Cognitive Intraindividual variability in injection drug use among Hispanic residents of Puerto Rico.

Despite a high prevalence of injection drug use (IDU) in Puerto Rico, little is known about how it affects neuropsychological functioning in this population. Investigations of intra-individual variability (IIV) have alluded to its utility as a potential indicator of neural decline. The purpose of this study was to characterize IIV among Hispanic residents of Puerto Rico who engage in IDU. Injectors and non-injectors completed the Neuropsí Atención y Memoria battery. Measures of IIV were calculated for the overall test battery (OTB), the three battery indices, and three domains of attention, memory, and executive functioning. The injector group showed significantly greater IIV than the non-injector group on all measures (OTB, indices, and individual domains). Additionally, injectors showed significantly higher IIV in the domain of executive functioning compared to other cognitive domains and battery indices. In contrast, non-injectors did not show any significant within-group differences on any IIV measures. The higher performance variability observed in the IDU group suggests a negative influence of IDU on cognition, with executive functioning being more susceptible to these effects. These findings support the need for continued investigations into the clinical application of IIV for diagnostic and prognostic purposes in the Hispanic IDU population.

Increased metamemory accuracy with practice does not require practice with metamemory.

Given that learners do not always predict their future memory performance accurately, there is a need to better understand how metamemory accuracy can be improved. Prior research suggests that one way to improve is practice-participants tend to become better at predicting their future memory performance over the course of multi-trial learning experiments. However, it is currently unclear whether such improvements result from participants having practised making metamemory judgements or whether comparable improvements occur even in their absence. This issue was investigated in three multi-trial, cued recall experiments wherein participants either did or did not receive practice making judgements of learning. Metamemory accuracy increased across study blocks but did so equally for the two groups. These results indicate that increased metamemory accuracy with practice is not due to participants having practised explicit metamemory monitoring but instead due to other factors associated with multi-trial learning such as retrieval practice and the availability of prior test performance as a metamemory cue.

Limited HIV-associated neuropathologies and lack of immune activation in sub-saharan African individuals with late-stage subtype C HIV-1 infection.

Although previous studies have suggested that subtype B HIV-1 proviruses in the brain are associated with physiological changes and immune activation accompanied with microgliosis and astrogliosis, and indicated that both HIV-1 subtype variation and geographical location might influence the neuropathogenicity of HIV-1 in the brain. The natural course of neuropathogenesis of the most widespread subtype C HIV-1 has not been adequately investigated, especially for people living with HIV (PLWH) in sub-Saharan Africa. To characterize the natural neuropathology of subtype C HIV-1, postmortem frontal lobe and basal ganglia tissues were collected from nine ART-naïve individuals who died of late-stage AIDS with subtype C HIV-1 infection, and eight uninfected deceased individuals as controls. Histological staining was performed on all brain tissues to assess brain pathologies. Immunohistochemistry (IHC) against CD4, p24, Iba-1, GFAP, and CD8 in all brain tissues was conducted to evaluate potential viral production and immune activation. Histological results showed mild perivascular cuffs of lymphocytes only in a minority of the infected individuals. Viral capsid p24 protein was only detected in circulating immune cells of one infected individual, suggesting a lack of productive HIV-1 infection of the brain even at the late-stage of AIDS. Notably, similar levels of Iba-1 or GFAP between HIV + and HIV- brain tissues indicated a lack of microgliosis and astrogliosis, respectively. Similar levels of CD8 + cytotoxic T lymphocyte (CTL) infiltration between HIV + and HIV- brain tissues indicated CTL were not likely to be involved within subtype C HIV-1 infected participants of this cohort. Results from this subtype C HIV-1 study suggest that there is a lack of productive infection and limited neuropathogenesis by subtype C HIV-1 even at late-stage disease, which is in contrast to what was reported for subtype B HIV-1 by other investigators.

Viral Epitope Scanning Reveals Correlation between Seasonal HCoVs and SARS-CoV-2 Antibody Responses among Cancer and Non-Cancer Patients.

