Prophylactic Effect of Probiotic Streptococcus Salivarius eK12 Against Recurrent Streptococcus Pyogenes Pharyngotonsillitis Infection in Pediatrics

Study to Investigate the Prophylactic Effect of Streptococcus Salivarius eK12 Against Recurrent Group A Streptococcal Pharyngotonsillitis in Children: A Randomized, Double-Blind, Parallel-Group Controlled Trial

Recurrent Group A Streptococcus (GAS) infections, primarily presenting as strep throat, are a significant health concern in pediatric populations, leading to symptoms like sore throat, fever, and swollen lymph nodes. GAS is highly contagious and can spread easily among children in close-contact environments like schools and daycare centers, often resulting in frequent re-infections. Managing these recurrent infections typically requires antibiotics, but reliance on antibiotics carries risks, including resistance development, gut microbiota disruption, and various side effects. Probiotic therapy, particularly with Streptococcus salivarius K12, has shown promise as a preventive approach, utilizing competitive exclusion and antimicrobial production to inhibit GAS growth in the oral cavity. Recent studies have advanced this with the development of S. salivarius eK12, a re-engineered strain that enhances efficacy against GAS by preventing the survival-promoting interactions between the probiotic and pathogenic bacteria. This modified strain, now registered as Bactoblis® EVOL in Italy (EU), holds potential as an effective version to reduce the incidence and severity of GAS infections without the drawbacks of antibiotic therapy.

Study Overview

• Status: Completed
• Conditions: Streptococcus Pyogenes Pharyngotonsillitis
• Intervention / Treatment: Dietary supplement: Probiotic Bactoblis® EVOL (Streptococcus salivarius eK12); Dietary supplement: Oral vit D3

Detailed Description

The primary aim of this clinical trial is to evaluate whether prophylactic use of S. salivarius eK12 can effectively reduce the frequency of strep throat episodes in children. By assessing the efficacy of S. salivarius eK12 supplementation, this study seeks to provide insights into its potential as a non-antibiotic preventive strategy for recurrent strep throat infections in pediatric populations.

• Study Type: Interventional
• Enrollment (Actual): 82
• Phase: Not Applicable

Contacts and Locations

Study Locations

• Pakistan
  • Balouchistan
    • Quetta, Balouchistan, Pakistan, 87300
      • Saboor Clinic
  • Khyber Pakhtunkhwa
    • Haripur, Khyber Pakhtunkhwa, Pakistan, 22620
      • Dr. Khan Afsar ENT Clinic
  • Sindh
    • Hyderābād, Sindh, Pakistan, 71000
      • CDF Hospital (Bilawal Medical College, LUMHS)
    • Hyderābād, Sindh, Pakistan, 71500
      • Dr. Allah Wadhayo Kallo Clinic (Shifa Hurmat Diagnostic and Research Centre)

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Child
• Accepts Healthy Volunteers: Yes

Description

Inclusion Criteria:

• Children aged 3-11 years.
• Recent history of documented recurrent episodes of Streptococcus pyogenes pharyngo- tonsillitis infection in the last 6 months, or 4 or more episodes in the last 12 months (S.

pyogenes infection being defined as: McIsaac score with clinical score ≥ 2 plus confirmation of S. pyogenes presence with rapid throat swab (RAD) method or McIsaac score =5.

• Willingness to comply with the study protocol and attend scheduled follow-up visits.
• Written informed consent obtained from the parent or legal guardian.

Exclusion Criteria:

• Children with known allergies or hypersensitivity to probiotics or any components of the study product.
• Current use of antibiotics or within the past 30 days.
• Used oral colonizing probiotics in the last 6 months.
• Presence of severe underlying medical conditions affecting the immune system or gastrointestinal tract.
• Participation in other clinical trials involving investigational drugs or interventions.
• Inability to adhere to the study requirements due to logistical or social reasons.
• History of rheumatic disorders, bronchospasm, severe asthma, or allergy requiring corticosteroids.
• Past tonsillectomy or an indication for adeno-tonsillectomy; or severe respiratory or systemic disorders
• Individuals immunocompromised or had a condition favouring recurrent Acute Otitis Media (AOM), including severe atopy, acquired or congenital immunodeficiency, cleft palate, craniofacial abnormalities, obstructive adenoids, a chronically ruptured eardrum, sleep apnoea, or tympanostomy tubes
• Children undergoing current pharmacological therapies to prevent recurrent respiratory infections.
• Any other medical condition or circumstance that, in the investigator's judgment, would compromise the safety of the participant or the integrity of the study.

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Prevention
• Allocation: Randomized
• Interventional Model: Parallel Assignment
• Masking: Quadruple

Number of Arms

2

Arms and Interventions

Participant Group / Arm	Intervention / Treatment
Experimental: Probiotic supplement group; Participants in this group will receive probiotic Streptococcus salivarius eK12 (Bactoblis® EVOL containing ca. 1 billion CFU of Streptococcus salivarius eK12/capsule) as daily one orally dissolving tablet (to be slowly dissolved in the mouth before sleeping at night time), plus 200 IU vit D3 (one tablet) for 3-months.	Dietary supplement: Probiotic Bactoblis® EVOL (Streptococcus salivarius eK12); Bactoblis® EVOL: containing one billion CFU of probiotic Streptococcus salivarius eK12; Dietary supplement: Oral vit D3; Oral vit D3 tablets (200 IU)
Active Comparator: Control group; Participants in this group will receive only 200 IU vit D3 as daily two orally dissolving tablets under the same dosing regimen.	Dietary supplement: Oral vit D3; Oral vit D3 tablets (200 IU)

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Recurrence of Streptococcus pyogenes pharyngotonsillitis infection	Number of episodes of Streptococcus pyogenes pharyngotonsillitis infection	6-months

