As per the first author’s personal clinical experience, human papilloma virus (HPV) recurrent warts are cured with anecdotal use of bOPV (G. Vuppalapati, unpublished data).

Based on this observation and evidences from the literature, a cross sectional study is conducted to test whether the incidence and mortality of COVID-19 is influenced by the type and schedule of polio vaccinations in the respective national immunization programs (NIPs) of a group of countries. The results showed that the incidence and mortality are significantly lower in the population of countries that are using bivalent oral polio vaccine (bOPV) in regular and supplementary immunization activities, needs confirmation through in vivo animal studies and clinical trials for the possible plausible beneficial NSEs towards COVID-19 protection before the 2 nd wave of pandemic ensues.

Study design and data collection
A cross-sectional observational study of the COVID-19 epidemiology was conceived as per STROBE statement. Number of reported COVID-19 cases and deaths per million population was derived from an open source https://ourworldindata.org/coronavirus (up to 9 th June 2020).
For the completeness and to minimize the bias in selection of countries, all the countries with onset of COVID-19 epidemic with maximum of 1 case per million population as on 1st of February 2020 were also included in the study. Stratification based on type and schedule of polio vaccine 58 countries which met the selection criteria were divided into three groups based on the schedule of polio vaccine used for children under five years.
Group 1 (G1): countries using IPV in their RI schedule also labelled as ‘IPV only’;
Group 2 (G2): countries using bOPV and IPV/fractional IPV(fIPV) on RI labelled as ‘IPV + bOPV – RI’;
Group 3 (G3): countries received supplementary polio immunization activities (SIA) with bOPV on their national immunization days (NIDs) twice a year until 2019 in addition to RI with or without IPV/fIPV and labelled as ‘bOPV-RI+SIA’.

Statistical analysis
Exploratory analysis of country-wise data on the incidence of COVID-19 cases with corresponding deaths per million population was performed with a view to find the correlation and impact of various polio vaccine interventions in these three groups of countries on the vulnerability or immunity to COVID-19 transmission at their community level. For each group, the data was summarized as median and IQR (Inter Quartile Range), as shown in table1. Data analysis was performed using version IBM SPSS 25.0. The data on various countries is presumed to reflect independent findings, and the background variables have no impact for the purpose of the study. To check the hypothesis, the group medians were compared by using Kruskal Wallis test (non-parametric method) by taking p < 0.05 as cut-off to flag significance and then compared with COVID-19 infection and mortality (per million) among these vaccine variable groups for a gross visual representation of all the countries on the World Maps as on 9 th June, 2020

RESULTS
The data of reported COVID-19 cases and deaths as on 9thJune 2020in countries with an epidemic onset during the month of February 2020was collected and these countries were divided into three groups, as summarized in the Methods section. The study shows the number of COVID-19 cases and deaths per million population in G1 countries with a median of 2202.61 cases and 361.46 deaths compared to G2 with an average of 854.15cases and 59.45 deaths, and G3 with a median of 129.12 cases and 9.00 deaths (table 1).
Countries that adopted only IPV showed a higher average incidence of cases and deaths per million population when compared to the countries that adopted IPV + bOPV-RI and bOPV-RI + SIA (fig 1A, 1B, 1C). The data on different countries are assumed to represent independent findings, and the background factors are assumed to have no influence. The group medians 6 are compared by using Kruskal Wallis test (non-parametric method) which takes p < 0.05 as a cut-off to flag significance. For cases (H = 19.857, p < 0.0001) and for deaths (H = 25.819, p < 0.0001), significant differences in median were observed. Hence intervention has a significant effect on the occurrence of cases and deaths. Pair-wise comparison among the three groups also revealed that each pair of interventions has a significant difference in
medians shown in table1. In each group, the distribution of the number of cases and deaths is found to be skewed and have outliers that limit the use of classical statistical tests for comparison, and therefore non-parametric approach is used in the analysis. Group wise distribution of COVID-19 cases and deaths per million population shows the extreme outliers (fig 1D). The upward whisker indicates positive skewness of data while the horizontal line inside each box indicates the corresponding group median. The mortality rate in G1 countries was proportional to the number of cases, with many countries depicting deaths above the median mortality rate (27.25per million). It is easy to see that the median number of cases (deaths) is far below the overall median in G3 (fig 1D). It is assumed that the data on different countries represent independent observations but do not follow a known statistical distribution. The study also confirmed the lowest risk of p<0.0001 infection and death in the G3 countries using Kruskal Wallis method. In G1 and G3 countries, there is a linear relationship between the death rate and the rate of cases per million as indicated by a high positive and significant value of Pearson’s correlation coefficient (r)(fig 1E). The epidemiological correlation showed that the population in G3 countries received bOPV in RI and SIA reported to show the least number of COVID-19 cases (fig 2A and 2B) and deaths (fig 2C and 2D) per million population when compared to other groups. Interestingly, inverse relationship was observed between incidence of COVID-19 and population density (fig2E and2F). The overall analysis converges to the hypothesis that the population in G3 countries may have exhibited stronger beneficial NSEs exhibited by the booster effect of SIA with bOPV to find significantly lower number of COVID-19 cases and deaths. Besides the effects of bOPV, there may be group effects due to various variables, known and unknown influencing factors occurring in each of these countries that are not considered for this analysis, but an important and life-saving hypothesis has been proposed that needs to be further evaluated through a systematic study.

