Study Design and setting
To be able to fulfill the objective of the proposed study, Prospective Randomized Controlled Trail design was employed at Malalai and Maiwand hospitals in Kabul city during May-October 2020.
The study population was comprised of preterm neonates (Infant up to 28 day old) with weight less than 2000 g.
Inclusion and exclusion criteria
Preterm neonates with gestational age ≤ 36weeks or weight less than 2000 g enrolled in the study. We excluded the neonates with visible or physically detected malformation, birth weight less than 1000gr, severe birth asphyxia, seriously ill baby and parent who declined consent.
Sampling strategy and sample size
Block randomized sampling was used to recruit the study participants. Stata 14 used for sample size calculation. A hypothesized mean increment of 3 g in daily weight gain for the probiotic group took in consideration. The preterm and low birth weight neonate grows about 6.8 ± 4.8 gr/day. With alpha error of 5% and power of 80%, the two-sample means test estimated a sample size of 84 neonates (42 baby for each group). Forty-two premature neonates managed by dual strain probiotic (Probiotic group), and the other forty-two premature neonates took under routine care (Control group).
Dual strain probiotic, mother breast milk, premature formula, nasogastric tube (NGT) and syringes were used for the feeding of preterm infants. Premature formulas and dual strain probiotic purchased by the research project. Pre Biomil was a premature formula that made by FASSKA in France. Gutcare a dual strain probiotic that each sachet containing 500 mg Bifidobacterium and Clostridium butyricum was a product of Searle, Pakistan.
Totally 93 preterm neonates were studied for the effectiveness of probiotic on the growth and feeding intolerance. Six patients did not meet the inclusion criteria due to less than 1000 g weight (3 neonates) and critical ill condition (3 neonates). From 87 patients, 43 preterm newborn babies received mother breast milk or formula feeding together with dual strain probiotic and the other 44 babies took non-probiotic fortified mother breast-feeding or formula feeding. There were one and two cases of loss to follow up in probiotic and control groups respectively (Figure-1).
In the probiotic group, forty-two neonates were feed by mother breast milk together with dual strain probiotic (Gutcare) half sachet (250 mg) dissolved in 1–3 ml milk twice daily. The probiotic-fortified milk started from day 4 and continued for 1-3weeks. In the control group (forty-two), non-probiotic fortified mother breast-feeding was the choice.
The routine management such as incubator care, fluid and electrolyte maintenance, expressed breast milk (EBM) and premature formula via NGT for infants less than 1800gr or gestational age ≤ 35weeks undertook for two mentioned groups. EBM initiated as trophic feeds of 10 to 20 mL/kg/day at 2 hourly intervals in hemodynamically stable infants. Feeds advanced by 20 mL/kg/day up to 150 ml/kg/day
Primary outcome measures were daily weight gain and feeding intolerance defined ≥ two episode of vomiting during last 6hr. Secondary outcome measures is neonatal death during hospital stay.
Main variables were
Daily weight gain was calculated as weight gain in gram during observation period divided by number of days.
Risk of feeding intolerance defined as ≥ two episodes of vomiting during last 6hr.
Gestational age as weeks, determined by LMP or antenatal maternal ultrasound or gestational age assessment by neonatal heel-toe distance.
Risk of neonatal death during hospital stay.
Data collection tools & Statistical analysis
Initially raw data were collected in data collection sheets and then entered in SPSS software for statistical analysis. The evaluation of efficacy performed by comparing the mean and RR with the consideration of p-value. To detect the significance level, the mean of normally and non-normally distributed data were compared by independent t-test and Mann-Whitney test, respectively. Chi-square test was performed for dichotomous data. As we accepted the alpha errors of 0.05, therefore, the p-value < 0.05 was significant.
The proposal of this RCT was registered in the Australian New Zealand Clinical Trials Registry (ANZCTR) with the registration number of ACTRN12620000538943. The department of Neonatology, Kabul University of Medical Science (KUMS) and Research committee of Ministry of Higher Education of Afghanistan approved the RCT proposal and final report on the basis of Helsinki Declaration. The consents of parent were taken.