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A Note on the Potential BCG Vaccination – COVID-19 Molecular Link

The goal of our report is to stimulate the sharply focused follow-up experimental studies of the Bacillus CalmetteGuérin (BCG) COVID-19 axis. The emerging data suggest that there is a previously unrecognized link between the past and current tuberculosis vaccine BCG (a live attenuated strain of Mycobacterium bovis) immunization policies (summarized in The BCG World Atlas, www.bcgatlas.org) and COVID-19. Mandatory BCG vaccination appears to alleviate the severity of SARS-CoV-2 pathology in infected patients [1]. Most probably, SARS-CoV-2, a betacoronavirus originated from either bats and/or pangolins [25], is a causative agent of COVID-19 pandemic. The most recent studies using publicly available data of COVID-19 in 199 countries/regions suggested that BCG vaccination may have hindered the overall spread of the virus or clinical manifestation of the disease or both [6]. As of today, there is no plausible molecular explanation of this phenomenon. We hypothesized that, in addition to a general immunity boost caused by BCG vaccination, there is a level of sequence homology between the viral and BCG proteins and that this homology contributes to the disease alleviation in BCGvaccinated patients relative to a non-immune cohort.


INTRODUCTION
The goal of our report is to stimulate the sharply focused follow-up experimental studies of the Bacillus Calmette-Guérin (BCG) -COVID-19 axis. The emerging data suggest that there is a previously unrecognized link between the past and current tuberculosis vaccine BCG (a live attenuated strain of Mycobacterium bovis) immunization policies (summarized in The BCG World Atlas, www.bcgatlas.org) and COVID-19. Mandatory BCG vaccination appears to alleviate the severity of SARS-CoV-2 pathology in infected patients [1]. Most probably, SARS-CoV-2, a betacoronavirus originated from either bats and/or pangolins [2][3][4][5], is a causative agent of COVID-19 pandemic. The most recent studies using publicly available data of COVID-19 in 199 countries/regions suggested that BCG vaccination may have hindered the overall spread of the virus or clinical manifestation of the disease or both [6]. As of today, there is no plausible molecular explanation of this phenomenon. We hypothesized that, in addition to a general immunity boost caused by BCG vaccination, there is a level of sequence homology between the viral and BCG proteins and that this homology contributes to the disease alleviation in BCGvaccinated patients relative to a non-immune cohort.

RESULTS
To elucidate a possible mechanism that links BCG vaccination with SARS-CoV-2, we determined the level of homology between the proteins encoded by the viral and BCG genomes using the blastx program. In contrast with other sequence regions of the bacteria and the virus, there was a *Address correspondence to this author at the Sanford Burnham Prebys Medical Discovery Institute, 10901 North Torrey Pines Rd., La Jolla, CA 92037, USA; Tel: 858-646-3100x3076; E-mail: pcieplak@sbpdiscovery.org substantial level of homology of the N-terminal region of M. bovis BCG lipoprotein lppD sequence (NAANTRLRHAG-GVAAAIARAGGPELQRESTE 208-238, numbering starts from the N-end of Uniprot A0A0H3M501_MYCBP) with the NAANVYLKHGGGVAGALNKATNNAMQVESDD sequence of macrodomain 1 (an ADP-ribosylhydrolase, PDB 6VXS; Fig. 1A, Fig. 2 left) [7,8] of the SARS-CoV-2 orf1a,b/nsp3 (identical and similar residues are in bold and italics, respectively). An additional intriguing homology was recorded between the C-terminal NNAAIVLQLPQ-GTTLPKGF (bat) or NNAATVLQLPQGTTLPKGF (human) region of the RNA-binding domain of the SARS-CoV-2 nucleocapsid N phosphoprotein; PDB 6VYO; Fig. 2 right) [9][10][11], which represents a recently identified SARS-CoV-2 B-cell epitope [12,13], and the sequence regions from the three different proteins in M. bovis BCG. These regions are as follows: the ATVLTLPNGKPLPK region of dehydrogenase/reductase, the ATVLQLPE region of putative drugtransport integral membrane protein and the ATFLQ-RNLPRGTT region of putative secreted protein (Fig. 1B). Partial overlap of these BCG sequences with the viral B-cell epitope implies that the BCG-specific memory B-cells may cross-recognize the viral proteins and that this crossrecognition may affect the virus proliferation and disease severity.
Because a 15-year long post-vaccination protection against tuberculosis is highly credible, this may explain a low clinical manifestation, if any, of COVID-19 in children in China, a country with universal and long-standing BCG vaccination, and, on the contrary, a higher clinical manifestation in certain European countries where BCG programs are no longer deployed. Accordingly, the vaccinated elderly with a low residual level of the cross-reactive memory B-cells may be more susceptible to the virus relative to the freshly vaccinated youth. On the other hand, the vaccinated elderly  [12], with the three sequences of M. bovis BCG proteins: dehydrogenase/reductase, the putative drug-transport integral membrane protein and the putative secreted protein.  is likely to be more resilient to SARS-CoV-2 than the nonvaccinated elderly of similar age. These associations support the notion that BCG vaccination may provide specific protection against SARS-CoV-2 and that the protection level may directly be proportional to the residual cross-reactive Bcells, in addition to the general immunity boost caused by BCG. Furthermore, we speculate that the administration of the Purified Protein Derivative (PPD)/the Mantoux tuberculin skin test normally used for the detection of M. tuberculosis infection may be feasible for screening patients at risk of COVID-19. The test results may then serve as a surrogate marker for the prediction of the viral disease morbidity and mortality as well as to generate valuable information for the subsequent epidemiological studies.

CONSENT FOR PUBLICATION
Not applicable.

FUNDING
None.

CONFLICT OF INTEREST
The authors declare no conflict of interest, financial or otherwise.