Seasonal coronaviruses (HCoVs) are known to contribute to cross-reactive antibody (Ab) responses against SARS-CoV-2. While these responses are predictable due to the high homology between SARS-CoV-2 and other CoVs, the impact of these responses on susceptibility to SARS-CoV-2 infection in cancer patients is unclear. To investigate the influence of prior HCoV infection on anti-SARS-CoV-2 Ab responses among COVID-19 asymptomatic individuals with cancer and controls without cancers, we utilized the VirScan technology in which phage immunoprecipitation and sequencing (PhIP-seq) of longitudinal plasma samples was performed to investigate high-resolution (i.e., epitope level) humoral CoV responses. Despite testing positive for anti-SARS-CoV-2 Ab in the plasma, a majority of the participants were asymptomatic for COVID-19 with no prior history of COVID-19 diagnosis. Although the magnitudes of the anti-SARS-CoV-2 Ab responses were lower in individuals with Kaposi sarcoma (KS) compared to non-KS cancer individuals and those without cancer, the HCoV Ab repertoire was similar between individuals with and without cancer independent of age, sex, HIV status, and chemotherapy. The magnitudes of the anti-spike HCoV responses showed a strong positive association with those of the anti-SARS-CoV-2 spike in cancer patients, and only a weak association in non-cancer patients, suggesting that prior infection with HCoVs might play a role in limiting SARS-CoV-2 infection and COVID-19 disease severity.

Antibody profiling and predictive modeling discriminate between Kaposi sarcoma and asymptomatic KSHV infection.

Protein-level immunodominance patterns against Kaposi sarcoma-associated herpesvirus (KSHV), the aetiologic agent of Kaposi sarcoma (KS), have been revealed from serological probing of whole protein arrays, however, the epitopes that underlie these patterns have not been defined. We recently demonstrated the utility of phage display in high-resolution linear epitope mapping of the KSHV latency-associated nuclear antigen (LANA/ORF73). Here, a VirScan phage immunoprecipitation and sequencing approach, employing a library of 1,988 KSHV proteome-derived peptides, was used to quantify the breadth and magnitude of responses of 59 sub-Saharan African KS patients and 22 KSHV-infected asymptomatic individuals (ASY), and ultimately to support an application of machine-learning-based predictive modeling using the peptide-level responses. Comparing anti-KSHV antibody repertoire revealed that magnitude, not breadth, increased in KS. The most targeted epitopes in both KS and ASY were in the immunodominant proteins, notably, K8.129-56 and ORF65140-168, in addition to LANA. Finally, using unbiased machine-learning-based predictive models, reactivity to a subset of 25 discriminative peptides was demonstrated to successfully classify KS patients from asymptomatic individuals. Our study provides the highest resolution mapping of antigenicity across the entire KSHV proteome to date, which is vital to discern mechanisms of viral pathogenesis, to define prognostic biomarkers, and to design effective vaccine and therapeutic strategies. Future studies will investigate the diagnostic, prognostic, and therapeutic potential of the 25 discriminative peptides.

Pancreatic cancer tumor microenvironment is a major therapeutic barrier and target.

Pancreatic Ductal Adenocarcinoma (PDAC) is projected to become the 2nd leading cause of cancer-related deaths in the United States. Limitations in early detection and treatment barriers contribute to the lack of substantial success in the treatment of this challenging-to-treat malignancy. Desmoplasia is the hallmark of PDAC microenvironment that creates a physical and immunologic barrier. Stromal support cells and immunomodulatory cells face aberrant signaling by pancreatic cancer cells that shifts the complex balance of proper repair mechanisms into a state of dysregulation. The product of this dysregulation is the desmoplastic environment that encases the malignant cells leading to a dense, hypoxic environment that promotes further tumorigenesis, provides innate systemic resistance, and suppresses anti-tumor immune invasion. This desmoplastic environment combined with the immunoregulatory events that allow it to persist serve as the primary focus of this review. The physical barrier and immune counterbalance in the tumor microenvironment (TME) make PDAC an immunologically cold tumor. To convert PDAC into an immunologically hot tumor, tumor microenvironment could be considered alongside the tumor cells. We discuss the complex network of microenvironment molecular and cellular composition and explore how they can be targeted to overcome immuno-therapeutic challenges.

Cannabis Use Associates With Reduced Proviral Burden and Inflammatory Cytokine in Tissues From Men With Clade C HIV-1 on Suppressive Antiretroviral Therapy.

