2. Materials and methods2.1. Study areaThe Taihu Basin (11930100–1215402600E, and 30701900–321405600N) is located in the Yangtze River Delta region, with atotal area of 36,895 km2 including a water area of approximately6134 km2. It is a subtropical region, the average annual temperatureis 15–17 C and the average annual precipitation is 1177 mm(Gao et al., 2011b; Yu et al., 2012; Zhao et al., 2011). The basin iscovered naturally by subtropical evergreen broadleaf forest, andthere are various topographical types, including hilly areas towardthe west, and low-lying flood plain in the northern and easternparts, which occupying approximately 83% of the basin (Gao andGao, 2012; Gao et al., 2012; Sun and Huang, 1993). There are morethan 200 rivers distributed in the basin, and 172 rivers or channelsare connected to Lake Taihu (Xu and Qin, 2005). The Taihu Basin isone of the most developed and highly populated areas of China,with a population of 58.79 million and an average populationdensity of nearly 1600 inhabitants/km2. This area contributes morethan 10% of the gross domestic product (GDP) of China andoccupies only 0.4% of the nation’s territory (2011), the average GDPper capita was 2.3 times the national average (Taihu BasinAuthority, 2012, 2013). Thus, these natural conditions and therapid development of the economy may potentially lead to waterquality degradation in the basin (Qin et al., 2010).According to hydrology and topography characteristics, theTaihu Basin was delineated into two level I aquatic ecoregions:ecoregion I1 (western hill aquatic ecoregion) and ecoregion I2(eastern plain aquatic ecoregion) (Gao et al., 2011ab; Gao and Gao,2010). Sample sites in ecoregion I1 were riffles dominated bywadeable streams, and sample sites in ecoregion I2 were poolsdominated by unwadeable rivers. Due to these differences,separate MMIs for ecoregion I1 (MMIW) and ecoregion I2 (MMIE)were developed.2.2. Sample collection and processingSample sites were randomly selected via a randomizedsystematic design with a spatial component (Herlihy et al.,2000). Samples were collected from 104 streams and rivers ofthe basin from October 16th to November 8th, 2012. Sites wereaffected by a range of potential human-induced disturbances, frommild to severe. According to level I aquatic ecoregions of the basin,30 sites were located in ecoregion I1, and 90 sites were located inecoregion I2 (Fig. 1).Triplicate measures of physico-chemical parameters weremade in the water at each station before benthic macroinvertebrateswere sampled. Temperature, pH, dissolved oxygen (DO),turbidity and electrical conductivity (EC) were measured using aportable multi-probe meter (Model YSI-6600 V2, YSI Inc., YellowSprings, Ohio, USA) at the center of each of the sampling reaches. Ateach site, 2 L of water was sampled in sterilized plastic bottles,stored in a portable refrigerator at <4 C, and later analyzed fortotal phosphorus (TP), total nitrogen (TN), chemical oxygendemand in manganese (CODMn), and ammonia nitrogen (NH4) inthe laboratory following Standard Methods for the Analysis ofWater and Wastewater (2002). In addition, GPS information wasdocumented with a GPSmap 60CSx (Garmin, Taiwan).We evaluated habitats qualitatively based on modifications ofpublished studies (Barbour et al., 1999; Zheng et al., 2007). At thesample sites, we assessed five parameters that included channelflow status, substrate composition, habitat complexity, bankstability, and width of the vegetated riparian zone in ecoregion I1,whereas channel flow status, channel alteration, water fluiditystatus, habitat complexity, and width of the vegetated riparianzone were assessed in ecoregion I2. Each parameter was scored aspoor (0–5), fair (6–10), good (11–15), or excellent (16–20), withthe final habitat quality index (HQI) ranging from 0 to 100(Appendix A). The land use pattern was quantified from a land usedatabase, which was based on Advanced Land Observing Satellite(ALOS) fused images (2.5 m resolution) of Taihu Basin (Gao andGao, 2012).Benthic macroinvertebrate samples were collected in eachreach, which consisted of a length of 100 m delineated around therandomly selected site. Sampling was conducted with a 30 cmdiameter D-frame net with a 500 mm mesh using the multihabitatapproach described by Barbour et al. (1999). A variety ofmicrohabitat types were sampled at each reach, including riffles,pools, banks, submerged and overhanging macrophytes, anddepositional areas. In the large rivers, only the littoral section wassampled. At each reach, 10 sampling units (30 cm 50 cm) weretaken and pooled for analysis. Samples containing large volumesof sand were sorted in the field by repeated washing of sedimentswith water using a 0.5 mm sieve to collect organisms. Allremaining materials were fixed with 7% buffered formaldehydesolution. In the laboratory, samples were sorted by hand in whiteenamel pans. Specimens were identified to the lowest taxonomiclevel possible, blotted until dry, and weighed with an electronicbalance (Sartorius BS-124, precision: 0.1 mg). Molluscs wereweighed with their shells. Individuals were counted andidentified to the lowest taxon possible, usually genus, throughuse of taxonomic keys (Cai et al., 1991; Liu et al., 1979; Morse et al.,
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