# A GLOBAL PERSPECTIVE ON VACCINES: PRIORITIES, CHALLENGES AND ONLINE INFORMATION

EDITED BY : Luciana Leite, Aldo Tagliabue, Rino Rappuoli and Odile Yvonne Leroy PUBLISHED IN : Frontiers in Immunology

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ISSN 1664-8714 ISBN 978-2-88963-533-7 DOI 10.3389/978-2-88963-533-7

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# A GLOBAL PERSPECTIVE ON VACCINES: PRIORITIES, CHALLENGES AND ONLINE INFORMATION

Topic Editors: Luciana Leite, Butantan Institute, Brazil Aldo Tagliabue, Institute for Genetic and Biomedical Research (IRGB), Italy Rino Rappuoli, GlaxoSmithKline, Italy Odile Yvonne Leroy, European Vaccine Initiative, Germany

Image: igorstevanovic/Shutterstock.com

Citation: Leite, L., Tagliabue, A., Rappuoli, R., Leroy, O. Y., eds. (2020). A Global Perspective on Vaccines: Priorities, Challenges and Online Information. Lausanne: Frontiers Media SA. doi: 10.3389/978-2-88963-533-7

# Table of Contents

*05 Editorial: A Global Perspective on Vaccines: Priorities, Challenges and Online Information*

Aldo Tagliabue, Luciana C. C. Leite, Odile Y. Leroy and Rino Rappuoli

*07 Assessing the Importance of Domestic Vaccine Manufacturing Centers: An Overview of Immunization Programs, Vaccine Manufacture, and Distribution*

Emma Rey-Jurado, Felipe Tapia, Natalia Muñoz-Durango, Margarita K. Lay, Leandro J. Carreño, Claudia A. Riedel, Susan M. Bueno, Yvonne Genzel and Alexis M. Kalergis

*23 Education in Vaccinology: An Important Tool for Strengthening Global Health*

Paul-Henri Lambert and Audino Podda

*27 Changing Priorities in Vaccinology: Antibiotic Resistance Moving to the Top*

Aldo Tagliabue and Rino Rappuoli

*36 Fake News or Weak Science? Visibility and Characterization of Antivaccine Webpages Returned by Google in Different Languages and Countries*

Nadia Arif, Majed Al-Jefri, Isabella Harb Bizzi, Gianni Boitano Perano, Michel Goldman, Inam Haq, Kee Leng Chua, Manuela Mengozzi, Marie Neunez, Helen Smith and Pietro Ghezzi

*48 Corrigendum: Fake News or Weak Science? Visibility and Characterization of Antivaccine Webpages Returned by Google in Different Languages and Countries*

Nadia Arif, Majed Al-Jefri, Isabella Harb Bizzi, Gianni Boitano Perano, Michel Goldman, Inam Haq, Kee Leng Chua, Manuela Mengozzi, Marie Neunez, Helen Smith and Pietro Ghezzi

*50 The US Military Commitment to Vaccine Development: A Century of Successes and Challenges*

Silvia Ratto-Kim, In-Kyu Yoon, Robert M. Paris, Jean-Louis Excler, Jerome H. Kim and Robert J. O'Connell

#### *117 Booster Vaccination With GVGH* Shigella sonnei *1790GAHB GMMA Vaccine Compared to Single Vaccination in Unvaccinated Healthy European Adults: Results From a Phase 1 Clinical Trial*

Odile Launay, Augustin G. W. Ndiaye, Valentino Conti, Pierre Loulergue, Antonella Silvia Sciré, Anais Maugard Landre, Pietro Ferruzzi, Naouel Nedjaai, Lena Dorothee Schütte, Joachim Auerbach, Elisa Marchetti, Allan Saul, Laura B. Martin and Audino Podda

*127 Infectious Disease Threats in the Twenty-First Century: Strengthening the Global Response*

David E. Bloom and Daniel Cadarette

# Editorial: A Global Perspective on Vaccines: Priorities, Challenges and Online Information

#### Aldo Tagliabue1 \*, Luciana C. C. Leite2 , Odile Y. Leroy 3 and Rino Rappuoli 4,5,6

1 Institute for Genetic and Biomedical Research, National Research Council, Monserrato, Italy, 2 Instituto Butantan, Saõ Paulo, Brazil, 3 European Vaccine Initiative, Heidelberg, Germany, 4 GlaxoSmithKline, Siana, Italy, 5 Toscana Life Sciences, Siena, Italy, 6 Imperial College, London, United Kingdom

Keywords: vaccines, antimicrobial resistance, vaccine hesitancy, vaccine education, technological platforms

#### **Editorial on the Research Topic**

#### **A Global Perspective on Vaccines: Priorities, Challenges and Online Information**

Vaccines are one of the most successful stories in global health. For 99.99% of mankind history, life expectancy has been <30 years, but in the last 300 years human life has increased by 55 years, of which 35 were gained in the last century. Vaccines are accountable for a significant part of this extraordinary result. Since the first successful vaccine for smallpox in 1796, vaccines for many other diseases have been generated, taking advantage of the developments of science. Smallpox has been totally eradicated, and polio could be soon eliminated. Many other effective vaccines have significantly reduced the incidence of diseases that have killed millions of people in the past. Paradoxically, the decreased impact of infectious diseases at the global level is making people think that vaccines are no longer necessary. One of the reasons behind the decline in vaccine confidence is that many people have become complacent, and we face now the phenomenon of vaccine hesitancy. Thus, we can say that vaccines are victims of their own success.

The topic "A global perspective on vaccines: priorities, challenges and online information" focuses on the most crucial issues in the vaccine field, with a view on the years to come.

Since infections travel the whole world with no borders, the war against microbes is a global one. In this context, the action of military forces in vaccine development (Ratto-Kim et al.) has been an important initiator, and the worldwide effort includes organizations that fight long-known diseases and, more challenging, emerging infections such as SARS, MERS, Ebola, and Zika (Marinho de Andrade Zanotto and Leite). The globalized modern way of life, with the increased number of travelers throughout continents, is aggravating the situation. It is extremely important to ensure preparedness and efficacy of vaccines leveraging on innovation.

**Research & Development** remain the core of the evolving field. The -omics revolution, combined with the power of the extremely fast-growing technologies that will rapidly involve artificial intelligence, is providing a large variety of new vaccine candidates. There is a great need for harmonization and simplification for the newly generated vaccines. To this end, new platforms are being developed that will allow for new generation vaccines with built-in adjuvanticity for preventing multiple infections in a single shot.

Safety and affordability are characteristics that must be ensured in modern vaccines. Affordability will allow for fast access to effective vaccines worldwide, in particular to populations in developing countries. In this view, promoting local manufacturing is a key element in vaccine affordability (Rey-Jurado et al.). Side effects must be minimized, including the mild ones, to ensure the full safety of preventive treatments. In parallel, the new vaccines should have the highest effectiveness in all populations, from people living in the clean industrialized world to the populations that are constantly facing large microbial burden in infection-endemic areas. In this

#### Edited and reviewed by:

Denise Doolan, James Cook University, Australia

> \*Correspondence: Aldo Tagliabue<EMAIL_ADDRESS>
#### Specialty section:

This article was submitted to Vaccines and Molecular Therapeutics, a section of the journal Frontiers in Immunology

> Received: 09 October 2019 Accepted: 15 October 2019 Published: 12 November 2019

#### Citation:

Tagliabue A, Leite LCC, Leroy OY and Rappuoli R (2019) Editorial: A Global Perspective on Vaccines: Priorities, Challenges and Online Information. Front. Immunol. 10:2556. doi: 10.3389/fimmu.2019.02556

**5**

view, information on the co-evolution of microbes with the human host, coming from genome-wide association studies, is opening new avenues to the understanding of the mechanisms of resistance to infections. Studying the resistant subjects will pose the basis to design unprecedented preventive and therapeutic approaches. Malaria is an important example for this approach and is offering the possibility to rethinking vaccine design (1, 2).

**Clinical trials** are already being designed to prove the feasibility of new technological approaches in naïve and previously primed subjects. The understanding of the mechanisms and timing of prime-boosting immunizations is important, also in light of the problems observed with a candidate Dengue vaccine (3). Some examples are provided in this topic (Rauch et al., Launay et al., Yao et al.).

In the last decades, a complex global health system against known and unknown infectious disease threats has arisen, encompassing various formal and informal networks of organizations that serve different stakeholders, have varying goals, modalities, resources, and accountability; operate at different regional levels (i.e., local, national, regional, or global); and cut across the public, private-for-profit, and private-not-forprofit sectors. Organizations such as the Bill and Melinda Gates Foundation, the Global Alliance for Vaccine Immunization, and the Coalition for Epidemic Preparedness Innovations are potent drivers that are transforming the vaccine world (Bloom and Cadarette).

Among the new global health challenges that we are facing, the most alarming is the growing inefficacy of antibiotics. The excessive and incorrect use of antibiotics has accelerated the generation of resistant pathogens, which in many cases show resistance to multiple drugs. The increasing inefficacy of current antibiotics is expected to cause 10 million deaths per year in the world by 2050 (Tagliabue and Rappuoli). Vaccines could be the solution to **Antimicrobial resistance** (AMR) and its impending death toll. Thus, AMR is revolutionizing vaccine R&D to the point that priorities are being re-evaluated, and vaccines are being developed for diseases that we have for

#### REFERENCES

long considered harmless because curable with antibiotics. Furthermore, scientists are exploring the potential protective role of vaccines beyond the classical induction of pathogenspecific adaptive humoral and cellular immunity. An increasing body of experimental data is now supporting the notion that vaccines can have non-specific protective effects (Uthayakumar et al.). The concept of innate immune memory is starting to be exploited for the design of personalized effective adjuvants in novel vaccination strategies.

The impressive scientific and technological advancements and the huge global health efforts need to be paralleled by worldwide initiatives in vaccine **Education**. Thus, the high-level training for present and future vaccinologists plays a fundamental role in broadening conceptual and applied knowledge in the vaccine field (Lambert and Podda). But vaccine education should not be limited to raising expert vaccinologists. It should also target the general public. As already mentioned, vaccine hesitancy is spreading and could become a major threat for vaccine effectiveness. Public perception and public acceptance will make the difference between successful control of infections and failure with consequent scourge propagation. The use of modern social media could be extremely important to spread and popularize the correct information, avoiding ideological and political exploitation (Arif et al.), thereby increasing acceptance and compliance. False information, such as the idea that vaccines cause autism, is difficult to eliminate, despite the solid epidemiological data against it (4). The impact of social media on our society is huge and rapidly transforming. But eliminating vaccine hesitancy is a moral imperative.

## AUTHOR CONTRIBUTIONS

AT wrote the manuscript. LL, OL, and RR critically revised it.

## ACKNOWLEDGMENTS

Thanks to Diana Boraschi for critical review and suggestions.

**Conflict of Interest:** LL is an employee of the Instituto Butantan, a government controlled institution engaged in vaccine development and trials. RR is head of the GSK Vaccine research and development.

The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Copyright © 2019 Tagliabue, Leite, Leroy and Rappuoli. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). 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.