DISCUSSION
The results from the present study indicated that countries adopting IPV only in their NIP exhibited a significantly higher average (median) incidence of COVID-19 cases as well as deaths when compared with bOPV-RI and bOPV-RI + SIA groups of countries. The mortality rate was changing with the incidence of COVID-19 in the IPV only group whereas it was constant and lower in the bOPV-RI + SIA group. At a macro level taking the country as a unit, there is evidence of significant differences in the incidence and mortality rates. These observations providing evidence that the live attenuated mucosal bOPV given in childhood vaccination program with significant spillover to nursing parents via feco-oral transmission may be instrumental in modulating immunity through induction of beneficial NSEs that resulted in reduced COVID-19 infection and death rates especially in the countries with high population and low sanitation standards. These NSEs could be due to activation of heterologous poly-specific T lymphocytes [3,13] or trained innate lymphoid cells (ILCs) or probably by other complex mechanisms yet to be established. There are enough epidemiological, experimental and clinical evidences for the vaccine induced heterologous immunity shown by cross reactive adaptive immune cells for a different viral pathogen than the intended one (for example, cross reactivity has been observed through induction of human immunodeficiency virus (HIV) antibodies after measles vaccination) and in addition to that, a network of cytokines, regulatory cells and trained innate immune cells contribute in manifesting heterologous immunity. The lowest risk observed in G3 populations could be due to the recent use of booster doses of bOPV in 2019-20 polio SIAs where bOPV being mucosal live vaccine would have helped to boost beneficial NSEs by priming the mucosa associated lymphoid tissue (MALT-specific immune responses against SARS-CoV-2. Due to the threat of tOPV (trivalent OPV) induced vaccine-associated paralytic poliomyelitis (VAPP) resulted in switch to bOPV globally in 2016 and eventually to only IPV after polio eradication as a part of World Health Organization (WHO)’s Endgame strategy of Global Polio Eradication Initiative (GPEI). But OPV is still in use in low-income countries rather than IPV for several reasons.[12] The other possible mechanisms for beneficial NSEs of live attenuated bOPV could be induction of mucosal innate immune responses involving preferential expression of several integrins and cell adhesion molecules associated with homing of lymphocytes to mucosal sites of immunization. Although, recent reports speculated the possibility of vaccine induced nonspecific effects using BCG, measles, childhood vaccination regimes in general for COVID-19 protection based on observational studies, they are yet to be confirmed through clinical trials to split the group effects (clinical trials are in progress to check the NSEs of BCG for COVID-19 protection in Australia), but no such study on the protective value of bOPV was reported. Route of mucosal vaccine administration and addition of adjuvants play a significant role in inducing effective mucosal innate immune response. Vaccination through sublingual route shown to induce heightened immune responses across various mucosal systems unlike the oral, rectal, nasal routes, suggesting administration of bOPV through sublingual route could be a better option for the clinical studies as the SARS CoV-2 is known to enter through oral, nasal and conjunctival mucosa.

CONCLUSIONS

A better understanding of NSEs induced by existing safe live attenuated vaccines and the possible immunological mechanisms that are responsible will help to exploit the beneficial heterologous immunity against COVID-19 and in the event of a pandemic of a new virus in future. The WHO Strategic Advisory Group of Experts (SAGE) on Vaccines and Immunization recently reviewed BCG, measles and DTP evidence for NSEs, and suggested further research into OPV and other live vaccines. The present statistical analysis indicated the reduction in the COVID-19 incidence and mortality in G2 and G3 countries which could be due to the exposure of OPV in the childhood vaccination program and to their nursing parents through fecal-oral route of transmission, more so in G3 countries due to bOPV SIAs. This scenario warrants large scale systematic epidemiological, clinical and immunological studies with bOPV to find out its prophylactic efficacy against COVID-19 in adults and the exact biological mechanisms that support the NSEs by bOPV. The possibility of beneficial NSEs by the combined use of other live vaccines in the childhood vaccination regime cannot be ruled out and may have to be considered for these studies. Questions regarding the longevity of these beneficial NSEs conferring heterologous and or trained immunity and the reasons for deleterious effects of IPV are to be answered to determine its clinical implications and for rational design of potential vaccination policies. Considering its proven safety, low cost, ready availability, ease of production and administration, we recommend immediate in vivo animal studies with bOPV to check for the heterologous adaptive and trained innate immune responses against unrelated pathogens such as SARS-CoV-2. Furthermore, clinical trials involving revaccination of immunocompetent healthcare personnel and adults with sublingual bOPV alone or combined with other live vaccines/safe adjuvants to boost beneficial NSEs and to confer maximum protection against COVID-19 are essential to reduce the global burden and for economic recovery until a specific and more effective vaccine becomes available. Such studies would also pave a way for paradigm shift in the field of immunology and approach to future pandemics.