Heterologous virus

All statistical analyses were conducted using GraphPad Prism 7. COVID vaccines have been shown to evoke a wide range of immune responses. The responses induced by individual vaccines are hard to compare because the studies have been done under different conditions. To enable direct comparison, we administered four different kinds of vaccines: inactivated vaccines INA manufacturer 1 , recombinant RBD vaccine rRBD manufacturer 2 , Ad5-vectored adenovirus vaccines rAd manufacturer 3 and mRNA vaccine mRNA manufacturer 4 in mice and studied the characteristics of the immune responses elicited by them under identical conditions.

Two weeks after the immunization, the level of binding antibody and neutralizing antibody NAb against live virus as well as pseudovirus were measured. A similar trend was found in NAb titres tested using pseudovirus. Humoral immune responses induced by different vaccine platforms. A Schematic representation of experimental protocol and immunization groups. Amongst, rAd vaccine showing the highest activation of T-cells.

Mice were sacrificed for measuring T cell responses. Mice were immunized intramuscularly at intervals of 14 days and sacrificed for NAb and binding antibody measurement 14 days after the last dose.

Humoral immune responses induced by homologous or heterologous prime boost regimens A. Schematic representation of experimental protocol and immunization groups. Blank control group were injected with PBS.

Similar results were obtained in pseudovirus NAb assay. However, there was no significant difference between the two groups both in NAb and binding antibody titres Figure 3 B—D.

Taken together, our results indicate that boosting inactivated vaccine with a dose of heterologous vaccine could further improve humoral immune responses both for NAb and binding IgG when compared with homologous vaccine.

Our results showed that the most recognized peptide pools originated from S1-RBD aa: Figure 4 , which was consistent with data shown in Figure 2. Thus, a heterologous prime boost strategy consisting of 2-doses of inactivated vaccine followed by one dose of either recombinant RBD, adenovirus vectored or mRNA could effectively improve the Spike-specific T cell responses.

To further define the Th subtype of systemic T cell immune responses induced by the administration of heterologous vaccines following two doses of inactivated vaccines, a Meso Scale Discovery Assay MSD was conducted.

Supernatants of splenic lymphocytes stimulated by four different spike peptide pools were collected for each regimen and pooled together. IL-2 levels were highly improved after peptide stimulation in all vaccinated groups from 0. For IL, a modest increase was observed in vaccinated groups compared with blank control. However, there was no significant difference in the levels of IL secretion between those seven vaccination regimens.

Reports on comparison of the immunogenic properties of COVID vaccines studied under similar conditions using identical methods are sparse. In this study, we describe the immunogenic characteristics of vaccines developed by four different platforms. Previously, Locci et al. The differences in the results can be partly attributed to the dosage. Moreover, the delivery system, antigen construction, and adjuvant used may also lead to the different results.

These results highlight the pitfalls of comparison of vaccines where the studies were not performed under identical conditions. Heterologous prime-boost strategy has been pioneered one decade ago and proved to be effective in previous studies on vaccines against emerging virus such as HIV-1 [ 25 ], influenza [ 24 ] and SARS-CoV-2 [ 26 , 27 ]. In this study, we tested the 3-doses regimen for inactivated vaccine manufacturer 1 in a mouse model in this study. Our result showed that the 3doses regimen improved the NAb level slightly than 2-doses regimen but not significantly.

This is consistent with the reported phase 1—2 clinical trial data of inactivate vaccine developed by Wuhan institute of sinopharm, which demonstrated that 3-doses regimen GMT of did not significantly improve the NAb titres compared with 2-doses regimen GMT [ 13 ]. While, heterologous prime-boost with two-doses of inactivated vaccine followed by rRBD, rAd or mRNA all elicited significantly higher NAb levels than 2-doses regimen of single inactivated vaccine Figure 3 B.

Interestingly, one-dose of mRNA we tested in this study in a mouse model did not showed any superiority in NAb levels against one-dose of inactivated vaccine as shown in Figure 1 B. Thus, The NAb responses were amplified when vaccinated with heterologous vaccines, and heterologous prime-boost could be an effective alternative approach to break the immunity bottleneck caused by homologous prime-boost.

