The primary purpose of this study was to survey a sample of infantry soldiers to determine their preferred backpack load distribution. The second objective was to examine the effect of backpack load position, walking speed and surface grade on the physiological responses of infantry soldiers. In the first phase of the study, analysis of 500 questionnaires indicated that backpack item arrangement was typically determined by convenience and habit. About 33.6% and 30.6% of the soldiers reported placing heavier items in the lower and higher positions, respectively. In the second experimental phase of the study, five male infantry soldiers who had completed basic training (including 30 km tactical road march) each performed eight treadmill walking bouts at one of two speeds (i.e., 3.2 and 6.4 km/h) and two surface grades (0% and 6%) carrying 15% of their body weight in a backpack with two chambers (upper and lower). Respiratory frequency, minute ventilation, oxygen consumption and heart rate were recorded using the Quark pulmonary system. The MANOVA results indicate that the effect of walking speed was strongly significant for all physiological indices. In addition, as might be expected, walking on a 6% grade was associated with higher oxygen consumption and heart rate compared to level going. In terms of load position, the mean respiratory frequency was significantly higher where the load was carried in the upper position. Furthermore, there was a significant interaction between load position and walking grade on oxygen consumption. Where participants were walking at zero incline, there was no significant difference in mean oxygen consumption. By contrast, the mean oxygen consumption was significantly higher where loads were carried in the upper position and negotiating the 6% grade. Previous research has found that, ideally, load carriage should complement stability, bringing the load's center of gravity as close to the body as possible and making use of the larger muscles. However, carrying heavy loads close to the trunk can affect lung function. Thus, load placement is an important factor in physiological response to load carriage, and optimum choice of upper or lower position when distributing items in a backpack may be dependent on the walking grade. Relevance to industry The need to carry heavy loads for long distances is common to a range of human endeavor, with physical transport used by soldiers, various types of workers and recreational hikers. Load distribution and walking gradient are important factors in terms of the efficiency of load carriage and should be taken into consideration in both the design and loading of backpacks. Thus, the findings of the present study will be relevant to infantry soldiers, mountain climbers or of manual material handling (MMH) in industry, providing practical implications for tasks associated with exposure to uneven walking/working surfaces.