**8. Conclusions**

*Microorganisms*

**7. Generalization of the research**

**36**

**Figure 8.**

*The general scheme of isolation, purification, and identification of mesophilic sulfate-reducing bacteria.*

Moore W.E. found SRB for the first time in people's feces and identified it as *Desulfomonas pigra* [53], which subsequently is reclassified as *Desulfovibrio piger* [16]. The described bacterial strains are similar to that of Moore et al. [53] except for the G–C content of the DNA, which is 64 mol%. Obligate anaerobic, sulfate-reducing, non-saccharolytic, non-proteolytic, nonspore-forming, and Gram-negative bacteria that are straight and vibrio-like and have rounded ends (0.8–1.0 × 2.5–10.0 μm) [10]. These microorganisms use lactate, pyruvate, ethanol, and hydrogen as electron donors for sulfate reduction. They oxidize lactate and pyruvate incompletely to acetate. The optimum temperature for growth is +37°C. Growth is not affected by 20% bile. Colonies on anaerobic blood agar are translucent, 2 mm in diameter, circular, and non-hemolytic. Cells contain desulfoviridin and cytochrome *c*3. These bacteria isolated from human specimens (feces, peritoneal fluids, and intra-abdominal collections). The type strain, isolated from human feces, is ATCC 29098 [10].

Taking into consideration all research described in the chapter, it is necessary to generalize that isolation of mesophilic SRB from environmental samples (water, soil, swamp, etc.) and intestinal samples can be similar, although swamps and feces are required to determine dry matter of the samples. It is important to purify a positive sample of SRB from other satellite microorganisms such as *Clostridium*, *Bacteroides*, *Pseudomonas*, etc. With this aim, obtained SRB mixed cultures should be 3–5 times repassed to the modified Postgate agar medium with sulfite which inhibit the growth other microorganisms. Ability to growth of SRB mixed culture with high sulfite concentration allows to eliminate (purify SRB) from other microorganisms which can be in close interactions with SRB. The cultivation conditions depend on sample from where it is isolated. The key criteria for identification based on physiological and biochemical characteristics are the morphology of bacterial

The methods of sample selections from water, soil, swamp, and feces of humans or animals and from biopsy material and the process of SRB isolation and purifications are similar, although cultivation conditions may differ. Identification based on physiological and biochemical properties is a complex process, and many other factors must be considered. For this identification, Bergey's Manuals are recommended to be used. The sequence analysis of the 16S rRNA gene should confirm the identification process based on physiological and biochemical properties.

It is of vital importance to obtain new strains of the SRB from various ecotopes and identify them and study their growth and physiological and biochemical properties. Aside from that, the process of dissimilatory sulfate reduction by SRB and the production of hydrogen sulfide should be investigated in order to clarify the etiological role of these bacteria in the nature and in the development of various diseases.

### **Acknowledgements**

This study was supported by Grant Agency of the Masaryk University (MUNI/A/0902/2018).
