Ian N. Moore

Prophylaxis With a Middle East Respiratory Syndrome Coronavirus (MERS-CoV)-Specific Human Monoclonal Antibody Protects Rabbits From MERS-CoV Infection.

Abstract With >1600 documented human infections with Middle East respiratory syndrome coronavirus (MERS-CoV) and a case fatality rate of approximately 36%, medical countermeasures are needed to prevent and limit the disease. We examined the in vivo efficacy of the human monoclonal antibody m336, which has high neutralizing activity against MERS-CoV in vitro. m336 was administered to rabbits intravenously or intranasally before infection with MERS-CoV. Prophylaxis with m336 resulted in a reduction of pulmonary viral RNA titers by 40–9000-fold, compared with an irrelevant control antibody with little to no inflammation or viral antigen detected. This protection in rabbits supports further clinical development of m336.

Acidic chitinase primes the protective immune response to gastrointestinal nematodes

Acidic mammalian chitinase (AMCase) is known to be induced by allergens and helminths, yet its role in immunity is unclear. Using AMCase-deficient mice, we show that AMCase deficiency reduced the number of group 2 innate lymphoid cells during allergen challenge but was not required for establishment of type 2 inflammation in the lung in response to allergens or helminths. In contrast, AMCase-deficient mice showed a profound defect in type 2 immunity following infection with the chitin-containing gastrointestinal nematodes Nippostrongylus brasiliensis and Heligmosomoides polygyrus bakeri. The impaired immunity was associated with reduced mucus production and decreased intestinal expression of the signature type 2 response genes Il13, Chil3, Retnlb, and Clca1. CD103+ dendritic cells, which regulate T cell homing, were also reduced in mesenteric lymph nodes of infected AMCase-deficient mice. Thus, AMCase functions as a critical initiator of protective type 2 responses to intestinal nematodes but is largely dispensable for allergic responses in the lung.

Severity of Clinical Disease and Pathology in Ferrets Experimentally Infected with Influenza Viruses Is Influenced by Inoculum Volume

ABSTRACT Ferrets are a valuable model for influenza virus pathogenesis, virus transmission, and antiviral therapy studies. However, the contributions of the volume of inoculum administered and the ferrets respiratory tract anatomy to disease outcome have not been explored. We noted variations in clinical disease outcomes and the volume of inoculum administered and investigated these differences by administering two influenza viruses (A/California/07/2009 [H1N1 pandemic] and A/Minnesota/11/2010 [H3N2 variant]) to ferrets intranasally at a dose of 106 50% tissue culture infective doses in a range of inoculum volumes (0.2, 0.5, or 1.0 ml) and followed viral replication, clinical disease, and pathology over 6 days. Clinical illness and respiratory tract pathology were the most severe and most consistent when the viruses were administered in a volume of 1.0 ml. Using a modified micro-computed tomography imaging method and examining gross specimens, we found that the right main-stem bronchus was consistently larger in diameter than the left main-stem bronchus, though the latter was longer and straighter. These anatomic features likely influence the distribution of the inoculum in the lower respiratory tract. A 1.0-ml volume of inoculum is optimal for delivery of virus to the lower respiratory tract of ferrets, particularly when evaluation of clinical disease is desired. Furthermore, we highlight important anatomical features of the ferret lung that influence the kinetics of viral replication, clinical disease severity, and lung pathology. IMPORTANCE Ferrets are a valuable model for influenza virus pathogenesis, virus transmission, and antiviral therapy studies. Clinical disease in ferrets is an important parameter in evaluating the virulence of novel influenza viruses, and findings are extrapolated to virulence in humans. Therefore, it is highly desirable that the data from different laboratories be accurate and reproducible. We have found that, even when the same virus was administered at similar doses, different investigators reported a range of clinical disease outcomes, from asymptomatic infection to severe weight loss, ocular and nasal discharge, sneezing, and lethargy. We found that a wide range of inoculum volumes was used to experimentally infect ferrets, and we sought to determine whether the variations in disease outcome were the result of the volume of inoculum administered. These data highlight some less explored features of the model, methods of experimental infection, and clinical disease outcomes in a research setting.

Receptor specificity does not affect replication or virulence of the 2009 pandemic H1N1 influenza virus in mice and ferrets.

Human influenza viruses predominantly bind α2,6 linked sialic acid (SA) while avian viruses bind α2,3 SA-containing complex glycans. Virulence and tissue tropism of influenza viruses have been ascribed to this binding preference. We generated 2009 pandemic H1N1 (pH1N1) viruses with either predominant α2,3 or α2,6 SA binding and evaluated these viruses in mice and ferrets. The α2,3 pH1N1 virus had similar virulence in mice and replicated to similar titers in the respiratory tract of mice and ferrets as the α2,6 and WT pH1N1 viruses. Immunohistochemical analysis determined that all viruses infected similar cell types in ferret lungs. There is increasing evidence that receptor specificity of influenza viruses is more complex than the binary model of α2,6 and α2,3 SA binding and our data suggest that influenza viruses use a wide range of SA moieties to infect host cells.