# Assessing the Importance of Domestic Vaccine Manufacturing Centers: An Overview of Immunization Programs, Vaccine Manufacture, and Distribution

*Emma Rey-Jurado1 , Felipe Tapia2 , Natalia Muñoz-Durango1 , Margarita K. Lay3 , Leandro J. Carreño4 , Claudia A. Riedel5 , Susan M. Bueno1 , Yvonne Genzel2 and Alexis M. Kalergis1,6\**

*1Millennium Institute on Immunology and Immunotherapy, Departamento de Genética Molecular y Microbiología, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile, Santiago, Chile, 2Max Planck Institute for Dynamics of Complex Technical Systems, Magdeburg, Germany, 3Departamento de Biotecnología, Facultad de Ciencias del Mar y Recursos Biológicos, Universidad de Antofagasta, Antofagasta, Chile, 4Millennium Institute on Immunology and Immunotherapy, Programa de Inmunología, Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Santiago, Chile, 5Millennium Institute on Immunology and Immunotherapy, Departamento de Ciencias Biológicas, Facultad de Ciencias Biológicas y Facultad de Medicina, Universidad Andrés Bello, Santiago, Chile, 6Departamento de Endocrinología, Facultad de Medicina, Pontificia Universidad Católica de Chile, Santiago, Chile*

#### *Edited by: Aldo Tagliabue,*

*ALTA, Italy*

#### *Reviewed by:*

*Francesco Berlanda Scorza, Program for Appropriate Technology in Health, United States Vito Di Cioccio, GSK Vaccines Institute for Global Health, Italy*

> *\*Correspondence: Alexis M. Kalergis<EMAIL_ADDRESS>
#### *Specialty section:*

*This article was submitted to Vaccines and Molecular Therapeutics, a section of the journal Frontiers in Immunology*

*Received: 11 November 2017 Accepted: 04 January 2018 Published: 18 January 2018*

#### *Citation:*

*Rey-Jurado E, Tapia F, Muñoz-Durango N, Lay MK, Carreño LJ, Riedel CA, Bueno SM, Genzel Y and Kalergis AM (2018) Assessing the Importance of Domestic Vaccine Manufacturing Centers: An Overview of Immunization Programs, Vaccine Manufacture, and Distribution. Front. Immunol. 9:26. doi: 10.3389/fimmu.2018.00026*

Vaccines have significantly reduced the detrimental effects of numerous human infectious diseases worldwide, helped to reduce drastically child mortality rates and even achieved eradication of major pathogens, such as smallpox. These achievements have been possible due to a dedicated effort for vaccine research and development, as well as an effective transfer of these vaccines to public health care systems globally. Either public or private institutions have committed to developing and manufacturing vaccines for local or international population supply. However, current vaccine manufacturers worldwide might not be able to guarantee sufficient vaccine supplies for all nations when epidemics or pandemics events could take place. Currently, different countries produce their own vaccine supplies under Good Manufacturing Practices, which include the USA, Canada, China, India, some nations in Europe and South America, such as Germany, the Netherlands, Italy, France, Argentina, and Brazil, respectively. Here, we discuss some of the vaccine programs and manufacturing capacities, comparing the current models of vaccine management between industrialized and developing countries. Because local vaccine production undoubtedly provides significant benefits for the respective population, the manufacture capacity of these prophylactic products should be included in every country as a matter of national safety.

Keywords: vaccine manufacturing, immunization programs, vaccine distribution, vaccine shortages, good manufacturing practices

## INTRODUCTION

The incidence of numerous infectious diseases that are life threatening to humans has drastically declined since the development of safe and effective vaccines and the implementation of global vaccination programs worldwide. In fact, the variola virus, which caused smallpox disease that killed millions of individuals throughout history, was successfully eradicated from Earth during the

**7**

1980s (1), due to a worldwide immunization campaign against this major pathogen. Moreover, poliovirus, which severely affects the health of children with lifelong disabling consequences, has almost been eradicated from the world. Since 1999 very few cases of polio disease have been reported, probably due to two of the three poliovirus types. Indeed, the goal of the World Health Organization (WHO) is to achieve the eradication of polio during 2018. Therefore, millions of human lives have been saved by means of the implementation of national immunization programs in all countries, and the demand for new prophylactics to protect against infectious diseases is constantly growing. Although vaccine manufacturing is usually associated with biopharmaceutical companies, some public and academic institutions also produce these prophylactic formulations (2). Despite the existence of those manufacturers aiming at increasing vaccine availability, shortage of these products has taken place several times causing that not enough doses were available in some countries.

In this article, we attempt to comprehensively discuss the WHO current recommendations for routine immunization and some of the national immunization programs. Further, we associate such vaccination programs to the global vaccine manufacture and distribution capabilities, focusing in some industrialized and developing countries. The comparison between these two types of nations was done to point out key management differences among them, when aiming at guaranteeing prophylaxis against serious infectious diseases in their populations. In addition, we also examined the dependency on foreign vaccine supply of some countries, classifying them according to their capacity to supply the local demand with domestic facilities or *via* importation from other states.

## VACCINES CURRENTLY RECOMMENDED BY THE WHO

According to the WHO, children should be immunized with bacille Calmette–Guerin (BCG), diphtheria-tetanus-acellular pertussis (DTaP), MMR (combines Mumps, Measles, and Rubella), and vaccines to prevent Hepatitis B, poliovirus, *Haemophilus influenza*e type B (Hib), several serotypes of *Streptococcus pneumoniae*, rotavirus, and papillomavirus (3). In addition to these vaccines for children, the influenza vaccine is also recommended to be administered in certain susceptible groups, such as pregnant women, healthcare workers, children aged 6–59 months and the elderly (>65 years old) (3). Furthermore, the coverage of routine Expanded Program on Immunization (EPI), which includes vaccines against tuberculosis (TB), diphtheria, tetanus, and pertussis, polio and measles, varies from country to country (**Figure 1**).

## Vaccination for Poliomyelitis: An Example of a Nearly Eradicated Disease

Although poliomyelitis cases decreased greatly in 1988, 74 cases of this disease were reported in 2015. The majority of them occurred in Pakistan and in Afghanistan. Therefore, the goal proposed by the WHO is to eradicate poliomyelitis by 2018. Poliomyelitis is an infection caused by poliovirus that affects the human nervous system (4). The trivalent attenuated oral polio vaccine (tOPV), which includes the types 1, 2, and 3, has been used since the beginning of the 1960s. However, due to the polio type 2 vaccine components were pointed out as the infectious source leading to a large number of cases of vaccinederived polioviruses, global initiatives have suggested to switch from the trivalent to a bivalent polio vaccine. Such vaccine only includes type 1 and 3 viruses (5). Interestingly, the wild type poliovirus type 2 has not been reported since 1999 and was declared eradicated in September 2015. Besides, the poliovirus type 3 has not been detected since 2012 and the poliovirus type 1 is likely the only strain remaining in circulation.

As an additional effort to keep population protected against all types of poliovirus during the eradication program, the WHO instructed to include at least one dose of the inactivated polio vaccine (IPV) in the sequential shift from the tOPV toward the dOPV (6). The IPV is composed by the three types of poliovirus, which is intramuscularly administered. Clinical trials in children have shown that this vaccine is an excellent booster and capable of enhancing the mucosal immune response in primed subjects (4, 7).The future goal is to shift from dOPV to IPV at the time when type 1 and 3 polioviruses were eradicated.

## Vaccination for Respiratory Diseases: TB, Pneumonia, and Influenza

A different vaccine type, the BCG vaccine, has been used in over a billion people since 1921 to prevent TB (8). Although not able to induce a strong protective immunity in adults, the BCG vaccine has been shown to protect against meningitis TB disease in children (9). However, the BCG vaccine currently is not utilized in children from countries with low rates of TB incidence, such as the USA, Spain, Australia, Norway, Canada, and England (10). In those countries, the BCG vaccine is only recommended for those children showing a negative tuberculin skin test and that are continually exposed to adults with untreated or ineffectively treated for TB disease. Further, BCG vaccination is also recommended for health care workers in settings of frequent exposure to TB patients (11). Furthermore, because the BCG vaccine derives from attenuated bacteria passaged in the 1960s, the large number of passages affecting the banks available today has led to multiple genetic changes in the bacilli. Several studies supported the notion that this genetic divergence could be responsible for the variant protective capacity against TB shown by the various BCG vaccine strains (8, 12). Thus, an efficient BCG vaccine that provides full protection is still required. The major BCG manufacturers prequalified by the WHO are the Staten Serum Institute (Denmark), Serum Institute of India Ltd., Japan BCG Laboratory, and Intervax Ltd. (Canada). In addition, some Asian and Eastern-European countries possess their own locally-produced BCG vaccine, such as China (China National Biotec Group), Serbia (Torlak Institute), and Vietnam (IVAC) (8).

Bacteria-caused pneumonia, due to infection with various serotypes of *S. pneumoniae* (Pneumococcal disease) and Hib display a high rate of morbidity and mortality worldwide, although nowadays, the majority of the deaths take place in sub-Saharan Africa and Asia (13). Both pneumococcal and Hib vaccines are recommended by the WHO (3). However, not all

countries include these vaccines in their national immunization programs and, for instance, the public health systems of some South Asian countries do not use them at all (**Figure 1**). Thus, whereas these vaccines were introduced in the 1990s in most industrialized countries, still these prophylactics are not funded by public health systems in some developing countries, such as South Asian nations. Consequently, still 18/100,000 and 26/10,000 cases of Hib were reported in children younger than 5 years old in Vietnam and in China, respectively. To handle these high incidence rates, organizations including the Global Alliance for Vaccine and Immunization (GAVI) and the United Nations International Children's Emergency Fund have financed pneumococcal and Hib vaccines to provide coverage for developing countries (14). Several studies conclusively have supported the notion that public health systems should add these vaccines to their national immunization programs with their own funding, in every developing country. Thus, adopting these measures, the incidence of these major infectious diseases could be reduced (15, 16). Further, some GAVI-supported countries experienced a transition from GAVI-derived support to a fully self-financed Hib vaccination program. Thereby, strategic immunization plans are required to provide vaccines to their population (17).

Viral respiratory diseases generated by the influenza virus causes low rates of mortality but high rates of morbidity worldwide every year (18–20). This seasonal disease is mainly caused by two types of influenza viruses: A and B (21). The influenza A virus displays a high rate of variation causing frequent antigenic changes, in a process known as antigen drift. For this reason, the influenza vaccine confers only limited-time protection (up to 2 years) and it is necessary to reformulate and manufacture new influenza vaccines every year. Influenza vaccination is recommended by the WHO for high-risk individuals, including children, pregnant women, healthcare workers, the elderly and individuals suffering from chronic conditions, such as asthma, diabetes and heart disease (3). Further, organizations such as the American Academy of Pediatrics recommend the seasonal influenza vaccination for children of 6 months and older (22). However, the coverage of this vaccine still remains low despite the influenza vaccination strategies, including government involvement and national programs (23). Importantly, pandemic influenza H1N1 emerged in 2009, affecting mainly children and the elderly. The global number of deaths during the first 12 months of virus circulation was reported from 151,700 to 575,400 people (24). Moreover, the older age groups presented higher severity in postpandemic influenza outbreaks (25).

## Vaccination to Prevent Diphtheria, Tetanus, and Whopping Cough

Another vaccine of global relevance is the DTaP (14). This vaccine protects against three severe infectious diseases: diphtheria, tetanus, and pertussis. First, diphtheria is caused by *Corynebacterium diphtheria*, which produces pharyngeal infection, myocarditis, polyneuropathy, and systemic toxicity (26). Second, tetanus is caused by *Clostridium tetani* and the typical symptoms include muscle spam and contraction (26). Finally, pertussis, also known as whopping cough, is caused by *Bortedella pertussis*, which can produce loss of weight, subconjunctival hemorrhages, and syncope (26). Currently these three diseases circulate in the population worldwide and the highest rates are observed in children from countries with low vaccination coverage, especially in developing countries (27–29). However, and despite high vaccination coverage, several outbreaks have recently been reported in industrialized countries (30). For instance, an outbreak in the USA was reported in 2012, resulting in 48,277 cases of pertussis (31). According to the United States Center for Disease Control and Prevention (CDC), DTaP protects from whopping cough in 7 out of 10 vaccinated subjects, while it efficiently protects against the severe illness. In fact, the introduction of DTaP vaccine in the USA reduced from 100,000 to 32,000 cases of pertussis per year. Despite these good results, DTaP could fail to provide long-lasting protection in humans (31). It is important to indicate that the WHO estimates that there still are about 16 million cases of pertussis and 30,000 of diphtheria per year worldwide, being the highest rates in India (32). Therefore, these infections are still an important public health burden that requires close monitoring.

## Vaccination to Prevent Cervical Cancer

The nine-valent human papillomavirus vaccine (HPV) is recommended for routine vaccination of girls at age of 9 or 10 years old to confer protection against cervical cancer caused by the HPV (33). This new vaccine is significantly more expensive as compared to the other vaccines. Thereby, although it is highly recommended vaccine, not all children are being immunized to prevent this cancer (33). Despite the fact that the first HPV vaccine was available in 2006, today only two biopharmaceutical companies manufacture this vaccine (33). A study performed in France showed 95.93% effectiveness for the HPV vaccine in sexually active young women (34). Despite such effectiveness, a strong parent refusal remains in several countries to vaccinate children against HPV due to safety and effectiveness concerns, as reported in a survey in the USA (35).

## Vaccination to Prevent Diarrheal Diseases

Another recent vaccine included in the immunization programs of several industrialized and developing countries is the one to prevent rotavirus-infections (3). This virus is one the most common causes of severe gastroenteritis with diarrhea-related hospitalizations in children worldwide, which shows in particular high mortality rates in developing countries (36). The WHO has recommended that this vaccine should be included in all national immunization programs, being strongly recommended for countries showing a high mortality rate in children under 5 years old due to severe dehydrating diarrhea (37). Nowadays, an increasing number of countries, such as the USA and Germany have incorporated the rotavirus vaccine in their national immunization programs. A meta-analysis conducted on individuals of Europe, North America and Latin America showed that this vaccine has an efficacy of 53% against rotavirus infections, 73% against rotavirus-related hospitalizations, and 74% against severe diarrhea episodes (38).