Higher IgG2a always mediated strong cell-mediated cytotoxicity ADCC effect and opsonophagocytosis by macrophages [ 32 , 33 ], which might provide an alternative approach eliminating virus. Thus, a heterologous boost with adenovirus vectored vaccine after 2 doses of inactivated vaccine might confer greater protection against SARS-CoV-2 infections. The strong T cell responses induced by virus vectored and nucleic acid vaccines might benefit from the cytoplasm expression of antigen and the improving of MHC-I presentation process.

Actually, as we described in a previous report, heterologous prime with one dose of inactivated or recombinant RBD vaccine followed by adenovirus vectored vaccine elicited significantly higher T cell responses than two doses of inactivated or recombinant RBD vaccines [ 26 ].

Our results indicate that the combination of exogenous antigen with an endogenous antigen might effectively improve the targeted-antigen specific T cell responses, while the repeated use of exogenous antigens as boosters is not that effective. Safety issues arising out of activation of high numbers of CTLs, especially in people with known immune-related disorders and conditions should be given due consideration while scheduling heterologous vaccination strategies evoking strong T-cell responses.

Multiplex cytokine analysis for homologous and heterologous prime-boost regimens. Lymphocytes isolated from mice kept on different immunization regimens were stimulated with 4 spike peptide pools. Supernatants were pooled by different groups or different peptide pools were collected. In summary, we have characterized the humoral and cellular responses evoked by four COVID vaccine platforms in mouse model.

A heterologous prime-boost strategy consisting of 2 doses of inactivated vaccine followed by either a recombinant subunit, adenovirus vectored or mRNA vaccine increased NAb antibody titres and Th1-type T cell responses. Detailed mechanisms of these effects of BCG vaccination are still unclear. Role of trained innate immunity in providing heterologous immunity upon BCG vaccination has also been demonstrated in humans 26 , 40 , The diphtheria-tetanus-pertussis DTP vaccine shows excellent protection against the three targeted diseases, however, it has been shown that mortality in females but not males increases from other infectious diseases 45 , It was suggested that immunization with DTP may deregulate the female immune system so that subsequent unrelated infections are fought inefficiently, whereas immunization with BCG or measles vaccine subsequent to DTP may circumvent the harmful effect of DTP Therefore, the non-specific effects of vaccines must be thoroughly studied with respect to sex differences.

Potential mechanisms for heterologous effects of vaccines may include cross-reactivity between shared epitopes of unrelated pathogens, trained immunity in innate cells such as natural killer NK , natural killer T cells NKT and monocytes, modulation of type 1, type 17, regulatory and memory T cells, cytokine responses, and modulation of mean concentration of antibodies as well as cross-reactive antibodies Nevertheless, the exact contribution of each of these potential mechanisms in producing the observed heterologous effects of vaccines remains to be delineated.

Heterologous immunity has been shown commonly among closely related pathogens, e. It has been suggested that the history of exposure to various microbial infections and the resulting changes in the memory T cell repertoires determine the existence of a cross-reactivity network in each individual, and therefore cross-reactivity against multiple epitopes may be observed in an individual Heterologous immunity has been experimentally shown in mice by Welsh and Selin, who have reported that some levels of protection against vaccinia virus VV infection is obtained in mice that have been earlier exposed to infections with lymphocytic choriomeningitis virus LCMV , murine cytomegalovirus MCMV , Pichinde virus PV , or influenza A virus IAV 1 , 17 , 49 , However, VV infection does not cross-protect against any of the tested heterologous pathogens and does not show reciprocal heterologous immunity Our studies have demonstrated unexpected cross-reactivity between adenoviruses and Hepatitis C virus Most of these examples of heterologous immunity have demonstrated a role for cross-reactive T cells, but other mechanisms may also be contributing toward the overall effect.

With the induction of cross-reactive T cells, besides enhanced or inhibited clearance of a virus, T cell immunodominance patterns may be altered resulting in unusual skewing of T cell repertoires It has been shown that heterologous virus challenge may lead to expansion of cross-reactive narrowly focused T cell repertoire and viral escape, whereas homologous viral challenge may allow expansion of more oligoclonal T cell responses 17 , It has also been suggested that memory T cells generated after an infection have a lower activation threshold and may be activated by the bystander effect local cytokines independent of TCR signaling, contributing significantly to heterologous immunity Further, it must be highlighted that in most instances, heterologous immunity is not nearly as effective as specific immunity but may be sufficient to alter the otherwise severe course of a heterologous infection and mortality.

