Tuberculosis disease is associated with elevated plasma ribonuclease activity in antiretroviral treatment-naïve people with HIV.

BACKGROUND: The role of ribonucleases in tuberculosis (TB) among people with HIV (PWH) is unknown. We explored ribonuclease activity in plasma from PWH with and without TB. METHODS: Participants were identified from a cohort of treatment-naïve PWH in Ethiopia who had been classified for TB disease (HIV+/TB + or HIV+/TB-). Ribonuclease activity in plasma was investigated by quantification of synthetic spike-in RNAs using sequencing and qPCR, and by a specific ribonuclease activity assay. Quantification of ribonuclease 1, 2, 3, 6, 7 and T2 proteins was performed by ELISA. Ribonuclease activity and protein concentrations were correlated with markers of TB and HIV disease severity and with concentrations of inflammatory mediators. RESULTS: Ribonuclease activity was significantly higher in plasma of HIV+/TB + (n = 51) compared to HIV+/TB- (n = 78), causing reduced stability of synthetic spike-in RNAs. concentrations of ribonucleases 2, 3 and T2 were also significantly increased in HIV+/TB + compared to HIV+/TB-. Ribonuclease activity was correlated with HIV viral load, and inversely correlated with CD4 count, mid-upper arm circumference and body mass index. Moreover, ribonuclease activity correlated with concentrations of interleukin-27, kynurenine/tryptophan ratio and procalcitonin. CONCLUSION: PWH with TB disease have elevated plasma ribonuclease activity, which is also associated with HIV severity and systemic inflammation.

HIV-2 mediated effects on target and bystander cells induce plasma proteome remodeling.

Despite low or undetectable plasma viral load, people living with HIV-2 (PLWH2) typically progress toward AIDS. The driving forces behind HIV-2 disease progression and the role of viremia are still not known, but low-level replication in tissues is believed to play a role. To investigate the impact of viremic and aviremic HIV-2 infection on target and bystander cell pathology, we used data-independent acquisition mass spectrometry to determine plasma signatures of tissue and cell type engagement. Proteins derived from target and bystander cells in multiple tissues, such as the gastrointestinal tract and brain, were detected at elevated levels in plasma of PLWH2, compared with HIV negative controls. Moreover, viremic HIV-2 infection appeared to induce enhanced release of proteins from a broader range of tissues compared to aviremic HIV-2 infection. This study expands the knowledge on the link between plasma proteome remodeling and the pathological cell engagement in tissues during HIV-2 infection.

HIV-2 Neutralization Sensitivity in Relation to Co-Receptor Entry Pathways and Env Motifs.

HIV-2, compared to HIV-1, elicits potent and broadly neutralizing antibodies, and uses a broad range of co-receptors. However, both sensitivity to neutralization and breadth of co-receptor use varies between HIV-2 isolates, and the molecular background is still not fully understood. Thus, in the current study, we have deciphered relationships between HIV-2 neutralization sensitivity, co-receptor use and viral envelope glycoprotein (Env) molecular motifs. A panel of primary HIV-2 isolates, with predefined use of co-receptors, was assessed for neutralization sensitivity using a set of HIV-2 Env-directed monoclonal antibodies and co-receptor indicator cell lines. Neutralization sensitivity of the isolates was analysed in relation target cell co-receptor expression, in addition to amino acid motifs and predicted structures of Env regions. Results showed that HIV-2 isolates were more resistant to neutralizing antibodies when entering target cells via the alternative co-receptor GPR15, as compared to CCR5. A similar pattern was noted for isolates using the alternative co-receptor CXCR6. Sensitivity to neutralizing antibodies appeared also to be linked to specific Env motifs in V1/V2 and C3 regions. Our findings suggest that HIV-2 sensitivity to neutralization depends both on which co-receptor is used for cell entry and on specific Env motifs. This study highlights the multifactorial mechanisms behind HIV-2 neutralization sensitivity.

A Stronger Innate Immune Response During Hyperacute Human Immunodeficiency Virus Type 1 (HIV-1) Infection Is Associated With Acute Retroviral Syndrome.