Protective efficacy of influenza group 2 hemagglutinin stem-fragment immunogen vaccines

The stem of the influenza A virus hemagglutinin (HA) is highly conserved and represents an attractive target for a universal influenza vaccine. The 18 HA subtypes of influenza A are phylogenetically divided into two groups, and while protection with group 1 HA stem vaccines has been demonstrated in animal models, studies on group 2 stem vaccines are limited. Thus, we engineered group 2 HA stem-immunogen (SI) vaccines targeting the epitope for the broadly neutralizing monoclonal antibody CR9114 and evaluated vaccine efficacy in mice and ferrets. Immunization induced antibodies that bound to recombinant HA protein and viral particles, and competed with CR9114 for binding to the HA stem. Mice vaccinated with H3 and H7-SI were protected from lethal homologous challenge with X-79 (H3N2) or A/Anhui/1/2013 (H7N9), and displayed moderate heterologous protection. In ferrets, H7-SI vaccination did not significantly reduce weight loss or nasal wash titers after robust 107 TCID50 H7N9 virus challenge. Epitope mapping revealed ferrets developed lower titers of antibodies that bound a narrow range of HA stem epitopes compared to mice, and this likely explains the lower efficacy in ferrets. Collectively, these findings indicate that while group 2 SI vaccines show promise, their immunogenicity and efficacy are reduced in larger outbred species, and will have to be enhanced for successful translation to a universal vaccine.Influenza: Developing a universal vaccine for influenza A group 2Progress has been made towards a universal vaccine targeting the ‘group 2’ subtype of influenza A virus. Today’s flu vaccines target the ‘head’ section of the virus’ hemagglutinin protein; however, this section acquires mutations which require the reformulation of vacccines. In this paper, a vaccine candidate designed to focus an immune response against the more stable protein ‘stem’ is described by a team of scientists led by Kanta Subbarao of the United States’ National Institutes of Health and Raghavan Varadarajan of the Indian Institute of Science. The candidate vaccine offered moderate protection to mice but did not provide significant antiviral effects when tested in ferrets. The authors suggest that, while their approach shows promise, improvement is needed before it could be translated into vaccines against human influenza infection.

Lethal CD4 T Cell Responses Induced by Vaccination Against Staphylococcus aureus Bacteremia

Multiple candidate vaccines against Staphylococcus aureus infections have failed in clinical trials. Analysis of a recent prematurely halted vaccine trial revealed increased mortality rates among vaccine recipients in whom postsurgical S. aureus infection developed, emphasizing the potential for induction of detrimental immune responses and the need to better understand the requirements for protective immunity against S. aureus. These failures of single-antigen vaccines have prompted ongoing development of multicomponent vaccines to target the multitude of S. aureus virulence factors. In the current study, we used lethally irradiated S. aureus as a model multicomponent vaccine and showed that vaccination of mice decreased survival in a bacteremia challenge model. These deleterious effects were due to a CD4 T-cell-dependent interferon γ response and could be prevented by inhibiting development of this response during vaccination. Our results identify the potential for vaccination to induce pathological immune responses, and they have implications for recent vaccine failures and the design of future staphylococcal vaccines.

Mechanism of splenic cell death and host mortality in a Plasmodium yoelii malaria model

Malaria is a fatal disease that displays a spectrum of symptoms and severity, which are determined by complex host-parasite interactions. It has been difficult to study the effects of parasite strains on disease severity in human infections, but the mechanisms leading to specific disease phenotypes can be investigated using strains of rodent malaria parasites that cause different disease symptoms in inbred mice. Using a unique mouse malaria model, here we investigated the mechanisms of splenic cell death and their relationship to control of parasitemia and host mortality. C57BL/6 mice infected with Plasmodium yoelii nigeriensis N67C display high levels of pro-inflammatory cytokines and chemokines (IL-6, IFN-γ, TNF-α, CXCL1, and CCL2) and extensive splenic damage with dramatic reduction of splenic cell populations. These disease phenotypes were rescued in RAG2−/−, IFN-γ−/−, or T cell depleted mice, suggesting IFN-γ and T cell mediated disease mechanisms. Additionally, apoptosis was one of the major pathways involved in splenic cell death, which coincides with the peaks of pro-inflammatory cytokines. Our results demonstrate the critical roles of T cells and IFN-γ in mediating splenic cell apoptosis, parasitemia control, and host lethality and thus may provide important insights for preventing/reducing morbidity associated with severe malaria in humans.

