Effects of Nasal-spraying LiveSpo Navax in Treatment of Influenza Virus in Children
Condition: Acute Respiratory Tract Infections
MeSH terms: Infections; Influenza, Human; Respiratory Tract Infections
Study type: Interventional

Summary: 
      Acute Respiratory Tract Infections (ARTIs) in children are common diseases, with influenza
      virus types A and B is one of the main causes of serious symptoms in young children. Although
      an influenza vaccine is available, influenza vaccination requires annual injections, which
      made it difficult for young children to get effectively immunized. Moreover, antiviral
      nucleotide drugs including Tamiflu (oseltamivir) are unsafe for young children and are
      recommended only for high-risk patients. Probiotics have emerged as promising safe candidates
      for supportive treatment of ARTIs and reduction of antibiotic dependence in recent years.
      Here, investigators propose that direct spraying of probiotics into the nose can be a fast
      and effective symptomatic treatment for ARTIs due to influenza virus.

      The aim of the study about to evaluate the effectiveness of nasal-spraying probiotics
      containing spores of two bacterial strains, Bacillus subtilis and Bacillus clausii in
      preventing and supporting the treatment of children having acute respiratory symptoms due to
      influenza infection.

      Study Population: sample size is 70. Description of Sites: the study is carried out at
      Vietnam National Children's Hospital.

      Description of Study Intervention: totally 70 eligible patients are divided randomly into 2
      groups (n = 35/group each): Patients in Control group received the routine treatment and
      three times per day 0.9% NaCl physiological saline while the patients in the Navax group
      received three times per day LiveSpo Navax in addition to the same standard of care
      treatment. The standard treatment regimen is 2-5 days but can be extended further depending
      on the severity of the patient's respiratory failure.

      Study duration: 15 months
    

Detailed description: 
      Influenza virus types A and B is the common virus that causes Acute Respiratory Tract
      Infections (ARTIs) with a high risk of bronchiolitis for young children and infants.
      Influenza-infection symptoms range from mild fever, cough, runny nose, and wheezing to severe
      symptoms such as fast pulse, fast breathing, and respiratory failure. In a systematic review
      and meta-analysis of the global burden of seasonal influenza respiratory infections in young
      children, they estimated that 28.000 - 111.500 children under the age of 5 die as a result of
      influenza infections in 2008, of which 99% of these deaths occurred in developing countries.
      Influenza vaccination requires annual injections, which made it difficult for young children
      to get effectively immunized. Moreover, antiviral nucleotide drugs including Tamiflu
      (oseltamivir) are unsafe for young children and are recommended only for high-risk patients.
      Probiotics have emerged as promising safe candidates for supportive treatment of ARTIs and
      reduction of antibiotic dependence in recent years. However, the use of oral administrative
      probiotics as functional foods is effective only for mild symptoms and not applicable for
      Acute RTIs (ARTIs). Here, investigators propose that direct spraying of probiotics into the
      nose can be a fast and effective symptomatic treatment for ARTIs.

      The objective was to investigate the symptomatic treatment effects of probiotic product
      LiveSpo Navax, as a liquid-suspension form containing Bacillus spores of safe B. subtilis
      ANA4 and B. clausii ANA39 strains, in children having acute respiratory diseases caused by
      influenza virus, investigators evaluation of improved efficacy and reduced treatment time of
      LiveSpo Navax in influenza-infected children; and measurement of changes in influenza viral
      load, co-infectious bacterial concentrations, and major cytokine indicators in the
      nasopharyngeal samples before and after 2 days using LiveSpo Navax.

      Methods: A randomized, blind, and controlled clinical trial is conducted. The patient's
      parents are required to provide the following information about their children: full name,
      sex, age, obstetric history, vaccination history, and antibiotic use history… After informed
      consent, 70 patients with ARTIs due to influenza virus will be randomized into 2 groups (n =
      35/group): the control group (named "Control" group) uses 0.9% NaCl physiological saline and
      an experimental group (named the "Navax" group) use the probiotics LiveSpo Navax. The patient
      is given a coded spray in the form of a blind sample to ensure the objectivity of the study.
      The clinical follow-up will be 5 days, nasopharyngeal samples will be collected at day 0 and
      day 2 to evaluate potential reductions in viral load and co-infection bacteria, as well as
      modulation of overreacted cytokine release and the presence of probiotic spores in the
      patient's nasal mucosa.

