Melatonin treatment enhances the efficiency of mice immunization with Venezuelan equine encephalomyelitis virus TC-83

To determine whether treatment with melatonin (MLT) improves the efficiency of immunization against Venezuelan equine encephalomyelitis (VEE) virus, mice were vaccinated with TC-83 VEE virus and treated daily with MLT (1 or 5 mg/kg) starting 3 days before immunization, until 10 days after. IgM antibody titers were determined at days 7, 14, and 21 post-immunization. IL-10 levels were assayed at day 14 postvaccination. Treatment with MLT increased antibody titers 14 days after the immunization. IL-10 levels also increased with MLT treatment (1 and 5 mg/kg). Mice were challenged with live VEE virus at day 21 postimmunization, and viral titers were plaque assayed in chicken embryo fibroblasts 4 days after the infection. Following this challenge brain virus levels were significantly reduced. The results suggest that MLT treatment enhances the efficiency of mice immunization against VEE virus.     

Intranasal type I interferon treatment is beneficial only when administered before clinical signs onset in the SARS-CoV-2 hamster model

Impaired type I interferons (IFNs) production or signaling have been associated with severe COVID-19, further promoting the evaluation of recombinant type I IFNs as therapeutics against SARS-CoV-2 infection. In the Syrian hamster model, we show that intranasal administration of IFN-α starting one day pre-infection or one day post-infection limited weight loss and decreased viral lung titers. By contrast, intranasal administration of IFN-α starting at the onset of symptoms three days post-infection had no impact on the clinical course of SARS-CoV-2 infection. Our results provide evidence that early type I IFN treatment is beneficial, while late interventions are ineffective, although not associated with signs of enhanced disease.     

Neuraminidase inhibitors for treatment of human and avian strain influenza: A comparative modeling study

Treatment of seasonal influenza viral infections using antivirals such as neuraminidase inhibitors (NAIs) has been proven effective if administered within 48 h post-infection. However, there is growing evidence that antiviral treatment of infections with avian-derived strains even as late as 6 days post-infection (dpi) can significantly reduce infection severity and duration. Using a mathematical model of in-host influenza viral infections which can capture the kinetics of both a short-lived, typical, seasonal infection and a severe infection exhibiting sustained viral titer, we explore differences in the effects of NAI treatment on both types of influenza viral infections. Comparison of our model's behavior against experimental data from patients naturally infected with avian strains yields estimates for the times at which patients were infected that are consistent with those reported by the patients, and estimates of drug efficacies that are lower for patients who died than for those who recovered. In addition, our model suggests that the sustained, high, viral titers often seen in more severe influenza virus infections are the reason why antiviral treatment delayed by as much as 6 dpi will still lead to reduced viral titers and shortened illness. We conclude that NAIs may be an effective and beneficial treatment strategy against more severe strains of influenza virus characterized by high, sustained, viral titers. We believe that our mathematical model will be an effective tool in guiding treatment of severe influenza viral infections with antivirals.     

Comparison of Monkeypox Viruses Pathogenesis in Mice by In Vivo Imaging

Monkeypox viruses (MPXV) cause human monkeypox, a zoonotic smallpox-like disease endemic to Africa, and are of worldwide public health and biodefense concern. Using viruses from the Congo (MPXV-2003-Congo-358) and West African (MPXV-2003-USA-044) clades, we constructed recombinant viruses that express the luciferase gene (MPXV-Congo/Luc+and MPXV-USA-Luc+) and compared their viral infection in mice by biophotonic imaging. BALB/c mice became infected by both MPXV clades, but they recovered and cleared the infection within 10 days post-infection (PI). However, infection in severe combined immune deficient (SCID) BALB/c mice resulted in 100% lethality. Intraperitoneal (IP) injection of both MPXV-Congo and MPXV-Congo/Luc+resulted in a systemic clinical disease and the same mean time-to-death at 9 (±0) days post-infection. Likewise, IP injection of SCID-BALB/c mice with MPXV-USA or the MPXV-USA-Luc+, resulted in similar disease but longer (P<0.05) mean time-to-death (11±0 days) for both viruses compared to the Congo strains. Imaging studies in SCID mice showed luminescence in the abdomen within 24 hours PI with subsequent spread elsewhere. Animals infected with the MPXV-USA/Luc+had less intense luminescence in tissues than those inoculated with MPXV-Congo/Luc+, and systemic spread of the MPXV-USA/Luc+virus occurred approximately two days later than the MPXV-Congo/Luc+. The ovary was an important target for viral replication as evidenced by the high viral titers and immunohistochemistry. These studies demonstrate the suitability of a mouse model and biophotonic imaging to compare the disease progression and tissue tropism of MPX viruses.     

