Reduced Mortality among COVID-19 ICU Patients after Treatment with HemoClear Convalescent Plasma in Suriname.

Convalescent plasma is a promising therapy for coronavirus disease 2019 (COVID-19), but its efficacy in intensive care unit (ICU) patients in low- and middle-income country settings such as Suriname is unknown. Bedside plasma separation using the HemoClear device made convalescent plasma therapy accessible as a treatment option in Suriname. Two hundred patients with severe SARS-CoV-2 infection requiring intensive care were recruited. Fifty eight patients (29%) received COVID-19 convalescent plasma (CCP) treatment in addition to standard of care (SOC). The CCP treatment and SOC groups were matched by age, sex, and disease severity scores. Mortality in the CCP treatment group was significantly lower than that in the SOC group (21% versus 39%; Fisher's exact test P = 0.0133). Multivariate analysis using ICU days showed that CCP treatment reduced mortality (hazard ratio [HR], 0.35; 95% confidence interval [CI], 0.18 to 0.66; P = 0.001), while complication of acute renal failure (creatinine levels, >110 mol/L; HR, 4.45; 95% CI, 2.54 to 7.80; P < 0.0001) was independently associated with death. Decrease in chest X-ray score in the CCP treatment group (median -3 points, interquartile range [IQR] -4 to -1) was significantly greater than that in the SOC group (median -1 point, IQR -3 to 1, Mann-Whitney test P = 0.0004). Improvement in the PaO2/FiO2 ratio was also significantly greater in the CCP treatment group (median 83, IQR 8 to 140) than in the SOC group (median 35, IQR -3 to 92, Mann-Whitney P = 0.0234). Further research is needed for HemoClear-produced CCP as a therapy for SARS-CoV-2 infection together with adequately powered, randomized controlled trials. IMPORTANCE This study compares mortality and other endpoints between intensive care unit COVID-19 patients treated with convalescent plasma plus standard of care (CCP), and a control group of patients hospitalized in the same medical ICU facility treated with standard of care alone (SOC) in a low- and middle-income country (LMIC) setting using bedside donor whole blood separation by gravity (HemoClear) to produce the CCP. It demonstrates a significant 65% survival improvement in HemoClear-produced CCP recipients (HR, 0.35; 95% CI, 0.19 to 0.66; P = 0.001). Although this is an exploratory study, it clearly shows the benefit of using the HemoClear-produced CCP in ICU patients in the Suriname LMIC setting. Additional studies could further substantiate our findings and their applicability for both LMICs and high-income countries and the use of CCP as a prepared readiness method to combat new viral pandemics.

Relation of phenotype evolution of HIV-1 to envelope V2 configuration.

Biological variability of human immunodeficiency virus type-1 (HIV-1) is involved in the pathogenesis of acquired immunodeficiency syndrome (AIDS). Syncytium-inducing (SI) HIV-1 variants emerge in 50 percent of infected individuals during infection, preceding accelerated CD4+ T cell loss and rapid progression to AIDS. The V1 to V2 and V3 region of the viral envelope glycoprotein gp120 contained the major determinants of SI capacity. The configuration of a hypervariable locus in the V2 domain appeared to be predictive for non-SI to SI phenotype conversion. Early prediction of HIV-1 phenotype evolution may be useful for clinical monitoring and treatment of asymptomatic infection.

Macrophage-tropic variants initiate human immunodeficiency virus type 1 infection after sexual, parenteral, and vertical transmission.

Macrophage-tropic, non-syncytium-inducing, HIV-1 variants predominate in the asymptomatic phase of infection and may be responsible for establishing infection in an individual exposed to the mixture of HIV-1 variants. Here, genotypical and phenotypical characteristics of virus populations, present in sexual, parenteral, or vertical donor-recipient pairs, were studied. Sequence analysis of the V3 domain confirmed the presence of a homogeneous virus population in recently infected individuals. Biological HIV-1 clones were further characterized for syncytium inducing capacity on the MT2 cell line and for macrophage tropism as defined by the appearance of proviral DNA upon inoculation of monocyte-derived macrophages. Both sexual and parenteral transmission cases revealed a selective outgrowth in the recipient of the most macrophage-tropic variant(s) present in the donor. In three out of five vertical transmission cases, more than one highly macrophage-tropic virus variant was present in the child shortly after birth, suggestive of transmission of multiple variants. In three primary infection cases, homogeneous virus populations of macrophage-tropic, non-syncytium-inducing variants were present prior to seroconversion, thus excluding humoral immunity as the selective pressure in favour of macrophage-tropic variants. These observations may have important implications for vaccine development.

