Primary Viremia Research Articles

Viral modulation of type II interferon increases T cell adhesion and virus spread

During primary varicella zoster virus (VZV) infection, infected lymphocytes drive primary viremia, causing systemic dissemination throughout the host, including the skin. This results in cytokine expression, including interferons (IFNs), which partly limit infection. VZV also spreads from skin keratinocytes to lymphocytes prior to secondary viremia. It is not clear how VZV achieves this while evading the cytokine response. Here, we show that VZV glycoprotein C (gC) binds IFN-γ and modifies its activity, increasing the expression of a subset of IFN-stimulated genes (ISGs), including intercellular adhesion molecule 1 (ICAM1), chemokines and immunomodulatory genes. The higher ICAM1 protein level at the plasma membrane of keratinocytes facilitates lymphocyte function-associated antigen 1-dependent T cell adhesion and expression of gC during infection increases VZV spread to peripheral blood mononuclear cells. This constitutes the discovery of a strategy to modulate IFN-γ activity, upregulating a subset of ISGs, promoting enhanced lymphocyte adhesion and virus spread.

Safety surveillance for PrEP in pregnant and breastfeeding women.

The risk of HIV acquisition is higher during pregnancy and postpartum than other times. Newly acquired maternal HIV infection associated with high primary viraemia, substantially increases the risk of vertical HIV transmission. Pre-exposure prophylaxis (PrEP) reduces the risk of HIV acquisition. Currently available products include oral tenofovir/emtricitabine (TDF/FTC) and tenofovir alafenamide (TAF)/FTC), long-acting cabotegravir (CAB-LA) and the dapivirine ring (DVR). All except oral TDF/FTC have limited safety data available for use in pregnant and breastfeeding women. The safety of new PrEP agents for pregnant women and the fetus, infant and child, either exposed in utero or during breastfeeding is an ongoing concern for health care workers and pregnant and breastfeeding women, particularly as the safety risk appetite for antiretroviral (ARV) agents used as PrEP is lower in pregnant and breastfeeding women who are HIV-uninfected, compared to women living with HIV taking ARVs as treatment. With the widespread rollout of TDF/FTC among pregnant women in South Africa and other low-middle income countries (LMIC) and the potential introduction of new PrEP agents for pregnant women, there is a need for safety surveillance systems to identify potential signals of risk to either the mother or fetus, measure the burden of such a risk, and where appropriate, provide specific reassurance to PrEP users. Safety data needs to be collected across the continuum of the product life cycle from pre-licensure into the post-marketing period, building a safety profile through both passive and active surveillance systems, recognising the strengths and limitations of each, and the potential for bias and confounding. Pharmacovigilance systems that aim to assess the risk of adverse birth outcomes in pregnant women exposed to PrEP and other agents need to consider the special requirements of pregnancy epidemiology to ensure that the data derived from surveillance are sufficiently robust to inform treatment policies. Here we review the known safety profiles of currently available PrEP candidates in women of child-bearing potential, pregnancy and breastfeeding and discuss pragmatic approaches for such surveillance in HIV-endemic LMICs.

Features of the pathogenesis, epidemiology, diagnostics, approaches to the treatment and prevention of small pox, the current state of the problem in Ukraine and the world