BACKGROUND: Human immunodeficiency virus 1 (HIV-1) tissue reservoirs remain the main obstacle against an HIV cure. Limited information exists regarding cannabis's effects on HIV-1 infections in vivo, and the impact of cannabis use on HIV-1 parenchymal tissue reservoirs is unexplored. METHODS: To investigate whether cannabis use alters HIV-1 tissue reservoirs, we systematically collected 21 postmortem brain and peripheral tissues from 20 men with subtype C HIV-1 and with suppressed viral load enrolled in Zambia, 10 of whom tested positive for cannabis use. The tissue distribution and copies of subtype C HIV-1 LTR, gag, env DNA and RNA, and the relative mRNA levels of cytokines IL-1ß, IL-6, IL-10, and TGF-ß1 were quantified using PCR-based approaches. Utilizing generalized linear mixed models we compared persons with HIV-1 and suppressed viral load, with and without cannabis use. RESULTS: The odds of tissues harboring HIV-1 DNA and the viral DNA copies in those tissues were significantly lower in persons using cannabis. Moreover, the transcription levels of proinflammatory cytokines IL-1ß and IL-6 in lymphoid tissues of persons using cannabis were also significantly lower. CONCLUSIONS: Our findings suggested that cannabis use is associated with reduced sizes and inflammatory cytokine expression of subtype C HIV-1 reservoirs in men with suppressed viral load.

Co-infection and co-localization of Kaposi sarcoma-associated herpesvirus and Epstein-Barr virus in HIV-associated Kaposi sarcoma: a case report.

Kaposi sarcoma (KS), a multifocal vascular neoplasm frequently observed in HIV-positive individuals, primarily affects the skin, mucous membranes, visceral organs, and lymph nodes. KS is associated primarily with Kaposi sarcoma-associated herpesvirus (KSHV) infection. In this case report, we present a rare occurrence of co-infection and co-localization of KSHV and Epstein-Barr virus (EBV) in KS arising from the conjunctiva, which, to our knowledge, has not been reported previously. Immunohistochemistry (IHC), DNA polymerase chain reaction (PCR), and EBV-encoded RNA in situ hybridization (EBER-ISH) were utilized to demonstrate the presence of KSHV and EBV infection in the ocular KS lesion. Nearly all KSHV-positive cells displayed co-infection with EBV. In addition, the KS lesion revealed co-localization of KSHV Latency-Associated Nuclear Antigen (LANA) and EBV Epstein Barr virus Nuclear Antigen-1 (EBNA1) by multi-colored immunofluorescence staining with different anti-EBNA1 antibodies, indicating the possibility of interactions between these two gamma herpesviruses within the same lesion. Additional study is needed to determine whether EBV co-infection in KS is a common or an opportunistic event that might contribute to KS development and progression.

Comparative polar and lipid plasma metabolomics differentiate KSHV infection and disease states.

BACKGROUND: Kaposi sarcoma (KS) is a neoplastic disease etiologically associated with infection by the Kaposi sarcoma-associated herpesvirus (KSHV). KS manifests primarily as cutaneous lesions in individuals due to either age (classical KS), HIV infection (epidemic KS), or tissue rejection preventatives in transplantation (iatrogenic KS) but can also occur in individuals, predominantly in sub-Saharan Africa (SSA), lacking any obvious immune suppression (endemic KS). The high endemicity of KSHV and human immunodeficiency virus-1 (HIV) co-infection in Africa results in KS being one of the top 5 cancers there. As with most viral cancers, infection with KSHV alone is insufficient to induce tumorigenesis. Indeed, KSHV infection of primary human endothelial cell cultures, even at high levels, is rarely associated with long-term culture, transformation, or growth deregulation, yet infection in vivo is sustained for life. Investigations of immune mediators that distinguish KSHV infection, KSHV/HIV co-infection, and symptomatic KS disease have yet to reveal consistent correlates of protection against or progression to KS. In addition to viral infection, it is plausible that pathogenesis also requires an immunological and metabolic environment permissive to the abnormal endothelial cell growth evident in KS tumors. In this study, we explored whether plasma metabolomes could differentiate asymptomatic KSHV-infected individuals with or without HIV co-infection and symptomatic KS from each other. METHODS: To investigate how metabolic changes may correlate with co-infections and tumorigenesis, plasma samples derived from KSHV seropositive sub-Saharan African subjects in three groups, (A) asymptomatic (lacking neoplastic disease) with KSHV infection only, (B) asymptomatic co-infected with KSHV and HIV, and (C) symptomatic with clinically diagnosed KS, were subjected to analysis of lipid and polar metabolite profiles RESULTS: Polar and nonpolar plasma metabolic differentials were evident in both comparisons. Integration of the metabolic findings with our previously reported KS transcriptomics data suggests dysregulation of amino acid/urea cycle and purine metabolic pathways, in concert with viral infection in KS disease progression. CONCLUSIONS: This study is, to our knowledge, the first to report human plasma metabolic differentials between in vivo KSHV infection and co-infection with HIV, as well as differentials between co-infection and epidemic KS.