The widespread overlap in heterologous immunity between these different viruses suggests that cross-reactivity among pathogens is prevalent. It was further demonstrated that humans also have cross-reactive T cells and antibodies similar to data obtained in mice. Overall, the results demonstrated that cross-reactive flavivirus immunity can provide enhanced protection to a heterologous infection. There are also multiple examples of heterologous immunity between bacteria and viruses.

Herpes virus infection in mice with MCMV and Murine c-Herpesvirus, has been demonstrated to induce protective immunity against bacterial pathogens such as Listeria monocytogenes and Yersinia pestis A number of other heterologous effects of BCG appear to be related to its effect on innate immunity 58 and are not described here.

It has been demonstrated that a fungal species Candida albicans hyphal wall protein Hyr1p shares significant structural homology to a bacteria species Acinetobacter baumannii cell surface protein, and active with rHyr1p or passive with anti-Hyr1p antibodies immunization of mice protects them from systemic infection with A. Most experimental studies of heterologous immunity have used animals mice immunized or challenged with a pathogen followed by determining the immune response or protection against an unrelated organism.

Heterologous immunity is rather difficult to demonstrate in humans due to continuous exposure with a number of pathogens, in comparison to inbred mice raised in a controlled laboratory environment.

Furthermore, due to constant exposure to various pathogens, the memory T cell pool of an individual is also constantly changing. In an adult human, cross-reactive T cells represent a pool of cells ready to respond to a new pathogen. The quality and quantity of these cells are ultimately dependent upon an individual's immune history resulting from previous infections.

A2 positive HCV-seronegative donors. Low dose exposure or acute clearance of HCV of the cohort was excluded in this study, and their origin from previous heterologous infections was suggested Infection with Dengue virus DENV in humans can sometimes lead to dengue hemorrhagic fever and shock syndrome.

This severe immunopathology following DENV infection has been associated with re-exposure of individuals immune to one strain serotype of DENV with another strain. It has been demonstrated that cross-reactive non-neutralizing antibody can bind to viruses without inactivating them and enhance the infection of macrophages that bear Fc receptors for those antibodies Furthermore, extensive T cell cross-reactivity occurs between different serotypes of DENV and a T cell response to the second dengue virus infection may induce CD8 T cells that have a higher affinity to the previously encountered Dengue virus, dampening the T cell response to the second virus.

Therefore, both antibody-dependent immune enhancement and cross-reactive, low affinity T cell responses may combine to exacerbate the disease pathology Previous exposure to pathogens leads to induction of innate and adaptive immune responses, which result in establishing a substantial pool of memory B and T cells long after the pathogen insult has been eliminated.

However, these also form a pool of ready-to-respond cross-reactive cells with low stimulation requirements. However, their precise contribution to success or failure of infection with these pathogens is still not clear Figure 2. Impact of specific vs.

I Depicts the results of specific immunity. Similarly, exposure or infection with pathogen D will stimulate and expand T cells specific against D and protect against re-infection with pathogen D but not against subsequent infections with pathogens E and F.

II Demonstrates the consequences of heterologous immunity. Upon administration with vaccine against A, a broad repertoire of cross-reactive T cells will be activated and expanded, and an individual would be protected against subsequent exposure with pathogen A, but also protected against pathogens B and C to some extent. Similarly, exposure or infection with pathogen D will induce cross-reactive T cells and protect against re-infection with pathogen D, but may also protect against subsequent infection with pathogens E and F.

Therefore, the exposure history of an individual may modulate the outcome of future infections with multiple pathogens. As an example, infection with HCV presents an interesting scenario. Induction of immune responses has been shown to be delayed in infected individuals, but it is not clear if some pre-existing cross-reactive immunity plays a role in spontaneous clearance of an acute infection In seronegative individuals, HCV clearance has been reported after acute infection In some patients with persistent infection, spontaneous resolution of infection and seroconversion to antibody negative status has been found Possible factors to explain this include a prior undetectable HCV exposure followed by clearance resulting in the induction of T cells reactive against HCV antigens, b abortive infection, or c occult infection with HCV.