## Vaccination to Prevent Typhoid Fever

Typhoid fever is a life-threatening systemic disease caused by human adapted *Salmonella enterica* serovars, such as Typhi, Paratyphi A, Paratyphi B, and Paratyphi C (39, 40). These are Gram negative enterobacteria that infect humans by contamination of food and water supplies, causing disseminated infections that compromise internal organs, such as spleen, liver, bone marrow, and blood (39, 41). The incidence of these diseases is low in industrialized countries (less than 10 cases per 100,000) and high in developing countries, specifically in Asia and in Africa (more than 100 per 100,000) (42–44). Importantly, a significant increase in *S.* paratyphi A has been reported in the last years in Asian countries, reporting up to 44-fold increase in the period 2007–2013 in Cambodia (45). Currently, there are three licensed vaccines to prevent typhoid fever: The Vivotif®, Typbar®, and the Typhim V® vaccines. The Vivotif® is a live attenuated vaccine approved by the FDA for use in humans, based on the Ty21a strain, which was generated in the 1970 by chemical mutagenesis (46). This vaccine was previously produced and distributed by Crucell Switzerland It Ltd., but recently the American company PaxVax has acquired the license for this product. This vaccine is provided as a lyophilized formulation (in capsules) and used orally to promote mucosal immunity against these bacteria. A large clinical study performed in Chile showed that the rate of protection after three immunizations was 69% (47). In contrast, the Typhim Vi® and Typbar® are inactivated vaccines consisting of the Vi capsular polysaccharide, which are produced by Sanofi Pasteur and Bharat Biotech, respectively (48). The Typhim Vi® vaccine is administered intramuscularly and confers an antibody-based protection (49). The rate of protection for this vaccine is close to 75% (50). Further, those vaccines do not confer cross-immune protection against *S.* Paratyphy A, for which does not exist a licensed vaccine available to prevent disease caused by this bacterium (51). Because of the immune memory conferred by both vaccines are very limited, their inclusion in immunization programs has not been recommended. However, the use of this vaccine has been encouraged by the WHO, especially when sanitation measures are threatened, for instance during natural disasters that impair the accessibility to clean water. Nevertheless, due to the growing emergence of antibiotic-resistant strains of *S.* Typhi in developing countries like India, the permanent use of these vaccines, as well as the generation of improved ones, would be highly appropriate to apply in their populations (52).

## Vaccination to Prevent Meningitis

Meningitis is an inflammation of the membranes covering the brain and spinal cord known as meninges, which can be caused by viral, bacterial or fungal infection, but also by due to noninfectious causes, as it has been reported (53). The main bacterial agents responsible for this disease are *S. pneumoniae,* Hib, and *N. meningitidis*, which could be prevented by available vaccines (54). Meningococcal disease has been reported worldwide, but largest epidemics have affected mainly sub-Saharan African countries, known as the "meningitis belt" having 430 million of high-risk population (53, 55).

According to the recent report in May 2017 by the CDC (56), there are two types of meningococcal vaccines available in the USA. The first vaccine is based on bacterial conjugates: Menactra® and Menveo®, both conferring protection against A, C, W, and Y meningococcal serogroups. The second is a serogroup B recombinant meningococcal vaccine: Bexsero® and Trumenba®. An additional vaccine, named MenAfriVac® (produced by the Serum Institute of India Private Ltda.), confers protection against *N. meningitidis* serogroup A (Nm A), which is the most prevalent in the African "meningitis belt" (55). The MenAfriVac® vaccine was a result of collaborative efforts between the WHO and the PATH in the Meningitis Vaccine Project, with the purpose of developing a vaccine against the specific agent affecting importantly the health of the African population, presenting a low-cost manufacturing and being independent of the cold chain distribution (57, 58). Since the national routine immunization strategic plan started in 2010, the incidence of Nm A meningitis fell from 0.27 per 100,000 in 2004–2010 to 0.02 per 100,000 in 2011–2013 (59). According to the recent WHO weekly record, 19 of the 26 countries belonged to the African "meningitis belt" have shown a sustained decreased incidence for Nm A cases, which means a reduction by at least 57% of the meningitis burden in that area (55). Also, clinical trials demonstrated that MenAfriVac® decreases carriage rates in immunized populations and provides herd immunity probably because of the high antibody titers observed during the development and safety testing of the vaccine (60, 61). Due to the national immunization program for this vaccine was a success, Ghana and Sudan currently include the MenAfriVac® in their routine immunization schedule (55). Despite the significant decrease in the prevalence on Nm A, it is important to highlight the necessity to continue with immunization programs to guarantee protection against different serogroups (62). Further, experts alert of the possible serogroup replacement, following application of massive immunization programs (63). In fact, in 2015 an epidemic with a novel strain of *N. meningitidis* serogroup C was reported in Niger and Nigeria. In addition, in 2016 the principal *N. meningitidis* serogroup W was found in Ghana and Togo, although with a low number of cases (55). For that reason, the continuous research in this area is a central challenge toward elimination of meningococcal meningitis epidemics in Africa.

## VACCINE TYPES, MANUFACTURING PROCEDURES, AND CURRENT RESEARCH ON MANUFACTURING STATUS

## Types of Vaccines and Manufacturing Procedures

Vaccines can be classified as live-attenuated, inactivated, sub units, recombinant, conjugated, toxoids, or DNA, according to the final preparation of the microorganism or antigen (64). Liveattenuated and inactivated microorganisms cover the major fraction of licensed vaccines for use in humans. Smallpox, BCG, yellow fever, polio, chickenpox, rotavirus, typhoid fever (Ty21a vaccine), and influenza are examples of licensed vaccines produced with live attenuated microorganisms (8, 65–67).  Only few vaccines are produced using recombinant technologies (hepatitis B virus, influenza, HPV) or *via* purification of partial components of a microorganism [*S.* Typhi Vi capsular polysaccharide, diphtheria, tetanus, pneumococcus, meningococcus, Hib, pertussis toxoid, and anthrax protective antigen (PA)] (73, 74). However, there has been an increased interest in the usage of these technologies in the past years (75).

There are different methods of vaccines production, which include isolation of microorganisms from either infected tissues (e.g., smallpox), bacteria growth in fermenters (e.g., vaccines for TB, typhoid fever, plague, whooping cough, diphtheria, tetanus, pneumococcus, meningococcus, pertussis, anthrax), isolation from virus grown in cell cultures (e.g., polio, chickenpox, rotavirus, hepatitis A virus (HAV), influenza) or isolation from virus grown in eggs (e.g., influenza, yellow fever). For the case of bacteria grown in fermenters, is not the microorganism itself that is used for the vaccine elaboration, rather some of its components from cell-free filtrates (e.g., vaccines for tetanus, pertussis, anthrax). For example, the anthrax vaccine adsorbed consists in the PA purified from filtrates by precipitation with alum, which also serves as an adjuvant (76).

An interesting change in the way of manufacturing has been recently carried out for influenza vaccines, which has been produced for more than 50 years in embryonated chicken eggs (77). However, GlaxoSmithKline (GSK) and Seqirus are currently producing influenza virus using cell culture technology in bioreactors (approved by the FDA in 2012) to generate new licensed influenza vaccines (78). Likewise, the Kaketsuken vaccine company is working on the development of a cell culturebased process, using the EB66 cell line, to elaborate a vaccine for pandemic flu, which is currently under clinical studies (79, 80). More recently, the Protein Sciences Corporation received approval for commercialization of a licensed novel influenza vaccine consisting of purified recombinant hemagglutinin antigens expressed in insect cell cultures (81). Similar efforts are in progress toward the development of cell culture-based yellow fever vaccines using Vero cell cultures in microcarriers (82). For anthrax, a plant-derived recombinant protective antigen has been developed as a vaccine, which is currently under evaluation in clinical trials (76, 83).

Thus, cell culture technologies, together with the enhancement of upstream and downstream processes, will bring production efficiencies to a next level as compared to the eggbased technology, and will increase manufacturing speed and capacities, thereby avoiding the shortage of these vaccines in the future (84, 85).

### Vaccine Research in the Industry versus the Academia

Vaccine portfolios in many pharmaceutical companies have decreased in the last decades due to the cost and time involved for vaccine development, which are much more costly and time consuming to develop than other drugs (86). However, pharmaceutical companies as well as academic institutions are continuously investing in vaccine research. For example, the number of vaccines in development has increased about twofold, according to a study comprising the 1995–2008 period in the USA (87). This fact can be explained, in part, by the advancement of alternative technologies, such as baculovirus-based recombinant vaccines, virus-like particles, viral vectors and RNA or DNA vaccines (74, 88–93). Moreover, with a world population projected to be of 10 billion by 2050, a 90% of it is estimated to live in developing countries (United Nations projection) (94). Thus, the subsequent increase in the vaccine market from USD 25 billion by today to USD 100 billion by 2025, will continue to encourage vaccine research and development (95).

Many research groups in academic institutions have made considerable efforts on vaccine discovery and research, but only few of them have been able to move forward into the development vaccine process. A reduced technology transfer efficiency may be due to difficulties on establishing private-academy license agreements (LA) (96). Indeed, Public-Private-Partnerships (PPP) has shown to be relevant for some vaccine developments, such as for the prototype of HIV vaccine (97). Thus, these LA and PPP enable the implementation of new and improved vaccines in high-tech centers before a product is transferred into the market. Another factor is the requirement of facilities with Good Manufacturing Practicing (GMP) certification and high-quality personnel to develop vaccine production processes. The staff capacities and facilities to investigate, develop and manufacture vaccines are key to respond rapidly to the global emergencies, such as the recent Ebola outbreak (98).

The increase of vaccine manufacturers has impacted on the global market, allowing to lower the prices of vaccines and to improve the global demand. Further, partnerships, such as GAVI Alliance, UNICEF, and the WHO have also been key for enhancing that kind of vaccine production in developing countries (99). As example, the new vaccine manufacturing countries such Brazil, the Russian Federation, India, China, and South Africa (known as BRICS) play a substantial and increasing role in the global vaccine market. These countries not only produce traditional vaccines at competitive low costs and under the WHO-prequalified standards, but they also generate innovative products due to current strategic alliances with multinational corporations (99, 100). The most successful case of this strategic alliance, is the Bio-Manguinho plant in Brazil, that will be producing an affordable measles and rubella vaccine with the support of the Bill & Melinda Gates Foundation together with the Brazilian Ministry of Health (100). An arising number of pharmaceuticals along with the NIH are interested in enlarging the number of vaccines manufactured in those institutes, which in turn involved discussion of the agreement of the Trade Related Aspects of Intellectual Property Rights.

## DIVERSITY OF IMMUNIZATION PROGRAMS WORLDWIDE: REGIONAL EXAMPLES AND THE GAP BETWEEN INDUSTRIALIZED AND DEVELOPING COUNTRIES

Worldwide, the diversity in national immunization programs is extensive, therefore the list of vaccines included and distributed in each country shows significant differences (**Figure 1**). Furthermore, the vaccination plan for the USA might even be different depending on the state, while in Europe the immunization plans have significant differences among the countries belonging to the European Union (**Figure 1**). On the other hand, there are variations in the financing mechanisms for vaccine production within Europe. For instance, the National Health System funds the rotavirus vaccine in Germany, but not in Spain. Other vaccines, such as the live attenuated Japanese encephalitis, cholera, and yellow fever vaccines are recommended only in some Asian countries, such as in India and in Thailand. Furthermore, meningococcal C conjugate vaccines are included in the National Health System of Australia, Chile, and Spain, but not in those of Asian countries like in India. Another example of diversity on immunization schedule is the BCG vaccine against TB. This vaccine is being administered only in some countries in Europe, Asia, Africa, and South America, but it is not administered in industrialized countries such as in the USA (14). **Table 1** summarizes the differences of the immunization programs between seven countries, including industrialized and developing countries (101, 102).

