2.1. Sample collection, analyses of environmental parameters and prokaryote abundances Samples were collected from surface at 5 stations along a transect from the Pearl River estuary to the oceanic region of the NSCS in August 2009 and January 2010 (Fig. 1, Table 1). Seawater samples were collected using Niskin bottles (12 L) attached to a conductivity, temperature, and depth (CTD) rosette multi-sampler. Environmental parameters (nitrate (NO3 −), phosphate (PO4 3−), dissolved oxygen, and chlorophyll a (Chl a) concentrations) were measured as described in Kong et al. (2011) (Table 1). Temperature was measured by the CTD. Picoplankton cell densities were quantified using flow cytometry (FCM) following the methods described in Chen et al. (2012) and Liu et al. (2014). At each station, 1 L of seawater was filtered successively through a 3.0 mm and 0.22 mm (47 mm) polycarbonate membranes (PALL Corporation). Samples retained on 0.22 mm membranes were stored at 80 °C immediately after filtration, and used for DNA extraction.