As the largest tributary of the Heilong River, the Songhua River flows about 1,434 kilometers from the Changbai Mountains through the Jilin and Heilongjiang provinces. It occupies a significant position in the ecological and commercial fields in Northeast China. As it suffers from complex contaminant conditions, comprehensive environmental improvement and management of the Songhua River is vital. Thus, a relatively accurate monitoring and investment system is essential to supply basic data for decision and policy making. In this study, we aim to analyze the bacterial concentration and diversity, along with their relationship to various environmental factors in the river, for the purpose of a brief evaluation towards the microbial condition in the Songhua River. Representative sampling points were chosen from upstream to downstream of the Songhua River. A total of fifteen water samples and ten sludge samples were collected. The bacterial concentration was assessed by both traditional heterotrophic plate counts (HPC) and a novel flow cytometric (FCM) technique. HPC used to be widely utilized as an approach to detect the cultivable bacterial communities; however, it has been under discussion due to its non-negligible drawbacks and the development of new alternative tools. Results show that the order of magnitude of cultivable cells counted by HPC is about 10⁶ while the order of magnitude of total cells counted by flow cytometry is about 10⁴. It shows that traditional heterotrophic plant counts might easily neglect those cells that are uncultivable but possess physiological properties. Also, flow cytometry exhibits several advantages compared with HPC, for instance, more rapid rates and greater sensitivity. Regression analysis of environmental factors and microorganism concentrations was made based on four main factors: temperature, pH, dissolved oxygen, and conductivity. Results show that pH has the most significant effect towards the total cell count in each sample. The molecular technique using 16S rRNA profiles generated by PCR-DGGE was used in order to determine the variation in bacterial community structure of water and sludge samples from the Songhua River. Nested PCR and touch down PCR were utilized for the purpose of generating more DNA from samples, especially aquatic samples. Similarity analysis, cluster analysis and statistical analysis were performed according to the DGGE gels and the data output by Quantity One. The results showed that the bacterial diversity in water samples has an obvious discrimination between upper and lower zones, and the demarcation point is near the area of Harbin city. However, bacterial diversity in the sludge samples showed no significant differences along the river basin. The Shanno-Wiener Index, Uniformity Index and richness of water and sludge samples were calculated according to the DGGE profiles. The results showed that the bacterial diversity and richness decreases from downstream to upstream. The present study provides fundamental data for bacterial concentration and diversity, together with influencing environmental factors, which could be used to support the future discrimination between functional zones in the whole Songhua River Basin.