However, the possibility of cross-reactive T cells showing such responses cannot be excluded, especially since our studies have demonstrated robust cross-reactivity between commonly found adenoviruses Ad5 and HCV 52 , The intriguing example of HCV demonstrates the complexity in determining the role of heterologous immunity in pathogenesis or protection from an infection, but undoubtedly indicates its significant role in natural immunity against a pathogen.

These microbiota are enriched at mucosal barriers such as gut and respiratory mucosa. Microbiota play an integral role in immune and metabolic homeostasis in an individual and can also contribute to control or prevent infection with a pathogen Microbiota can either directly inhibit infection with pathogens by competing for an available niche, or indirectly inhibit infection through immune-mediated mechanisms Microbiota can also either allow the persistence of peripheral memory T cells or induction of regulatory T cells to induce tolerance to some pathogenic antigens.

It has been suggested that the variety of antigens derived from the members of the microbiota can prime and lead to a diverse repertoire of memory T and B cells. These, in turn, can demonstrate enhanced immunity to a newly exposed pathogen through cross-reactivity Microbiota-mediated induction of pre-existing antibodies against HIV gp has been shown to be detrimental to the induction of neutralizing antibodies against HIV Env gp This was seen in recipients of a DNA primed—rAd5 boosted HIV-1 vaccine in clinical trials 85 where a microbiota-mediated heterologous immune responses had a negative effect on vaccine efficacy.

The most successful human vaccines so far have been live attenuated viral or bacterial pathogens such as Measles, BCG, Poliovirus oral polio vaccine , and Smallpox virus. These vaccines tend to induce life-long immunity.

The successes of these vaccines led to their application as delivery vectors for an unrelated antigen in their makeup. Ad are commonly used as vectors to deliver transgenes in gene therapy and vaccines 87 — In addition to respiratory exposure, Ad have been shown to remain in the human gut for extended periods, making them akin to members of gut microbiota Consequently, rare human Ad and Ad of different animal species such as from cattle and chimpanzees have also been tested as vaccine vectors 91 , Although it has been shown that neutralizing antibodies don't cross-recognize Ad of different serotype or species, T cell cross-reactivity between different Ad has been demonstrated in humans Due to the presence of conserved T cell epitopes in hexons, cross-reactivity among divergent serotypes from chimpanzees and humans has been observed.

By virtue of T cells being cross-reactive, these pools of Ad-specific T cells that are present universally in humans may provide an efficient source of effector T cells to target heterologous pathogens that regularly infect people. Our studies have demonstrated unexpected and surprising homologies between peptides of HCV antigens and Ad antigens, and robust cross-reactive cellular and humoral immunity between Ad and HCV Intriguingly, the nature of the transgene antigen had a significant impact on the levels of cross-reactive immunity induced against HCV antigens It is possible that Ad also has cross-reactivity with other pathogens, but this has not been explored yet.

These observations shed light on another rather unstudied aspect of vaccine vectors and support the notion that heterologous immunity induced by vaccine vectors may lead to significant heterologous immunity against another pathogen possibly influencing the natural course of infection with that pathogen. Consequently, non-specific effects of vaccine vectors must also be examined thoroughly.

Specificity and memory are the two traits of the adaptive immune system exploited for the development and application of vaccines. It is expected that administration of a vaccine would induce vaccine-antigen-specific T and B cell memory responses, which upon exposure to the corresponding pathogen, would rapidly and specifically lead to prevention or clearance of the infection.

Experimental and clinical evidence as well as theoretical constructs have clearly demonstrated that heterologous immunity or cross-reactivity of adaptive immune cells is not an isolated or accidental phenomenon, but rather a fundamental attribute of adaptive immunity, forming an integral part of the host defense system against pathogens under natural conditions.

Mobilizing cross-reactive adaptive immunity by a heterologous therapeutic vaccine may be an ultimate strategy to induce effective protective immunity in such chronic disease conditions.