Introduction. Monkey pox is an anthropozoonous disease caused by orthopoxvirus, characterized by fever, general intoxication and skin rash, in some cases it can lead to fatal consequences; an infectious disease covered by international health regulations (IHR). The causative agent of the disease is classified as a biological agent - agent of a biological weapon. Most of the monkeypox cases associated with the 2022 outbreak are in the Western Hemisphere. Purpose: study of data on etiological, pathogenetic, epidemiological aspects, modern approaches to the diagnosis, treatment and prevention of monkeypox, the state of the problem in Ukraine and the world. Materials and methods: to perform the task, a review of publications was conducted in electronic databases of medical publications PubMed Medline of the US National Library of Medicine (NLM), Cochrane Database of Systematic Reviews, CDC (Centers for Disease Control and Prevention) website of the federal agency of the Ministry of Health USA. Results: The second generation ACAM2000 and third generation JYNNEOS™ monkeypox vaccines are currently available. Both vaccines are able to create immunity against monkeypox. Post-exposure vaccination can help prevent or reduce the severity of monkeypox. ACAM2000 is a single-dose vaccinia vaccine licensed by the FDA for the prevention of smallpox. Full immunity is achieved 4 weeks after vaccination. FDA expanded access to investigational protocol allows use of ACAM2000 for monkeypox immunization during an outbreak. The immunogenicity and protective efficacy of ACAM2000 is equivalent to that of the first generation Dryvax® smallpox vaccine. Smallpox vaccine virus is used as a live attenuated virus in the smallpox vaccine. Cases of encephalitis and Guillain-Barré syndrome following smallpox vaccination show that the incidence varies not only from place to place, but also from year to year in the same place. For the purposes of containment of the outbreak, prevention to reduce the risk of transmission from person to person, immediate detection of new cases and surveillance measures are necessary. Health care workers who provide care to patients with suspected or confirmed monkeypox who work with their specimens should use standard precautions to control infection. Specimens taken from animals or humans suspected of having monkeypox should be handled in equipped laboratories by trained personnel. According to WHO recommendations, samples taken from patients should be appropriately labeled, prepared for transportation using triple packaging. To prevent the further spread of monkeypox through the animal trade, captive animals potentially infected with smallpox must be isolated from other animals and quarantined for 30 days. Conclusions: 1.The monkeypox virus, which used to be endemic to certain regions of Africa, is now a global concern, with cases increasingly being reported in regions of the Western Hemisphere. 2. In connection with the fact that transmission from person to person most often occurs through airborne droplets or through direct contact with the mucous secretions of an infected person, social distancing and contact tracing are advisable. 3. In the last 6 months, cases of monkeypox have been confirmed among middle-aged people, which can be explained by the loss of cross-immunity from the smallpox vaccine. 4. The monkeypox virus multiplies in the cytoplasm and matures, creating primary viremia. Manifestations of a severe course of monkeypox are bronchopneumonia, respiratory distress syndrome, encephalitis, chorioretinal scars on the retina, etc. To minimize the risk of complications, it is advisable to provide timely treatment. 5. Medical organizations around the world are focused on understanding how cases of monkeypox are increasingly occurring in countries in the Western Hemisphere. 6. The study of etiotropic methods of treating monkey pox is promising.

Heparin: A simplistic repurposing to prevent SARS-CoV-2 transmission in light of its in-vitro nanomolar efficacy

The world is currently facing a novel coronavirus (SARS-CoV-2) pandemic. The greatest threat that is disrupting the normal functioning of society is the exceptionally high species independent transmission. Drug repurposing is understood to be the best strategy to immediately deploy well-characterized agents against new pathogens. Several repurposable drugs are already in evaluation for determining suitability to treat COVID-19. One such promising compound includes heparin, which is widely used in reducing thrombotic events associated with COVID-19 induced pathology. As part of identifying target-specific antiviral compounds among FDA and world-approved libraries using high-throughput virtual screening (HTVS), we previously evaluated top hits for anti-SARS-CoV-2 activity. Here, we report results of highly efficacious viral entry blocking properties of heparin (IC50 = 12.3 nM) in the complete virus assay, and further, propose ways to use it as a potential transmission blocker. Exploring further, our in-silico analysis indicated that the heparin interacts with post-translational glycoconjugates present on spike proteins. The patterns of accessible spike-glycoconjugates in open and closed states are completely contrasted by one another. Heparin-binding to the open conformation of spike structurally supports the state and may aid ACE2 binding as reported with cell surface-bound heparan sulfate. We also studied spike protein mutant variants' heparin interactions for possible resistance. Based on available data and optimal absorption properties by the skin, heparin could potentially be used to block SARS-CoV-2 transmission. Studies should be designed to exploit its nanomolar antiviral activity to formulate heparin as topical or inhalation-based formulations, particularly on exposed areas and sites of primary viremia e.g. ACE2 rich epithelia of the eye (conjunctiva/lids), nasal cavity, and mouth.

Brief Report: Understanding Preferences for HIV Care Among Patients Experiencing Homelessness or Unstable Housing: A Discrete Choice Experiment.