Selective inhibition of syncytium-inducing and nonsyncytium-inducing HIV-1 variants in individuals receiving didanosine or zidovudine, respectively.

By studying changes in the clonal composition of HIV-1 populations during the first weeks of zidovudine (ZDV) treatment before the development of ZDV resistance-conferring mutations, we demonstrated previously a selective inhibition of nonsyncytium-inducing (NSI) HIV-1, even when present as coexisting population in individuals also harboring syncytium-inducing (SI) HIV-1. In this study, we observed the opposite in individuals receiving didanosine (ddI) treatment. In these individuals (n = 7) a median -0.98 log change (range -1.55-0.08) in infectious cellular SI load was observed, whereas the coexisting NSI load was only minimally affected (median -0.15 log, range -1.27-0.50; P = 0.03). The virus phenotype-dependent treatment responses were independent of the clonal composition of HIV-1 populations at baseline. Individuals treated with a combination of ZDV and ddI revealed an equal decline of both NSI and SI infectious cellular load (n = 4; NSI: median -1.55 log, range -2.19 to -1.45; SI: median -1.47 log, range -1.81 to -0.86; P = 0.56). To test the hypothesis that the previously reported optimal activation of ZDV and ddI in activated and resting T cells, respectively, in combination with the differential T cell tropism of NSI and SI HIV-1 is the basis for the observed virus phenotype specific efficacy of nucleoside analogs, we studied the effect of treatment with a protease inhibitor that does not require intracellular activation. Individuals receiving ritonavir (n = 4) indeed showed equal declines in NSI and SI infectious cellular load (NSI: median -2.37 log, range -2.59 to -2.16; SI: median -2.82 log, range -3.14 to -2.50; P = 0.25). Our data suggest HIV-1 phenotype as an additional parameter in the design of optimal treatment regimens.

Resistance of surface-dried virus to common disinfection procedures.

It is believed that surface-dried viruses can remain infectious and may therefore pose a threat to public health. To help address this issue, we studied 0.1 N NaOH and 0.1% hypochlorite for their capacity to inactivate surface-dried lipid-enveloped (LE) [human immunodeficiency virus (HIV), bovine viral diarrhoea virus (BVDV) and pseudorabies virus (PRV)] and non-lipid-enveloped [NLE; canine parvovirus (CPV) and hepatitis A virus (HAV)] viruses in a background of either plasma or culture medium. In addition, 80% ethanol was tested on surface-dried LE viruses. Without treatment, surface-dried LE viruses remained infectious for at least one week and NLE viruses for more than one month. Irrespective of the disinfectant, inactivation decreased for viruses dried in plasma, which is more representative of viral contaminated blood than virus in culture medium. Inactivation by all disinfectants improved when preceded by rehydration, although the infectivity of CPV actually increased after rehydration and disinfection may thus be overestimated in the absence of rehydration. This is the first comprehensive study of five important (model) viruses in a surface-dried state showing persistence of infectivity, resistance to three commonly used disinfectants and restoration of susceptibility after rehydration. Our results may have implications for hygiene measurements in the prevention of virus transmission.

Efficient inhibition of both syncytium-inducing and non-syncytium-inducing wild-type HIV-1 by lamivudine in vivo.

