.

We investigated the current molecular epidemiological status of HIV-1 in Mongolia, a country with very low incidence of HIV-1 though with rapid expansion in recent years. HIV-1 pol (1065 nt) and env (447 nt) genes were sequenced to construct phylogenetic trees. The evolutionary rates, molecular clock phylogenies, and other evolutionary parameters were estimated from heterochronous genomic sequences of HIV-1 subtype B by the Bayesian Markov chain Monte Carlo method. We obtained 41 sera from 56 reported HIV-1-positive cases as of May 2009. The main route of infection was men who have sex with men (MSM). Dominant subtypes were subtype B in 32 cases (78%) followed by subtype CRF02_AG (9.8%). The phylogenetic analysis of the pol gene identified two clusters in subtype B sequences. Cluster 1 consisted of 21 cases including MSM and other routes of infection, and cluster 2 consisted of eight MSM cases. The tree analyses demonstrated very short branch lengths in cluster 1, suggesting a surprisingly active expansion of HIV-1 transmission during a short period with the same ancestor virus. Evolutionary analysis indicated that the outbreak started around the early 2000s. This study identified a current hot spot of HIV-1 transmission and potential seed of the epidemic in Mongolia. Comprehensive preventive measures targeting this group are urgently needed.
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http://dx.doi.org/10.1089/AID.2010.0196DOI Listing
October 2011

Outbreak of infections by hepatitis B virus genotype A and transmission of genetic drug resistance in patients coinfected with HIV-1 in Japan.

J Clin Microbiol 2011 Mar 19;49(3):1017-24. Epub 2011 Jan 19.

Department of Infection and Immunology, Clinical Research Center, Nagoya Medical Center, 4-1-1 Sannomaru, Nakaku, Nagoya 4600001, Japan.

The major routes of hepatitis B virus (HBV) infection in Japan has been mother-to-child transmission (MTCT) and blood transfusion. However, HBV cases transmitted through sexual contact are increasing, especially among HIV-1-seropositive patients. To understand the molecular epidemiology of HBV in HBV/HIV-1 coinfection, we analyzed HBV genotypes and HIV-1 subtypes in HBV/HIV-1-coinfected patients at Nagoya Medical Center from 2003 to 2007. Among 394 HIV-1-infected Japanese men having sex with men (MSM) who were newly diagnosed during the study period, 31 (7.9%) tested positive for the hepatitis B virus surface antigen. HBV sequence analyses were successful in 26 cases, with 21 (80.7%) and 5 (19.3%) cases determined as genotypes A and C, respectively. Our finding that HBV genotype A was dominant in HIV-1-seropositive patients alerts clinicians to an alternative outbreak of HBV genotype A in the HIV-1-infected MSM population and a shift in HBV genotype from C to A in Japan. The narrow genetic diversity in genotype A cases suggests that genotype A has been recently introduced into the MSM population and that sexual contacts among MSM were more active than speculated from HIV-1 tree analyses. In addition, we found a lamivudine resistance mutation in one naïve case, suggesting a risk of drug-resistant HBV transmission. As genotype A infection has a higher risk than infection with other genotypes for individuals to become HBV carriers, prevention programs are urgently needed for the target population.
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http://dx.doi.org/10.1128/JCM.02149-10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3067677PMC
March 2011

Trends in transmitted drug-resistant HIV-1 and demographic characteristics of newly diagnosed patients: nationwide surveillance from 2003 to 2008 in Japan.

Antiviral Res 2010 Oct 6;88(1):72-9. Epub 2010 Aug 6.

National Hospital Organization, Nagoya Medical Center, Clinical Research Center, 4-1-1 Sannomaru, Naka-ku, Nagoya 4600001, Japan.

The emergence and transmission of drug-resistant human immunodeficiency virus-1 (HIV-1) compromises antiretroviral treatment for HIV-1. Thus, testing for drug resistance is recommended at diagnosis and before initiating highly active antiretroviral treatment. We conducted an epidemiological study enrolling newly diagnosed patients between 2003 and 2008 in our nationwide surveillance network. In the 6-year study period, the prevalence of drug-resistant HIV-1 among 2573 patients, consisting mainly of Japanese men in their late-30s and infected through male-to-male sexual contacts, followed an increasing trend from 5.9% (16/273) in 2003 to 8.3% (50/605) in 2008. Nucleoside reverse transcriptase inhibitor-associated mutations predominated in each year, with T215 revertants being the most abundant. The predictive factor for drug-resistant HIV-1 transmission was subtype B (OR=2.36; p=0.004), and those for recent HIV-1 infection were male gender (OR=3.79; p=0.009), MSM behavior (OR=1.67; p=0.01), Japanese nationality (OR=2.31; p=0.008), and subtype B (OR=5.64; p<0.05). Continued activities are needed to raise awareness of the risks of HIV-1 infection and complications of drug-resistant strains. Continued surveillance is also needed to understand trends in the HIV-1 epidemic.
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http://dx.doi.org/10.1016/j.antiviral.2010.07.008DOI Listing
October 2010

Protection of macaques with diverse MHC genotypes against a heterologous SIV by vaccination with a deglycosylated live-attenuated SIV.