Homelessness and unstable housing (HUH) negatively impact care outcomes for people living with HIV (PLWH). To inform the design of a clinic program for PLWH experiencing HUH, we quantified patient preferences and trade-offs across multiple HIV-service domains using a discrete choice experiment (DCE). We sequentially sampled PLWH experiencing HUH presenting at an urban HIV clinic with ≥1 missed primary care visit and viremia in the last year to conduct a DCE. Participants chose between 2 hypothetical clinics varying across 5 service attributes: care team "get to know me as a person" versus not; receiving $10, $15, or $20 gift cards for clinic visits; drop-in versus scheduled visits; direct phone communication to care team versus front-desk staff; and staying 2 versus 20 blocks from the clinic. We estimated attribute relative utility (ie, preference) using mixed-effects logistic regression and calculated the monetary trade-off of preferred options. Among 65 individuals interviewed, 61% were >40 years old, 45% White, 77% men, 25% heterosexual, 56% lived outdoors/emergency housing, and 44% in temporary housing. Strongest preferences were for patient-centered care team [β = 3.80; 95% confidence interval (CI): 2.57 to 5.02] and drop-in clinic appointments (β = 1.33; 95% CI: 0.85 to 1.80), with a willingness to trade $32.79 (95% CI: 14.75 to 50.81) and $11.45 (95% CI: 2.95 to 19.95) in gift cards/visit, respectively. In this DCE, PLWH experiencing HUH were willing to trade significant financial gain to have a personal relationship with and drop-in access to their care team rather than more resource-intensive services. These findings informed Ward 86's "POP-UP" program for PLWH-HUH and can inform "ending the HIV epidemic" efforts.

Primary Epstein-Barr Virus Infection With Marked Cholestasis, Warm Autoimmune Hemolytic Anemia, and Pancytopenia

Abstract Primary Epstein-Barr virus (EBV) infection may rarely present with cholestatic hepatitis or hemolytic anemia. We share a case of EBV-associated infectious mononucleosis presenting with a unique triad of findings comprising cholestatic hepatitis with marked hyperbilirubinemia, warm autoimmune hemolytic anemia, and pancytopenia. Early diagnostic challenges included negative Monospot heterophile testing. Additional testing revealed EBV serologies consistent with acute primary infection, EBV viremia and bone marrow biopsy showing EBV-encoded RNA–positive cells. A tapered prednisone course was administered for warm autoimmune hemolytic anemia. Improvement in symptoms and laboratory findings occurred over 3 weeks.

HUBUNGAN IMUNOGLOBULIN M (IgM) DAN IMUNOGLOBULIN G (IgG) DENGAN DERAJAT KEPARAHAN INFEKSI DENGUE DI RSUD Dr. M. HAULUSSY DAN RS SUMBER HIDUP AMBON PERIODE 2018

The dengue virus that has been injected through the bite of the Aedes mosquito will spread through the blood and cause primary viremia. This situation will stimulate the body's immune system to eliminate dengue infection, including the secretion of immunoglobulin M (IgM) and immunoglobulin G (IgG). Different immune system mechanisms will have an impact on the severity of dengue infection. This study aims to determine the relationship between IgM and IgG with the severity of dengue infection patients in RSUD. Dr. M. Haulussy and RS. Sumber Hidup Ambon in 2018. This research is an analytic cross-sectional approach that uses secondary data. The number of samples was 26 patients with purposive sampling. The result showed primary infections were at mild severity, while the secondary infection was found all severity. The analytic concluded there is no relation between IgM and IgG with the severity of dengue infection (p=0,237).

Early Treatment of Coxsackievirus B3-Infected Animals With Soluble Coxsackievirus-Adenovirus Receptor Inhibits Development of Chronic Coxsackievirus B3 Cardiomyopathy.