OBJECTIVE: To determine the effect of lamivudine (3TC) on syncytium-inducing (SI) and non-SI (NSI) HIV-1 populations in vivo. DESIGN: Responses in virus load and 3TC resistance in 40 3TC-treated subjects were analysed in relation to the presence or absence of SI HIV-1 variants. METHODS: Peripheral blood samples were collected at regular intervals from 40 HIV-1-infected subjects during 3TC treatment. Virus isolates obtained at the start of treatment were typed for SI-capacity by coculture with MT-2 cells. Changes in levels of viral RNA in plasma were determined by quantitative reverse transcriptase PCR. The relative amount of wild-type and mutant virus at codon 184, associated with HIV resistance to 3TC, was determined using a primer-guided nucleotide incorporation assay after amplification of part of the reverse transcriptase gene. In five subjects the frequency of productively infected CD4+ cells with SI or NSI variants was determined in relation to codon 184 genotype. RESULTS: Twenty-six subjects harboured only NSI variants at baseline, whereas 14 subjects also harboured SI variants. Although baseline plasma viral RNA load and CD4 cell counts were different between the two groups, no differences in the response to 3TC therapy were observed for these parameters. In-depth analysis of five subjects showed that the kinetics of virus load changes and emergence of 3TC resistance mutations were similar in plasma and cells, and comparable for the SI and NSI populations present in one individual. CONCLUSIONS: These data show that, in contrast to didanosine and zidovudine, the pressure exerted by 3TC is similar for SI and NSI M184 populations.

Temporal relationship between elongation of the HIV type 1 glycoprotein 120 V2 domain and the conversion toward a syncytium-inducing phenotype.

The second and third variable domains (V2 and V3) of the human immunodeficiency virus type 1 (HIV-1) gp120 envelope molecule have been shown to be determinants of syncytium-inducing (SI) capacity. Previously we have reported evidence that increased length of the V2 domain and duplication or relocation of potential N-linked glycosylation sites in V2 might be used as prognostic markers for evolution toward an SI phenotype. Here, we used a PCR assay that discriminates a 6-nucleotide difference in the length of the V2 domain, with a sensitivity of 1 elongated V2 domain when present in a background of 125 to 625 short V2 domains. Analysis of DNA isolated directly from PBMCs from 11 HIV-1-infected individuals prior to SI phenotype conversion revealed, however, that the usefulness of this PCR for V2 length polymorphism as predictive marker for SI phenotype evolution is limited. The strong association as observed in our previous study between elongation of the V2 domain and an SI phenotype prompted us to expand our first analysis. An extremely significant correlation was observed between V2 length and virus phenotype for samples obtained at about the moment of SI conversion, but not for samples obtained 3 to 35 months after SI phenotype conversion, suggesting that changes in V2 may be only transiently required to allow SI phenotype evolution. This possibly only transient nature of V2 elongation may explain the discrepancy between results by our group and others.

Schistosome circulating anodic antigen in serum of individuals infected with Schistosoma japonicum from the Philippines before and after chemotherapy with praziquantel.

The presence of the schistosome circulating anodic antigen (CAA) in serum of patients infected with Schistosoma japonicum from The Philippines has been investigated using an enzyme-linked immunosorbent assay (ELISA). Serum samples were tested from 48 patients who excreted S. japonicum eggs, 9 individuals with a negative stool examination, and 20 controls with both a negative stool and a negative circumoval precipitin test. No false positive result was detected for the unequivocally negative controls. CAA could be demonstrated in 72.9% of the egg-excreting patients. A positive correlation between parasite burden (eggs per gram of faeces) and antigen level (CAA titre) was found (Spearman's rho = 0.48, P < 0.001, n = 48). Four of 18 sera from the egg-negative individuals were positive in the ELISA. In view of the fact that anti-worm antibodies were also detected in these 4 sera, those reactions suggest active infection not detected by stool examination. In serum from patients treated with praziquantel, a significant drop in CAA titre was seen within 5 d after treatment (Wilcoxon's chi T = -2.23, P = 0.0258, n = 21). In conclusion, the detection of CAA by ELISA in S. japonicum infection can give valuable information in both individual diagnosis and therapeutic drug monitoring, as well as in epidemiological studies or disease control programmes.

Viral safety of C1-inhibitor NF.