PLoS One 2010 Jul 20;5(7):e11678. Epub 2010 Jul 20.

AIDS Research Center, National Institute of Infectious Diseases, Shinjuku-ku, Tokyo, Japan.

HIV vaccine development has been hampered by issues such as undefined correlates of protection and extensive diversity of HIV. We addressed these issues using a previously established SIV-macaque model in which SIV mutants with deletions of multiple gp120 N-glycans function as potent live attenuated vaccines to induce near-sterile immunity against the parental pathogenic SIVmac239. In this study, we investigated the protective efficacy of these mutants against a highly pathogenic heterologous SIVsmE543-3 delivered intravenously to rhesus macaques with diverse MHC genotypes. All 11 vaccinated macaques contained the acute-phase infection with blood viral loads below the level of detection between 4 and 10 weeks postchallenge (pc), following a transient but marginal peak of viral replication at 2 weeks in only half of the challenged animals. In the chronic phase, seven vaccinees contained viral replication for over 80 weeks pc, while four did not. Neutralizing antibodies against challenge virus were not detected. Although overall levels of SIV specific T cell responses did not correlate with containment of acute and chronic viral replication, a critical role of cellular responses in the containment of viral replication was suggested. Emergence of viruses with altered fitness due to recombination between the vaccine and challenge viruses and increased gp120 glycosylation was linked to the failure to control SIV. These results demonstrate the induction of effective protective immune responses in a significant number of animals against heterologous virus by infection with deglycosylated attenuated SIV mutants in macaques with highly diverse MHC background. These findings suggest that broad HIV cross clade protection is possible, even in hosts with diverse genetic backgrounds. In summary, results of this study indicate that deglycosylated live-attenuated vaccines may provide a platform for the elucidation of correlates of protection needed for a successful HIV vaccine against diverse isolates.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0011678PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2907403PMC
July 2010

HLA-associated immune pressure on Gag protein in CRF01_AE-infected individuals and its association with plasma viral load.

PLoS One 2010 Jun 17;5(6):e11179. Epub 2010 Jun 17.

Department of Microbiology, Faculty of Science, Mahidol University, Bangkok, Thailand.

Background: The human leukocyte antigen (HLA)-restricted cytotoxic T-lymphocyte (CTL) immune response is one of the major factors determining the genetic diversity of human immunodeficiency virus (HIV). There are few population-based analyses of the amino acid variations associated with the host HLA type and their clinical relevance for the Asian population. Here, we identified HLA-associated polymorphisms in the HIV-1 CRF01_AE Gag protein in infected married couples, and examined the consequences of these HLA-selected mutations after transmission to HLA-unmatched recipients.

Methodology/principal Findings: One hundred sixteen HIV-1-infected couples were recruited at a government hospital in northern Thailand. The 1.7-kb gag gene was amplified and directly sequenced. We identified 56 associations between amino acid variations in Gag and HLA alleles. Of those amino acid variations, 35 (62.5%) were located within or adjacent to regions reported to be HIV-specific CTL epitopes restricted by the relevant HLA. Interestingly, a significant number of HLA-associated amino acid variations appear to be unique to the CRF01_AE-infected Thai population. Variations in the capsid protein (p24) had the strongest associations with the viral load and CD4 cell count. The mutation and reversion rates after transmission to a host with a different HLA environment varied considerably. The p24 T242N variant escape from B57/58 CTL had a significant impact on the HIV-1 viral load of CRF01_AE-infected patients.

Conclusions/significance: HLA-associated amino acid mutations and the CTL selection pressures on the p24 antigen appear to have the most significant impact on HIV replication in a CRF01_AE-infected Asian population. HLA-associated mutations with a low reversion rate accumulated as a footprint in this Thai population. The novel HLA-associated mutations identified in this study encourage us to acquire more extensive information about the viral dynamics of HLA-associated amino acid polymorphisms in a given population as effective CTL vaccine targets.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0011179PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2887364PMC
June 2010

Molecular evolutionary analysis of the influenza A(H1N1)pdm, May-September, 2009: temporal and spatial spreading profile of the viruses in Japan.

PLoS One 2010 Jun 10;5(6):e11057. Epub 2010 Jun 10.

Infectious Diseases Surveillance Center, National Institute of Infectious Diseases, Tokyo, Japan.