Germany is an example where vaccination is mostly voluntary with a reduced role of the state in the implementation of vaccination programs. Around 90% of the vaccines are given by private physicians and only the remaining small fraction of the vaccines is given by public institutions, schools or daycare centers (103). Massive school immunization programs are not mandatory, but the immunization status is checked at schools. This information is collected and documented by the Robert Koch Institute. The Berlin measles outbreak of 2015 and the death of a non-vaccinated infant raised the discussion as to whether vaccination in Germany must be mandatory (104). This discussion has been intensified considering that the Europe is confronting the largest immigration since the World War II. The collapse of national immunization programs in the countries undergoing political turmoil has led to children-disease outbreaks, which could have been prevented by vaccination. Moreover, refugees are susceptible to diseases due to overcrowding, physical and

#### Table 1 | National immunization programs of seven countries.

*Developed and developing countries were selected according to their geographical area and income. Orange: not funded by the public health system. Blue: funded by the public health system.*

*aDeveloping countries. bDepends on the region, this vaccine is included in the public health system.*

*cOnly vaccine against measles.*

*BCG, bacille Calmette–Guerin; HepB, hepatitis B virus; DTaP, diphtheria, tetanus, and pertussis; MMR, mumps, measles, and rubella; Hib, Haemophilus influenzae type B; HPV, human papillomavirus; JE, Japanese encephalitis live vaccine.*

psychological stress, malnutrition and low availability of sanitary systems. These health aspects and conditions constitute a serious threat to immigrants, as well as to international programs aimed at eradicating vaccine-preventable diseases. Recent studies of measles, mumps, rubella, and varicella seroprevalence in refugees in Germany have shown satisfactory immunity in adults but low seroprevalence in children, suggesting thorough and prompt vaccination of children entering Europe (105). The opposite has been found for hepatitis A immunity in refugees in Germany, where the high rate of HAV protection supports the thesis that the probability of large HAV outbreaks in current German refugee centers is low (71). Nevertheless, vaccination of refugees against HAV is highly recommended.

The immunization programs in the USA follow the CDC guidelines (106). In this country, as mentioned earlier, vaccine coverage differs widely among states, varying for instance with ≥2 doses of HAV from 41.2% in Mississippi to 72.8% in Nebraska (107). Recent nonmedical exemptions in immunization laws have prompted serious concerns about potential vaccine coverage weakening. However, after the recent outbreaks of vaccinepreventable diseases mandatory immunizations at entry-schools and primary care facilities have emerged. Indeed, those states that allow exemptions, including religious and philosophical reasons, have shown a significantly higher incidence of vaccinepreventable diseases, as compared to those states allowing less exceptions for vaccination (108). Interestingly, the coverage of vaccines in the USA will depend on the insurance plan of each individual. Accordingly to the CDC, the coverage of children aged 19–35 months was lower in those children uninsured or covered by public insurance programs, such as Medicaid, as compared to private insurance-covered kids (107). However, some the USA vaccine manufacturers and the National Vaccine Programs offer help to those people who cannot afford some vaccines, such as the one for HPV. Importantly, up to 32.9% of children of 19–35 months of age in the USA live below poverty level and can fail to receive all the required vaccines (107). To overcome this problem, the Vaccines for Children Program in the US offers free vaccines to children living in poverty (107).

In South America, the Pan-American Health Organization (OPS) provides a caring cooperation system, named the "Fondo Rotatorio," designed to obtain the vaccines recommended by the WHO at low prices (109). As for the case of Chile, the Public Health Institute and the Ministry of Health direct the Chilean National Immunization Program (CNIP) following international recommendations. Vaccines included in the CNIP are funded by the government and given to hospitals, family health centers and some schools in Chile. The introduction of the latest vaccines in the CNIP has significantly reduced the incidence of certain diseases, such as bacteria-caused pneumonia and cervical cancer. One example is the 10-valent pneumococcal vaccine, which was introduced in January 2011 and thereafter, the number of hospitalizations due to pneumonia were successfully reduced (110). Such effectiveness of the 10-valent pneumococcal vaccine has also been demonstrated in other South-American countries (111). In 2015, the Chilean government supported the introduction of the HPV vaccine in the CNIP and thereby, most of 9–10 years old girls have been vaccinated since then as a program to prevent cervical cancer.

Thus, each country has its own national immunization program (112), which in most cases includes vaccines that are sponsored by their public health systems reaching different levels of coverage (**Figure 2**). Nevertheless, many developing countries have difficulties to finance all the vaccines recommended by the WHO. As a result, different organizations have arisen to provide economical support to the developing countries requiring vaccines. For instance, the Global Vaccine Activation Plan (GVAP) has established itself the goal of reducing some vaccine-preventable diseases by 2020 (113). Moreover, most traditional vaccines are sold at lower prices to organizations, such as UNICEF and the Pan-American Health Organization to reach developing countries (14). Although global coverage has improved, in countries such as India, Nigeria, Pakistan and Indonesia, a low immunization coverage still exist (113). It is noteworthy that 35 of the 45 classified as lower-middle income countries by the World Bank Classification are not being supported by GVAP Alliance, thereby these countries are struggling to reach underused and new vaccines to immunize their children (14). Also, one of the GVAP goals was to eliminate the maternal and neonatal tetanus, measles, and rubella in 2014, but unfortunately this goal was not achieved (113). One of the main reasons for this failure has been the unstable political situation in some countries and the inefficient introduction of these vaccines in national immunization programs (113). Therefore, economical gaps still remain between industrialized and developing countries to accomplish efficient immunization programs for their children. With globalization, leading to increased and fast movements of goods and people traveling to all remote areas in the world, these differences in health protection can be a risk for outbreaks, epidemics or even worse, pandemics. Importantly, several organizations including the GAVI Alliance, UNICEF, the Bill & Melinda Gates Foundation, the United States National Institute of Allergies and Infectious Diseases, the WHO, together with governments and other institutions support the goals of the GVAP to reduce some vaccine-preventable diseases by 2020 (113).

## VACCINE MANUFACTURE AND DISTRIBUTION: STATUS OF ACADEMIC, PUBLIC, AND PRIVATE MANUFACTURING COMPANIES

#### Vaccine Production and Distribution

Although mainly private pharmaceutical companies have engaged in vaccine manufacturing and distribution, there are also successful efforts made by academic or public institutions to achieve this goal (**Table 2**). Vaccine manufacturing requires specific and expensive facilities with high scale production, and quality standards to ensure consistency and controlled elaboration of these products. This is typically achieved following the guidelines of the current (c) GMP in compliance with the local regulatory authorities. Therefore, most of the countries have contract agreements with specific cGMP-certified manufacturers to purchase the vaccines required for their populations. For example, the private sector is in charge of the 5–10% of the vaccines market in Asia (114).

The USA is one example of a country, in which both private and public sectors provide vaccines for their population (115). This is an advantage, because the public health system can choose from different sources and prices. The main pharmaceutical companies that produce and distribute vaccines around the world include GSK, the United Kingdom; Pfizer, the USA; Sanofi Pasteur, France; Merck & Co., the USA; Roche, France; Seqirus, Australia; Valneva SE, France (**Table 2**). In addition, emerging pharmaceutical companies, such as Astellas Pharma, Japan; Takeda, Japan, and AstraZeneca, United Kingdom currently invest in vaccine R&D. Other international companies, including the Serum Institute of India and the Bharat Biotech International supply vaccines to countries without local vaccine manufacture facility, such as Chile. Particularly, the Serum Institute of India is a state-owned vaccine manufacturing center that produces most of the vaccines recommended by the WHO including BCG, polio, Hib, DTaP, and MMR. Similarly, national public enterprises, including the Immunobiological Technology Guinhos (Bio-Manguinhos/Fiocruz) and the Butantan Institute supply most of the vaccines in Brazil (**Table 2**). Importantly, the two institutions previously mentioned supply about up to 83% of the Brazilian National Immunization Program demand, thereby reaching up to 179,855,000 national doses (116). A different situation can be found in Germany, where most of the vaccines are purchased from the private sector (90%) and 90% of them are financed by statutory insurance policies (117). The government provides the rest of the vaccines as part of special immunization programs. Recent studies have shown that no more than 0.47 and 0.25% of the German and Spanish healthcare budget, respectively, are addressed to vaccine production (117).

Due to the problems stated above, in the year 2000 an organization aimed to create alliances of vaccine manufacturers in developing countries was established. This organization, known as the Developing Countries Vaccine Manufactures Network (DCVMN), includes near 50 vaccine manufacturers in 17 developing countries in Latin America, Africa, the Middle East, and Asia (118–120). The companies that are members of this organization produce more than 40 different vaccines, including the ones recommended by the WHO including BCG, polio, Hib, DTaP, and MMR (**Table 2**) (118, 120). Although the DCVMN main goal is to provide a high quality (cGMP compliant) and sustainable supply of vaccines for developing countries, there are still not enough to provide the increasing demand of vaccines.

#### Vaccine Shortages

The coverage of the national immunization programs relies on the available supply of vaccines. Several countries have experienced vaccine shortages at some point, which have included BCG, Hib, DTaP, pneumococcal conjugate, MMR, meningococcal, yellow fever, and influenza vaccines (121, 122). As an example, Sanofi Pasteur, one of the major producers of BCG, the current

*BCG, bacille Calmette–Guerin; HepB, hepatitis B virus; DTP, diphtheria, tetanus, and pertussis; MMR, mumps, measles, and rubella; Hib, Haemophilus influenzae type B; HPV, human papillomavirus; JE, Japanese encephalitis live vaccine.*

vaccine for TB, experienced significant manufacturing problems during 2012 and 2014. As a result, distribution of this vaccine was seriously compromised in several countries (123). Indeed, approximately 16.5 million doses shortfall of BCG occurred at the end of 2015 was estimated, using mathematical models, to be associated with 7,433 excess of TB deaths worldwide (124). In 2015, short supplies for the meningococcal vaccine worldwide threatened the health of the population in Nigeria, a place where an important epidemic of meningitis took place (125). An additional example is the Hib boost vaccine, for which doses were not available in the USA from December 2007 to September 2009 (122). Moreover, several physicians have reported shortages of influenza vaccines, especially for high-risk populations in the USA during the years 2004–2005 (126). Further, Africa and the USA have also experienced shortages for the yellow fever vaccine during the last 2 years (127–129). Similarly, significant shortages of the pneumococcal conjugate vaccine occurred during the period 2003–2004, causing an important decrease of 10.6% of the coverage of >4 doses of the seven-valent pneumococcal conjugate vaccine in 16-month-old children (130). Likewise, such shortage issues have prompted the concern of elaborating protocols for ensuring availability of those vaccines for at least the high-risk populations (131). Because the pandemic of influenza is highly extensive, the demand for this vaccine worldwide is very high, causing sometimes problems of vaccine shortage (132, 133). This situation is particularly dramatic when pandemics on influenza arise, such as the H1N1 in 2009 (134).

Different reasons can explain disruptions of the vaccine supplies, such as vaccines that leave the market, problems in the production, loss of the GMP in manufacturing centers/companies, and changes in the formulation of vaccines (135). An important correlation is that fewer vaccine manufacture suppliers exist for one vaccine the larger the impact of supply shortage can have on the population (135). To solve the vaccine shortage in case of epidemics, global vaccine stockpiles have been established for vaccines, including smallpox, meningococcal, yellow fever, oral cholera, and pandemic influenza vaccines (136). Moreover, the challenge for institutions, such as the Brazilian government, is to make investments for local vaccine development and manufacturing to avoid international dependency and the threat of shortage (116).

## GLOBAL EMERGING DISEASES AND ANTIBIOTIC RESISTANCE

The Ebola, Zika, and influenza virus pandemics are examples of worldwide emergencies that have recently affected various regions of the planet. In 2009, the H1N1 influenza pandemic resulted in the highest number of cases in Mexico (134). In April of 2009, the first cases with severe respiratory disease started to be concentrated in the Federal District of Mexico's most populated area. The Mexico's National Institute of Respiratory Disease struggled with such situation to contain the propagation of the influenza virus (137). Months later, the H1N1 virus was spread to over 213 countries causing 16,226 deaths and the WHO declared it to be the first flu outbreak in the last 41 years (138). The H1N1 2009 pandemic was identified as a new influenza A subtype of swine origin, and consequently, at that moment no vaccines were available. After that outbreak, a vaccine was rapidly developed, include the 2009 H1N1 influenza virus antigen in order to protect against that virus (139). However, if new mutations arise resulting in a new pandemic subtype, then the available vaccine will be useless and again no vaccine will be accessible to protect against a potential new virulent strain with a high rate of mortality, such as seen with the previous H1N1 influenza A virus pandemics.

In 2014, West Africa experienced a devastating outbreak of Ebola and multiple countries were affected. In response to that situation, several countries and institutions such as the WHO and the CDC activated emergency operations to control the situation (98). Although the end of transmission of Ebola was reported in Liberia and in Guinea, still the WHO in Guinea, Liberia and Sierra Leone has still reported a total of 28,616 Ebola cases, with 11,310 deaths (140). Ebola virus is associated with hemorrhagic fever and is transmitted by corporal fluids. No vaccine or treatment is available for this virus; thereby efforts in that situation were to limit transmission of the disease.