Yes-associated protein-1 overexpression in ocular surface squamous neoplasia; a potential diagnostic marker and therapeutic target.

Yes-associated protein-1 (YAP-1) is a Hippo system transcription factor, which serves as an oncogene in squamous cell carcinoma, and several solid tumors when the Hippo pathway is dysregulated. Yet, the activity of YAP-1 in ocular surface squamous neoplasia (OSSN) has not been determined. Here, we investigate the relationship between YAP-1 overexpression and OSSN. Using a cross-sectional study design, we recruited 227 OSSN patients from the University Teaching Hospitals in Lusaka, Zambia. Immunohistochemistry was used to assess YAP-1 protein overexpression in tumor tissue relative to surrounding benign squamous epithelium. OSSN patient samples (preinvasive, n = 62, 27% and invasive, n = 165, 73%) were studied. One hundred forty-nine invasive tumors contained adjacent preinvasive tissue, bringing the total number of preinvasive lesions examined to 211 (62 + 149). There was adjacent benign squamous epithelium in 50.2% (114/227) of OSSN samples. Nuclear YAP- 1 was significantly overexpressed in preinvasive (Fisher's (F): p <.0001, Monte Carlo (MC): p <.0001) and invasive (F: p <.0001, MC: p <.0001) OSSN in comparison to adjacent benign squamous epithelium when analyzed for basal keratinocyte positive count, staining intensity, expression pattern, and Immunostaining intensity-distribution index. YAP-1 expression did not differ between preinvasive and invasive OSSN (p >.05), keratinizing and non- keratinizing cancer (p >.05), or between T1/T2 and T3/T4 stages in invasive tumors (p >.05). However, grade 2 and 3 tumors had significantly stronger nucleus YAP-1 overexpression intensity than grade 1 tumors (F: p = .0078, MC: p = .0489). By immunohistochemistry, we identified significant overexpression (upregulation of YAP-1 protein expression) in preinvasive and invasive OSSN lesions compared to neighboring benign squamous epithelium. YAP-1 expression was significantly higher in poorly and moderately differentiated invasive squamous cancer than in well-differentiated carcinomas. Overexpression of YAP-1 within the margin of preinvasive and invasive OSSN, but not in the neighboring normal epithelium, indicates that it plays a role in the development and progression of OSSN.

Memory and attention: A double dissociation between memory encoding and memory retrieval.

Research has found substantial negative effects of divided attention (DA) during encoding but less substantial effects when attention is divided during retrieval, an asymmetry which has been interpreted as indicating that different control processes or forms of attention are involved in encoding and retrieval (e.g., Chun & Johnson, 2011; Craik, Govoni, Naveh-Benjamin, & Anderson, 1996; Long, Kuhl, & Chun, 2018). The extant evidence, however, is not strong support for qualitative differences and might simply indicate differential sensitivity. The present experiments document a stronger, double dissociation by focusing on the Attentional Boost Effect (ABE) - a phenomenon in which the detection of targets in a secondary task enhances encoding of co-occurring stimuli. The dual-task interaction account proposes that the classical negative effects produced by dual-task interference are offset by a transient increase in externally-directed attention brought about by target detection. Since externally-directed attention is less valuable for retrieval processes, the ABE should result in a net negative effect when applied in the test phase because the dual-task interference would no longer be offset by the externally-directed boost occurring during target trials. Experiments 1, 2 and 4 confirmed the predictions by showing that test words paired with target stimuli were recognized significantly worse than test words paired with distractor stimuli. In contrast, Experiments 3 and 4 replicated the usual positive effects of the ABE with respect to encoding. We discuss these findings in light of recent theoretical proposals suggesting that encoding and retrieval processes are subserved by different forms of attention (external [perceptual] vs. internal [reflective]). Implications for the Transfer-Appropriate-Processing view of memory are also illustrated.