Background: In March 2009, pandemic influenza A(H1N1) (A(H1N1)pdm) emerged in Mexico and the United States. In Japan, since the first outbreak of A(H1N1)pdm in Osaka and Hyogo Prefectures occurred in the middle of May 2009, the virus had spread over 16 of 47 prefectures as of June 4, 2009.

Methods/principal Findings: We analyzed all-segment concatenated genome sequences of 75 isolates of A(H1N1)pdm viruses in Japan, and compared them with 163 full-genome sequences in the world. Two analyzing methods, distance-based and Bayesian coalescent MCMC inferences were adopted to elucidate an evolutionary relationship of the viruses in the world and Japan. Regardless of the method, the viruses in the world were classified into four distinct clusters with a few exceptions. Cluster 1 was originated earlier than cluster 2, while cluster 2 was more widely spread around the world. The other two clusters (clusters 1.2 and 1.3) were suggested to be distinct reassortants with different types of segment assortments. The viruses in Japan seemed to be a multiple origin, which were derived from approximately 28 transported cases. Twelve cases were associated with monophyletic groups consisting of Japanese viruses, which were referred to as micro-clade. While most of the micro-clades belonged to the cluster 2, the clade of the first cases of infection in Japan originated from cluster 1.2. Micro-clades of Osaka/Kobe and the Fukuoka cases, both of which were school-wide outbreaks, were eradicated. Time of most recent common ancestor (tMRCA) for each micro-clade demonstrated that some distinct viruses were transmitted in Japan between late May and early June, 2009, and appeared to spread nation-wide throughout summer.

Conclusions: Our results suggest that many viruses were transmitted from abroad in late May 2009 irrespective of preventive actions against the pandemic influenza, and that the influenza A(H1N1)pdm had become a pandemic stage in June 2009 in Japan.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0011057PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2883557PMC
June 2010

HIV-2 CRF01_AB: first circulating recombinant form of HIV-2.

J Acquir Immune Defic Syndr 2010 Jul;54(3):241-7

Department of Infection and Immunology, Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Japan.

Background: Five HIV-2-seropositive cases were recently identified in Japan, outside the HIV-2 endemic area of West Africa. To clarify the molecular epidemiology of HIV-2 in Japan, we analyzed sequences of these cases in detail.

Methods: HIV-2 genetic groups were determined by gag and env sequences. For suspected recombinant isolates, the genetic structure was determined by full-length genomic analyses. To understand the history and evolution of HIV-2 recombinant isolates, we estimated the time of most recent common ancestor by Bayesian Markov chain Monte Carlo method.

Results: Three isolates were determined as recombinants of groups A and B, and their mosaic genome structures were identical with that of 7312A, a recombinant isolate reported in 1990 from Côte d'Ivoire. Our 3 isolates and 7312A fulfilled the criteria for determining a circulating recombinant form (CRF). These isolates were verified by the Los Alamos HIV sequence database as the first CRF of HIV-2, HIV-2 CRF01_AB. The mean time of most recent common ancestor of CRF01_AB was estimated as between 1964 and 1973, several decades after the estimated emergence of HIV-2.

Conclusions: We recently identified HIV-2 CRF01_AB cases in Japan. This ectopic observation of the virus outside its original endemic area suggests an ongoing global spread of HIV-2 CRF01_AB.
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http://dx.doi.org/10.1097/QAI.0b013e3181dc98c1DOI Listing
July 2010

Isolation and characterization of a novel Borrelia group of tick-borne borreliae from imported reptiles and their associated ticks.

Environ Microbiol 2010 Jan 16;12(1):134-46. Epub 2009 Sep 16.

United Graduate School of Agricultural Science and Veterinary Science, Gifu University, Gifu, Japan.

The members of the genus Borrelia are transmitted by arthropods and known to be infectious to vertebrates. Here we found isolates and DNAs belonging to the Borrelia turcica and unknown Borrelia species from imported reptiles and their ectoparasites. The Borrelia strains were isolated from blood and multiple organs of exotic tortoises, and were experimentally infectious to captive-bred tortoises. These findings suggest that these tortoises may be a candidate as the reservoir host of the Borrelia species. In this study, the Borrelia strains were also isolated from and/or detected in hard-bodied ticks, Amblyomma ticks and Hyalomma ticks. In some of these ticks, immunofluorescence imaging analysis revealed that the Borrelia had also invaded into the tick salivary glands. Accordingly, these ticks were expected to be a potential vector of the Borrelia species. Sequencing analyses of both housekeeping genes (flaB gene, gyrB gene and 16S rDNA gene) and 23S rRNA gene-16S rRNA gene intergenic spacer region revealed that these Borrelia strains formed a monophyletic group that was independent from two other Borrelia groups, Lyme disease Borrelia and relapsing fever Borrelia. From these results, the novel group of Borrelia comprises the third major group of arthropod-transmitted borreliae identified to date.
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http://dx.doi.org/10.1111/j.1462-2920.2009.02054.xDOI Listing
January 2010

Net positive charge of HIV-1 CRF01_AE V3 sequence regulates viral sensitivity to humoral immunity.