Antibodies enhance the infection of phorbol-ester-differentiated human monocyte-like cells with coxsackievirus B4

Coxsackievirus B4 (CV-B4), in presence of antibodies and through a specific viral receptor CAR and Fcγ receptors II and III, can infect monocytes which results in interferon-α synthesis. The antibody-dependent enhancement of CV-B4 infection in the human monocytic-like THP-1 cell line has been investigated. The preincubation of CV-B4 with human plasma or human polyvalent immunoglobulins enhanced the infection of phorbol–myristate–acetate (PMA)-activated THP-1 cell cultures. CV-B4 replicated in these cells as demonstrated by the intracellular detection of infectious particles, viral protein VP1 (immunofluorescence), positive and negative viral RNA (RT-PCR). The viability of infected and control cell cultures was not different up to 20 days post-infection. Activated cell cultures inoculated with CV-B4 harbored intracellular RNA up to 14 days post-infection and produced IFNα that was detected by intracellular immunofluorescence staining as soon as 4 h post-infection with a maximum at 48 h post-infection and by RT-PCR all along the experiment. Together, these data demonstrate that PMA-activated THP-1 cells can be infected with CV-B4, can produce IFNα as a result of interactions between the virus, antibodies and specific receptors. This cellular model can be used to investigate further the mechanism and the result of the antibody-dependent enhancement of CV-B4 infection.     

Protective cellular responses to Burkholderia mallei infection

Burkholderia mallei is a Gram-negative bacillus causing the disease glanders in humans. During intraperitoneal infection, BALB/c mice develop a chronic disease characterised by abscess formation where mice normally die up to 70 days post-infection. Although cytokine responses have been investigated, cellular immune responses to B. mallei infection have not previously been characterised. Therefore, the influx and activation status of splenic neutrophils, macrophages and T cells was examined during infection. Gr-1⁺ neutrophils and F4/80⁺ macrophages infiltrated the spleen 5 h post-infection and an increase in activated macrophages, neutrophils and T cells occurred by 24 h post-infection. Mice depleted of Gr-1⁺ cells were acutely susceptible to B. mallei infection, succumbing to the infection 5 days post-infection. Mice depleted of both CD4 and CD8 T cells did not succumb to the infection until 14 days post-infection. Infected μMT (B cell) and CD28 knockout mice did not differ from wildtype mice whereas iNOS-2 knockout mice began to succumb to the infection 30 days post-infection. The data presented suggests that Gr-1⁺ cells, activated early in B. mallei infection, are essential for controlling the early, innate response to B. mallei infection and T cells or nitric oxide are important during the later stages of infection.     

Efficiency of viral entry determines the capacity of murine erythroleukemia cells to support persistent infections by mammalian reoviruses.

To determine mechanisms by which persistent viral infections are established and maintained, we initiated persistent infections of murine erythroleukemia (MEL) cells by using reovirus strains type 3 Abney and type 3 Dearing. Establishment of persistent reovirus infections of MEL cells was not associated with a significant cytopathic effect despite the presence of high titers of infectious virus in the cultures (>10(5) PFU/ml of culture lysate). Maintenance of persistently infected MEL-cell cultures was associated with coevolution of mutant viruses and cells. Mutant viruses produced greater yields than the parental wild-type (wt) strains in MEL cells cured of persistent infection and in cells treated with ammonium chloride, a weak base that blocks viral disassembly. Mutant cells supported growth of wt infectious subvirion particles, which are disassembly intermediates generated in vitro by treatment of virions with chymotrypsin, substantially better than growth of wt virions. These findings indicate that viral and cellular mutations selected during maintenance of persistently infected MEL-cell cultures affect acid-dependent proteolysis of virions during entry into cells. We also found that wt infectious subvirion particles produce greater yields than wt virions in wt MEL cells, which suggests that inefficient viral disassembly in MEL cells favors establishment of persistent infection. Therefore, steps in reovirus replication leading to viral disassembly appear to be critical determinants of the capacity of MEL cells to support both establishment and maintenance of persistent reovirus infections.     

