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Page 6 of 15 Kamilari et al. Microbiome Res Rep 2025;4:3 https://dx.doi.org/10.20517/mrr.2024.47

Figure 1. The relative abundance of the most abundant microbes identified at the species level (mostly including the family and the
genus level) based on 16S rRNA gene sequencing for (A) Cyprus, (B) Mytilini, and based on ITS gene loci sequencing for (C) Cyprus, (D)
Mytilini. The code (T) refers to traditional sausages, whereas the code (I) refers to industrial sausages.

Identification of microbial biomarkers
The LEfSe tool was used to determine if the relative representation of the detected microbial taxa was
disproportionately distributed among sausages produced in different areas where the LEfSe algorithm was
applied [Figure 2A and B]. Sausages from Mytilini were distinguished by a greater representation of the
bacteria Kocuria and the family Pasteurellaceae, as well as the undesirable fungi Alternaria, Pleosporaceae,
Aspergillus, Trichomonascus, Bulleribasidiaceae, and Wallemia. Cypriot sausages from Nicosia were
overrepresented by the species Leuconostoc mesenteroides, whereas those from Limassol by members of the
bacterial family Actinomycetaceae and by the fungal genera Neofusicoccum, Aureobasidium, Botrytis,
Pestalotiopsis, and Holtermanniella. Finally, traditional sausages from Pitsilia indicated an elevated relative
abundance of the fungi Glomerellaceae, Wallrothiella, and Vishniacozyma.

The traditional manufacturing conditions applied in Cypriot Pitsilia sausages contributed to a significantly
higher relative representation of Lactobacillus, including L. helveticus [Figure 2C]. The species
Staphylococcus succinus, and the genera Neofusicoccum, Botryosphaeriaceae, Aureobasidium, Aspergillus,
Xerochrysium, and Meyerozyma were also overrepresented [Figure 2C and D]. Industrially produced
sausages exhibited an increased relative abundance of Clostridium and Enterobacteriaceae, and the fungi
Leucosporidiaceae and Tausonia.