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Two Rhizoclonium strains thriving in contrasting spring types (slightly hot spring/ambient, shaded pool spring) and biogeographic areas (El-Farafra Oasis in the Western Desert of Egypt/River-Po Plain in Northern Italy) were studied in depth based on field and cultured material, bright-field and fluorescence microscopy, and phylogenetic molecular analyses (SSU and LSU rRNA gene sequences). This polyphasic approach revealed that the two Rh. strains clearly belonged to the Rhizoclonium clade but differed in some of their key diagnostic features. The Egyptian Rhizoclonium strain, isolated from the spring-fed (Ain El-Balad) agricultural ditch in El-Farafra Oasis, was described with the working name Rhizoclonium sp. 10.6m from a desert, slightly-hot spring’ based on its smaller cell diameter, ecological and phylogenetic molecular traits compared to allied morphospecies. Moreover, it was highlighted that the L/D ratio can be more than 3.0 within this group. The Italian Rhizoclonium strain, collected from the Fontanile Valle Re-shaded, pool ambient spring (Emilia-Romagna Region), was virtually identical to other strains identified as Rh. hieroglyphicum from China and Japan, due to its highly supported congruence in morphological characteristics and phylogenetic position. This study provided the first partial LSU and SSU rRNA gene sequences for European Rh. hieroglyphicum based on available literature. Phenotypic plasticity of rhizoid formation was also observed in both Rhizoclonium strains studied using culturing approaching techniques. Our investigation also confirms that a lot of work with a variety of approaches is still needed to assess the ecological preferences, morphological plasticity, and phylogenetics of freshwater Rhizoclonium taxa worldwide.