SOURCE:

Faculty: Biosences
Department: Applied Microbiology And Brewing

CONTRIBUTORS:

Victor-Aduloju, A. T.
Anyamene, C. O.

ABSTRACT:

Diabetic ulcers is one of the most common complications in diabetic patients. They are considered as a high-risk factor for lower-limb amputation. The present study was conducted to evaluate the wound-healing potentials of probiotics isolated from laboratory-produced ‘kunun-zaki’ on streptozotocin-induced diabetic rats. The ‘kunun-zaki’ is a slurry produced from millet and sorghum with a thickener (paddy rice and sweet potatoes) at different substitution levels (50:50%, 80:20%) were evaluated for their physicochemical, proximate, vitamins, mineral, sensory and microbial parameters. The isolation of probiotics was carried out using standard culture and biochemical techniques. These isolates were screened for their probiotic properties using different values of pH, temperature and sodium chloride (1-10% concentration). The antibacterial activity of the Lactobacillus strains was evaluated using agar well diffusion method against three pathogens (Pseudomonas aeruginosa, Staphylococcus haemolyticus and Klebsiella pneumoniae) isolated from a chronic ulcer. The Lactobacillus with the highest inhibitory zone were molecularly identified using polymerase chain reaction method. Seventy five Wistar female rats were divided into 5 groups and each group was sub-divided into 3 of 5 rats each (Group A- normal control, B- diabetic control, C- diabetic treated with Lactobacillus casei, D- diabetic treated with Lactobacillus plantarum and group E- diabetic treated with gentamicin) assigned to sub-groups is the pathogenic organisms to be treated. A full-thickness wound (2.0 cm) was made at the upper paravertebral of each rat and infected with the pathogenic organisms. After 24 h of injury, body weight, blood glucose level, percentage wound-healing and the total red blood cell and white blood cell counts of rats were examined. The pH of the samples ranged from 4.30–5.67 and the total soluble solids content ranged from 9.47% - 13.93%. ‘Kunun-zaki’ produced using paddy rice prepared at 50:50% (millet/sorghum) had 86.07% moisture content, 4.45% ash, 0.34% fiber value, 1.90% protein and 6.80% carbohydrate. The sweet potatoes based ‘kunun-zaki’ had highest vitamin C of 32.25 mg/100ml, 18.21mg/100ml vitamin E, 154.67mg/100ml magnesium and 245mg/100ml potassium. Furthermore, the sensory quality characteristics (flavour and overall acceptability) of the ‘kunun-zaki’ processed with paddy rice and paddy rice + sweet potatoes at 50:50% and 80:20% (millet and sorghum) had the highest scores of 6.35 and significantly different (P<0.05) from other samples. A total of eight Lactobacillus strains viz: Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus delbruekii, Lactobacillus lactis, Lactobacillus helveticus and Lactobacillus bulgaricus were isolated from eight ‘kunun-zaki’ samples. The Lactobacillus plantarum and Lactobacillus casei survived in a wide range of NaCl concentration (1-10%) and in pH 2. Lactobacillus plantarum isolated from 50:50 ‘kunun-zaki’ with paddy rice had the highest inhibitory zone against all the pathogenic organisms and is significantly different (P< 0.05) from other strains. The molecular identification revealed the Lactobacillus species with highest zones of inhibition to be Lactobacillus casei and Lactobacillus plantarum. The body weight was slightly increased in group A (control) but decreased in group B on day 7. The fasting blood glucose level of untreated diabetic rats (233mg/dL) was significantly higher than other groups on day 7. Lactobacillus plantarum significantly decreased all infected wound areas as compared to other groups and increased wound healing. The control group had the highest haemoglobin of 13.69g/dL on day 15. All the treatment groups showed non-significant change (P> 0.05) in the percentage of neutrophil in rats infected with Klebsiella pneumoniae in day 7, however, there was a significant increase (P< 0.05) in neutrophils, in diabetic control rats in day 15.This study has shown that the use of ‘kunun-zaki’ can serve as a natural probiotic drink. The isolated Lactobacillus plantarum has been shown to be the most effective probiotic which promotes diabetic ulcer healing.