Faculty Bibliography

In early life, susceptibility to invasive infection skews toward a small subset of microbes, whereas other pathogens associated with diseases later in life, including Streptococcus pneumoniae (Spn), are uncommon among neonates. To delineate mechanisms behind age-dependent susceptibility, we compared age-specific mouse models of invasive Spn infection. We show enhanced CD11b-dependent opsonophagocytosis by neonatal neutrophils improved protection against Spn during early life. The augmented function of neonatal neutrophils was mediated by higher CD11b surface expression at the population level due to dampened efferocytosis, which also resulted in more CD11b^hi "aged" neutrophils in peripheral blood. Dampened efferocytosis during early life could be attributed to the lack of CD169⁺ macrophages in neonates and reduced systemic expressions of multiple efferocytic mediators, including MerTK. On experimentally impairing efferocytosis later in life, CD11b^hi neutrophils increased and protection against Spn improved. Our findings reveal how age-dependent differences in efferocytosis determine infection outcome through the modulation of CD11b-driven opsonophagocytosis and immunity.

Small animal models have been a challenge for the study of SARS-CoV-2 transmission, with most investigators using golden hamsters or ferrets. Mice have the advantages of low cost, wide availability, less regulatory and husbandry challenges, and the existence of a versatile reagent and genetic toolbox. However, adult mice do not robustly transmit SARS-CoV-2. Here we establish a model based on neonatal mice that allows for transmission of clinical SARS-CoV-2 isolates. We characterize tropism, respiratory tract replication and transmission of ancestral WA-1 compared to variants Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Delta (B.1.617.2), Omicron BA.1 and Omicron BQ.1.1. We identify inter-variant differences in timing and magnitude of infectious particle shedding from index mice, both of which shape transmission to contact mice. Furthermore, we characterize two recombinant SARS-CoV-2 lacking either the ORF6 or ORF8 host antagonists. The removal of ORF8 shifts viral replication towards the lower respiratory tract, resulting in significantly delayed and reduced transmission in our model. Our results demonstrate the potential of our neonatal mouse model to characterize viral and host determinants of SARS-CoV-2 transmission, while revealing a role for an accessory protein in this context.

Macrophages facilitate critical functions in regulating pathogen clearance and immune homeostasis in tissues. The remarkable functional diversity exhibited by macrophage subsets is dependent on tissue environment and the nature of the pathological insult. Our current knowledge of the mechanisms that regulate the multifaceted counter-inflammatory responses mediated by macrophages remains incomplete. Here, we report that CD169+ macrophage subsets are necessary for protection under excessive inflammatory conditions. We show that in the absence of these macrophages, even under mild septic conditions, mice fail to survive and exhibit increased production of inflammatory cytokines. Mechanistically, CD169+ macrophages control inflammatory responses via interleukin-10 (IL-10), as CD169+ macrophage-specific deletion of IL-10 was lethal during septic conditions, and recombinant IL-10 treatment reduced lipopolysaccharide (LPS)-induced lethality in mice lacking CD169+ macrophages. Collectively, our findings show a pivotal homeostatic role for CD169+ macrophages and suggest they may serve as an important target for therapy under damaging inflammatory conditions.

Small animal models have been a challenge for the study of SARS-CoV-2 transmission, with most investigators using golden hamsters or ferrets ^1,2 . Mice have the advantages of low cost, wide availability, less regulatory and husbandry challenges, and the existence of a versatile reagent and genetic toolbox. However, adult mice do not transmit SARS-CoV-2 ³ . Here we establish a model based on neonatal mice that allows for transmission of clinical SARS-CoV-2 isolates. We characterize tropism, respiratory tract replication and transmission of ancestral WA-1 compared to variants alpha (B.1.1.7), beta (B.1.351), gamma (P.1), delta (B.1.617.2) and omicron (B.1.1.529). We identify inter-variant differences in timing and magnitude of infectious particle shedding from index mice, both of which shape transmission to contact mice. Furthermore, we characterize two recombinant SARS-CoV-2 lacking either the ORF6 or ORF8 host antagonists. The removal of ORF8 shifts viral replication towards the lower respiratory tract, resulting in significantly delayed and reduced transmission. Our results demonstrate the potential of our neonatal mouse model to characterize viral and host determinants of SARS-CoV-2 transmission, while revealing for the first time a role for an accessory protein this context.

Background: Coronavirus disease 2019 (COVID19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) remains a global health emergency even with effective vaccines and limited FDA-approved therapies. To limit mortality and morbidity across the spectrum of disease, the need for therapeutics remains critical. Galectin9 (gal9) is a beta-galactoside binding protein that modulates cell-cell and cell-matrix interactions. In response to SARS-CoV2 infection, it has been shown that circulating gal9 levels are elevated in patient sera with moderate to severe disease. Additionally, it has been reported that gal9 unexpectedly may competitively bind the host ACE2 receptor, potentially impeding viral entry. Therefore, we hypothesized that early recombinant gal-9 treatment post infection may prevent binding of the virus to susceptible host cells resulting in decreased severity of SARS-CoV2-associated disease.
Method(s): To determine the therapeutic potential of gal9 for treating COVID19, we infected K18-hACE2 transgenic mice intranasally with 10⁴ particle forming units (PFU) of SARS-CoV2. 6 hours post infection (hpi), mice were treated with a single dose of 30 ug of recombinant human gal9 (rhgal9) or PBS intraperitoneally and subsequently monitored 12 days for morbidity. Subgroups of mice were humanely euthanized at 2 and 5 days pi (dpi) for viral plaque assay, flow cytometry, and protein analysis from lung tissue and bronchial alveolar lavage (BAL).
Result(s): We found that mice treated with rhgal9 during the acute phase of infection exhibit improved survival compared to PBS treated animals (25%, p<0.0001). We found that at 5 dpi, rhgal9 treated mice exhibited enhanced viral clearance in the BAL but not in the lung parenchyma. Additionally, we found increased CD8 T cell (p<0.001) and decreased neutrophil (p<0.05) frequencies in the lung at 5 dpi. Finally, we found that BAL fluid had elevated levels of Type 1 Interferon [IFNa (p<0.01) and IFNb (p<0.01)] at 2 dpi and increased MyD88 proinflammatory cytokines [IL1a (p<0.05), IL1b (p<0.01), TNFa (p<0.05), and MIP1a (p<0.05) at 5 dpi.
Conclusion(s): Our study suggests that rhgal9 treatment may be potentially therapeutic for treating acute COVID19. Our data suggest that rhgal9 treatment in combination with other anti-inflammatory mediators may curtail damaging inflammation associated with SARS-CoV2 disease. Further studies are required to determine the optimal time, combination and duration of treatment pi to effectively target the gal9 pathways