We studied the efficacy of virus reduction by three process steps (polyethylene glycol 4000 (PEG) precipitation, pasteurization, and 15nm virus filtration) in the manufacturing of C1-inhibitor NF. The potential prion removing capacity in this process was estimated based on data from the literature. Virus studies were performed using hepatitis A virus (HAV) and human immunodeficiency virus (HIV) as relevant viruses and bovine viral diarrhea virus (BVDV), canine parvovirus (CPV) and pseudorabies virus (PRV) as model viruses, respectively. In the PEG precipitation step, an average reduction in infectious titer of 4.5log(10) was obtained for all five viruses tested. Pasteurization resulted in reduction of infectious virus of >6log(10) for BVDV, HIV, and PRV; for HAV the reduction factor was limited to 2.8log(10) and for CPV it was zero. Virus filtration (15nm) reduced the infectious titer of all viruses by more than 4.5log(10). The overall virus reducing capacity was >16log(10) for the LE viruses. For the NLE viruses CPV and HAV, the overall virus reducing capacities were >8.7 and >10.5log(10), respectively. Based on literature and theoretical assumptions, the prion reducing capacity of the C1-inhibitor NF process was estimated to be >9log(10).

Relation between changes in cellular load, evolution of viral phenotype, and the clonal composition of virus populations in the course of human immunodeficiency virus type 1 infection.

The relationship between the evolution of human immunodeficiency virus type 1 (HIV-1) biologic phenotype, changes in the proportion of infected peripheral blood mononuclear cells, and the relative contribution of non-syncytium-inducing (NSI) and syncytium-inducing (SI) HIV-1 variants to virus load was studied during the course of HIV-1 infection. In 65 HIV-1-infected subjects, the proportion of infected CD4 T cells was higher in persons who carried SI variants. Longitudinal studies revealed that the emergence of SI HIV-1 variants can occur at relatively low numbers of HIV-1-infected cells. Emergence of SI variants frequently coincided with an increase of virus load due to an expansion of both NSI and SI variants, although the contribution of SI viruses to the total virus population significantly increased with time after SI phenotype conversion. These data indicate that NSI to SI phenotype conversion, rather than resulting from high virus load, is part of the sequence of events that leads to increased virus load and CD4 cell depletion.

In vivo HIV-1 infection of CD45RA(+)CD4(+) T cells is established primarily by syncytium-inducing variants and correlates with the rate of CD4(+) T cell decline.

Switch from non-syncytium-inducing (NSI) to syncytium-inducing (SI) HIV type 1 (HIV-1) is associated with accelerated CD4(+) T cell depletion, which might partially be explained by higher virulence of SI variants compared with NSI variants. Because NSI and SI variants use different coreceptors for entry of target cells, altered tropism might offer an explanation for increased pathogenesis associated with SI HIV-1 infection. To investigate whether SI and NSI HIV-1 variants infect different CD4(+) T cell subsets in vivo, the distribution of SI and NSI variants over CD4(+) memory (CD45RA(-)RO(+)) and naive (CD45RA(+)RO(-)) cells was studied by using limiting dilution cultures. In contrast to NSI variants that were mainly present in CD45RO(+) cells, SI variants were equally distributed over CD45RO(+) and CD45RA(+) cells. Infection of memory cells by both NSI and SI HIV-1 and infection of naive cells primarily by SI HIV-1 corresponded closely with the differential cell surface expression of CXCR4 and CCR5. The frequency of SI-infected CD45RA(+) CD4(+) T cells, but not the frequency of NSI- or SI-infected CD45RO(+) CD4(+) T cells, correlated with the rate of CD4(+) T cell depletion. Infection of naive cells by SI HIV-1 may interfere with CD4(+) T cell production and thus account for rapid CD4(+) T cell depletion.

Pathogenesis of primary R5 human immunodeficiency virus type 1 clones in SCID-hu mice.

