New Zealand has the third highest rate of obesity within OECD countries with 8% of the adult population presenting type 2 diabetes and 25% living with pre-diabetes. Obesity and type 2 diabetes significantly increase the risk of cardiovascular disease and cancer and are critical modifiable risk factors associated with these conditions. In this study we determined whether relative loss of fat from visceral (compared to subcutaneous fat) depots can be manipulated by probiotics among participants with obesity and pre-diabetes who restricted caloric intake through intermittent fasting (IF). A randomized double-blind, two parallel arm study was performed in Auckland, New Zealand. Of the total 33 participants who were randomized, 26 completed the study, of whom 22 had magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS) data at baseline and 12 weeks after starting treatment. All participants commenced 12-week of intermittent fasting (IF) as 2 days per week of caloric restriction to 600-650 kcal/day (IF5:2) and were randomized 1:1 to either daily probiotic (Lacticaseibacillus rhamnosus HN001) or matching placebo during this period. subcutaneous adipose tissue (SCAT), visceral adipose tissue (VAT), liver fat (LF), and pancreatic fat (PF), as quantified through MRI/MRS, were compared in probiotic versus placebo groups. Changes in fat depots were also compared with changes in haemoglobin A1c (HbA1c) and body mass index (BMI.) Overall, the average weight loss ranged from 94.2 to 89.5 kg and was mild-positively correlated with percentage of VAT to SAT ratio (%V:S) (r = 0.215, (p = 0.392). There was a correlation with %V:S and improvements in HbA1c (r = -0.007, p = 0.973). There was no difference in %V:S between probiotic versus placebo groups at 12 week. Intermittent fasting resulted in a modest weight loss of 5% which, when combined with probiotics, is not able to better target visceral fat loss compared to placebo. There was a correlation between %V:S fat reduction and improvement in HbA1c in overweight people with pre-diabetes. Therefore, intermittent fasting is beneficial in reducing weight and improve biomarkers in obese pre-diabetic participants, however, the beneficial effects of probiotics need to be further investigated for proper conclusion.