Longitudinal Variations in Antibody Responses against SARS-CoV-2 Spike Epitopes upon Serial Vaccinations.

The COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) impacted healthcare, the workforce, and worldwide socioeconomics. Multi-dose mono- or bivalent mRNA vaccine regimens have shown high efficacy in protection against SARS-CoV-2 and its emerging variants with varying degrees of efficacy. Amino acid changes, primarily in the receptor-binding domain (RBD), result in selection for viral infectivity, disease severity, and immune evasion. Therefore, many studies have centered around neutralizing antibodies that target the RBD and their generation achieved through infection or vaccination. Here, we conducted a unique longitudinal study, analyzing the effects of a three-dose mRNA vaccine regimen exclusively using the monovalent BNT162b2 (Pfizer/BioNTech) vaccine, systematically administered to nine previously uninfected (naïve) individuals. We compare changes in humoral antibody responses across the entire SARS-CoV-2 spike glycoprotein (S) using a high-throughput phage display technique (VirScan). Our data demonstrate that two doses of vaccination alone can achieve the broadest and highest magnitudes of anti-S response. Moreover, we present evidence of novel highly boosted non-RBD epitopes that strongly correlate with neutralization and recapitulate independent findings. These vaccine-boosted epitopes could facilitate multi-valent vaccine development and drug discovery.

Immune profiling in Puerto Rican injection drug users with and without HIV-1 infection.

Antiretroviral therapy has been effective in suppressing HIV viral load and enabling people living with HIV to experience longer, more conventional lives. However, as people living with HIV are living longer, they are developing aging-related diseases prematurely and are more susceptible to comorbidities that have been linked to chronic inflammation. Coincident with HIV infection and aging, drug abuse has also been independently associated with gut dysbiosis, microbial translocation, and inflammation. Here, we hypothesized that injection drug use would exacerbate HIV-induced immune activation and inflammation, thereby intensifying immune dysfunction. We recruited 50 individuals not using injection drugs (36/50 HIV+) and 47 people who inject drugs (PWID, 12/47 HIV+). All but 3 of the HIV+ subjects were on antiretroviral therapy. Plasma immune profiles were characterized by immunoproteomics, and cellular immunophenotypes were assessed using mass cytometry. The immune profiles of HIV+/PWID-, HIV-/PWID+, and HIV+/PWID+ were each significantly different from controls; however, few differences between these groups were detected, and only 3 inflammatory mediators and 2 immune cell populations demonstrated a combinatorial effect of injection drug use and HIV infection. In conclusion, a comprehensive analysis of inflammatory mediators and cell immunophenotypes revealed remarkably similar patterns of immune dysfunction in HIV-infected individuals and in people who inject drugs with and without HIV-1 infection.

Upregulation of Cell Surface Glycoproteins in Correlation with KSHV LANA in the Kaposi Sarcoma Tumor Microenvironment.