Coxsackie-B-viruses (CVB) are frequent causes of acute myocarditis and dilated cardiomyopathy, but an effective antiviral therapy is still not available. Previously, we and others have demonstrated that treatment with an engineered sCAR-Fc (soluble coxsackievirus-adenovirus receptor fused to the carboxyl-terminus of human IgG) efficiently neutralizes CVB3 and inhibits the development of cardiac dysfunction in mice with acute CVB3-induced myocarditis. In this study, we analyzed the potential of sCAR-Fc for treatment of chronic CVB3-induced myocarditis in an outbred NMRI mouse model. NMRI mice were infected with the CVB3 strain 31-1-93 and treated with a sCAR-Fc expressing adeno-associated virus 9 vector 1, 3, and 7 days after CVB3 infection. Chronic myocarditis was analyzed on day 28 after infection. Initial investigations showed that NMRI mice develop pronounced chronic myocarditis between day 18 and day 28 after infection with the CVB3 strain 31-1-93. Chronic cardiac infection was characterized by inflammation and fibrosis as well as persistence of viral genomes in the heart tissue and by cardiac dysfunction. Treatment of NMRI mice resulted in a distinct reduction of cardiac inflammation and fibrosis and almost complete elimination of virus RNA from the heart by day 28 after infection. Moreover, hemodynamic measurement revealed improved cardiac contractility and diastolic relaxation in treated mice compared with mice treated with a control vector (mean±SD; maximal pressure, 81.9±9.2 versus 69.4±8.6 mm Hg, P=0.02; left ventricular ejection fraction, 68.9±8.5 versus 54.2±11.5%, P=0.02; dP/dtmax, 7275.2±1674 versus 4432.6±1107 mm Hg/s, P=0.004; dP/dtmin, -4046.9±776 versus -3146.3±642 mm Hg/s, P=0.046). The therapeutic potential of sCAR-Fc is limited, however, since postponed start of sCAR-Fc treatment either 3 or 7 days after infection could not attenuate myocardial injury. Early therapeutic employment of sCAR-Fc, initiated at the beginning of the primary viremia, inhibits the development of chronic CVB3-induced myocarditis and improves the cardiac function to a level equivalent to that of uninfected animals.

Zika virus: persisting tissue reservoirs and their potential for clinically relevant pathology

The rapid spread of Zika virus through the Caribbean and Americas appears associated with greater levels of congenital zika and Guillain-Barre syndromes than previously seen. In addition, new disease pathologies such as persisting sexual transmission and severe liver injury have been identified. With zika vaccines years away from licensing an understanding of potential long-term health consequences following adult infection needs to be gained, especially where organs from infected people may be used for transplantation. To model the pathology of human Zika virus infection Old World and New World non-human primates were sub-cutaneously infected with a Caribbean Zika virus isolate. Following termination during primary viremia or later time points post peripheral viral clearance FFPE tissue sections were analysed for Zika virus RNA (RNAscope) and host responses (immunohistochemistry). Zika virus RNA was detected in multiple tissues from 3dpc with low levels of viral RNA detectable by RNAscope through to 101 dpc. New World non-human primates retained higher levels of persisting virus within tissues, notably within spleen, small intestine, kidney, liver and genital tissues. Differing pathologies were observed with Zika virus detected clustered within kidney glomeruli and associated with liver inflammatory infiltrates within New World species. Such pathologies are being documented in adults with resolved primary infection symptoms. This includes clinical complications following solid organ transplantation where immune suppression may allow viral reactivation either from the patient or transplanted organ. In the absence of a protective vaccine the potential risks from persisting viral reservoirs needs to be understood to ensure appropriate clinical management.

Zika virus: out of sight but not out of the mind. Viral persistence and host responses within the central and peripheral nervous systems

Following the rapid spread of Zika virus infection through the Caribbean and Americas, model systems of Zika virus infection have been established to determine disease pathology and potential health impacts for those infected with this neurotropic flavivirus. NIBSC has established both Old World (rhesus and cynomolgus macaques) and New World (red-bellied tamarins) non-human primate disease models of human Zika virus infection. Animals were infected sub-cutaneously with the Caribbean Zika isolate PRVABC59 and pairs terminated during primary viremia (3 dpc) or post clearance of plasma viremia (42 and 101 dpc). FFPE sections from multiple tissues, including brain and peripheral nerves were analysed for Zika virus RNA (RNAscope), viral proteins and host responses (immunohistochemistry). All animals became infected and rapidly cleared high levels of peripheral viremia. Zika virus RNA was detected in multiple tissues from 3 dpc with low levels of viral RNA and viral NS1 protein remaining detectable through to 101 dpc. Viral RNA was associated with both glial and neuronal cells throughout the brain and schwann cell nuclei in peripheral nerves. Neuroinflammation (astrogliosis, microgliosis, peri-vascular cuffing) and increased levels of CD3+, CD8+ or CD45+ cells were present from 3 dpc through to 101 dpc. Disruption of MAP2 neuronal dendrite staining and peripheral nerve myelin basic protein staining was also observed. The continued detection of Zika virus within tissues 3 months post infection and in the absence of a detectable peripheral viremia raises questions regarding potential long term effects of this virus and associated inflammatory responses within the nervous systems.