Furthermore, as stated earlier, cross-reactive lymphocytes may not be the only mechanism behind heterologous immunity, and trained innate immunity could explain some of the observations of heterologous immunity. Therefore, it should also be pursued in future vaccine design and efficacy studies.

In an individual, T and B cell repertoires originate by random VDJ gene rearrangement, however, their cross-reactivity and ability to respond to various antigens in a host is shaped to a large extent by exposure history and microbiota, and therefore, cross-reactivity remains largely unpredictable at the individual level. Acknowledgment of the heterologous side of adaptive immunity does have important consequences on self-tolerance, autoimmunity, and vaccination strategies.

Potential positive heterologous effects of vaccines have been discussed in this article, but heterologous immunity can also have dire consequences upon cross-reacting to self-antigens. It is essential that future investigation of vaccine design must exploit the beneficial aspects of heterologous immunity and at the same time devise strategies to avoid the potentially harmful effects.

As stated earlier, cross-reactivity may vary among individuals and different sexes, and may largely remain unpredictable making application to vaccines which are usually population-based, not an easy task. In addition, recent understanding of the role of microbiota in immune homeostasis and induction of immune tolerance in a host has opened new avenues of investigation in vaccines.

Specifically, it has been shown that pathological precipitation of many of the autoimmune diseases e. Moreover, even in individuals with genetic susceptibility and the peripheral presence of potentially autoreactive T cells, modifications of gut microbiota may allow modulating the disease pathology Additionally, development of tools and techniques to predict cross-reactivity against both unrelated pathogens and self-antigens would aid in vaccine design and coverage of protection obtained by a vaccine.

National Center for Biotechnology Information , U. Front Immunol. Published online Nov 8. Author information Article notes Copyright and License information Disclaimer. This article was submitted to Vaccines and Molecular Therapeutics, a section of the journal Frontiers in Immunology.

Received Jul 25; Accepted Oct The use, distribution or reproduction in other forums is permitted, provided the original author s and the copyright owner s are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. This article has been cited by other articles in PMC. Abstract The central paradigm of vaccination is to generate resistance to infection by a specific pathogen when the vacinee is re-exposed to that pathogen.

Keywords: heterologous non-specific effects of vaccines, heterologous immunity, T cells, antibody, innate and adaptive immune response. Open in a separate window. Figure 1. Heterologous Immunity Across a Broad Range of Pathogens Heterologous immunity has been shown commonly among closely related pathogens, e.

Figure 2. Robert L. Kirsten E. Meagan E. Lisa A. Angela R. Hana M. Christina A. Judith M. Richard E. Mark J. Rebecca C. Robert W. Frenck, Jr. Angelica C. Tara M. Christine M. Janet I. Seema U. Clara P. Elizabeth R. Mehul S. Adrian B.

Sarah E. David C. Kathleen M. David S. Paul C. John H. National Center for Biotechnology Information , U. Version 2. Preprints have not been peer reviewed. Atmar , M. Lyke , M. Deming , M. D, Lisa A. Jackson , M. Branche , M. El Sahly , M. Rostad , M. Martin , M. Rupp , M. Mulligan , M.

Brady , M. Kottkamp , M. Babu , M. Posavad , Ph. Archer , M. Nayak , M. Dominguez Islas , Ph. D, Elizabeth R. Brown , Sc. Suthar , Ph. Juliana McElrath , M. McDermott , Ph. Montefiori , Ph. Neuzil , M. Stephens , M. Roberts , Ph. Beigel , M. El Sahly. Dominguez Islas.

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Associated Data Supplementary Materials Supplement 1. Abstract Background: While Coronavirus disease Covid vaccines are highly effective, breakthrough infections are occurring.

Results: individuals were enrolled: received mRNA, received Ad Conclusion: Homologous and heterologous booster vaccinations were well-tolerated and immunogenic in adults who completed a primary Covid vaccine regimen at least 12 weeks earlier.

Immunogenicity: Immunogenicity was assessed on Day 1 prior to booster vaccination and at Days 15 and 29 unavailable for all groups after boost. Statistical Analysis: Summaries of baseline characteristics including serological endpoints are reported for all enrolled participants.

Table 1.