The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected over 173.3 million people worldwide. Of these, a significant minority have been severe or critical, leading to over 3.7 million deaths worldwide. The resulting illness, called coronavirus disease 2019 (COVID-19), has led to a major global public health crisis; the largest humanity has seen in the last hundred years.
As intensive efforts are on to counter the virus, a new study by an international team of researchers reports the unexpected success of a bacterium in modulating the immune response in the airway and protecting the cells against infection with this virus.
The team’s findings have been published in the journal Pathogens.
SARS-CoV-2 enters the host cell via the angiotensin-converting enzyme 2 (ACE2), expressed on the cells of the respiratory epithelium and the lung. In severe cases, the illness is associated with extensive lung damage and acute respiratory distress syndrome (ARDS), and multi-organ injury, sometimes leading to death.
The severity of the illness is thought to be not only due to the viral cytopathic effects of the virus on the infected cells, but even more, by a hyperactive inflammatory response. The virus appears to inhibit the release of type I and III interferons (IFNs), as well as antiviral factors, promoting severe infection.
Moreover, infection-induced epithelial cell death leads to the release of inflammatory mediators and the migration of inflammatory immune cells into the injured airway tissues. All these release further chemokines and cytokines, and a vicious cycle of increasing inflammation is formed.
In trying to counteract these destructive processes, anti-inflammatory drugs and antivirals have been used, as well as immunomodulatory drugs. The new paper presents another possibility, that of using the respiratory microbiome to reduce the impact of the virus.
The role of the microbiome
A study of COVID-19 patients with a spectrum of severity showed no less than 60 bacterial operational taxonomic units (OTUs) were found only in SARS-CoV-2 patients, mostly from phylum Bacteroidota and Firmicutes. Moreover, Prevotella was the most common bacterial genus to be found only in severe COVID-19 patients, while Dolosigranum species were found in mild COVID-19 in inverse proportion to Prevotella.
Following up on this, the current researchers earlier showed a favorable impact of this bacterium on the innate immune response in the respiratory tract. When given intranasally, the bacterium D. pigrum 040417 not only cleared the virus faster but prevented lung damage due to inflammation. They also found that the beneficial effect of the bacterium was specific to the strain they used.
In mice, the nasal administration of D. pigrum 040417 modulated the innate immune response and improved resistance to both pneumococcal and respiratory syncytial virus (RSV) infections.
In the present paper, they used Calu-3 cells, a human lung epithelial cell line. When incubated with D. pigrum 040417, the epithelial cells in culture were not adversely affected. However, the production of IFN-β and IL-6 was increased, sparing CXCL8. This effect was not seen with D. pigrum 030918.
This experiment is based on the fact that double-stranded ribonucleic acid (dsRNA) is an intermediate during coronavirus replication and transcription. This is detected by cellular antiviral defenses in the respiratory epithelium, triggering cytokine release.
Specifically, pattern recognition receptors (PRRs) in the host recognize dsRNA from the virus, leading to the production of type I and III interferons, the primary antiviral defense. This triggers interferon-stimulated gene (ISG) expression, which in turn activates other antiviral systems.
The coronaviruses are able to hide from these defense systems. In fact, anti-dsRNA systems are less strongly activated by SARS-CoV-2 than with the Sindbis virus, but more than with the earlier Middle East respiratory syndrome coronavirus (MERS-CoV). This could mean that SARS-CoV-2 is not as good as other SARS-like viruses at escaping dsRNA-dependent immune pathways.
If so, the interferon pathway could be used to ramp up the early antiviral defenses to restrict viral replication with this virus.
Modulation of cytokine profile
After incubation with the bacterium D. pigrum, the cells showed an increase in the levels of IFN-β, IL-6 and CXCL8 at baseline. When subsequently stimulated by the Toll-like receptor 3 (TLR3) agonist, polyinosinic:polycytidylic acid poly(I:C), which corresponds to dsRNA, both control and treated cells showed a four- to five-fold increase in the levels of IFN-β and IL-6.
The chemokines CCL5 and CXCL10 were absent at baseline, but were produced following stimulation.
The increase in IFN-β and IL-6 was significantly greater for the cells pre-treated with D. pigrum 040417 at baseline and after stimulation, compared to controls. Conversely, the rises in CXCL8, CCL5 and CXCL10 concentrations were lower in the D. pigrum 040417-treated cells.
D. pigrum 030918 did not induce any change in cytokine levels compared to controls.
Reduced rate of SARS-CoV-2 replication
The researchers also found that SARS-CoV-2 replication within Calu-3 cells was slower after D. pigrum 040417 pre-treatment, accompanied by reduced LDH levels. LDH is a marker of cell damage. Again, D. pigrum 030918 failed to show any beneficial effect.
In untreated cells, the levels of IFN-β and IL-6 increased after infection at 48 hours post-infection, as did CXCL8, CCL5 and CXCL10. However, while the former remained at the same level after 72 hours, the latter continued to increase.
Following incubation with D. pigrum 040417, along with the delay in replication, the cytokine profile also showed a significant change. While IFN-β and IL-6 rose to higher levels, CXCL8 levels dropped at both 48 and 72 hours, while CCL5 and CXCL10 went down at 72 hours.
What are the implications?
For the first time, this study has shown that the nasal administration of D. pigrum 040417 modulates the innate immune response of the nasal epithelium to TLR3 stimulation by poly(I:C) and to SARS-CoV-2 infection. It may be that the presence of beneficial commensal bacteria changes the immunologic characteristics of the respiratory epithelium and thus increases their resistance to some pathogens.
The enhanced IFN-β production with D. pigrum 040417 may be responsible for the lower rate of replication of SARS-CoV-2, by indicating increased efficiency of dsRNA antagonism by the host cell immune pathways.
Late interferon responses are associated with intense inflammation and tissue damage. In fact, some studies have shown that SARS-CoV-2 induces inflammation at an early stage of infection, with high levels of several chemokines, including CXCL8.
High levels of these cytokines are characteristically found in severe or critical COVID-19, indicating a dysregulated inflammatory response at both respiratory and systemic levels. The reduction in CXCL8, CCL5 and CXCL10 in epithelial cells when pre-treated with D. pigrum 040417 might, perhaps, indicate that this bacterium could help prevent such inflammatory damage.
The study also points to the need to identify the most beneficial bacterial strains, as all do not have the same efficacy. In addition, it is noteworthy that D. pigrum 040417 had only a partial effect on viral replication. Thus, there is a need to determine which bacterial strains and genera work together to protect against infection or symptomatic disease.