Therapeutic potential of the methanolic extract of Lepidium sativum seeds on mice infected with Trypanosoma evansi

The present study aimed to investigate the therapeutic potential of the methanolic extract of Lepidium sativum seeds in mice experimentally infected with Trypanosoma evansi. A total of thirty-two male Swiss albino mice were randomly divided into four groups: the first group was the normal control, while the second, third and fourth groups were infected intraperitoneally with 1 × 10⁴ trypanosomes. The third and fourth groups were treated with 100 μl of Lepidium sativum seed extract (LSSE) at a dose of 200 mg/kg body weight intraperitoneally (infected + LSSEI) and orally (infected + LSSEO) respectively, once a day, for a period of four days.Parasitaemia was found to be significantly raised in the untreated infected group, reaching 2 × 10⁷ at day 4 post-infection, but was significantly reduced by 65.5% and 88% in the mice treated orally and intraperitoneally with LSSE, respectively. The erythrocyte count, HCT, haemoglobin content, leucocyte count and the percentage of lymphocytes was significantly reduced in the untreated infected group, while the treatment with LSSE returned these parameters to their pre-infection values. In addition, our study proved that LSSE provided protection against liver tissue damage and decreased the levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT). The present study also established that intraperitoneal injection of LSSE is more effective than oral administration in the treatment of trypanosome infection in mice. In conclusion, the infection caused haematological, biochemical and histological changes that were ameliorated following treatment with LSSE.     

Negative Regulation of Type I IFN Expression by OASL1 Permits Chronic Viral Infection and CD8+ T-Cell Exhaustion

The type I interferons (IFN-Is) are critical not only in early viral control but also in prolonged T-cell immune responses. However, chronic viral infections such as those of human immunodeficiency virus (HIV) and hepatitis C virus (HCV) in humans and lymphocytic choriomeningitis virus (LCMV) in mice overcome this early IFN-I barrier and induce viral persistence and exhaustion of T-cell function. Although various T-cell-intrinsic and -extrinsic factors are known to contribute to induction of chronic conditions, the roles of IFN-I negative regulators in chronic viral infections have been largely unexplored. Herein, we explored whether 2′–5′ oligoadenylate synthetase-like 1 (OASL1), a recently defined IFN-I negative regulator, plays a key role in the virus-specific T-cell response and viral defense against chronic LCMV. To this end, we infected Oasl1 knockout and wild-type mice with LCMV CL-13 (a chronic virus) and monitored T-cell responses, serum cytokine levels, and viral titers. LCMV CL-13-infected Oasl1 KO mice displayed a sustained level of serum IFN-I, which was primarily produced by splenic plasmacytoid dendritic cells, during the very early phase of infection (2–3 days post-infection). Oasl1 deficiency also led to the accelerated elimination of viremia and induction of a functional antiviral CD8 T-cell response, which critically depended on IFN-I receptor signaling. Together, these results demonstrate that OASL1-mediated negative regulation of IFN-I production at an early phase of infection permits viral persistence and suppresses T-cell function, suggesting that IFN-I negative regulators, including OASL1, could be exciting new targets for preventing chronic viral infection.     

Favipiravir treatment prolongs the survival in a lethal mouse model intracerebrally inoculated with Jamestown Canyon virus

