The Efficacy of Three Doses of Live Attenuated, Oral Rotavirus Vaccine 116E

Prior to study initiation, study clinics and offices will be set up in each of the enrollment sites. Awareness about the study will be created in the community. All categories of staff will be recruited and trained. CRFs, SOPs and registers will be developed. Regulatory and IRB approvals will be obtained. Permissions and clearances will be sought from the Government.

The study team will identify pregnant women, newborns and infants aged less than 8 weeks as potential participants. Information about the study will be shared with these families, and those willing to participate will be enrolled after obtaining consent and subsequent screening of the subject. Subjects will be enrolled in the study since 2 months + 29 days of age and will be given 3 doses of the Test Article/placebo along with childhood vaccines (which will include vaccines against diphtheria, pertussis, tetanus, Haemophilus influenzae B, Hepatitis B and IPV, and pneumococcal conjugated vaccine at 2 and 4 month of age). Participants vaccinated with a dose of vaccine from the National Immunization Program corresponding to their current age, in which more than 48 hours have passed since the last dose received, the subject will not be considered for the study. The Test Article will be liquid ORV 116E, 105.0ffu stored at 2-8°C. The Test Article/placebo will be co-administered with the childhood vaccines that are scheduled at the regular National Program of Immunization vaccination visits around 2 months, 4 months and 6 months of age. After each visit, a safety evaluation will be carried out on all participants. A card will be given to the caregivers for the daily record of temperature, diarrhea, vomiting, food refusal and/or irritability, for a period of 1 week. On day 3 and 7, the research team will contact the caregivers by telephone to note the general state of health of the infant and to collect the card. The subjects will be followed f or episodes of acute gastroenteritis every two weeks through contact by email or text message (+ 2 days), until 8 months + up to 14 days post 3 doses of vaccination. The expected duration of subjec participation is 13 months. In addition to the contacts at every two weeks follow-up, parents/primary caregiver will be advised to contact the study team whenever the subject has symptoms of gastroenteritis (GE; the operational definition of GE for this study will be the change in the habitual pattern of the stools, both in more frequent passage than is normal for the individual and/or in the decrease in stool consistency), signs or symptoms of suspected intussusception (IS, defined as stomach pain and vomiting that start suddenly and come and go several times per hour, and then later, bloody stools) or any other illness requiring hospital referral. If the caregiver report symptoms of GE, an "illness visit" will be activated. The caregiver will be asked to collect a stool specimen. The field worker will ensure that the parents/primary caregiver have stool containers available with them so that stool specimens can be collected whenever the enrolled subject has GE. In the next 24-48 hours, the stool sample will be processed with an ELISA rotavirus test. If the result is negative, the illness visit will be discontinued, and the episode will be registered. If the result is positive, a daily telephone follow-up will be carried out throughout the GE episode, up to 24 hours after the last altered stool. If during the illness visit any symptom/sign of dehydration and/or clinical severity is verified by the research team, the participant will be invited to be evaluated at the study center. During the illness visit the following variables will be collected daily by the caregiver in an illness visit card: stools number and consistency, vomiting episodes, temperature (based on subjective perception of the caregiver of potential fever, or at the same time point during the consecutive days, preferably between 2 and 8 PM), administration of ORS yes/no, number of administrations, and if possible, volume administered). If the child is hospitalized, the following information will be obtained from hospitalization charts: number and consistency of stool episodes, number of vomiting episodes, highest daily temperature, hydration evaluation, use of ORS and/or IV solutions (volume used and fluid description) duration of hospitalization, and any potential complication. If in the follow-up every two weeks of the participants, an episode that did not activate the illness visit of GE is reported or detected, the episode will be recorded and an attemp t will be made to collect as much data as possible retrospectively.

The study team will contact the parents/primary caregiver through mobile phone and collect the information of the illness visit card.

Out of the enrolled subjects, 150 from each group will constitute the "Immunogenicity Subset".

Visit 1 Baseline (vaccine at 2 Months of age + 29 days):

• The participant will be screened for eligibility based on vitals (heart rate and body temperature) and physical examination.
• If the participant is eligible (in good general health or stable preexisting disease as per the discretion of the Principal investigator).
• A study vaccine/placebo will be administered. Following vaccination, participants will remain at the study site for at least 30minutes of observation to record any immediate adverse event.
• Daily card will be given to the parent/primary caregiver.
• Blood sample (3 mL) will be collected for immunogenicity analysis in a subset of (n=300) participants prior to vaccination.

Visit 2 (vaccine at 4 Months of age, +29 days):

• Study participants will return to the OPD for vitals and physical examination (general and systemic examination). A study vaccine

/placebo will be administered.

• Following vaccination, participants will remain at the study site for at least 30minutes of observation to record any adverse event.
• Daily card will be collected and a new card will be given to the parent/primary caregiver.

Visit 3 (vaccine at approximately 6 Months of age, +29 days):

• Study participants will return to the OPD for vitals and physical examination (general and systemic examination). A study vaccine

/placebo will be administered.

• Following vaccination, participants will remain at the study site for at least 30minutes of observation to record any adverse event.
• Diary card will be collected and a new card will be given to the parent/primary caregiver. Visit 4 (1 Month after visit 3 ++5days): (only for immunogenicity group)
• Study participants (Subset) will return to the OPD for physical examination (general and systemic examination) and diary card will be collected.
• Blood sample (3 mL) will be collected for immunogenicity analysis in a subset of n=300.
• Diary card (Photograph) will be collected from the non-immunogenic participants. Blood samples: In the "Immunogenicity Subset", 3 ml blood specimens will be collected at baseline and 28 (+) 5 days after the third dose of the Test Article/placebo to assess the anti-rotavirus IgA antibody titer. Stool Specimens: The study team will attempt to collect a stool specimen for every GE episode, preferably within the first 2 days after episode onset . The stool specimen may be collected up to 7 days after the last day of diarrhea. The stool specimens will be sent to the laboratory. If the episode is a suspected intussusception or vaccine associated gastroenteritis that has occurred within four weeks after each dose of the Test Article/placebo, the specimen will be sent to the laboratory immediately for rotavirus testing and typing. For all ELISA RV positive samples, an aliquot of the stool specimen will be assessed for 22 enteropathogens by multiplex-PCR panel and to identify the genotype of the virus.

Inclusion criteria

• At least one parent(s) or legally acceptable representative's consent for participation and are able to understand study procedures
• Subjects aged at least 2 months at recruitment
• No plans to move in the next 12 months

Exclusion criteria

• Administration of rotavirus vaccine in the past
• Participants vaccinated with a dose of vaccine from the National Immunization Program corresponding to their current age, in which more than 48 hours have passed since the last dose received
• Presence of any illness requiring hospital referral (temporary exclusion)
• Known case of immunodeficiency disease, known HIV positive
• Known case of chronic gastroenteritis disease, chronic pulmonary disease, chronic renal disease, congenital heart disease (Stable with no on-going medication).
• Any other conditions which in the judgment of the investigator warrant exclusion (e.g. no exclusion criteria but seems 'ill', investigators suspects neglect)
• Diarrhea on the day of enrollment (temporary exclusion)
• A known sensitivity or allergy to any components of the study vaccines.
• Major congenital or genetic defect.
• Has received any immunoglobulin therapy and/or blood products since birth.
• History of chronic administration (defined as more than 14 days) of immunosuppressants including corticosteroids. Infants on inhaled or topical steroids may be permitted to participate in the study, at the discretion of the principal investigator.