HIV-associated epidemic Kaposi sarcoma (EpKS) remains one of the most prevalent cancers in sub-Saharan Africa despite the widespread uptake of anti-retroviral therapy and HIV-1 suppression. In an effort to define potential therapeutic targets against KS tumors, we analyzed previously published KS bulk tumor transcriptomics to identify cell surface biomarkers. In addition to upregulated gene expression (>6-fold) in the EpKS tumor microenvironment, biomarkers were selected for correlation with KSHV latency-associated nuclear antigen (LANA) expression. The cell surface glycoprotein genes identified were KDR, FLT4, ADAM12, UNC5A, ZP2, and OX40, as well as the endothelial lineage determinants Prox-1 and CD34. Each protein was evaluated for its expression and co-localization with KSHV LANA using multi-color immunofluorescence in KS tissues, KSHV-infected L1T2 cells, uninfected TIVE cells, and murine L1T2 tumor xenografts. Five surface glycoproteins (KDR, FLT4, UNC5A, ADAM12, and CD34) were associated with LANA-positive cells but were also detected in uninfected cells in the KS microenvironment. In vitro L1T2 cultures showed evidence of only FLT4, KDR, and UNC5A, whereas mouse L1T2 xenografts recapitulated human KS cell surface expression profiles, with the exception of CD34 and Prox-1. In KS tumors, most LANA-positive cells co-expressed markers of vascular as well as lymphatic endothelial lineages, suggesting KS-associated dedifferentiation to a more mesenchymal/progenitor phenotype.

Elevated iNOS and 3'-nitrotyrosine in Kaposi's Sarcoma tumors and mouse model.

Kaposi's Sarcoma (KS) is a heterogenous, multifocal vascular malignancy caused by the human herpesvirus 8 (HHV8), also known as Kaposi's Sarcoma-Associated Herpesvirus (KSHV). Here, we show that KS lesions express iNOS/NOS2 broadly throughout KS lesions, with enrichment in LANA positive spindle cells. The iNOS byproduct 3-nitrotyrosine is also enriched in LANA positive tumor cells and colocalizes with a fraction of LANA-nuclear bodies. We show that iNOS is highly expressed in the L1T3/mSLK tumor model of KS. iNOS expression correlated with KSHV lytic cycle gene expression, which was elevated in late-stage tumors (>4 weeks) but to a lesser degree in early stage (1 week) xenografts. Further, we show that L1T3/mSLK tumor growth is sensitive to an inhibitor of nitric oxide, L-NMMA. L-NMMA treatment reduced KSHV gene expression and perturbed cellular gene pathways relating to oxidative phosphorylation and mitochondrial dysfunction. These finding suggest that iNOS is expressed in KSHV infected endothelial-transformed tumor cells in KS, that iNOS expression depends on tumor microenvironment stress conditions, and that iNOS enzymatic activity contributes to KS tumor growth.

Antibody epitope profiling of the KSHV LANA protein using VirScan.

The humoral antibody response against Kaposi sarcoma-associated herpesvirus (KSHV) in infected individuals has been characterized demonstrating the latency-associated nuclear antigen (LANA) as the most antigenic KSHV protein. Despite the antigenicity of the protein, specific LANA epitopes have not been systematically characterized. Here, we utilized a bacteriophage T7 library, which displays 56-amino acid KSHV LANA peptides with 28-amino acid overlap (VirScan), to define those epitopes in LANA targeted by antibodies from a cohort of 62 sub-Saharan African Kaposi sarcoma (KS) patients and 22 KSHV-infected asymptomatic controls. Intra- and inter-patient breadth and magnitude of the anti-LANA responses were quantified at the peptide and amino acid levels. From these data, we derived a detailed epitope annotation of the entire LANA protein, with a high-resolution focus on the N- and C-termini. Overall, the central repeat region was highly antigenic, but the responses to this region could not be confidently mapped due to its high variability. The highly conserved N-terminus was targeted with low breadth and magnitude. In a minority of individuals, antibodies specific to the nuclear localization sequence and a portion of the proline-rich regions of the N-terminus were evident. In contrast, the first half of the conserved C-terminal domain was consistently targeted with high magnitude. Unfortunately, this region was not included in LANA partial C-terminal crystal structures, however, it was predicted to adopt predominantly random-coil structure. Coupled with functional and secondary structure domain predictions, VirScan revealed fine resolution epitope mapping of the N- and C-terminal domains of LANA that is consistent with previous antigenicity studies and may prove useful to correlate KSHV humoral immunity with pathogenesis.

Employing Respondent Driven Sampling (RDS) to recruit people who inject drugs (PWID) and other hard-to-reach populations during COVID-19: Lessons learned.