BackgroundJamestown Canyon virus (JCV) is a mosquito-borne orthobunyavirus that causes acute febrile illness, meningitis, and meningoencephalitis, primarily in North American adults. Currently, there are no available vaccines or specific treatments against JCV infections.Methodology/Principal findingsThe antiviral efficacy of favipiravir (FPV) against JCV infection was evaluated in vitro and in vivo in comparison with that of ribavirin (RBV) and 2’-fluoro-2’-deoxycytidine (2’-FdC). The in vitro inhibitory effect of these drugs on JCV replication was evaluated in Vero and Neuro-2a (N2A) cells. The efficacy of FPV in the treatment of JCV infection in vivo was evaluated in C57BL/6J mice inoculated intracerebrally with JCV, as per the survival, viral titers in the brain, and viral RNA load in the blood. The 90% inhibitory concentrations (IC₉₀) of FPV, RBV, and 2’-FdC were 41.0, 61.8, and 13.6 μM in Vero cells and 20.7, 25.8, and 8.8 μM in N2A cells, respectively. All mice infected with 1.0×10⁴ TCID₅₀ died or were sacrificed within 10 days post-infection (dpi) without treatment. However, mice treated with FPV for 5 days [initiated either 2 days prior to infection (−2 dpi–2 dpi) or on the day of infection (0 dpi–4 dpi)] survived significantly longer than control mice, administered with PBS (p = 0.025 and 0.011, respectively). Moreover, at 1 and 3 dpi, the virus titers in the brain were significantly lower in FPV-treated mice (0 dpi–4 dpi) versus PBS-treated mice (p = 0.002 for both 1 and 3 dpi).Conclusions/SignificanceAlthough the intracerebral inoculation route is thought to be a challenging way to evaluate drug efficacy, FPV inhibits the in vitro replication of JCV and prolongs the survival of mice intracerebrally inoculated with JCV. These results will enable the development of a specific antiviral treatment against JCV infections and establishment of an effective animal model.     

Subacute SARS-CoV-2 replication can be controlled in the absence of CD8+ T cells in cynomolgus macaques

SARS-CoV-2 infection presents clinical manifestations ranging from asymptomatic to fatal respiratory failure. Despite the induction of functional SARS-CoV-2-specific CD8⁺ T-cell responses in convalescent individuals, the role of virus-specific CD8⁺ T-cell responses in the control of SARS-CoV-2 replication remains unknown. In the present study, we show that subacute SARS-CoV-2 replication can be controlled in the absence of CD8⁺ T cells in cynomolgus macaques. Eight macaques were intranasally inoculated with 10⁵ or 10⁶ TCID₅₀ of SARS-CoV-2, and three of the eight macaques were treated with a monoclonal anti-CD8 antibody on days 5 and 7 post-infection. In these three macaques, CD8⁺ T cells were undetectable on day 7 and thereafter, while virus-specific CD8⁺ T-cell responses were induced in the remaining five untreated animals. Viral RNA was detected in nasopharyngeal swabs for 10–17 days post-infection in all macaques, and the kinetics of viral RNA levels in pharyngeal swabs and plasma neutralizing antibody titers were comparable between the anti-CD8 antibody treated and untreated animals. SARS-CoV-2 RNA was detected in the pharyngeal mucosa and/or retropharyngeal lymph node obtained at necropsy on day 21 in two of the untreated group but undetectable in all macaques treated with anti-CD8 antibody. CD8⁺ T-cell responses may contribute to viral control in SARS-CoV-2 infection, but our results indicate possible containment of subacute viral replication in the absence of CD8⁺ T cells, implying that CD8⁺ T-cell dysfunction may not solely lead to viral control failure.     

Targeted Deletion of FGL2 Leads to Increased Early Viral Replication and Enhanced Adaptive Immunity in a Murine Model of Acute Viral Hepatitis Caused by LCMV WE

Mounting effective innate and adaptive immune responses are critical for viral clearance and the generation of long lasting immunity. It is known that production of inhibitory factors may result in the inability of the host to clear viruses, resulting in chronic viral persistence. Fibrinogen-like protein 2 (FGL2) has been identified as a novel effector molecule of CD4⁺CD25⁺ Foxp3⁺ regulatory T (Treg) cells that inhibits immune activity by binding to FCγRIIB expressed primarily on antigen presenting cells (APC). In this study, we show that infection of mice with Lymphocytic Choriomeningitis Virus WE (LCMV WE) leads to increased plasma levels of FGL2, which were detected as early as 2 days post-infection (pi) and persisted until day 50 pi. Mice deficient in FGL2 (fgl2^−/−) had increased viral titers of LCMV WE in the liver early p.i but cleared the virus by day 12 similar to wild type mice. Dendritic cells (DC) isolated from the spleens of LCMV WE infected fgl2^−/− had increased expression of the DC maturation markers CD80 and MHC Class II compared to wild type (fgl2^+/+). Frequencies of CD8⁺ and CD4⁺ T cells producing IFNγ in response to ex vivo peptide re-stimulation isolated from the spleen and lymph nodes were also increased in LCMV WE infected fgl2 ^−/− mice. Increased frequencies of CD8⁺ T cells specific for LCMV tetramers GP₃₃ and NP₃₉₆ were detected within the liver of fgl2^−/− mice. Plasma from fgl2^−/− mice contained higher titers of total and neutralizing anti-LCMV antibody. Enhanced anti-viral immunity in fgl2^−/− mice was associated with increased levels of serum alanine transaminase (ALT), hepatic necrosis and inflammation following LCMV WE infection. These data demonstrate that targeting FGL2 leads to early increased viral replication but enhanced anti-viral adaptive T & B cell responses. Targeting FGL2 may enhance the efficacy of current anti-viral therapies for hepatotropic viruses.     