BACKGROUND: Acute retroviral syndrome (ARS) is associated with human immunodeficiency virus type 1 (HIV-1) subtype and disease progression, but the underlying immunopathological pathways are poorly understood. We aimed to elucidate associations between innate immune responses during hyperacute HIV-1 infection (hAHI) and ARS. METHODS: Plasma samples obtained from volunteers (≥18.0 years) before and during hAHI, defined as HIV-1 antibody negative and RNA or p24 antigen positive, from Kenya, Rwanda, Uganda, Zambia, and Sweden were analyzed. Forty soluble innate immune markers were measured using multiplexed assays. Immune responses were differentiated into volunteers with stronger and comparatively weaker responses using principal component analysis. Presence or absence of ARS was defined based on 11 symptoms using latent class analysis. Logistic regression was used to determine associations between immune responses and ARS. RESULTS: Of 55 volunteers, 31 (56%) had ARS. Volunteers with stronger immune responses (n = 36 [65%]) had increased odds of ARS which was independent of HIV-1 subtype, age, and risk group (adjusted odds ratio, 7.1 [95% confidence interval {CI}: 1.7-28.8], P = .003). Interferon gamma-induced protein (IP)-10 was 14-fold higher during hAHI, elevated in 7 of the 11 symptoms and independently associated with ARS. IP-10 threshold >466.0 pg/mL differentiated stronger immune responses with a sensitivity of 84.2% (95% CI: 60.4-96.6) and specificity of 100.0% (95% CI]: 90.3-100.0). CONCLUSIONS: A stronger innate immune response during hAHI was associated with ARS. Plasma IP-10 may be a candidate biomarker of stronger innate immunity. Our findings provide further insights on innate immune responses in regulating ARS and may inform the design of vaccine candidates harnessing innate immunity.

Aerosolization and recovery of viable murine norovirus in an experimental setup.

Noroviruses are the major cause for viral acute gastroenteritis in the world. Despite the existing infection prevention strategies in hospitals, the disease continues to spread and causes extensive and numerous outbreaks. Hence, there is a need to investigate the possibility of airborne transmission of norovirus. In this study, we developed an experimental setup for studies on the infectivity of aerosolized murine norovirus (MNV), a model for the human norovirus. Two aerosol generation principles were evaluated: bubble bursting, a common natural aerosolization mechanism, and nebulization, a common aerosolization technique in laboratory studies. The aerosolization setup was characterized by physical and viral dilution factors, generated aerosol particle size distributions, and the viral infectivity after aerosolization. We found a lower physical dilution factor when using the nebulization generator than with the bubble bursting generator. The viral dilution factor of the system was higher than the physical dilution; however, when comparing the physical and viral dilution factors, bubble bursting generation was more efficient. The infectivity per virus was similar using either generation principle, suggesting that the generation itself had a minor impact on MNV infectivity and that instead, the effect of drying in air could be a major reason for infectivity losses.

Alternative biomarkers for classification of latent tuberculosis infection status in pregnant women with borderline Quantiferon plus results.

Borderline interferon-gamma (IFN-γ) results (near the cut-off level 0.35 IU/ml) occur in QuantiFERON (QFT) assays. We investigated the performance of alternative biomarkers for classification of latent tuberculosis infection (LTBI) status in pregnant women with borderline QFT IFN-γ responses. Pregnant women (n = 96) were identified from a cohort study in Ethiopia, based on QFT-Plus IFN-γ results (QFT-low: <0.20 IU/ml, n = 33; QFT-borderline: 0.20-0.70 IU/ml, n = 31; QFT-high: >0.70 IU/ml, n = 32), including 12 HIV-positive individuals in each group and with 20 HIV-negative non-pregnant women from the same cohort with QFT IFN-γ <0.20 IU/ml as controls. Concentrations of 8 markers (IL-1ra, IL-6, IL-8, IP-10, MCP-1, MCP-2, osteopontin and resistin) were measured in whole blood QFT supernatants, stimulated separately with TB1 and TB2 antigens. K-nearest neighbor analysis (KNN) was used to classify participants with regard to likelihood of LTBI. Concentrations of MCP-2, IP-10 and IL-1ra were higher in QFT-borderline compared to QFT-low participants in both antigen stimulations (p < 0.001). KNN classification indicated high likelihood of LTBI in 13/31 (42%) women with QFT-borderline IFN-γ results. MCP-2, IP-10 and IL-1ra expressed in whole blood after TB antigen stimulation may be considered as alternative biomarkers for classification of LTBI status in pregnant women with borderline QFT IFN-γ results.

Quantification of HIV-2 DNA in Whole Blood.

Time to AIDS infection is longer with HIV-2, compared to HIV-1, but without antiretroviral therapy both infections will cause AIDS-related mortality. In HIV-2 infection, monitoring of antiretroviral treatment (ART) efficacy is challenging since a large proportion of HIV-2-infected individuals displays low or undetectable plasma RNA levels. Hence, quantification of cellular DNA load may constitute an alternative method for monitoring ART efficacy. Moreover, sensitive HIV-2 DNA quantification protocols are also important for the characterization of the HIV-2 reservoirs, and ultimately for the development of HIV-2 cure strategies. We have developed a sensitive and robust HIV-2 DNA quantification protocol based on whole blood as DNA source, including normalization of leukocyte cell numbers using parallel quantification of the single copy porphobilinogen deaminase gene. The specificity and sensitivity of the assay was 100%. The limit of detection was 1 copy and limit of quantification was 5 copies. When applying this protocol to HIV-2 infected, it was found that HIV-2 viral DNA was detectable in individuals in whom viral RNA was undetectable or under quantification level. Thus, this method provides a sensitive approach to HIV-2 DNA viral quantification from whole blood of HIV-2 infected patients.