      Real-time PCR for detection of microorganism in nasopharyngeal samples: semi-quantitative
      assays for measuring changes in influenza viral load and co-infection bacterial
      concentrations is conducted by the real-time RT-PCR/PCR routine protocol which has been
      standardized under ISO 15189:2012 criteria and used in Vietnam National Children's Hospital.
      Detection of B. subtilis ANA4 and B. clausii ANA39 are also conducted by real-time PCR SYBR
      Green that has been standardized under ISO 17025: 2017 standard and routinely in the Key
      Laboratory of Enzyme and Protein Technology, VNU University of Science.

      ELISA assays for cytokine levels: pro-inflammatory cytokines levels (pg/mL) including
      interleukin (IL-6, IL-8) and TNF-alpha are quantified using an enzyme-linked immunosorbent
      assay kit (ELISA) according to the manufacturer's instructions.

      During treatment, patients are monitored daily for typical clinical symptoms of Flu-induced
      respiratory tract infections, including runny nose, fever, dry rales, moist rales, body
      temperature (oC), oxymetry (SpO2) (%), pulse (beats/min), and breath (beats/min) until
      discharged. The patients' health conditions are observed by doctors and nurses, and their
      pieces of information are filled into medical records. During this study, parents' patients
      are asked to abstain from consumption for their children of other probiotics, either via
      nasal spray or oral administration, and refrain from cleaning nose for their children with
      other 0.9% NaCl physiological saline sprayers.

      Data collection and statistical analysis: individual medical records are collected, and the
      patient's information is then gathered and systematized in a data set. The efficacy of
      LiveSpo Navax is evaluated and compared to 0.9% NaCl physiological saline based on the
      following clinical and sub-clinical criteria obtained in Navax and Control groups: (i) the
      symptomatic-relieving day; (ii) the reduction levels (2^△Ct) of influenza viral load and
      co-infection bacteria concentrations. △Ct for target genes is calculated as Ct (threshold
      cycle) at day 2 - Ct at day 0 while Ct of internal control is adjusted to be equal among all
      samples; (iii) the reduction levels of IL-6, IL-8, and TNF-alpha cytokines. The tabular
      analysis is performed on dichotomous variables using the χ2 test or Fisher's exact test when
      the expected value of any cell is below five. Continuous variables are compared using either
      the Wilcoxon test, t-test, or the Mann-Whitney test when data are not normally distributed.
      The correlations among the variables are assessed by Spearman's correlation analysis.
      Statistical and graphical analyses are performed on GraphPad Prism v8.4.3 software (GraphPad
      Software, CA, USA). The significance level of all analyzes is set at p < 0.05. P-values.

      Expected outcomes: (i) LiveSpo Navax alleviates influenza-infection symptoms about 30% more
      effectively, as indicated by 90% of patients using LiveSpo Navax (Navax group) are
      symptom-free at day 2-5 of intervention depending on symptoms, compared to 60% of patients in
      Control group; (ii) Patients in Navax group has more significant reductions in influenza
      viral load (>10 fold) than patients in Control group at day 2 of intervention.
    

Intervention name(s): LiveSpo Navax; 0.9% NaCl physiological saline
Intervention type(s): Combination Product; Drug
Intervention description(s): In Vietnam, LiveSpo Navax is manufactured as a Class-A medical device product (Product declaration No.210001337/PCBA-HN) under manufacturing standards approved by Hanoi Health Department, Ministry of Health, Vietnam (Certificate No YT117-19) and ISO 13485:2016.; Nasal-spraying 0.9% NaCl physiological saline is prepared by extracting 5 mL from 0.9% NaCl intravenous infusion 500 mL PP bottle (B.Braun, Germany, product declaration No. VD-32732-19), and then pouring it into the same opaque plastic spraying 10 mL-bottle that is used for LiveSpo Navax.
Other intervention name(s): Registration number: No.210001337/PCBA-HN; Registration number: VD-32723-19
Allocation: Randomized
Intervention model: Parallel Assignment
Intervention model description: Blind, randomized, and controlled clinical trial
Masking: Triple (Participant, Care Provider, Investigator)
Primary purpose: Treatment
Keywords: Influenza Virus (Flu); Acute Respiratory Tract Infections (ARTIs); Children; Nasal-spraying probiotics; Viral load; Co-infection bacteria; Cytokines; Bacillus spores