Revisions for Safety evaluation of Bacillus clausii AO1125 for its use as a probiotic in humans | Registro Público Cubano de Ensayos Clínicos

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Safety evaluation of Bacillus clausii AO1125 for its use as a probiotic in humans

General information

Section to complete general information about the trial: scientific and public title, protocol identifiers, sponsors and Source(s) of Monetary or Material Support.

Scientific title:

Safety evaluation of Bacillus clausii AO1125 for its use as a probiotic in humans”.

Secondary indentifying numbers:

Not applicable

Issuing authority of the secondary identifying numbers:

Not applicable

Source(s) of monetary or material support:

Biocollective Company, Denver Colorado USA

Authorization for beginning

Section to complete information about the regulatory approval of clinical trial: regulatory agency name, approval date and reference number in the agency.

Regulatory instance to authorize the initiation of the study:

Only approved by Ethics Committees

Principal investigator

Section to complete information about Email address, telephone number and postal address of the Principal Investigator.

First name:

Gissel

Last name:

García Menéndez

Medical Specialty :

Graduate in Biochemistry, Master in Virology, PhD in Sciences

Affiliation:

Hermanos Ameijeiras Clinical Surgical Hospital

Postal address:

San Lazaro 701 corner to Belascoain

City:

Havana

Country:

Cuba

Zip Code:

10300

Telephone:

+53-78761632

Email address:

gisselgarcia@infomed.sld.cu

gisselgarcia2805@gmail.com

Clinical sites to participate

Section to complete the data related to the clinical sites involved in the trial: site and responsible investigator for every site.

Countries of recruitment:

Cuba

Clinical sites:

Not applicable

Recruitment status

Section to complete information about the recruitment status and the date of first enrolment subject

Recruitment status:

Complete

Date of first enrollment:

05/09/2023

Health condition and Intervention

Section to complete information about the primary medical condition(s) or problem(s) studied and, a characteristics of the intervention(s).

Health condition(s) or Problem(s) studied:

healthy volunteers

Health condition(s) code:

Healthy Volunteers

Healthy Aging

Intervention(s):

Group I (Experimental) One capsule of Bacillus clausii AO1125 (2.5 x109 bacteria / capsule mixed with the following excipients: calcium phosphate, magnesium stearate and colloidal silica), orally, fasting for 2 months . Group II (Control). Placebo; same appearance and application scheme as the Experimental product.

Intervention code:

Probiotics

Dietary Supplements

Placebos

Intervention keyword:

Bacillus clausii AO1125

Outcomes and Timepoint

Section to complete information about primary and secondary outcomes including. It includes the metric or method of measurement used and, the time point for every outcome.

Primary outcome(s):

1.- Adverse effect. It will be measured taking into account: the presence of the adverse effect: (It does not appear to be an adverse effect, Possible adverse effect, Similar to an adverse effect, Adverse effect). Measurement time: at the end of the study, day 60. 2.- Intensity of the Adverse Effect. (Mild (Does not produce restrictions in daily activities), Moderate (Produces a partial restriction of daily activities), Severe (Inability to carry out daily activities)). Measurement time: at the end of the study, day 60.

Key secondary outcomes:

Hematological variables, clinical chemistry, hemodynamic parameters and calculation of muscle mass Clinical Chemistry: -Liver enzymes (UKAT/L) -creatinine (umolL/L) -Urea (mmol/L) Hematological parameters Blood glucose (mmol/L), Leukocytes (x 109/L) Red blood cells (x 1012/L) Hemoglobin (g/dL) Hematocrit (%) MCV (fL) MCHC (g/dL) RDW (%) platelets (x 109/L) MPV (fL) Neutrophils (%) Lymphocytes (%) Monocytes (%) Eosinophils (%) Basophils (%) Neutrophils (x 109/L) Lymphocytes (x 109/L) Monocytes (x 109/L) Eosinpophils (x 109/L) Basophils (x 109/L) Hemodynamic Parameters Systolic blood pressure (mmHg). Diastolic blood pressure (mmHg) Heart rate (bpm) Calculation of muscle mass Muscle mass (MM) will be calculated for men and women using the following formulas using height and weight. For Men: MM=(0.32810*Weight)+(0.33929*height)-29.5336 For Women: MM=(0.29569*weight)+(0.41813*height)-43.2933 Measurement time: All the variables will be measured at the beginning, day 0; and at the end of the study, day 60.

Selection criterias

Section to complete information about the inclusion and exclusion criteria for participant selection, including age and gender.