Oxidative stress in mice infected with Angiostrongylus cantonensis coincides with enhanced glutathione-dependent enzymes activity

This study aimed to estimate reactive oxygen species (ROS) production, antioxidants activity, and biomarkers level of oxidative damage to protein and DNA in the cerebrospinal fluid (CSF) of C57BL/6 mice infected with Angiostrongylus cantonensis. The mean ROS concentration in the CSF of infected mice increased gradually, and the increase in ROS in CSF became statistical significance at days 12-30 post-infection compared to that before infection (P < 0.001), and then ROS returned to normal level at day 45 after infection. In parallel with the increase in ROS in the CSF, infected mice showed similar of changes in reduced glutathione (GSH), glutathione reductase (GR), glutathione peroxidase (GPx), and glutathione S-transferase (GST) as that in ROS in the CSF. GSH, GR, GPx, and GST in the CSF of infected mice were all significantly higher than they were before infection during days 12-30 post-infection. However, protein carbonyl content and 8-hydroxy-2′-deoxyguanosine, biomarkers of oxidative damage to protein and DNA, respectively, were also significantly higher in the CSF of infected mice during this period. These results suggest that oxidative stress occur in the cells of central nervous system of mice infected with A. cantonensis during days 12-30 after infection due to ROS overproduction in CSF despite the increase in antioxidants during this period.     

The multi-functional reovirus σ3 protein is a virulence factor that suppresses stress granule formation and is associated with myocardial injury

The mammalian orthoreovirus double-stranded (ds) RNA-binding protein σ3 is a multifunctional protein that promotes viral protein synthesis and facilitates viral entry and assembly. The dsRNA-binding capacity of σ3 correlates with its capacity to prevent dsRNA-mediated activation of protein kinase R (PKR). However, the effect of σ3 binding to dsRNA during viral infection is largely unknown. To identify functions of σ3 dsRNA-binding activity during reovirus infection, we engineered a panel of thirteen σ3 mutants and screened them for the capacity to bind dsRNA. Six mutants were defective in dsRNA binding, and mutations in these constructs cluster in a putative dsRNA-binding region on the surface of σ3. Two recombinant viruses expressing these σ3 dsRNA-binding mutants, K287T and R296T, display strikingly different phenotypes. In a cell-type dependent manner, K287T, but not R296T, replicates less efficiently than wild-type (WT) virus. In cells in which K287T virus demonstrates a replication deficit, PKR activation occurs and abundant stress granules (SGs) are formed at late times post-infection. In contrast, the R296T virus retains the capacity to suppress activation of PKR and does not mediate formation of SGs at late times post-infection. These findings indicate that σ3 inhibits PKR independently of its capacity to bind dsRNA. In infected mice, K287T produces lower viral titers in the spleen, liver, lungs, and heart relative to WT or R296T. Moreover, mice inoculated with WT or R296T viruses develop myocarditis, whereas those inoculated with K287T do not. Overall, our results indicate that σ3 functions to suppress PKR activation and subsequent SG formation during viral infection and that these functions correlate with virulence in mice.     