Development of Clade-Specific and Broadly Reactive Live Attenuated Influenza Virus Vaccines against Rapidly Evolving H5 Subtype Viruses

ABSTRACT We have developed pandemic live attenuated influenza vaccines (pLAIVs) against clade 1 H5N1 viruses on an Ann Arbor cold-adapted (ca) backbone that induced long-term immune memory. In 2015, many human infections caused by a new clade (clade 2.2.1.1) of goose/Guangdong (gs/GD) lineage H5N1 viruses were reported in Egypt, which prompted updating of the H5N1 pLAIV. We explored two strategies to generate suitable pLAIVs. The first approach was to modify the hemagglutinin gene of a highly pathogenic wild-type (wt) clade 2.2.1.1 virus, A/Egypt/N03434/2009 (Egy/09) (H5N1), with its unmodified neuraminidase (NA) gene; this virus was designated Egy/09 ca. The second approach was to select a low-pathogenicity avian influenza H5 virus that elicited antibodies that cross-reacted with a broad range of H5 viruses, including the Egypt H5N1 viruses, and contained a novel NA subtype for humans. We selected the low-pathogenicity A/duck/Hokkaido/69/2000 (H5N3) (dk/Hok/00) virus for this purpose. Both candidate vaccines were attenuated and immunogenic in ferrets, inducing antibodies that neutralized homologous and heterologous H5 viruses with different degrees of cross-reactivity; Egy/09 ca vaccine antisera were more specific for the gs/GD lineage viruses but did not neutralize recent North American isolates (clade 2.3.4.4), whereas antisera from dk/Hok/69 ca-vaccinated ferrets cross-reacted with clade 2.3.4.4 and 2.2.1 viruses but not clade 1 or 2.1 viruses. When vaccinated ferrets were challenged with homologous and heterologous H5 viruses, challenge virus replication was reduced in the respiratory tract. Thus, the two H5 pLAIV candidates are suitable for clinical development to protect humans from infection with different clades of H5 viruses. IMPORTANCE In response to the continuing evolution of H5N1 avian influenza viruses and human infections, new candidate H5 live attenuated vaccines were developed by using two different approaches: one targeted a specific circulating strain in Egypt, and the other was based on a virus that elicits broadly cross-reactive antibodies against a wide range of H5 viruses. Both candidate vaccines were immunogenic and exhibited protective efficacy in ferrets. Our study permits a comparison of the two approaches, and the data support the further development of both vaccine viruses to optimally prepare for the further spread of clade 2.2.1 or 2.3.4.4 viruses.

Exposure of rhesus monkeys to cowpox virus Brighton Red by large-particle aerosol droplets results in an upper respiratory tract disease.

We previously demonstrated that small-particle (0.5-3.0 µm) aerosol infection of rhesus monkeys (Macaca mulatta) with cowpox virus (CPXV)-Brighton Red (BR) results in fulminant respiratory tract disease characterized by severe lung parenchymal pathology but only limited systemic virus dissemination and limited classic epidermal pox-like lesion development (Johnson et al., 2015). Based on these results, and to further develop CPXV as an improved model of human smallpox, we evaluated a novel large-particle aerosol (7.0-9.0 µm) exposure of rhesus monkeys to CPXV-BR and monitored for respiratory tract disease by serial computed tomography (CT). As expected, the upper respiratory tract and large airways were the major sites of virus-induced pathology following large-particle aerosol exposure. Large-particle aerosol CPXV exposure of rhesus macaques resulted in severe upper airway and large airway pathology with limited systemic dissemination.

First-in-human topical microbiome transplantation with Roseomonas mucosa for atopic dermatitis

The underlying pathology of atopic dermatitis (AD) includes impaired skin barrier function, susceptibility to Staphylococcus aureus skin infection, immune dysregulation, and cutaneous dysbiosis. Our recent investigation into the potential role of Gram-negative skin bacteria in AD revealed that isolates of one particular commensal, Roseomonas mucosa, collected from healthy volunteers (HVs) improved outcomes in mouse and cell culture models of AD. In contrast, isolates of R. mucosa from patients with AD worsened outcomes in these models. These preclinical results suggested that interventions targeting the microbiome could provide therapeutic benefit for patients with AD. As a first test of this hypothesis in humans, 10 adult and 5 pediatric patients were enrolled in an open-label phase I/II safety and activity trial (the Beginning Assessment of Cutaneous Treatment Efficacy for Roseomonas in Atopic Dermatitis trial; BACTERiAD I/II). Treatment with R. mucosa was associated with significant decreases in measures of disease severity, topical steroid requirement, and S. aureus burden. There were no adverse events or treatment complications. We additionally evaluated differentiating bacterial metabolites and topical exposures that may contribute to the skin dysbiosis associated with AD and/or influence future microbiome-based treatments. These early results support continued evaluation of R. mucosa therapy with a placebo-controlled trial.