Velogenic newcastle disease virus tissue tropism and pathogenesis of infection in chickens by application of in situ PCR, immunoperoxase staining and HE staining

Limited deep studies are available in the field of early stages of pathogenesis of Newcastle disease virus (NDV) infection and tissue tropism of NDV. In this study, 24 specific pathogen free (SPF) chickens of white leghorn breed were infected with Newcastle disease (ND) by intranasal administration of 10⁵ 50% EID50/0.1 mL of velogenic NDV (vNDV). A second group of 15 chickens were kept as a control group. Chickens were monitored every day to record clinical signs. Infected chickens were euthanized by cervical dislocation at successive times, namely at hours (hrs) 2, 4, 6, 12, days 1, 2, 4, and 6 post-inoculation (pi). Whereas, control group chickens were euthanized on days 0, 1, 2, 4, and 6 pi. Tissues of brain, trachea, lung, caecal tonsil, liver, kidney, spleen, heart, proventriculus, intestine, and thymus were collected, fixed in 10% buffered formalin, embedded in paraffin, and sectioned. HS staining, immunoperoxidase staining (IPS) and in situ PCR were applied. It was concluded that at hr 2 pi, virus seemed to be inclined to trachea and respiratory tract. Meanwhile, it attacked caecal tonsils, intestine and bursa of Fabricus. While primary viraemia was ongoing, virus created footing in kidney and thymus. At hr 4 pi, proventriculus, liver, and spleen were attacked. However, at hr 6 pi, brain and heart were involved. Secondary viraemia probably started as early as hr 12 pi since all collected tissues were positive. Tissue tropism was determined in trachea, caecal tonsil, liver, bursa of Fabricius, intestine, proventriculus, lung, spleen, thymus, kidney, heart, and brain.

Analysis of the immunological biomarker profile during acute Zika virus infection reveals the overexpression of CXCL10, a chemokine linked to neuronal damage.

BACKGROUNDInfection with Zika virus (ZIKV) manifests in a broad spectrum of disease ranging from mild illness to severe neurological complications and little is known about Zika immunopathogenesis.OBJECTIVESTo define the immunologic biomarkers that correlate with acute ZIKV infection.METHODSWe characterized the levels of circulating cytokines, chemokines, and growth factors in 54 infected patients of both genders at five different time points after symptom onset using microbeads multiplex immunoassay; comparison to 100 age-matched controls was performed for statistical analysis and data mining.FINDINGSZIKV-infected patients present a striking systemic inflammatory response with high levels of pro-inflammatory mediators. Despite the strong inflammatory pattern, IL-1Ra and IL-4 are also induced during the acute infection. Interestingly, the inflammatory cytokines IL-1β, IL-13, IL-17, TNF-α, and IFN-γ; chemokines CXCL8, CCL2, CCL5; and the growth factor G-CSF, displayed a bimodal distribution accompanying viremia. While this is the first manuscript to document bimodal distributions of viremia in ZIKV infection, this has been documented in other viral infections, with a primary viremia peak during mild systemic disease and a secondary peak associated with distribution of the virus to organs and tissues.MAIN CONCLUSIONSBiomarker network analysis demonstrated distinct dynamics in concurrence with the bimodal viremia profiles at different time points during ZIKV infection. Such a robust cytokine and chemokine response has been associated with blood-brain barrier permeability and neuroinvasiveness in other flaviviral infections. High-dimensional data analysis further identified CXCL10, a chemokine involved in foetal neuron apoptosis and Guillain-Barré syndrome, as the most promising biomarker of acute ZIKV infection for potential clinical application.