On the other hand, according to the CDC, most of the Zika virus cases have been reported in many countries of South America, Africa, Asia, and the USA (141). This virus is transmitted by a mosquito-borne (*Aedes aegypti*) and symptoms include mild fever, headache, arthralgia, myalgia, non-purulent conjunctivitis, and a pruritic maculopapular rash (142). However, the most concerning effect that has been associated with Zika virus is the prenatal microcephaly (143).

According to the WHO-vaccine pipeline tracker, vaccines against AIDS, malaria, enteric pathogens, including human norovirus, the respiratory syncytial virus, Zika virus, Dengue virus, and pulmonary TB are in different stages of development. Some of these diseases, such as AIDS or pulmonary TB have been a concerning problem, since for several years have not been obtained a definitive cure or an efficient vaccine to prevent them. In addition, other diseases, such as the ones caused by the Zika virus, have had emergency problems that have required a rapid response. One prompt response strategy for the past Ebola outbreaks has been the use of anti-Ebola antibodies from the blood of disease survivors. Therefore, strategies with monoclonal antibodies to treat Ebola are currently being studied (144). Moreover, research on nanoparticles, adenovirus-based, modified Vaccinia Ankara-based, and recombinant-rabies vaccines against Ebola are ongoing, even in phase I, II and III of clinical trials (98, 145). From Ebola vaccines in clinical trials so far, the most advanced one is a recombinant vesicular stomatitis virus–Zaire Ebola virus (rVSV-ZEBOV) vaccine that has been licensed to Merck and recently, showed to be effective in susceptible individuals (146). On the other hand, strategies such as adenovirus-based recombinant vaccines and cell culture-derived inactivated vaccines using BHK and Vero cells are under research for Zika virus vaccine development (147). Despite the research ongoing about Zika and Ebola viruses, or other common and fastidious viruses such as respiratory syncytial virus and human norovirus, no vaccines or efficient treatment are still available. Thus, high technology centers are urgently needed to provide a solution to these problems and offer a rapid response to global health emergency states.

As emerging diseases, microorganisms with multiple resistances to antimicrobial agents have been reported in the past years. Bacteria resistance to the available antimicrobial agents, such as *Klebsiella pneumoniae*, *Staphylococcus aureus*, *Escherichia coli*, and *Mycobacterium tuberculosis* have alarmed health care worldwide for their resistance to antimicrobial agents (148–151). Furthermore, availability of an effective therapy for patients infected with those microorganisms is limited and more research and development is needed (152). Despite policies concerning the use of antimicrobials and the development of new drugs, it is urgent to increase the vaccine manufacturing capacity to prevent the spreading of these infections with multiple antibiotic resistance (153).

## CONCLUDING REMARKS

There is no doubt that many diseases have been prevented due to the implementation of extensive vaccination programs. Domestic health public systems worldwide are committed to increase vaccination coverage for the population through national immunization programs. Thus, the WHO recommends to immunize children with BCG, DTaP, MMR, and vaccines to prevent hepatitis B, poliovirus, Hib, several serotypes of *S. pneumoniae*, rotavirus, and HPV. However, not all these vaccines are included in the national immunization programs of most countries. Not only the problem is the inclusion of some vaccines in local programs of immunizations but also the cost associated with its production, implementation, and delivery are part of the barriers. In this line, it is important to highlight the effort of some organizations such the WHO, the PATH, the GAVI Alliance, the UNICEF, the Bill & Melinda Gates Foundation, among others, to include as much population as possible in these immunization global strategies. Furthermore, shortages around the world have taken places during the past years, which have underscored the necessity to improving the capacities and infrastructure to produce and distribute vaccines. It is important to underscore the role played by new countries manufacturing vaccines, which include Brazil, the Russian Federation, India, China and South Africa (a group known as BRICS). Such local production has contributed to ensuring access to traditional vaccines and to maintaining the stability of immunization programs in developing countries. Also, an important gap between industrialized and developing countries prevails in this field. Further, Ebola, Zika, influenza virus pandemics, and antimicrobial resistance have raised alarms, questioning whether we are prepared to control rapidly and efficiently viral pandemics worldwide.

## AUTHOR CONTRIBUTIONS

ERJ, FT, NMD, and AK wrote the manuscript; ML, LC, SB, CR, and YG reviewed the manuscript; and AK reviewed and approved the version to be published. All authors listed have made substantial and intellectual contribution to the work.

## REFERENCES

## FUNDING

This work was supported by the Millenium Institute on Immunology and Immunotherapy from Chile (P09/016-F for AMK), CONICYT/FONDECYT POSTDOCTORADO No. 3160249, FONDECYT grants number: 1150862, 1070352, 1050979, 1040349, 1100926, 1110397, 1131012, 1140010, 1140011, 1150862, 1150173, 1161525, 1160695, 1140011, 3140455, 1160336 and 1170964. Biomedical Research Consortium (BMRC 13CTI21526-P4 for AK and 13CTI21526-P5 for SB). CRP-ICGEBCRP/CHI14-01. FONDEF grant D11I1080. PMI-UCH 1301. We thank to the Max Planck Society for providing the financial support for the PhD project of FT.

years old in Karachi, Pakistan, and Kolkata, India. *Clin Vaccine Immunol* (2014) 21(5):661–6. doi:10.1128/CVI.00791-13

the 2003–2004 pneumococcal conjugate vaccine shortage. *Pediatrics* (2008) 22(4):e835–40. doi:10.1542/peds.2008-1092

153. World Health Organization. *Global Action Plan on Antimicrobial Resistance*. WHO report. Geneva: World Health Organization (2015).

**Conflict of Interest Statement:** The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

*Copyright © 2018 Rey-Jurado, Tapia, Muñoz-Durango, Lay, Carreño, Riedel, Bueno, Genzel and Kalergis. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor 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.*

# Education in Vaccinology: An Important Tool for Strengthening Global Health

#### *Paul-Henri Lambert1 and Audino Podda2 \**

*1University of Geneva, Geneva, Switzerland, 2GSK Vaccines Institute for Global Health, Siena, Italy*

Over the past 20 years, education of scientists and public health professionals in Vaccinology has increased dramatically. There are now many international, regional, and national courses that provide education in vaccinology. The proliferation of these courses and the high number of applications submitted demonstrate the increasing and continuous need for improved education in this field since, generally, comprehensive vaccinology training is not offered to medical and/or biological sciences students as part of their Universities courses and consequently there is insufficient knowledge of vaccine topics among health-care providers. Multidisciplinary vaccinology courses have not only educational purposes but they may also contribute to strengthening the development, testing, and use of vaccines, which remain the most efficient tool for infectious disease prevention. The courses available have a varied focus and prioritize topics based on the trainees' different levels of professional exposure and requirements. Overall, they might be classified in two key categories: (i) courses targeting students who, after their university studies in Medicine, Biology, etc., develop a strong interest in vaccines, would like to learn more about the various aspects of vaccinology, and potentially develop a career in this field (postgraduate courses); (ii) courses targeting postdoctoral professionals, who already have a sufficiently broad knowledge of vaccinology, but would like to develop stronger skills to be able to play a leading role in decision-making for vaccine development (advanced professional courses). Both postgraduate and professional courses are available and are based on comprehensive curricula. In the future, particular attention should be paid to include in the training curricula topics that might help vaccine development, efficient and sustainable vaccine introduction through epidemiologically sound vaccination programs, and best practices to address associated challenges, including vaccine hesitancy which could become a threat to successful implementation of vaccination programs, particularly in developed countries. In addition, it appears that the next phase of vaccinology training could benefit from a global and more structured platform that could facilitate exchanges and collaboration and amplify the current capacity for disseminating vaccine education for future vaccinology leaders around the world. This would be favored by synergizing the efforts currently devoted to vaccinology education. To initiate this process of analysis and systematization, a multinational effort is needed.

*Edited by:*

*Aldo Tagliabue, Istituto di Ricerca Genetica e Biomedica (IRGB), Italy*

#### *Reviewed by:*

*David J. M. Lewis, Imperial College Healthcare NHS Trust, United Kingdom Anna Prugnola, GlaxoSmithKline, Belgium*

> *\*Correspondence: Audino Podda<EMAIL_ADDRESS>
#### *Specialty section:*

*This article was submitted to Vaccines and Molecular Therapeutics, a section of the journal Frontiers in Immunology*

*Received: 21 March 2018 Accepted: 07 May 2018 Published: 24 May 2018*

#### *Citation:*

*Lambert P-H and Podda A (2018) Education in Vaccinology: An Important Tool for Strengthening Global Health. Front. Immunol. 9:1134. doi: 10.3389/fimmu.2018.01134*

Keywords: training, vaccinology, education, global health, vaccines

## INTRODUCTION

Education in vaccinology is an important priority to strengthen development, testing and use of vaccines, which remain the most efficient tool for the prevention of infectious diseases both in developed and developing countries. The several courses available worldwide today have a different focus and curricula are tailored to the trainees' different levels of professional exposure and requirements (1, 2). Overall, they might be classified in two key categories: (i) courses for more junior scientists who, after completion of their biological studies at University, would like to know more about vaccines and vaccinology and might potentially develop a career in this field (postgraduate courses); (ii) courses for experienced scientists who already have a quite good knowledge of vaccinology and are ready to develop a deeper competence to lead vaccine development projects at various levels of responsibility and to actively participate in strategic groups deciding on vaccination policies at national, regional, or international levels (advanced professional courses).

## POSTGRADUATE COURSES IN VACCINOLOGY

Some of the disciplines representing the fundamental scientific background for efficiently working in a vaccine development environment, such as clinical aspects of infectious diseases, microbiology, immunology, epidemiology, biostatistics, and others, are regularly taught in University courses; however, most often these courses do not have a focus on the whole vaccine development process or on the public health context for the introduction of new vaccines and rarely are these disciplines presented with a multidisciplinary and holistic approach (3). In addition, theoretical teaching is not enough, and there is also a need for practicebased training and exposure to vaccine development-orientated activities. This is particularly true for disciplines that are not usually taught in university courses, such as Pharmacovigilance, Regulations, and Ethics in vaccine R&D studies and, particularly, aspects related to animal and human research. In this regard, internships within an experienced project team are an opportunity not only to allow young scientists to learn day-by-day vaccine development work but also to introduce them into the dynamics of a scientific community working together toward a common goal.

Multidisciplinary vaccinology courses are an important priority particularly for scientists from developing countries where vaccines have significantly contributed to the dramatic decrease in the number of deaths, due to infectious diseases, particularly in children below 5 years (4). However, in these countries, almost five million children still die every year and many of these deaths are due to vaccine preventable diseases. Therefore, there is a huge need not only for new vaccines against diseases mostly affecting developing countries, for which a vaccine is not yet available, but also for significant efforts and resources to introduce in Africa, Asia, and in general in low- and middleincome countries (LMIC) vaccines that are already available to children of developed nations. Development and introduction of new vaccines in these countries is obviously dependent on availability of locally generated data, particularly the highquality clinical data needed by regulatory authorities for vaccine registration and by WHO for vaccine pre-qualification. An essential requirement to make this happen is to have a cohort of well-trained scientists from developing countries who have a clear understanding of the whole process behind vaccine development and subsequent vaccine distribution. With these capabilities, local scientists may become active players and efficiently implement the various activities needed for registration of new vaccines and then support post-licensure vaccine introduction in the context of country tailored immunization campaigns. Therefore, vaccinology courses for scientists from developing countries should include classes on epidemiology and clinical development, but also education on public health systems operations, cold chain logistics, and vaccine distribution. Given the challenges associated with such extensive vaccinology training, identification of suitable candidates for the training activities is really key. Participants may have different educational backgrounds and different R&D experience; therefore, well thought selection criteria based on a grading system should be established upfront to make sure that selected candidates can get the most from the training activities.