Antioxidant metabolism in pearl millet genotypes during compatible and incompatible interaction with downy mildew pathogen

In this study, the ascorbic acid content, lipid peroxidation product, reactive oxygen generation and scavenging enzyme activities were determined in pearl millet [Pennisetum glaucum (L.) R.Br.] leaves. These parameters were analysed at two stages: (i) pre-infection [45 days after sowing (DAS)] and (ii) post-infection [7 days after infection (DAI), i.e. 57 DAS]. Lipid peroxidation product (malondialdehyde content) was recorded higher in compatible interaction at pre-infection stage while it was increased in incompatible interaction at post-infection stage. Resistant genotypes had higher ascorbic acid content at both the stages of analysis. Superoxide dismutase (SOD) activity was higher in susceptible genotypes at pre-infection but after infection it was found to be higher in resistant genotypes. Ascorbate peroxidase, catalase (CAT) and lipoxygenase activities were higher in resistant genotypes at both the stages of analysis. Native PAGE isozyme banding pattern of SOD, CAT, APX and esterase showed some inducible band(s) due to disease infection.     

Avian Influenza Infection Alters Fecal Odor in Mallards

Changes in body odor are known to be a consequence of many diseases. Much of the published work on disease-related and body odor changes has involved parasites and certain cancers. Much less studied have been viral diseases, possibly due to an absence of good animal model systems. Here we studied possible alteration of fecal odors in animals infected with avian influenza viruses (AIV). In a behavioral study, inbred C57BL/6 mice were trained in a standard Y-maze to discriminate odors emanating from feces collected from mallard ducks (Anas platyrhynchos) infected with low-pathogenic avian influenza virus compared to fecal odors from non-infected controls. Mice could discriminate odors from non-infected compared to infected individual ducks on the basis of fecal odors when feces from post-infection periods were paired with feces from pre-infection periods. Prompted by this indication of odor change, fecal samples were subjected to dynamic headspace and solvent extraction analyses employing gas chromatography/mass spectrometry to identify chemical markers indicative of AIV infection. Chemical analyses indicated that AIV infection was associated with a marked increase of acetoin (3-hydroxy-2-butanone) in feces. These experiments demonstrate that information regarding viral infection exists via volatile metabolites present in feces. Further, they suggest that odor changes following virus infection could play a role in regulating behavior of conspecifics exposed to infected individuals.     

Use of a Recombinant Vaccinia Virus Expressing Interferon Gamma for Post-Exposure Protection against Vaccinia and Ectromelia Viruses

Post-exposure vaccination with vaccinia virus (VACV) has been suggested to be effective in minimizing death if administered within four days of smallpox exposure. While there is anecdotal evidence for efficacy of post-exposure vaccination this has not been definitively studied in humans. In this study, we analyzed post-exposure prophylaxis using several attenuated recombinant VACV in a mouse model. A recombinant VACV expressing murine interferon gamma (IFN-γ) was most effective for post-exposure protection of mice infected with VACV and ectromelia virus (ECTV). Untreated animals infected with VACV exhibited severe weight loss and morbidity leading to 100% mortality by 8 to 10 days post-infection. Animals treated one day post-infection had milder symptoms, decreased weight loss and morbidity, and 100% survival. Treatment on days 2 or 3 post-infection resulted in 40% and 20% survival, respectively. Similar results were seen in ECTV-infected mice. Despite the differences in survival rates in the VACV model, the viral load was similar in both treated and untreated mice while treated mice displayed a high level of IFN-γ in the serum. These results suggest that protection provided by IFN-γ expressed by VACV may be mediated by its immunoregulatory activities rather than its antiviral effects. These results highlight the importance of IFN-γ as a modulator of the immune response for post-exposure prophylaxis and could be used potentially as another post-exposure prophylaxis tool to prevent morbidity following infection with smallpox and other orthopoxviruses.     

Influenza virus inoculum volume is critical to elucidate age‐dependent mortality in mice

The elderly exhibit increased mortality to influenza viral infection for unclear reasons. Mice are frequently used to model how aging impacts disease. Several studies have shown that aged mice exhibit an increased mortality to influenza virus, but two recent studies demonstrated the opposite. These two studies administered the virus intranasally in 20 µL, whereas the other studies used a viral inoculum in at least 30 µL. To determine whether the volume of the inoculum could explain the conflicting reports, we infected young and aged mice via intranasal instillation of 40 µL or 20 µL containing 1 x 10⁴ plaque‐forming units (PFU) of H1N1 influenza virus. We found that intranasal administration of 40 µL but not 20 µL of inoculum resulted in age‐dependent mortality in mice. Compared to aged mice infected with 40 µL inoculum, those infected with 20 µL inoculum showed reduced levels of live virus and IFN‐β in the lung 3 days postinfection. Furthermore, aged mice administered 40 µL of Evans blue intranasally displayed increased dye retention in their bronchoalveolar lavage fluid compared to those administered 20 µL of Evans blue. Our data demonstrate that the inoculating volume of virus is critical for adequate delivery of influenza virus to the lung and thus for efficient infection of aged mice. These findings shed light on discrepant results in the literature regarding aged mice and influenza infection, and establish that mice can be used to examine how aging impacts the response to this biomedically important infection.     