Gender:

Male/Female

Minimum age:

19 years

Maximum age:

none

Inclusion criteria:

1.- Healthy individuals, over 18 years of age and will have no distinction of sex or skin color. 2.- They give their consent to participate in the study. 3.- Individuals who have hematological, clinical chemistry and hemodynamic parameters in normal ranges

Exclusion criteria:

Individuals suffering from any comorbidity. and who show alterations in their hematological, clinical or hemodynamic parameters.

Type of population:

Adults

Type of participant:

Healthy volunteers

Study design

Section to complete information about the characteristics of the study design.

Type study:

Interventional

Purpose:

Other

Other purpose:

This is a study that will evaluate the probiotic strain Bacillus clausii AO1125 in healthy individuals, which has anti-inflammatory properties, so its use will have an impact on intestinal health as well as on improving the immunity of individuals.

Allocation:

Randomized controlled trial

Masking:

Double Blind

Control group:

Placebo

Study design:

Parallel

Phase:

1

Target sample size:

100

Contact for public queries

Section to complete information about Email address, telephone number and postal address of the contact who will respond to general queries, including information about current recruitment status

First Name:

Gissel

Last Name:

Garcia Menendez

Specialty:

Graduate in Biochemistry, Master in Virology, PhD in Sciences

Affiliation:

Hermanos Ameijeiras Clinical Surgical Hospital

Postal Address:

San Lazaro 701 corner to Belascoain

City:

Havana

Country:

Cuba

Zip Code:

10300

Telephone:

+53-78761632

Email :

gisselgarcia@infomed.sld.cu

gisselgarcia2805@gmail.com

Contact for scientific queries

Section to complete information about Email address, telephone number and postal address of the contact who will respond to scientific queries.

First Name:

Gissel

Last Name:

Garcia Menendez

Specialty:

Graduate in Biochemistry, Master in Virology, PhD in Sciences

Affiliation:

Hermanos Ameijeiras Clinical Surgical Hospital

Postal Address:

San Lazaro 701 corner to Belascoain

City:

Havana

Country:

Cuba

Zip Code:

10300

Telephone:

+53-78761632

Email :

gisselgarcia@infomed.sld.cu

gisselgarcia2805@gmail.com

Research Ethics Committees

Section to complete the data related to the review ethics committees.

Name of Research Ethics Committees:

Hermanos Ameijieras Clinical Surgery Hospital

Status of evaluation:

Approved

Status of evaluation date of Ethic Committee:

15/02/2023

Postal address of Ethic Committee :

San Lazaro 701 corner to Belascoain, Havana, ZC 10300, Cuba

Telephone:

+53-78761000

Email:

hha@infomed.sld.cu

About study completion

Section to complete the data related to the study completion.

Final enrolment number:

100

Study completion date:

10/01/2024

Results Study

Section to complete the data related to the summarized results.

Participant flow:

Participants were randomly assigned to either the A. clausii AO1125 group or the pla- group, with 50 individuals allocated to each group. The sample size calculation was based on an expected adverse event rate of 20.0% in the AO1125 group and 0.5% in the placebo group, with a Type I error rate of 5% (alpha = 0.05) and a Type II error rate of 20% (80% power). A 10% anticipated dropout rate was also incorporated into the calculation, resulting in a minimum requirement of 41 participants per group. However, it was determined to recruit 50 participants per group, providing a buffer to accommo-date any unanticipated withdrawals. Randomization was conducted using the EpiData 3.1 software , which ensured unbiased allocation to treatment and placebo groups. Blinding was strictly maintained by dispensing the capsules in identical packaging, preventing both participants and study personnel from distinguishing between the active treatment and the placebo. Following randomization and baseline screening, the final cohort comprised 50 par-ticipants in the AO1125 group and 49 in the placebo group, reflecting one exclusion post-randomization

Baseline characteristics:

Demographic Variables A. clausii AO1125 Cohort (n = 50) Placebo Cohort (n = 49 ) p Value Number % Number % Sex Female 29 58 31 63.3 0.896 a Male 21 42 18 36.7 Age (years) Median ± SD 50.06 ± 12.87 44.43 ± 14.74 0.185 b BMI Median ± SD 24.01 ± 3.65 23.63 ± 4.17 0.44 b

Outcome measures:

Primary Outcome:The study involved a total of 99 individuals, 49 in the placebo group and 50 in the A. clausii group. Most individuals, 83.8% (n = 83), did not show any adverse effects (AE). Secondary Outcome: To determine the averages of each chemical variable, the Student’s t-test for inde-pendent samples was employed. On the baseline day, the following variables were sta-tistically significant in both study groups: Total protein (p < 0.01), albumin (p = 0.008), Cholesterol (p = 0.042), Bilirubin D (p = 0.01), MHC (p = 0.03), and eosinophils (Eo) (p = 0.036). Additionally, MPV showed statistical significance only on day 60 (p = 0.034). Sig-nificant differences between the Alkalihalobacillus clausii group and the placebo group between baseline and day 60 were confirmed solely for GGT, both at baseline (p = 0.02) and on day 60 (p = 0.03). Student’s t-test for paired samples was applied to the A. clausii group to evaluate changes between baseline and day 60. The following parameters decreased significantly: Glycemia (p < 0.01), Creatinine (p < 0.01), and Urea (p = 0.016). However, there were sig-nificant increases in ALAT (p = 0.02) and ASAT (p < 0.01). A similar analysis in the placebo group revealed significant increases in Urea (p = 0.012), GGT (p = 0.026), Hematocrit (HTC) (p = 0.001), and Mean Corpuscular Volume (MCV) (p < 0.01). Significant reductions were observed in ALAT (p = 0.002), ASAT (p < 0.01), Cholesterol (p < 0.01), Total Bilirubin (p = 0.002), and Direct Bilirubin (p = 0.001). Bioimpedance was measured using weight and Body Mass Index (BMI). The t-test for both independent and paired samples did not reveal significant differences between the study groups in these measures The survey questions related to the physical and emotional health status of the study subjects were analyzed (SF-36, Questions 1, 6, 7, and 11). In both groups, the be-havior of the answers to these questions was very similar. Both the placebo group and the treatment group maintained similar responses at both times (Kappa~1, p = 0.000). The significant p values represent that the agreement between the responses of the two moments is high in both evaluated groups

Adverse events:

The study involved a total of 99 individuals, 49 in the placebo group and 50 in the A. clausii group. Most individuals, 83.8% (n = 83), did not show any adverse effects (AE) .Of these, 74% belonged to the group treated with the probiotic (n = 37) and 93.9% to the placebo group (n = 46). Significant differences were observed between the treated group and placebo according to the development of AE related to participants in the study (p = 0.007).Among the participants who received the A. clausii probiotic capsule, 13 individuals (26%) experienced mild adverse events (AEs). The reported adverse events included gastrointestinal pain or discomfort (n = 3), gas (n = 10), headache (n = 1), constipation (n = 3), and diarrhea (n = 1). The study cohort consisted of 60 females and 39 males. Adverse events were ob-served in 8 females (8/60) and 8 males (8/39). A moderate-intensity adverse event was reported in 1 female from the placebo group (7%), who developed edema, which coin-cided with a diagnosis of dengue infection. This moderate AE was likely associated with the arbovirus rather than the intervention. None of the participants reported limitations in their daily activities, which is consistent with the mild intensity of the events. The adverse effects diminished over the course of the treatment

Summary study:

Abstract: (1) Background: Alkalihalobacillus clausii AO1125 is a Gram-positive, motile, spore-forming bacterium with potential as a probiotic due to its broad-spectrum antimicrobial ac-tivity, inhibiting pathogens like Listeria monocytogenes, Staphylococcus aureus, and Clostridium difficile, as well as anti-rotavirus activity. Its resilience in gastrointestinal conditions suggests benefits for gut health. This study evaluates the safety and probiotic potential of A. clausii AO1125. (2) Meth-ods: Genome annotation identified genes linked to probiotic traits such as stress resistance, gut colonization, immune modulation, and antimicrobial production. The genome was screened for antibiotic resistance genes using CARD, bacteriocin clusters using BAGEL4, and virulence factors via VFDB. Cytotoxicity was assessed on Vero cells and erythrocytes, and a Phase I, double-blind, placebo-controlled clinical trial was conducted with 99 healthy volunteers (50 AO1125, 49 placebo). (3) Results: Genomic analysis confirmed minimal antibiotic resistance genes and the absence of virulence factors, supporting safety. A. clausii AO1125 showed no pathogenicity, cytotoxicity, or hemolytic activity and was well-tolerated in clinical settings, with mild, transient abdominal gas as the most common adverse event. (4) Conclusions: The safety profile and genetic basis for probiotic and antimicrobial properties support A. clausii AO1125 as a promising probiotic candidate for gastrointestinal health, warranting further clinical research.

Registration and Update

Section to complete information about the name of Primary Registry, date of registration and the unique ID number assigned by the registry (RPCEC).

Primary registry:

RPCEC

Unique ID number:

RPCEC00000429

Date of Registration in Primary Registry:

07/08/2023

Record Verification Date:

2025/12/12

Next update date:

2026/12/12

Link to the spanish version:

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Safety evaluation of Bacillus clausii AO1125 for its use as a probiotic in humans

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