We studied the replication and cytopathicity in SCID-hu mice of R5 human immunodeficiency virus type 1 (HIV-1) biological clones from early and late stages of infection of three patients who never developed MT-2 cell syncytium-inducing (SI; R5X4 or X4) viruses. Several of the late-stage non-MT-2 cell syncytium-inducing (NSI; R5) viruses from these patients depleted human CD4(+) thymocytes from SCID-hu mice. Earlier clones from the same patients did not deplete CD4(+) thymocytes from SCID-hu mice as well as later clones. We studied three R5 HIV-1 clones from patient ACH142 in greater detail. Two of these clones were obtained prior to the onset of AIDS; the third was obtained following the AIDS diagnosis. In GHOST cell infection assays, all three ACH142 R5 HIV-1 clones could infect GHOST cells expressing CCR5 but not GHOST cells expressing any of nine other HIV coreceptors tested. Furthermore, these patient clones efficiently infected stimulated peripheral blood mononuclear cells from a normal donor but not those from a homozygous CCR5Delta32 individual. Statistical analyses of data obtained from infection of SCID-hu mice with patient ACH142 R5 clones revealed that only the AIDS-associated clone significantly depleted CD4(+) thymocytes from SCID-hu mice. This clone also replicated to higher levels in SCID-hu mice than the two earlier clones, and a significant correlation between viral replication and CD4(+) thymocyte depletion was observed. Our results indicate that an intrinsic property of AIDS-associated R5 patient clones causes their increased replication and cytopathic effects in SCID-hu mice and likely contributes to the development of AIDS in patients who harbor only R5 quasispecies of HIV-1.

Evolution of syncytium-inducing and non-syncytium-inducing biological virus clones in relation to replication kinetics during the course of human immunodeficiency virus type 1 infection.

To investigate the temporal relationship between human immunodeficiency virus type 1 (HIV-1) replicative capacity and syncytium-inducing (SI) phenotype, biological and genetic characteristics of longitudinally obtained virus clones from two HIV-1-infected individuals who developed SI variants were studied. In one individual, the emergence of rapidly replicating SI and non-syncytium-inducing (NSI) variants was accompanied by a loss of the slowly replicating NSI variants. In the other subject, NSI variants were always slowly replicating, while the coexisting SI variants showed an increase in the rate of replication. Irrespective their replicative capacity, the NSI variants remained present throughout the infection in both individuals. Phylogenetic analysis of the V3 region showed early branching of the SI variants from the NSI tree. Successful SI conversion seemed a unique event since no SI variants were found among later-stage NSI variants. This was also confirmed by the increasing evolutionary distance between the two subpopulations. At any time point during the course of the infection, the variation within the coexisting SI and NSI populations did not exceed 2%, indicating continuous competition within each viral subpopulation.

Interaction of pseudomonas aeruginosa with epithelial cells: identification of differentially regulated genes by expression microarray analysis of human cDNAs.

Pseudomonas aeruginosa is an opportunistic pathogen that plays a major role in lung function deterioration in cystic fibrosis patients. To identify critical host responses during infection, we have used high-density DNA microarrays, consisting of 1,506 human cDNA clones, to monitor gene expression in the A549 lung pneumocyte cell line during exposure to P. aeruginosa. We have identified host genes that are differentially expressed upon infection, several of which require interaction with P. aeruginosa and the expression of the major subunit of type IV pili, PilA. Differential expression of genes involved in various cellular functions was identified, and we selected the gene encoding the transcription factor interferon regulatory factor 1 (IRF-1) for further analysis. The levels of the IRF-1 transcript increased 3- to 4-fold in A549 cells after adherence by P. aeruginosa. A similar increase of IRF-1 mRNA was observed in A549 cells exposed to wild-type P. aeruginosa when compared to an isogenic, nonpiliated strain. However, this difference was abolished when serum was present during the incubation of bacteria. Exposure of A549 cells to purified P. aeruginosa lipopolysaccharide did not result in a significant increase in IRF-1 mRNA. Although the P. aeruginosa-induced increased IRF-1 expression depends on the presence of bacterial adhesin, our findings do not preclude the possibility that other bacterial products are responsible for IRF-1 activation, which is enhanced by bacterial adherence to cells. These data show that microarray technology can be an important tool for studying the complex interplay between bacterial pathogens and host.

Transmission of zidovudine-resistant human immunodeficiency virus type 1 variants following deliberate injection of blood from a patient with AIDS: characteristics and natural history of the virus.