Hepatic immunopathology during occult hepacivirus re-infection

Despite drug advances for Hepatitis C virus (HCV), re-infections remain prevalent in high-risk populations. Unfortunately, the role of preexisting viral immunity and how it modulates re-infection is unclear. GBV-B infection of common marmosets is a useful model to study tissue immune responses in hepacivirus infections, and in this study we re-challenged 4 animals after clearance of primary viremia. Although only low-to-absent viremia was observed following re-challenge, GBV-B viral RNA was detectable in liver, confirming re-infection. Microscopic hepatic lesions indicated severe-to-mild lymphocyte infiltration and fibrosis in 3 out of 4 animals. Further, GBV-B-specific T cells were elevated in animals with moderate-to-severe hepatopathology, and up to 3-fold increases in myeloid dendritic and activated natural killer cells were observed after infection. Our data indicate that occult hepacivirus re-infections occur and that new liver pathology is possible even in the presence of anti-hepacivirus T cells and in the absence of high viremia.

Intracerebroventricular and Intravascular Injection of Viral Particles and Fluorescent Microbeads into the Neonatal Brain.

In the study on the pathogenesis of viral encephalitis, the infection method is critical. The first of the two main infectious routes to the brain is the hematogenous route, which involves infection of the endothelial cells and pericytes of the brain. The second is the intracerebroventricular (ICV) route. Once within the central nervous system (CNS), viruses may spread to the subarachnoid space, meninges, and choroid plexus via the cerebrospinal fluid. In experimental models, the earliest stages of CNS viral distribution are not well characterized, and it is unclear whether only certain cells are initially infected. Here, we have analyzed the distribution of cytomegalovirus (CMV) particles during the acute phase of infection, termed primary viremia, following ICV or intravascular (IV) injection into the neonatal mouse brain. In the ICV injection model, 5 µl of murine CMV (MCMV) or fluorescent microbeads were injected into the lateral ventricle at the midpoint between the ear and eye using a 10-µl syringe with a 27 G needle. In the IV injection model, a 1-ml syringe with a 35 G needle was used. A transilluminator was used to visualize the superficial temporal (facial) vein of the neonatal mouse. We infused 50 µl of MCMV or fluorescent microbeads into the superficial temporal vein. Brains were harvested at different time points post-injection. MCMV genomes were detected using the in situ hybridization method. Fluorescent microbeads or green fluorescent protein expressing recombinant MCMV particles were observed by fluorescent microscopy. These techniques can be applied to many other pathogens to investigate the pathogenesis of encephalitis.

Biphasic CD8+ T-Cell Defense in Simian Immunodeficiency Virus Control by Acute-Phase Passive Neutralizing Antibody Immunization.

Identifying human immunodeficiency virus type 1 (HIV-1) control mechanisms by neutralizing antibodies (NAbs) is critical for anti-HIV-1 strategies. Recent in vivo studies on animals infected with simian immunodeficiency virus (SIV) and related viruses have shown the efficacy of postinfection NAb passive immunization for viremia reduction, and one suggested mechanism is its occurrence through modulation of cellular immune responses. Here, we describe SIV control in macaques showing biphasic CD8(+) cytotoxic T lymphocyte (CTL) responses following acute-phase NAb passive immunization. Analysis of four SIVmac239-infected rhesus macaque pairs matched with major histocompatibility complex class I haplotypes found that counterparts receiving day 7 anti-SIV polyclonal NAb infusion all suppressed viremia for up to 2 years without accumulating viral CTL escape mutations. In the first phase of primary viremia control attainment, CD8(+) cells had high capacities to suppress SIVs carrying CTL escape mutations. Conversely, in the second, sustained phase of SIV control, CTL responses converged on a pattern of immunodominant CTL preservation. During this sustained phase of viral control, SIV epitope-specific CTLs showed retention of phosphorylated extracellular signal-related kinase (ERK)(hi)/phosphorylated AMP-activated protein kinase (AMPK)(lo) subpopulations, implying their correlation with SIV control. The results suggest that virus-specific CTLs functionally boosted by acute-phase NAbs may drive robust AIDS virus control. In early HIV infection, NAb responses are lacking and CTL responses are insufficient, which leads to viral persistence. Hence, it is important to identify immune responses that can successfully control such HIV replication. Here, we show that monkeys receiving NAb passive immunization in early SIV infection strictly control viral replication for years. Passive infusion of NAbs with CTL cross-priming capacity resulted in induction of functionally boosted early CTL responses showing enhanced suppression of CTL escape mutant virus replication. Accordingly, the NAb-infused animals did not show accumulation of viral CTL escape mutations during sustained SIV control, and immunodominant CTL responses were preserved. This early functional augmentation of CTLs by NAbs provides key insights into the design of lasting and viral escape mutation-free protective immunity against HIV-1 infection.

