Combustion is known to be affected by variations in the air-fuel mixture concentration, residual gas concentration, turbulent kinetic energy, ignition, etc. However, because each of these factors is related to cycle-to-cycle variations, their effects on combustion variation are unclear. The purpose of this study was to clarify the influences of the air-fuel mixture distribution near the spark plug and variation in the relative position of the ignition on the combustion variation. A 4-cylinder port injection gasoline engine was used as the test engine, and the combustion variation was investigated by measuring the cylinder pressure and air-fuel ratio (A/F) near the spark plug for each cycle using a micro-Cassegrain sensor for each cylinder. The air-fuel mixture distribution was calculated using a Reynolds averaged Navier-Stokes simulation, and the spatial region of the high ignition probability was determined from the gas flow velocity. Then, the combustion progress when ignition occurred at each point of the region was calculated, and was compared with the measured combustion variation. As a result, the frequency distribution of the A/F within the ignition position variation region was correlated with the measured A/F for each cycle. Based on the combustion calculation results when the ignition position was changed, the difference that occurred in the growth rate of the initial flame kernel was related to the difference in the air-fuel mixture concentration at the ignition timing. These results suggested that variations in the relative positions of the air-fuel mixture distribution and ignition may strongly affect combustion variation.