PLoS One 2008 Sep 12;3(9):e3206. Epub 2008 Sep 12.

Department of Public Health, Yokohama City University School of Medicine, Kanagawa, Japan.

The third variable region (V3) of the human immunodeficiency virus type 1 (HIV-1) envelope gp120 subunit participates in determination of viral infection coreceptor tropism and host humoral immune responses. Positive charge of the V3 plays a key role in determining viral coreceptor tropism. Here, we examined by bioinformatics, experimental, and protein modelling approaches whether the net positive charge of V3 sequence regulates viral sensitivity to humoral immunity. We chose HIV-1 CRF01_AE strain as a model virus to address the question. Diversity analyses using CRF01_AE V3 sequences from 37 countries during 1984 and 2005 (n = 1361) revealed that reduction in the V3's net positive charge makes V3 less variable due to limited positive selection. Consistently, neutralization assay using CRF01_AE V3 recombinant viruses (n = 30) showed that the reduction in the V3's net positive charge rendered HIV-1 less sensitive to neutralization by the blood anti-V3 antibodies. The especially neutralization resistant V3 sequences were the particular subset of the CCR5-tropic V3 sequences with net positive charges of +2 to +4. Molecular dynamics simulation of the gp120 monomers showed that the V3's net positive charge regulates the V3 configuration. This and reported gp120 structural data predict a less-exposed V3 with a reduced net positive charge in the native gp120 trimer context. Taken together, these data suggest a key role of the V3's net positive charge in the immunological escape and coreceptor tropism evolution of HIV-1 CRF01_AE in vivo. The findings have molecular implications for the adaptive evolution and vaccine design of HIV-1.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0003206PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2527523PMC
September 2008

Molecular epidemiology of the heterosexual HIV-1 transmission in Kunming, Yunnan Province of China suggests origin from the local IDU epidemic.

AIDS Res Hum Retroviruses 2005 Nov;21(11):977-80

Laboratory of Molecular Virology and Epidemiology, AIDS Research Center, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Tokyo 162-8640, Japan.

Molecular epidemiological investigation was conducted among injecting drug users (IDUs) (n = 11) and heterosexuals (n = 15) in Kunming, Yunnan Province of China. HIV-1 genotypes were determined based on the nucleotide sequences of 2.6-kb gag-RT region. The distribution of genotypes among IDUs was as follows: CRF07_BC (5/11) and CRF08_BC (5/11); subtype B' (1/11). Similarly, a majority of Kunming heterosexuals (14/15) were infected with CRF07_BC (4/15), CRF08_BC (6 /15), or subtype B' (4/15), known to predominate among IDUs in China. This contrasts with trends in the coastal regions of China and surrounding southeastern Asian countries, where CRF01_AE predominates among heterosexuals. The heterosexual HIV-1 epidemic in Kunming thus appears to derive from the local IDU epidemic. Of note, subtype B' was the most prevalent strain among heterosexuals before 1997, while CRF07_BC and CRF08_BC became predominant in 2002, indicating a transition of HIV-1 genotype distribution between the early and the more recent samples from Kunming heterosexuals.
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http://dx.doi.org/10.1089/aid.2005.21.977DOI Listing
November 2005

Heterosexual transmission of novel CRF01_AE and subtype B recombinant forms of HIV type 1 in northern Thailand.

AIDS Res Hum Retroviruses 2005 Aug;21(8):734-8

Thai National Institute of Health, Department of Medical Sciences, Ministry of Public Health, Nonthaburi, Thailand.

The increased proportion of CRFO1_AE/subtype B recombinant infections among injecting drug users raised a concern that such recombinant forms may also spread in a heterosexual population in Thailand. Using the BootScan method, we reanalyzed pol gene sequences among 114 heterosexually infected individuals in northern Thailand, who were tested for a drug-resistance genotype between July 2000 and July 2001. Two individuals were suspected of carrying a recombinant HIV-1. Thus we analyzed a nearly full-length HIV genome in the two individuals and their spouses. An identical recombinant form of CRF01_AE and subtype B was found in one couple, indicating that this recombinant virus was heterosexually transmitted. Interestingly, this recombinant form had multiple breakpoints in the core protein of Gag and both infected individuals had a high CD4+ cell count without antiretroviral therapy. CRFO1/subtype B recombinant forms exist in a heterosexual population in northern Thailand. Some recombinant virus may be associated with a slow rate of HIV disease progression.
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http://dx.doi.org/10.1089/aid.2005.21.734DOI Listing
August 2005