Persistent and Compartmentalised Disruption of Dendritic Cell Subpopulations in the Lung following Influenza A Virus Infection

Immunological homeostasis in the respiratory tract is thought to require balanced interactions between networks of dendritic cell (DC) subsets in lung microenvironments in order to regulate tolerance or immunity to inhaled antigens and pathogens. Influenza A virus (IAV) poses a serious threat of long-term disruption to this balance through its potent pro-inflammatory activities. In this study, we have used a BALB/c mouse model of A/PR8/34 H1N1 Influenza Type A Virus infection to examine the effects of IAV on respiratory tissue DC subsets during the recovery phase following clearance of the virus. In adult mice, we found differences in the kinetics and activation states of DC residing in the airway mucosa (AMDC) compared to those in the parenchymal lung (PLDC) compartments. A significant depletion in the percentage of AMDC was observed at day 4 post-infection that was associated with a change in steady-state CD11b⁺ and CD11b⁻ AMDC subset frequencies and significantly elevated CD40 and CD80 expression and that returned to baseline by day 14 post-infection. In contrast, percentages and total numbers of PLDC were significantly elevated at day 14 and remained so until day 21 post-infection. Accompanying this was a change in CD11b⁺and CD11b⁻ PLDC subset frequencies and significant increase in CD40 and CD80 expression at these time points. Furthermore, mice infected with IAV at 4 weeks of age showed a significant increase in total numbers of PLDC, and increased CD40 expression on both AMDC and PLDC, when analysed as adults 35 days later. These data suggest that the rate of recovery of DC populations following IAV infection differs in the mucosal and parenchymal compartments of the lung and that DC populations can remain disrupted and activated for a prolonged period following viral clearance, into adulthood if infection occurred early in life.     

Virulence-Dependent Alterations in the Kinetics of Immune Cells during Pulmonary Infection by Mycobacterium tuberculosis

A better understanding of the kinetics of accumulated immune cells that are involved in pathophysiology during Mycobacterium tuberculosis (Mtb) infection may help to facilitate the development of vaccines and immunological interventions. However, the kinetics of innate and adaptive cells that are associated with pathogenesis during Mtb infection and their relationship to Mtb virulence are not clearly understood. In this study, we used a mouse model to compare the bacterial burden, inflammation and kinetics of immune cells during aerogenic infection in the lung between laboratory-adapted strains (Mtb H37Rv and H37Ra) and Mtb K strain, a hyper-virulent W-Beijing lineage strain. The Mtb K strain multiplied more than 10- and 3.54-fold more rapidly than H37Ra and H37Rv, respectively, during the early stage of infection (at 28 days post-infection) and resulted in exacerbated lung pathology at 56 to 112 days post-infection. Similar numbers of innate immune cells had infiltrated, regardless of the strain, by 14 days post-infection. High, time-dependent frequencies of F4/80^-CD11c⁺CD11b^-Siglec-H⁺PDCA-1⁺ plasmacytoid DCs and CD11c^-CD11b⁺Gr-1^int cells were observed in the lungs of mice that were infected with the Mtb K strain. Regarding adaptive immunity, Th1 and Th17 T cells that express T-bet and RORγt, respectively, significantly increased in the lungs that were infected with the laboratory-adapted strains, and the population of CD4⁺CD25⁺Foxp3⁺ regulatory T cells was remarkably increased at 112 days post-infection in the lungs of mice that were infected with the K strain. Collectively, our findings indicate that the highly virulent Mtb K strain may trigger the accumulation of pDCs and Gr1^intCD11b⁺ cells with the concomitant down-regulation of the Th1 response and the maintenance of an up-regulated Th2 response without inducing a Th17 response during chronic infection. These results will help to determine which immune system components must be considered for the development of tuberculosis (TB) vaccines and immunological interventions.