An example of this approach is given by the Master in Vaccinology and Pharmaceutical Clinical Development of the University of Siena (5), which one of us, AP, contributed to set up and implement. This course, a collaborative effort between academia and vaccine industry, particularly tailored for young physicians from developing countries, is an 18-month program, combining theoretical and practical training. The theoretical teaching component includes 10 modules in the key vaccinology disciplines (Public Health and Vaccine Development Process; Immunology and Preclinical Research; Manufacturing and Quality Control Processes; Infectious Diseases and Vaccine Prevention; Clinical Development Methodology, Biostatistics and Clinical Data Management; Pharmacovigilance; Epidemiology, Health Systems and Economics; Good Clinical Practices, Clinical Quality Assurance and Clinical Trial Operations; Regulatory Affairs; Policies and Recommendations for Vaccines in the World) and, in addition, parallel educational seminars for personal and professional development. This extensive theoretical training is supplemented by a 7-month training, at the University of Siena and within different departments of the sponsors and collaborative institutions, followed by investigational site training. Finally, the value of this course to the students is maximized by a faculty including worldwide experts from well-known international universities, supranational organizations, and vaccine industry (5).

## ADVANCED COURSES IN VACCINOLOGY

In the past 20 years, there was an explosive development and introduction of new vaccines that have or may have a considerable impact on public health strategies. As a result, there is now an increasing need for experts with a broad understanding of major issues in vaccinology. This need exists as well in industry, including major players and subject matter experts, as in academia and public health. In fact, it is of critical importance for decision

makers in industry to understand the needs and the determinants that will influence the use of a given new vaccine in various country settings. Similarly, experts involved in public health strategy and in decisions related to the introduction of a new vaccination program at national, regional, or international levels must know key issues in the development process, essential safety considerations, limitations of the manufacturing process, and vaccine-related economic issues, e.g., cost-effectiveness. Managing real or alleged post-licensure safety issues is of critical importance. These aspects are also of great concern for academic professionals involved in training scientists with a potential role in vaccine development or monitoring.

A good example of this type of training is the Advanced Course of Vaccinology, ADVAC, which one of us, PHL, contributed to set up and implement. This course, organized on an annual basis since 2000 by University of Geneva and Fondation Mérieux, at Veyrierdu-Lac (France), in partnership with WHO, Johns Hopkins SPH & CDC and support from the European Commission and the Bill and Melinda Gates Foundation (6). At inception, it was aiming at filling major gaps in global vaccination strategies: (i) a lack of scientists with a broad vision of issues related to vaccines and immunization, (ii) a lack of qualified decision makers to identify priority targets in vaccinology, and (iii) a lack of qualified policy makers for deciding on the introduction of new vaccines in vaccination programs. Since 2000, 18 courses have been organized, gathering in total 1,070 participants from over 100 countries (**Figure 1**). To ensure a maximal impact, it appeared of particular importance to select highly motivated candidates, likely to have soon increasing responsibilities. It was also critical for appropriate networking to maintain a course format allowing the mixing of people with diverse professional backgrounds and diverse geographic origin: 41% came from high-income countries, 42% from LMIC, and 17% from industry (**Figure 1**). The ADVAC curriculum is providing a broad view of the various aspects of vaccinology: (1) priority targets for vaccine R&D, (2) understanding vaccine-induced immune responses, (3) new vaccine approaches, (4) clinical assessment of vaccine efficacy, (5) vaccine safety and regulatory aspects, (6) decision-making process for introduction of new vaccines, (7) defining optimal vaccination strategies, and (8) dealing with real or alleged adverse effects. The success of these courses is certainly dependent on the quality of the lecturers who are all top level vaccinologists on the international scene. However, a key factor is the interactive nature of all sessions, particularly in small groups or during group exercises including role play sessions and informal debates. The concurrent evaluation of training sessions is particularly helpful to adjust the level of training to the needs of the students. A follow-up program for ADVAC alumni has proven effective to maintain and increase the network of vaccinologists that is resulting from the initial training effort.

#### DISCUSSION

Several postgraduate and advanced professional courses are available for training of junior and senior scientists, interested to deepen their respective knowledge in vaccinology. As shown by the examples mentioned in this review, some of these courses have already good multidisciplinary curricula; however, looking at the challenges and gaps that still limit the expansion and the sustainability of vaccination programs, there are a number of topics that should be more deeply addressed in future trainings.

An important gap toward expansion of vaccination in developing countries, particularly in Africa, is the lack of a sufficient manufacturing capacity that could enable local development and production of new vaccines, thus making vaccination programs sustainable in most of LMIC once GAVI support is over. Among other factors, development of local manufacturing capacity is affected by lack of a well-trained and competent pool of local scientists and technicians who could reliably support technical operations ranging from technology transfer activities to development, formulation, manufacturing, quality control, and release of vaccines.

Recently, some training courses on these aspects have been organized by WHO, also in collaboration with both public and private institutions (7, 8). In addition, as part of the ADITEC project funded by the European Commission (9), the WHO and the University of Lausanne organized various theoretical and practical 1-week courses in "Adjuvants and vaccine formulations" with the objective of training students on production, purification, characterization, and control of recombinant antigens, and on methods of preparation of adjuvants, including oil-in-water emulsions and aluminum gels, their formulation with antigens, and quality control of the resulting vaccines. Outcome of these technical trainings was excellent, based on the feedback received, and similar initiatives should be more frequently organized and offered to fruition in the future.

Anti-vaccination sentiments are heterogeneous beliefs, commonly defined as vaccine hesitancy, may represent an important cause of reduced vaccination coverage, both in developing and in developed countries, and sometimes may lead to recrudescence of infectious diseases for which vaccines have been available for a long time (10). Despite their unquestionable contribution to the reduction of morbidity and mortality from infectious diseases and, more in general, to an increased level of public health worldwide, for several reasons, mostly unfounded, vaccines have been associated to negative perceptions about their safety and, consequently, a growing sense of mistrust is associated with their use and should be properly addressed. Adequate education of health-care professionals is of paramount importance to address and reduce parental anxiety, concerns, and fears and therefore vaccinology trainings should more and more include well-documented sessions on vaccine safety. Similarly important is that vaccinologists are appropriately educated also on the potential side effects of vaccination, including identification and quantification of risks, so that, providing balanced and respectful information, they may contribute to re-establishment of trust (11).

An alarmingly high number of emerging bacterial infections are caused by the increasing anti-microbial resistance (AMR) worldwide and they may play an even worse effect on global morbidity and mortality in the near future (12). This is largely due to excessive and often inappropriate use of new antibiotics in medical practice and to the poorly controlled antibiotic use in animal food industry. Education of vaccinology scientists on the achieved reduction of antibiotics use and AMR by vaccination, with tangible benefits going beyond the non-vaccinated populations, through herd immunity,

#### REFERENCES

might push toward development of new future vaccines having also AMR reduction in their target product profile. This might also lead, on the one side, to better quantify the magnitude of antimicrobial use and of AMR for a given vaccine preventable disease and, on the other side, to select more appropriate vaccine candidates, including vaccines against highly resistant serotypes of the pathogen and/or virulence factors relevant for resistance acquisition.

Some other aspects deserve more and more attention in future vaccinology trainings; they include (i) preclinical and clinical vaccine assessment in LMIC, (ii) financing of vaccination programs, (iii) vaccine delivery, (iv) vaccine introduction strategies, and (v) vaccine regulations.

## CONCLUSION

Our vision of the future of vaccinology, and associated medical and social impacts, is that more and more scientists will be required for the implementation of all aspects of the vaccinology lifecycle process, from vaccine research to optimal vaccine use in the field. Therefore, the importance of appropriately developing the technical skills of next generation vaccinologists is paramount, best initiatives currently devoted to vaccinology education should join forces and, with a multinational effort, a global and structured platform for future training of vaccine scientists around the world should be developed. To achieve this goal, a global commitment to provide continuous education and training is needed from all stakeholders, including Academia, Industry, and Public Health Institutions, with the ultimate objective of ensuring sustainability of life saving vaccination programs at the global level.

## AUTHOR CONTRIBUTIONS

Both authors contributed equally to this manuscript.

Available from: http://www.who.int/phi/vaccines\_workforce\_Dec2011/en/ (Accessed: March 07, 2018).

**Conflict of Interest Statement:** AP is an employee of the GSK group of companies and is actively involved in several vaccine development programs and vaccinology education initiatives. P-HL has no potential conflicts to disclose.

The reviewer AP declared a shared affiliation, though no other collaboration, with one of the authors AP to the handling Editor.

*Copyright © 2018 Lambert and Podda. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner 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.*

# Changing Priorities in vaccinology: Antibiotic Resistance Moving to the Top

#### *Aldo Tagliabue1 \* and Rino Rappuoli 2*

*1 Institute for Genetic and Biomedical Research, CNR, Cagliari, Italy, 2GSK Vaccines, Siena, Italy*

Antimicrobial resistance (AMR) is currently the most alarming issue for human health. AMR already causes 700,000 deaths/year. It is estimated that 10 million deaths due to AMR will occur every year after 2050. This equals the number of people dying of cancer every year in present times. International institutions such as G20, World Bank, World Health Organization (WHO), UN General Assembly, European Union, and the UK and USA governments are calling for new antibiotics. To underline this emergency, a list of antibiotic-resistant "priority pathogens" has been published by WHO. It contains 12 families of bacteria that represent the greatest danger for human health. Resistance to multiple antibiotics is particularly relevant for the Gram-negative bacteria present in the list. The ability of these bacteria to develop mechanisms to resist treatment could be transmitted with genetic material, allowing other bacteria to become drug resistant. Although the search for new antimicrobial drugs remains a top priority, the pipeline for new antibiotics is not promising, and alternative solutions are needed. A possible answer to AMR is vaccination. In fact, while antibiotic resistance emerges rapidly, vaccines can lead to a much longer lasting control of infections. New technologies, such as the high-throughput cloning of human B cells from convalescent or vaccinated people, allow for finding new protective antigens (Ags) that could not be identified with conventional technologies.  Selected antibodies expressed by recombinant DNA techniques can be used for passive immunization in animal models and tested for protection. Antibodies providing the best protection can be employed to identify new Ags and then used for generating highly specific recombinant Fab fragments. Co-crystallization of Ags bound to Fab fragments will allow us to determine the structure and characteristics of new Ags. This structure-based Ag design will bring to a new generation of vaccines able to target previously elusive infections, thereby offering an effective solution to the problem of AMR.

Keywords: antibiotic resistance, vaccination, reverse vaccinology, human immunology, public health

#### "MICROBES MAKETH MAN"

Modifying the old saying "Manners maketh man" (reported by William Horman in *The Vulgaria* written in 1519), a few years ago the magazine The Economist published in its Leaders section a comment regarding the new vision of the interaction between microbes and man (1). The new saying clearly indicates that microbes have determined in many ways the evolution of the human

#### *Edited by:*

*Lee Mark Wetzler, Boston University, United States*

#### *Reviewed by:*

*Giampiero Pietrocola, University of Pavia, Italy Ali Ellebedy, Washington University in St. Louis, United States*

*\*Correspondence: Aldo Tagliabue<EMAIL_ADDRESS>
#### *Specialty section:*

*This article was submitted to Vaccines and Molecular Therapeutics, a section of the journal Frontiers in Immunology*

*Received: 18 March 2018 Accepted: 30 April 2018 Published: 30 May 2018*

#### *Citation:*

*Tagliabue A and Rappuoli R (2018) Changing Priorities in Vaccinology: Antibiotic Resistance Moving to the Top. Front. Immunol. 9:1068. doi: 10.3389/fimmu.2018.01068*

**27**

species. At a first level, bacteria, viruses, fungi and archaea have literally become a part of us, forming the so called microbiota. A recent study has defined more precisely the number of bacteria present in our body, which is in the order of 39 trillion cells (2). Since the estimated number of human cells in the body (about 84% of which are red blood cells) is in the order of 30 trillion, the ratio between bacterial and human cells is about 1.3. The numbers may vary significantly from person to person and could change significantly with each defecation, ranging from 30 to 50 trillion in each individual. Women may also have a higher ratio of bacterial vs. human cells, because they have fewer red blood cells. This evaluation does not take into account fungi, viruses, and archaea, which all make up the human microbiota and would increase the ratio of microbes to human cells. Thus, we can consider ourselves like superorganisms, in which microbes do many jobs in exchange for the raw materials and the shelter their host provides. This alone shows how closely host and microbiota have co-evolved.

But microbes are also part of a living universe outside us, and often they act as parasites able to regulate the human life span. Over the entireness of the three million years of our species' evolution, life expectancy has always been between 25 and 35 years until very recently, and infections have been the main regulators of our life span. By learning from observation of nature, human beings progressively improved their living conditions to the point that about 250 years ago life expectancy started to increase. In 1900, mankind had already reached a life expectancy of approximately 50 years (3). Nowadays, a child born in a high-income country can expect to live 85 years. The additional 35 years of life that we gained during the last century are substantially due to the conquest of infectious diseases, which used to kill 50% of people before the age of 20. These were viral diseases such as smallpox, rabies, measles, rubella, mumps, and bacterial infections such as diphtheria, tetanus, typhoid fever, and cholera (4). This result has been achieved primarily by improved hygiene, but also by treatment of infectious diseases with antibiotics and by their prevention throughout vaccination. As negative control of this important result, we have to consider the poor areas of our planet, where hygiene, vaccines, and antibiotics are not properly used even today. As a consequence, in these areas infections still represent a major cause of mortality, maintaining life expectancy below 50 years.