Macrophages are known to mediate anti-helminth responses, but it remains uncertain which subsets are involved or how macrophages actually kill helminths. Here, we show rapid monocyte recruitment to the lung after infection with the nematode parasite Nippostrongylus brasiliensis. In this inflamed tissue microenvironment, these monocytes differentiate into an alveolar macrophage (AM)-like phenotype, expressing both SiglecF and CD11c, surround invading parasitic larvae, and preferentially kill parasites in vitro. Monocyte-derived AMs (Mo-AMs) express type 2-associated markers and show a distinct remodeling of the chromatin landscape relative to tissue-derived AMs (TD-AMs). In particular, they express high amounts of arginase-1 (Arg1), which we demonstrate mediates helminth killing through L-arginine depletion. These studies indicate that recruited monocytes are selectively programmed in the pulmonary environment to express AM markers and an anti-helminth phenotype.

Respiratory failure is associated with increased mortality in COVID-19 patients. There are no validated lower airway biomarkers to predict clinical outcome. We investigated whether bacterial respiratory infections were associated with poor clinical outcome of COVID-19 in a prospective, observational cohort of 589 critically ill adults, all of whom required mechanical ventilation. For a subset of 142 patients who underwent bronchoscopy, we quantified SARS-CoV-2 viral load, analysed the lower respiratory tract microbiome using metagenomics and metatranscriptomics and profiled the host immune response. Acquisition of a hospital-acquired respiratory pathogen was not associated with fatal outcome. Poor clinical outcome was associated with lower airway enrichment with an oral commensal (Mycoplasma salivarium). Increased SARS-CoV-2 abundance, low anti-SARS-CoV-2 antibody response and a distinct host transcriptome profile of the lower airways were most predictive of mortality. Our data provide evidence that secondary respiratory infections do not drive mortality in COVID-19 and clinical management strategies should prioritize reducing viral replication and maximizing host responses to SARS-CoV-2.

Mounting a robust immune response against SARS-CoV-2 requires neutralization as well as effector T cell functions. In this issue of Cell Host Microbe, Tauzin et al. characterize the humoral and T cell responses after a single dose of BNT162b2 mRNA vaccine in individuals with or without previous exposure to SARS-CoV-2.

Increasing age is the strongest predictor of risk of COVID-19 severity and mortality. Immunometabolic switch from glycolysis to ketolysis protects against inflammatory damage and influenza infection in adults. To investigate how age compromises defense against coronavirus infection, and whether a pro-longevity ketogenic-diet (KD) impacts immune-surveillance, we developed an aging model of natural murine beta coronavirus (mCoV) infection with mouse hepatitis virus strain-A59 (MHV-A59). When inoculated intranasally, mCoV is pneumotropic and recapitulates several clinical hallmarks of COVID-19 infection. Aged mCoV-A59-infected mice have increased mortality and higher systemic inflammation in the heart, adipose tissue and hypothalamus, including neutrophilia and loss of γδ T cells in lungs. Activation of ketogenesis in aged mice expands tissue protective γδ T cells, deactivates the NLRP3 inflammasome and decreases pathogenic monocytes in lungs of infected aged mice. These data establish harnessing of the ketogenic immunometabolic checkpoint as a potential treatment against coronavirus infection in the aged.

Despite mounting evidence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) engagement with immune cells, most express little, if any, of the canonical receptor of SARS-CoV-2, angiotensin-converting enzyme 2 (ACE2). Here, using a myeloid cell receptor-focused ectopic expression screen, we identified several C-type lectins (DC-SIGN, L-SIGN, LSECtin, ASGR1, and CLEC10A) and Tweety family member 2 (TTYH2) as glycan-dependent binding partners of the SARS-CoV-2 spike. Except for TTYH2, these molecules primarily interacted with spike via regions outside of the receptor-binding domain. Single-cell RNA sequencing analysis of pulmonary cells from individuals with coronavirus disease 2019 (COVID-19) indicated predominant expression of these molecules on myeloid cells. Although these receptors do not support active replication of SARS-CoV-2, their engagement with the virus induced robust proinflammatory responses in myeloid cells that correlated with COVID-19 severity. We also generated a bispecific anti-spike nanobody that not only blocked ACE2-mediated infection but also the myeloid receptor-mediated proinflammatory responses. Our findings suggest that SARS-CoV-2-myeloid receptor interactions promote immune hyperactivation, which represents potential targets for COVID-19 therapy.