We describe the development and persistence of severe immunodeficiency in a previously healthy young woman shortly after she was deliberately injected with blood that was drawn from a patient with AIDS. The heterogenous populations of human immunodeficiency virus (HIV) in donor and recipient were as closely related as those reported in previous cases of proven transmission. The relatively large proportion of non-syncytium-inducing (NSI) clones in the recipient suggests a selective but not complete suppression of syncytium-inducing (SI) viruses. The continuous presence of SI viruses might explain the severe immunosuppression that persisted once the recipient seroconverted. A codon 215 mutation (indicative of zidovudine resistance) was present in SI and NSI clones of the donor and in NSI clones of the recipient. The relative increase in codon 215 resistance mutation in the absence of zidovudine therapy was secondary to the increase in NSI clones. Findings in this case suggest that qualities of an inoculum and/or the route of transmission are important determinants in the subsequent clinical course of HIV disease.

Serum levels of circulating anodic antigen and circulating cathodic antigen detected in mice infected with Schistosoma japonicum or S. mansoni.

Serum concentrations of circulating anodic antigen (CAA) and circulating cathodic antigen (CCA) were studied in mice infected with either Schistosoma japonicum or S. mansoni cercariae. Sera from uninfected mice were negative for both antigens. CAA was detectable in the S. japonicum-infected mice as early as at 2 weeks post-infection (p.i.), and levels were higher in these animals than in the S. mansoni-infected group during the full study period. At the moment of perfusion, 10 weeks p.i., a median of 9 and 29 worms, respectively, were recovered from the S. japonicum- and S. mansoni-infected mice, and the median CAA levels were 326 and 27 ng/ml, respectively. In contrast, CCA levels were much lower in the S. japonicum-infected group (27 ng/ml) as compared with the S. mansoni-infected mice (282 ng/ml). These results suggest an important difference between S. japonicum and S. mansoni infections in CAA and CCA production and/or clearance and indicate a significant role for CAA in the diagnosis of human schistosomiasis japonicum.

Changes in cellular virus load and zidovudine resistance of syncytium-inducing and non-syncytium-inducing human immunodeficiency virus populations under zidovudine pressure: a clonal analysis.

Zidovudine treatment preferentially benefits persons with only non-syncytium-inducing (NSI) human immunodeficiency virus type 1 (HIV-1) variants. To understand this differential efficacy, changes in cellular virus load, clonal composition of HIV-1 populations, and development of resistance-conferring reverse transcriptase mutations were studied in 17 persons initiating zidovudine therapy. Zidovudine treatment resulted in larger and more sustained decreases in cellular virus load in persons with NSI variants only compared with persons also carrying syncytium-inducing (SI) variants. Although the former group had a delayed emergence of resistance mutations, differences in initial responses between the 2 groups were independent of the emergence of resistance mutations. Changes in virus load in subjects also carrying SI variants were due mainly to loss of coexisting NSI virus. Resistance mutations emerged at similar rates in both coexisting variants. Data suggest that mechanisms other than drug resistance are necessary to completely explain the phenotype-dependent benefit of zidovudine.

Temporal relationship between human immunodeficiency virus type 1 RNA levels in serum and cellular infectious load in peripheral blood.

Cross-sectional analysis of 252 paired serum and peripheral blood mononuclear cell (PBMC) samples derived from 54 human immunodeficiency virus type 1 (HIV-1)-infected persons revealed a correlation between HIV-1 RNA load in serum and infectious load in peripheral CD4 T cells after 18 months of follow-up and before an AIDS diagnosis (Pearson's correlation coefficient [r(p)] = .71, P < .001) and during antiviral treatment (r[p] = .78, P < .001). To gain insight into the temporal relationship between both measures of virus load, longitudinally obtained samples from 23 persons with various clinical courses (slow or rapid disease progression, long-term survival) and 22 persons undergoing antiviral therapy (zidovudine or didanosine, or both, or ritonavir) were analyzed. In general, the kinetics of changes in both measures of virus load were similar in the natural course of infection (78% of study participants) and during treatment (82% of participants). These findings suggest that PBMC and serum represent closely related, if not the same, viral compartments.