Background: Respondent Driven Sampling (RDS) is an effective sampling strategy to recruit hard-to-reach populations but the impact of the COVID-19 pandemic on the use of this strategy in the collection of data involving human subjects, particularly among marginalized and vulnerable populations, is not known. Based on an ongoing study using RDS to recruit and study the interactions between HIV infection, injection drug use, and the microbiome in Puerto Rico, this paper explores the effectiveness of RDS during the pandemic and provided potential strategies that could improve recruitment and data collection. Results: RDS was employed to evaluate its effectiveness in recruiting a group of people who inject drugs (PWID) and controls (N = 127) into a study in the midst of the COVID-19 pandemic. The participants were distributed among three subsets: 15 were HIV+ and PWID, 58 were HIV- PWID, and 54 were HIV+ and not PWID. Findings: Results show that recruitment through peer networks using RDS was possible across all sub-groups. Yet, while those in the HIV+ PWID sub-group managed to recruit from other-sub groups of HIV- PWID and HIV+, this occurred at a lower frequency. Conclusion: Despite the barriers introduced by COVID-19, it is clear that even in this environment, RDS continues to play a powerful role in recruiting hard-to-reach populations. Yet, more attention should be paid at how future pandemics, natural disasters, and other big events might affect RDS recruitment of vulnerable and hard-to-reach populations.

Subtype C HIV-1 reservoirs throughout the body in ART-suppressed individuals.

Subtype B HIV-1 reservoirs have been intensively investigated, but reservoirs in other subtypes and how they respond to antiretroviral therapy (ART) is substantially less established. To characterize subtype C HIV-1 reservoirs, we implemented postmortem frozen, as well as formalin fixed paraffin embedded (FFPE) tissue sampling of central nervous system (CNS) and peripheral tissues. HIV-1 LTR, gag, envelope (env) DNA and RNA was quantified using genomic DNA and RNA extracted from frozen tissues. RNAscope was used to localize subtype C HIV-1 DNA and RNA in FFPE tissue. Despite uniform viral load suppression in our cohort, PCR results showed that subtype C HIV-1 proviral copies vary both in magnitude and tissue distribution, with detection primarily in secondary lymphoid tissues. Interestingly, the appendix harbored proviruses in all subjects. Unlike subtype B, subtype C provirus was rarely detectable in the CNS, and there was no detectable HIV-1 RNA. HIV-1 RNA was detected in peripheral lymphoid tissues of 6 out of 8 ART-suppressed cases. In addition to active HIV-1 expression in lymphoid tissues, RNAscope revealed HIV RNA detection in CD4-expressing cells in the appendix, suggesting that this tissue was a previously unreported potential treatment-resistant reservoir for subtype C HIV-1.

Systematic post-mortem analysis of brain tissue from an HIV-1 subtype C viremic decedent revealed a paucity of infection and pathology.

Whether the human brain is a robust reservoir for HIV-1 subtype C has yet to be established. We aimed to determine whether HIV-1 subtype C infection can be detected in the brain tissue of a viremic individual at post-mortem and whether the viral burden was differential between different brain regions. This study reports a 38-year-old Zambian female decedent with severe wasting who was on Atripla for antiretroviral therapy. The cause of death was determined to be HIV/AIDS end-stage disease. The QuantStudio 3 Real-Time PCR System analyzed formalin-fixed paraffin-embedded tissue DNA from a systematic sampling of the entire left-brain hemisphere. Plasma and cerebral spinal fluid HIV-1 RNA loads were 576,123 and 14,962 copies/mL, respectively. The lymph node DNA viral load was 2316 copies per 106 cells. Two hundred and six (96.3%) tissue blocks had amplifiable DNA. HIV-1 viral DNA was detected in 35.9% of the blocks, the highest in the basal ganglia (66.7%) and the frontal lobe (46%). Overall, HIV detection was random, with low viral copies detected by quantitative polymerase chain reaction (qPCR); the lowest was observed in the occipital (median, IQR, range) 0.0 [0.0-0.0], 0.0-31.3, and the highest in the basal ganglia (mean ± SD, range, 125.1149.5, 0.0-350.0). Significant differences in HIV-1 DNA distribution were observed between the occipital versus parietal (p = 0.049), occipital versus frontal (p = 0.019), occipital versus basal ganglia (p = 0.005), cerebellum versus frontal (p = 0.021), cerebellum versus basal ganglia (p = 0.007), and temporal versus frontal (p = 0.034).