The U.S. Geological Survey, in cooperation with the Houston-Galveston Area Council and the Galveston Bay Estuary Program under the authority of the Texas Commission on Environmental Quality, did a study in 2007 to assess the variation in biotic assemblages (benthic macroinvertebrate and fish communities) and stream-habitat data with sampling strategy and method in tidal segments of Highland Bayou and Marchand Bayou in Galveston County. Data were collected once in spring and once in summer 2007 from four stream sites (reaches) (short names Hitchcock, Fairwood, Bayou Dr, and Texas City) of Highland Bayou and from one reach (short name Marchand) in Marchand Bayou. Only stream-habitat data from summer 2007 samples were used for this report. Additional samples were collected at the Hitchcock, Fairwood, and Bayou Dr reaches (multisample reaches) during summer 2007 to evaluate variation resulting from sampling intensity and location. Graphical analysis of benthic macroinvertebrate community data using a multidimensional scaling technique indicates there are taxonomic differences between the spring and summer samples. Seasonal differences in communities primarily were related to decreases in the abundance of chironomids and polychaetes in summer samples. Multivariate Analysis of Similarities tests of additional summer 2007 benthic macroinvertebrate samples from Hitchcock, Fairwood, and Bayou Dr indicated significant taxonomic differences between the sampling locations at all three reaches. In general, the deepwater samples had the smallest numbers for benthic macroinvertebrate taxa richness and abundance. Graphical analysis of species-level fish data indicates no consistent seasonal difference in fish taxa across reaches. Increased seining intensity at the multisample reaches did not result in a statistically significant difference in fish communities. Increased seining resulted in some changes in taxa richness and community diversity metrics. Diversity increases associated with increased electrofishing intensity were relatively consistent across the two multisample electrofishing reaches (Hitchcock and Fairwood). Differences in the physical characteristics of the Highland and Marchand Bayou reaches are largely the result of the differences in channel gradient and position in the drainage network or watershed of each reach. No trees were observed on the bank adjacent to the five transects at either the Bayou Dr or Texas City reaches. Riparian vegetation at the more downstream Fairwood, Bayou Dr, and Texas City reaches was dominated by less-woody and more-herbaceous shrubs, and grasses and forbs, than at the more upstream Hitchcock and Marchand reaches. The width of the vegetation buffer was variable among all reaches and appeared to be more related to the extent of anthropogenic development in the riparian zone rather than to natural changes in the riparian buffer. Four additional transects per reach were sampled for habitat variables at Hitchcock, Fairwood, and Bayou Dr. Medians of most stream-habitat variables changed with increased sampling intensity (addition of two and four transects to the standard five transects), although none of the differences in medians were statistically significant. All habitat quality index values for the five reaches scored in the intermediate category. Increasing sampling intensity did not change the habitat quality index score for any of the reaches.