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Protection against viral infection	Number of incidences of viral throat infection	6-months
Protection against Acute Otitis Media infection	Number of incidences of Acute Otitis Media infection	6-months
Protection against respiratory infection	Number of incidences of Tracheitis infection	6-months
Protection against Rhinitis	Number of incidences of Rhinitis	6-months
Protection against flu	Number of incidences of flu	6-months
Protection against Enteritis	Number of incidences of Enteritis infection	6-months
Protection against Stomatitis	Number of incidences of Stomatitis infection	6-months
Severity of Group A Streptococcal (GAS) Symptoms	Severity of GAS-associated symptoms assessed using a 10-point Visual Analogue Scale (VAS)	6-months
Use of Antibiotics	Number of antibiotic courses prescribed during the 6-month study period.	6-months
Persistence of Streptococcus salivarius eK12 in saliva after washout	Post hoc exploratory ancillary analysis, conceived after study completion and unblinding, evaluating the presence of the administered Streptococcus salivarius eK12 strain in saliva at the end of the 6-month study period, approximately 3 months after discontinuation of eK12 supplementation. Saliva samples will be examined using a strain-specific microbiological or molecular method capable of distinguishing eK12 from the parental K12 strain. The outcome will be reported as the proportion of participants with detectable eK12 and, if a validated quantitative method is used, the salivary abundance of eK12. This analysis was not a prespecified outcome of the original clinical trial.	Month 6, approximately 3 months after discontinuation of eK12 supplementation.

Collaborators and Investigators

• Sponsor: Liaquat University of Medical & Health Sciences
• Collaborators: Università degli Studi dell'Insubria

Publications and helpful links

General Publications

• Di Pierro F, Colombo M, Giuliani MG, Danza ML, Basile I, Bollani T, Conti AM, Zanvit A, Rottoli AS. Effect of administration of Streptococcus salivarius K12 on the occurrence of streptococcal pharyngo-tonsillitis, scarlet fever and acute otitis media in 3 years old children. Eur Rev Med Pharmacol Sci. 2016 Nov;20(21):4601-4606.
• Di Pierro F, Colombo M, Zanvit A, Rottoli AS. Positive clinical outcomes derived from using Streptococcus salivarius K12 to prevent streptococcal pharyngotonsillitis in children: a pilot investigation. Drug Healthc Patient Saf. 2016 Nov 21;8:77-81. doi: 10.2147/DHPS.S117214. eCollection 2016.
• Di Pierro F, Colombo M, Zanvit A, Risso P, Rottoli AS. Use of Streptococcus salivarius K12 in the prevention of streptococcal and viral pharyngotonsillitis in children. Drug Healthc Patient Saf. 2014 Feb 13;6:15-20. doi: 10.2147/DHPS.S59665. eCollection 2014.
• Di Pierro F, Donato G, Fomia F, Adami T, Careddu D, Cassandro C, Albera R. Preliminary pediatric clinical evaluation of the oral probiotic Streptococcus salivarius K12 in preventing recurrent pharyngitis and/or tonsillitis caused by Streptococcus pyogenes and recurrent acute otitis media. Int J Gen Med. 2012;5:991-7. doi: 10.2147/IJGM.S38859. Epub 2012 Nov 30.
• Di Pierro F, Veeraraghavan G, Kalaiselvan K, Ashif B, Jeyakumar L, Gopalakrishnan G, Cazzaniga M, Bertuccioli A, Tanda ML, Zerbinati N, Ujjan I, Bugti AA, Bano A, Gull Y, Mumtaz N, Khan A. In Vitro and In Vivo Safety Evaluation of Streptococcus salivarius eK12, a Genetically Modified Dietary Probiotic Derived from the Oral Probiotic S. salivarius K12. J Microbiol Biotechnol. 2025 Dec 9;35:e2509016. doi: 10.4014/jmb.2509.09016.
• Di Pierro F, Bugti AA, Bano A, Kara M, Ujjan I, Mumtaz N, Gull Y, Cazzaniga M, Bertuccioli A, Tanda ML, Zerbinati N, Hameed S, Khan A. Study to evaluate the safety and tolerability of Streptococcus salivarius eK12, a genetically modified strain derived from the oral probiotic S. salivarius K12: Results from a randomized, double-blind, placebo-controlled, parallel-group clinical trial. Front Nutr. 2025 Dec 8;12:1701611. doi: 10.3389/fnut.2025.1701611. eCollection 2025.

Helpful Links

• Related Info

Study record dates

Study Major Dates

• Study Start (Actual): December 1, 2025
• Primary Completion (Actual): July 31, 2026
• Study Completion (Actual): July 31, 2026

Study Registration Dates

• First Submitted: April 13, 2024
• First Submitted That Met QC Criteria: April 13, 2024
• First Posted (Actual): April 17, 2024

Study Record Updates

• Last Update Posted (Actual): August 21, 2026
• Last Update Submitted That Met QC Criteria: August 20, 2026
• Last Verified: August 1, 2026

More Information

Terms related to this study

• Other Study ID Numbers: LUMHS/REC/-496/11.11.2024

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: YES
• IPD Plan Description: Deidentified individual participant data underlying the results reported in the published article, together with the corresponding data dictionary.
• IPD Sharing Time Frame: Beginning 3 months and ending 5 years after publication of the article.
• IPD Sharing Access Criteria: Data will be made available to researchers whose methodologically sound proposals are approved by the study investigators, for the purposes specified in the approved proposal and after execution of a data-use agreement. Requests should be directed to the corresponding author at pchiarmaria@gmail.com.
• IPD Sharing Supporting Information Type: SAP

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No