A display substrate and a display device are provided. The display substrate has a plurality of sub-pixels, the plurality of sub-pixels include first sub-pixels, second sub-pixels, and third sub-pixels; the first sub-pixels and the third sub-pixels are disposed alternately along a row direction to form a plurality of first pixel rows, and the first sub-pixels and the third sub-pixels in a same column among the plurality of first pixel rows are disposed alternately, the second sub-pixels are disposed side by side along the row direction to form a plurality of second pixel rows; and the display substrate includes a base substrate, a pixel driving circuit layer, a pixel definition layer, and a black matrix layer; the black matrix layer includes a plurality of first openings and a plurality of second openings.