#### DISCOVERING A GREAT TOOL AGAINST MICROBES

Antibiotics are an important example of how man can learn from nature to improve his own living conditions. The first observation that microbes are able to produces substances capable of killing pathogens came from the Italian scientist Vincenzio Tiberio in 1895, who showed the antibacterial activity of a natural substance produced by molds (5). Then Ernest Duchesne in France published in his doctorate thesis presented in 1897 (6) that the mold *Penicillium glaucum* possesses antibacterial properties. It was Alexander Flaming in 1928 that succeeded in definitely identifying the world's first antibiotic. It was a substance isolated from the mold *Penicillium notatum*, defined as benzylpenicillin (penicillin G) (7). The first industrial production came however after Howard Florey and Ernst Boris Chain continued the work of Flaming in Oxford. Thus, after the Pearl Harbor attack in 1941 a mass production could be initiated. By 1944, enough penicillin was produced to treat the wounded soldiers in the Allied forces. The 1945 Nobel Prize in Physiology and Medicine was assigned to Flaming, Florey and Chain. The era of antibiotics had begun, and several other molecules produced by microbes followed penicillin. With antibiotics, mankind could claim an historical success in the eternal war against pathogens. However, it was soon clear that antibiotics were not the definitive weapon.

In a book published in 1975, Stanley Falkow wrote that "*we owe to chemotherapy (antibiotics) the debt of reducing the high mortality rate of many bacterial infections*" and to hygiene and vaccines the debt of preventing them, however "*in helping to solve some of the problems of infectious diseases, chemotherapy has created some problems of its own*" (8). The problem created by antibiotics was the generation of bacterial strains resistant to multiple antibiotics, an event reported for the first time in 1956, with the isolation in Japan of a strain of *Shigella flexneri* resistant to streptomycin, tetracycline, chloramphenicol, and sulfonamides.

Today, antimicrobial resistance (AMR) has grown out of proportion, and many pathogenic bacteria are resistant to multiple antibiotics, including *Neisseria gonorrhoeae*, *Staphylococcus aureus*, *Pseudomonas aeruginosa*, *Salmonella typhi*, *Shigella*, *Escherichia coli*, *Acinetobacter*, *Proteus*, *Klebsiella*, *Serratia*, *Streptococcus pneumoniae*, *Mycobacterium tuberculosis*, *Vibrio cholerae*, *Helicobacter pylori*, and others. In a few cases, bacteria became resistant to most of the available antibiotics and are on the verge of becoming untreatable. As a consequence, AMR is perhaps the most alarming emerging problem of infectious diseases. Globally, AMR already causes 700,000 deaths/year, and the forecast is that in 2050 it will cause 10 million deaths/year, higher than the 8.2 million deaths caused by cancer today. As an example we can look at *S. pneumoniae*, also known as pneumococcus, a human pathogen that is the major cause of community-acquired pneumonia, bacterial meningitis, bacteremia, and otitis media (9, 10). In the past, most strains of *S. pneumoniae* were sensitive to penicillin, whereas today penicillin resistance goes from 5 up to 60% in various parts of the world. Thus, with time old antibiotics become less effective or lose efficacy, making the search for new molecules with different mechanisms of action a priority (11).

Alarming documents, calling for action and asking for new antibiotics, have been issued by governments such as those of the UK and USA, by the European Union (EU), and by international organizations such as the World Health Organization (WHO), the United Nations General Assembly, and the World Bank and the G20. The interest in fighting the increase in AMR has intensified, and new incentives for research and development of new drugs have been deployed. In 2016, about 500 million US\$ have been allocated to new and existing initiatives aiming to accelerate the development of new antibiotics1 . For example, the Innovative Medicines Initiative (the biggest public–private program in biomedical science of the EU Commission) funded several projects defined as *New Drugs*

1Boston Consulting Group, Federal Ministry of Health. *Breaking Through the Wall: a Call for Concerted Action on Antibiotics Research and Development* (2017). Available from: http://www.bcg.de/documents/file219507.pdf (Accessed: May 21, 2018).

*for Bad Bugs*2 . Other initiatives include CARB-X, a collaboration between US and UK partners3 , and the Global Antibiotic Research and Development Partnership, a collaboration between the Drugs for Neglected Diseases *initiative* and the WHO4 . It is also interesting a German proposal for a Global Union for Antibiotics Research and Development (GUARD), aimed at funding and coordinating a facility for antibiotics research and development.

On February 27th 2017, the WHO published a document5 , which we partially report hereafter: "*This is the first ever list of antibiotic-resistant "priority pathogens", a catalog of 12 families of bacteria that pose the greatest threat to human health. The list highlights in particular the threat of Gram-negative bacteria that are resistant to multiple antibiotics. These bacteria have built-in abilities to find new ways to resist treatment and can pass along genetic material that allows other bacteria to become multi-drug-resistant. The WHO list is divided into three categories according to the urgency of intervention, of critical, high and medium priority. The most critical group includes multidrug resistant bacteria that pose a particular threat in hospitals, nursing homes and among patients whose care requires devices (such as ventilators and blood catheters). The group encompass Acinetobacter, Pseudomonas and various Enterobacteriaceae (including Klebsiella, E. coli, Serratia, and Proteus). These bacteria can cause severe and often deadly infections such as bloodstream infections and pneumonia, and have become resistant to a large number of antibiotics, including carbapenems and third generation cephalosporins (the best available antibiotics for treating multi-drug resistant bacteria). The second and third tiers in the list—the high and medium priority categories— include other increasingly drugresistant bacteria that cause more common diseases, such as Neisseria gonorrhoeae (the agent of gonorrhea) and Salmonella (causing food poisoning). This WHO action intends to promote initiatives of basic science and advanced R&D from both publicly funded agencies and the private sector, aiming to discover new antibiotics*."

The WHO text continues as follows:

*"Tuberculosis (TB) was not included in the list, although its resistance to traditional treatment has been growing in recent years, because TB is targeted by other dedicated programs. However, we must remember that TB now kills more people than any other pathogen (1.8 million in 2015), and it is therefore a most urgent priority. Other bacteria that are not included in WHO list, such as Group A and group B Streptococcus and Chlamydia, have low levels of resistance to existing treatments and do not currently pose a significant public health threat, but there is a risk that with time also these pathogens may become resistant. The list was developed in collaboration with the Division of Infectious Diseases at the University of Tübingen, Germany, using a multi-criteria decision analysis technique vetted by a group of international experts.*  *The criteria for selecting pathogens in the list were: a) how deadly the infections they cause are; b) whether their treatment requires long hospital stays; c) how frequently they are resistant to existing antibiotics when people in communities catch them; d) how easily they are transmitted between animals, from animals to humans, and from person to person; e) whether they can be prevented (e.g., through good hygiene and vaccination); f) how many treatment options remain; and g) whether new antibiotics to treat them are already in the R&D pipeline."*

The WHO document strongly underlines the need for new treatments. Thus, the search for new antimicrobial drugs is and must remain a great priority. However, it is important to realize that the pipeline for new antibiotics is not very promising, thereby making unlikely that the problem will be solved along this line (12). On the other hand, another tool that, together with antibiotics, contributed to conquer and eliminate many infectious diseases, i.e., vaccines, have a very promising pipeline thanks to the new technologies (3). Thus, vaccines have the possibility to make a big contribution to the control of AMR.

#### RESISTANCE TO ANTIBIOTICS AND VACCINES

The analysis of how vaccines and antibiotics contributed to conquering infectious diseases during the last century was originally published by the group of one of the authors of this paper (13), and more recently re-analyzed in depth by Kennedy and Read [(14), **Figure 1**]. This analysis shows that resistance to antibiotics inevitably emerges every time that a new antibiotic is introduced, starting a process of selection in the target bacteria that will eventually make that antibiotic useless. The consequence is that there is a continuous need of a fresh supply of novel antibiotics, to maintaining effectiveness of the therapeutic treatment. This strategy worked very well up to 1970s, when the identification of new antibiotics was abundant. However, since then the pipeline for new antibiotics has been drying out, and we have not been able to discover new classes of antibiotics (11). In marked contrast, Kennedy and Read show that we can use vaccines for a long time, generating no or very little resistance (14). Thus, vaccines can control infections over a long period of time without becoming obsolete. This occurs because vaccines work prophylactically and prevent the start of infections, while drugs work therapeutically on an ongoing infection in which bacteria proliferate and mutate, allowing the drug to select the resistant variants.

Furthermore, drugs are targeting few metabolic pathways on the pathogens, whereas vaccines induce a protective immune response against multiple antigenic targets. It can be concluded that selection has fewer opportunities to act upon vaccination than with antibiotic treatment. Still, both vaccines and antibiotics are very important in the control of infections. **Table 1** reports the major differences in the mode of action of vaccines and antibiotics and provides information that can guide us to take advantage of their strengths and to minimize their weaknesses. From this comparison, it is evident that antibiotics are the only life-saving tool

2http://www.imi.europa.eu/content/nd4bb (Accessed: May 21, 2018).

3http://www.carb-x.org/about (Accessed: May 21, 2018).

4http://www.dndi.org/wp-content/uploads/2016/03/GARDP\_Briefer\_Document. pdf (Accessed: May 21, 2018).

5http://www.who.int/mediacentre/news/releases/2017/bacteria-antibiotics-needed/ en/ (Accessed: May 21, 2018).

Figure 1 | Time to detection of resistance of human pathogens to antimicrobials (in red) and to vaccines (in green). Black X symbols indicate insurgence of resistance, with lines starting at product introduction (yellow stars; except for smallpox vaccination that began much earlier; with modifications from Ref. (14) with the permission of the publisher).

that we can use during acute bacterial infections, although their often improper or excessive use is causing bacterial resistance in a continuously increasing fashion. The availability of vaccines to control infections may allow us to decrease the use of antibiotics and to generate less AMR. This will permit a more efficient use of existing and new antibiotics during acute infections. As shown in **Figure 1**, there are, however, some cases in which resistance evolved after vaccination. This can be due to several causes, such as the fact that vaccines can protect from disease but may not have the capability to completely prevent pathogen colonization and transmission, as in the case of the acellular pertussis vaccine, or it can be caused by serotype replacement after vaccination with vaccines not including all serotypes, as in the case of the vaccines against *S. pneumoniae*. Thus, even for vaccines the search for better protective antigens (Ag) is very important, particularly for antibiotic-resistant infections.

In May 2016, a group of experts coordinated by the economist Jim O'Neill published a comprehensive report entitled "*Tackling drug-resistant infections globally*" (11). Among many indications to prevent the increasing global problem of AMR, an entire section was devoted to vaccines. Hereafter we report part of the section "*We must reduce the demand for antimicrobial so the*  *current stock of drugs last longer*," and in particular in the point Intervention 6 entitled "*Promote development and use of vaccines and alternatives*":

*"Vaccines can prevent infections and therefore decrease the demand for therapeutic treatments, reducing the use of antimicrobials thereby slowing the rise of drug resistance. Thus, vaccines should be eligible for the same incentives applied for antibiotic development. In particular, it is recommended 1) to use existing vaccines in humans and animals; 2) to renew impetus for early-stage research; 3) to sustain a viable market for vaccines."*

Similarly, the WHO document on priority pathogens stresses that the role of vaccines in the global AMR crisis remains of great importance.

#### PROMOTING THE DISCUSSION ABOUT VACCINES AS A REMEDY TO AMR

Since 2004, the year of 100th anniversary of the foundation of the Serology and Vaccinology Institute Achille Sclavo, a forum for the discussion of the most important issues of the vaccine world takes place every year in Siena, Italy. Each annual meeting aimed to analyze the state-of-the-art of important themes in the field of vaccines and expand the vision for the years to come. The participation of excellent speakers and expert discussants ensured the high quality of the meetings, whose conclusions were published in international journals (15–24). The meetings were organized close to a popular event, the horse race named "Palio di Siena," for which the town is worldwide famous. Thus, those meetings were called the Palio Meetings. More recently, with the acquisition of the vaccine company in Siena by GSK, the venue of the meeting was moved to other locations in USA and Europe, but the traditional name was maintained. The topic of the meetings can vary but the mission is always based on one or more of the following pillars: (1) must define the state-of-the-art of cutting edge topics related to infectious diseases; (2) must advocate science policies to promote progress and improvement in human health; and (3) must be a strategic forum aimed to build new initiatives. **Table 2** reports the topics of the Palio Meetings during the years.

