**6. Changes of microbial community composition in the sewage sludge and soil upon sludge treatment and application**

The SS is characterized by a great microbial diversity, which may vary depending on the origin of sewage, its treatment, and industrial activity. Microbial activity in SS, transformation by-products, and residues may impact soil quality if SS is used as fertilizer/amendment [64]. The number of different groups of indicator microorganisms in 1 g of raw SS (wet) on average is 102 –103 for Salmonella (bacteria), Enteroviruses (viruses), Giardia (protozoa), and Ascaris (helminths), while 106 – for bacteria *Escherichia coli* [56, 65].

#### **6.1 Microbial community structure in the raw and treated sewage sludge**

Many factors modulate microbial community structure within SS, which may change from autotrophic to heterotrophic bacteria depending on the effluent source. According to Nascimento et al. and Nielsen et al. [64, 66], Proteobacteria phylum (21–65%) is predominant in municipal SS. This phylum was primarily dominated by Betaproteobacteria that represents bacteria involved in organic matter degradation and nutrient cycling. Bacteroidetes, Acidobacteria, and Chloroflexi were among the less prominent species. Our recent experiments have also revealed Proteobacteria to dominate in the raw SS (60.17% reads), which consisted of 16.40%, 29.18%, and 12.33% of Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria, respectively. At the genus level, the most abundant were Streptomyces (5.68%) and Pseudomonas (3.48%) (**Figure 5A**) [47].

Considerable changes in the microbial community structure of SS occur during biological treatment. Proteobacteria and Bacteroidetes were the most abundant in aerobic and anaerobic conditions, respectively [67, 68]. As reported by Rimkus et al. [47],

#### **Figure 5.**

*Relative abundance of microorganisms in sewage sludge. A—at the genus level with relative abundance ≥1% (29% from the total reads); B—Salmonella enterica and Escherichia coli in the sewage sludge before and after the 16-day incubation with different carbon amendments. Methods are according to Rimkus et al. [47].*

addition of three types of carbon sources (faba bean straw, wheat straw, and woodchip pellets) to the raw SS resulted in considerable changes in microbial community structure after 16 days of aerobic incubation. In particular, abundance of Firmicutes increased from 5% in the raw SS to 35–50% in the treated samples. In turn, abundance of Proteobacteria decreased from 62% in the raw SS to 32–45% in the treated samples. Yet, the SS incubation without C amendment resulted in a remarkable increase in virus abundance (i.e., 0.34% reads) [47]. The relative abundance of *Salmonella enterica* and *Escherichia coli* has been increased in the treated sludges, as compared to the raw SS (**Figure 5B**).
