Bacterial and fungal profiling of Maltese sheep cheese with amplicon metabarcoding

Abstract

This study aimed to investigate the seasonal variability of bacterial and fungal communities in raw sheep milk and Ġbejna from the Maltese Islands using 16S and ITS-based NGS. Firmicutes and Proteobacteria emerged as dominant bacterial phyla, with Lactococcus lactis, Hafnia alvei, and Streptococcus among the most prevalent species, while L. lactis were consistently abundant regardless of conditions in all samples. Fungal profiling revealed a predominance of Basidiomycota and Ascomycota, with Cutaneotrichosporon curvatus enriched in milk, while Trichosporon insectorum and Kluyveromyces lactis were dominant in dried cheeses. Alpha diversity (Chao1, ACE, Shannon) for fungal microbiota showed no significant seasonal variation, while bacterial microbiota exhibited significantly higher richness in summer than in winter samples. Additionally, beta diversity metrics indicated that cheese microbiomes were strongly shaped by geographical origin rather than sample type. The presence of potential spoilage or opportunistic bacteria and fungi highlights the need for improved hygiene control and fermentation conditions during cheese production. This is the first study to characterise the microbial diversity of Maltese ġbejna using NGS, revealing seasonal and regional variations that contribute to its cultural and economic value.