It was therefore almost mandatory that the subject of the 2017 Palio Meeting should be on the growing emergency caused by antibiotics failure, with the title "*Prioritizing vaccines to fight antimicrobial resistance*." This event shortly followed a meeting organized by David Salisbury on 2017 March 29–30 at the Chatham House, in London (25). The London meeting provided a clear consensus that vaccines complement the actions of antibiotics and can contribute to control, reduce, and sometimes eliminate diseases caused by AMR pathogens, more than any other intervention. Thus, the main scope of the 2017 Palio Meeting was to build on the conclusions of the London meeting, and posed the question of how can we make vaccines achieve their full potential and become one of the top tools to tackle AMR. Indeed, the discussion led to conclude that there is a need to make stronger, more evidence-based cases supporting the importance of vaccines in AMR prevention (24).

One example is represented by vaccines against the main strains of *S. pneumoniae*, vaccines that have reduced pneumonia cases in the first decade of this century and in parallel have decreased the number of infections resistant to front-line antibiotics (26). The introduction in 2009 in South Africa of a pneumococcal vaccine achieved an analogous result. Furthermore, it is interesting to note that the high use of antibiotics, prescribed to treat opportunistic bacterial infections in people weakened by flu, is prevented when flu vaccines are employed. There is a

need to make public the data generated by vaccine companies on vaccine effectiveness against AMR, and to continuously monitor the circulation of resistant bacterial strains. The discussion is continuing, and ideas and actions are becoming better defined (27). A global strategic effort to develop a portfolio of vaccines that target AMR is becoming mandatory.

### EVOLUTION IN VACCINE RESEARCH

Why vaccines are becoming an advantageous weapon to curb AMR? This is because their effectiveness in preventing infections has hugely improved, as a consequence of the enormous technological developments of the last two decades. Since the introduction of Jenner's vaccine against smallpox in 1798, the field of vaccines has steadily progressed, but in the last years vaccine development has enormously benefited from the -omics approaches. Thus, new potential vaccine candidates can be discovered in much shorter time than in the past, when vaccines have been developed more empirically (3).

The new techniques of genome sequencing introduced in the late 1990 completely changed the process for discovering novel vaccine Ags. The "*reverse vaccinology*" approach showed that, starting from sequence information, it is possible to discover the protective Ags without handling the microbes (28). A recently licensed vaccine against meningococcus B is the first vaccine produced with reverse vaccinology (29). During the last decade, vaccine design was further potentiated by new technologies, leading to an approach that has been named "*reverse vaccinology 2.0*" (30). As summarized in **Figure 2**, this approach takes advantage of human immunology for designing optimal vaccine Ags.

Ag design"). The new Ag can be developed with the best formulation or delivery system to then be tested in humans (from Ref. (30) with permission of the publisher).

Thanks to better knowledge in handling human B cells and by selecting the most favorable donors, it is now possible to produce highly specific recombinant monoclonal antibodies (mAbs) and also their Ag-binding fragments (Fabs) (31). Further analysis by structural biology approaches brings to 3D studies of the target Ags complexed with the Fabs. It is also possible to discover the protective epitopes capable of inducing broadly neutralizing Abs (32–34). Furthermore, new computational approaches have allowed to obtain completely novel immunogens capable of inducing protection (35).

In viral infections, new structure-based powerful vaccine molecules have been already designed by screening human mAbs from convalescent people and obtaining the molecular structure of Ags and Ag–Ab complexes, as described earlier. Examples are the identification of the pentamer as key Ag for cytomegalovirus (CMV), and of the pre-fusion Ag of respiratory syncytial virus (RSV).

Until recently, the most promising CMV vaccine candidate was a recombinant form of the fusion protein gB. However, the human trial of gB combined with a potent adjuvant showed only moderate efficacy, and therefore the vaccine development was put on hold. Later, isolation of human mAbs from people previously exposed to CMV demonstrated that the most potent CMV neutralizing antibodies were not recognizing gB, but a complex Ag made by five proteins (pentamer). A recombinant form of the pentamer induced neutralizing antibodies that are orders of magnitude more potent than those induced by gB. The new Ag is a very promising candidate for a CMV vaccine and will soon undergo human trials (35).

In the case of RSV, it was possible to obtain a humanized mAb, palivizumab, that binds to an epitope present in the F protein in both the pre-fusion (pre-F) and post-fusion (post-F) conformation. Initial studies to develop an RSV vaccine were mainly focused on the use of the post-F protein that, unlike pre-F, is highly stable both as soluble Ag and when displayed onto virus-like particles. However, experimental vaccines based on whole virus, live attenuated virus, or post-F protein have failed to yield appropriate levels safety or efficacy. The scenario changed when the isolation and characterization of human neutralizing mAbs elicited by natural infection showed that the majority of antibodies are specific for the pre-F form of the protein and failed to cross-react with the post-F conformation. A structure-based design of a stabilized RSV pre-F protein was eventually obtained by complementing the crystal structure of the pre-F protein complexed with a highly neutralizing antibody with the neutralizing studies. The designed pre-F protein (DS-Cav1) could induce neutralizing antibodies 10–15 times more potent than those elicited by previous vaccines and is presently being tested in human trials (36, 37).

We can conclude that interrogation of human antibody responses can allow us to identify pathogen epitopes that are more likely to be protective and that are difficult to discover by conventional technologies. So far, isolation of human mAbs has been successfully used to identify viral Ags that could not be discovered by conventional technologies. On the other hand, there are no data yet regarding the identification of new Ags for antibacterial vaccines. After the proof-of-concept obtained with viral Ags, it would be very important to apply the same approach to the identification of novel bacterial Ags. This will allow us to design innovative vaccines against antibiotic-resistant pathogens, thereby effectively tackling the most pressing global health emergency.

#### CONCLUSION

It is time to consider how to find an effective solution to fight antibiotic resistance, and win this battle in the never-ending war against pathogenic microorganisms. As discussed, the human species has experienced an impressive prolongation of its life expectancy and improvement in life conditions, due to hygiene, antibiotics, and vaccines. Now, one of these pillars has weakened to the point that it will affect some important medical methodologies, first of all important surgeries, but also immunosuppressive chemotherapy and consequently organ transplantation, a great success of the medicine of our era. Again, infectious diseases could severely reduce our life span, as we will not be able to survive important medical treatments or even accidental wounds. A possible solution in sight is that of developing a combined preventive-therapeutic approach, in which vaccines will be one of the two arms and chemotherapy the other one. There are reasons to believe that the combination of the two approaches will result in an overall success.

The main reason is the difference in the mode of action between vaccines and antibiotics. First of all, vaccines on their own are rarely capable to generate resistance. Another critical difference between antibiotics and vaccines is the rate of discovery of new effective molecules. In the past, new antibiotics were identified and regularly reached the clinic, particularly in the three decades after 1950. Since then, however, very few new molecules have been introduced in the clinical use. An opposite situation occurred in the case of vaccines, which have been developed at an increasing speed. As for today, 22 new vaccines became available since 1980. This is a consequence of the introduction of new technologies, such as recombinant DNA, that led to the generation of new synthetic sequences. Therefore, we have obtained a great reduction of the incidence of bacterial meningitis (caused by *Haemophilus influenzae*, *S. pneumoniae*, and *Neisseria meningitidis*) thanks to a new generation of very effective conjugated vaccines, generated by chemical technologies for covalently linking bacterial polysaccharides to proteins. More recently, genomic sequencing opened the access to a higher level in vaccine design, since it made possible to predict the thousands of proteins encoded by bacterial genes, and to identify those likely exposed on the cell surface, in search of new vaccine candidates. This approach, defined as "reverse vaccinology" has resulted in a first important protein vaccine against meningococcus B that is now in use worldwide (29). Finally, a better understanding of the mechanisms regulating the induction of a protective immune response has opened the possibility to introduce, in vaccine formulations, new moieties that can make them more effective. These substances are defined with the general name of adjuvants.

The difference in the mechanisms of action of vaccines and antibiotics is enormous.  What makes them effective is their capability to reach and poison targets across the strong barrier of the bacterial cell wall and avoid being ejected by potent efflux pumps. Any biochemical modification of the target microorganism can make the antibiotic inefficient. Among billions of bacteria present during an infection, such modifications can stochastically arise frequently. Vaccines are molecules with the capacity to evoke in the host a protective activity against infections. They do so by interacting with the immune system of the host, a system that during evolution has developed sophisticated mechanisms to recognize and destroy any kind of "danger" agents, essentially by distinguishing molecules that are different from self. The human immune system can potentially recognize any Ag in the universe, even those never encountered before, thanks to its complex gene rearrangement mechanisms. Thus, it is quite obvious that the potential of vaccines to protect us is extraordinary. And the more we learn about our immune system, the better we can design strategies and develop tools to protect our health from infections. For instance, in some cases it is now possible to cure established infections by administering the patient with specific antibodies produced in the lab with new technologies. In a way, these anti-infective antibodies could be considered as a new kind of antibiotic family, even though much more expensive.

For the time being, the strategy of combing antibiotics and vaccines remains the most sustainable option, which can allow us to avoid in an affordable way the AMR threat. We wish to stress again how serious this threat is, as we expect AMR will cause 10 million deaths/year from 2050.

Before embarking in complex combination studies, it is important to further investigate the role that existing vaccines could have against resistant infections. Indeed, there are some indications that an unconventional use of existing vaccines could provide important advantages. For instance, in New Zealand a new vaccine against meningitis B was introduced few years after the meningitis B outbreak of the end of the 1990. The vaccine was still produced with traditional techniques and was composed by bacterial outer membrane vesicles. Recent studies in the population vaccinated with this vaccine revealed protection against gonorrhea, a sexually transmitted infection induced by *N. gonorrhoeae* that is becoming resistant to antibiotics (38). The reason of this protection is likely due to the fact that bacteria causing meningitis and gonorrhea are genetically related. Furthermore, the current evidence shows that existing pneumococcal vaccines reduce AMR, due to the fact they prevent infection thereby reducing the carriage and transmission of antibiotic-resistant bacteria. Another example is the influenza vaccine that indirectly reduces the incidence of

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fever and sickness, thereby minimizing the use and, more often, the misuse of antibiotics.

The increasing AMR is one of the several alarming signals of the profound effects that human activities can have on our world and life on it. How can we try to solve the problem of infections that are resistant to antibiotics? The most critical point is the difficulty in obtaining new antibiotics. As already discussed, the classical approach does not work anymore, thus we need to devise a completely novel approach.

Passive immunization, i.e., the administration of immune antibodies, could perhaps be a solution, if we can solve the issue of sustainability. Nowadays, immune antibodies can be produced only in low amounts and with high costs. At the beginning of the last century, immune antibodies able to neutralize bacterial toxins were produced in big animals and largely employed, and contributed to building an industrial sector ("serum" institutes) that evolved in today's vaccine industry. We hope that the new technologies will allow us to revive the serology concept and make it a new tool against infection.

A revolutionary approach would be to make the bacteria living within or on us, our microbiota, to become our allies in fighting the infections. As already mentioned, a large component of our body is bacteria (over 50%). An increasing number of studies indicate that gut microbiota is influencing our health and pathological conditions (39). Intestinal microbes can influence host energy metabolism (40), intestinal epithelial proliferation (41), and immune responses (42). It has been shown that the microbiota composition could influence vaginosis (43), obesity (44), inflammatory bowel disease (45), functional bowel disorders (46), allergies (47), and other diseases. An increasing number of studies suggest we can educate our microbiota to combat metabolic and chronic diseases. Could it be also the case in fighting infections?

Several candidate vaccines are in development pipelines since the last few years. For sure the emergency that we are facing will change the health priorities, and vaccines against antibioticresistant bacterial strains will move to the top.

## AUTHOR CONTRIBUTIONS

AT wrote the manuscript. RR critically revised it.

## ACKNOWLEDGMENTS

The authors would like to thank Diana Boraschi (IBP-CNR, Napoli, Italy) for the careful editing of the manuscript.

**Conflict of Interest Statement:** AT declares the lack of any commercial or financial relationships that could be construed as a potential conflict of interest. RR declares to be an employee of GSK Vaccines, Siena, Italy.