Comparison of Monkeypox Virus Clade Kinetics and Pathology within the Prairie Dog Animal Model Using a Serial Sacrifice Study Design.

Monkeypox virus (MPXV) infection of the prairie dog is valuable to studying systemic orthopoxvirus disease. To further characterize differences in MPXV clade pathogenesis, groups of prairie dogs were intranasally infected (8 × 103 p.f.u.) with Congo Basin (CB) or West African (WA) MPXV, and 28 tissues were harvested on days 2, 4, 6, 9, 12, 17, and 24 postinfection. Samples were evaluated for the presence of virus and gross and microscopic lesions. Virus was recovered from nasal mucosa, oropharyngeal lymph nodes, and spleen earlier in CB challenged animals (day 4) than WA challenged animals (day 6). For both groups, primary viremia (indicated by viral DNA) was seen on days 6–9 through day 17. CB MPXV spread more rapidly, accumulated to greater levels, and caused greater morbidity in animals compared to WA MPXV. Histopathology and immunohistochemistry (IHC) findings, however, were similar. Two animals that succumbed to disease demonstrated abundant viral antigen in all organs tested, except for brain. Dual-IHC staining of select liver and spleen sections showed that apoptotic cells (identified by TUNEL) tended to colocalize with poxvirus antigen. Interestingly splenocytes were labelled positive for apoptosis more often than hepatocytes in both MPXV groups. These findings allow for further characterization of differences between MPXV clade pathogenesis, including identifying sites that are important during early viral replication and cellular response to viral infection.

PD-1 Analysis on CD28−CD27− CD4 T Cells Allows Stimulation-Independent Assessment of CMV Viremic Episodes in Transplant Recipients

Expression of the inhibitory receptor programmed death 1 (PD-1) on cytomegalovirus (CMV)-specific CD4 T cells defines a phenotype associated with CMV viremia in transplant recipients. Moreover, CD28−CD27− double negativity is known as a typical phenotype of CMV-specific CD4 T cells. Therefore, the co-expression of inhibitory receptors on CD28−CD27− CD4 T cells was assessed as a rapid, stimulation-independent parameter for monitoring CMV complications after transplantation. Ninety-three controls, 67 hemodialysis patients and 81 renal transplant recipients were recruited in a cross-sectional and longitudinal manner. CMV-specific CD4 T cell levels quantified after stimulation were compared to levels of CD28−CD27− CD4 T cells. PD-1 and cytotoxic T lymphocyte–associated antigen 4 (CTLA-4) expression on CD28−CD27− CD4 T cells were related to viremia. A percentage of ≥0.44% CD28−CD27− CD4 T cells defined CMV seropositivity (93.3% sensitivity, 97.1% specificity), and their frequencies correlated strongly with CMV-specific CD4 T cell levels after stimulation (r = 0.73, p < 0.0001). Highest PD-1 expression levels on CD28−CD27− CD4 T cells were observed in patients with primary CMV viremia and reactivation (p < 0.0001), whereas CTLA-4 expression was only elevated during primary CMV viremia (p < 0.05). Longitudinal analysis showed a significant increase in PD-1 expression in relation to viremia (p < 0.001), whereas changes in nonviremic patients were nonsignificant. In conclusion, increased PD-1 expression on CD28−CD27− CD4 T cells correlates with CMV viremia in transplant recipients and may serve as a specific, stimulation-independent parameter to guide duration of antiviral therapy.

