Aims: This study aimed at determining whether oral administration of a probiotic strain, Lactobacillus casei strain Shirota (LcS), can improve insulin resistance, which is the underlying cause of obesity-associated metabolic abnormalities, in diet-induced obesity (DIO) mice. Methods and Results: DIO mice were fed a high-fat diet without or with 0 center dot 05% LcS for 4 weeks and then subjected to an insulin tolerance test (ITT) or oral glucose tolerance test (OGTT). Oral administration of LcS not only accelerated the reduction in plasma glucose levels during the ITT, but also reduced the elevation of plasma glucose levels during the OGTT. In addition, plasma levels of lipopolysaccharide-binding protein (LBP), which is a marker of endotoxaemia, were augmented in the murine models of obese DIO, ob/ob, db/db and KK-Ay and compared to those of lean mice. LcS treatment suppressed the elevation of plasma LBP levels in DIO mice, but did not affect intra-abdominal fat weight. Conclusions: LcS improves insulin resistance and glucose intolerance in DIO mice. The reduction in endotoxaemia, but not intra-abdominal fat, may contribute to the beneficial effects of LcS. Significance and Impact of the Study: This study suggests that LcS has the potential to prevent obesity-associated metabolic abnormalities by improving insulin resistance.