Repressive Effect of Primary Virus Replication on Superinfection Correlated with Gut-Derived Central Memory CD4+ T Cells in SHIV-Infected Chinese Rhesus Macaques

A possible mechanism of susceptibility to superinfection with simian-human immunodeficiency virus (SHIV)-1157ipd3N4 was explored in twelve SHIVSF162P3-infected Chinese rhesus macaques. Based on the kinetics of viral replication for the second infecting virus following SHIV-1157ipd3N4 inoculation, the monkeys were divided into two groups: those relatively resistant to superinfection (SIR) and those relatively sensitive to superinfection (SIS). We found that superinfection-resistant macaques had high primary viremia, whereas superinfection-sensitive macaques had low primary viremia, suggesting that primary SHIVSF162P3 infection with a high viral-replication level would repress superinfection with a heterologous SHIV-1157ipd3N4. Although no correlation of protection against superinfection with virus-specific CD4+ T cell or CD8+ T cell immune responses from gut was observed prior to superinfection, superinfection susceptibility was strongly correlated with CD4+ Tcm cells from gut both prior to the second infecting virus inoculation and on day 7 after superinfection, but not with CD4+ Tem cells from gut or with CD4+ Tcm cells from peripheral blood and lymph node. These results point to the important roles of gut-derived CD4+ Tcm cells for the study of the mechanisms of protection against superinfection and the evaluation of the safety and efficacy of vaccines and therapies against acquired immune deficiency syndrome (AIDS).

Infectious feline herpesvirus detected in distant bone and tendon following mucosal inoculation of specific pathogen-free cats

Emerging evidence suggests that cats infected with feline herpesvirus-1 (FHV-1) may experience a brief viremic phase. The objective of this study was to determine whether natural routes of FHV-1 inoculation could result in viremic transmission of infectious virus to connective tissues (cortical bone, tendon). Three specific pathogen-free cats were experimentally inoculated with FHV-1 via a combined mucosal (oronasal, ocular) route. Cats were euthanized at the peak of clinical signs to aseptically harvest tissues (cortical bone, tendon, trachea/tongue) for co-culture with a susceptible cell line to promote spread of infectious virus. Viral infection of Crandall-Rees feline kidney cells was microscopically visualized by cytopathic effect (CPE). Additionally, co-culture DNA was extracted either at the point of CPE or 16 days of culture without evidence of CPE, to amplify FHV-1 glycoprotein B gene using real-time PCR. Infectious virus was detected in distant cortical bone (two cats, moderate to severe clinical signs) and tendon (one cat, severe clinical signs). Direct infection of mucosal (trachea, tongue) tissues also was confirmed in these two cats. In contrast, all co-cultured tissues from the third cat (mild clinical signs) were negative for FHV-1 by CPE and PCR. Results of this study demonstrated that early primary FHV-1 viremia may be distributed to distant connective tissues.

Suppression of the Interferon and NF-κB Responses by Severe Fever with Thrombocytopenia Syndrome Virus

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease characterized by high fever, thrombocytopenia, multiorgan dysfunction, and a high fatality rate between 12 and 30%. It is caused by SFTS virus (SFTSV), a novel Phlebovirus in family Bunyaviridae. Although the viral pathogenesis remains largely unknown, hemopoietic cells appear to be targeted by the virus. In this study we report that human monocytes were susceptible to SFTSV, which replicated efficiently, as shown by an immunofluorescence assay and real-time reverse transcription-PCR. We examined host responses in the infected cells and found that antiviral interferon (IFN) and IFN-inducible proteins were induced upon infection. However, our data also indicated that downregulation of key molecules such as mitochondrial antiviral signaling protein (MAVS) or weakened activation of interferon regulatory factor (IRF) and NF-κB responses may contribute to a restricted innate immunity against the infection. NSs, the nonstructural protein encoded by the S segment, suppressed the beta interferon (IFN-β) and NF-κB promoter activities, although NF-κB activation appears to facilitate SFTSV replication in human monocytes. NSs was found to be associated with TBK1 and may inhibit the activation of downstream IRF and NF-κB signaling through this interaction. Interestingly, we demonstrated that the nucleoprotein (N), also encoded by the S segment, exhibited a suppressive effect on the activation of IFN-β and NF-κB signaling as well. Infected monocytes, mainly intact and free of apoptosis, may likely be implicated in persistent viral infection, spreading the virus to the circulation and causing primary viremia. Our findings provide the first evidence in dissecting the host responses in monocytes and understanding viral pathogenesis in humans infected with a novel deadly Bunyavirus.