Antibacterial, Antifungal, and Cytotoxic Effects of Endophytic Streptomyces Species Isolated from the Himalayan Regions of Nepal and Their Metabolite Study

<b>Background/Objectives:</b> Recently, antimicrobial-resistant pathogens and cancers have emerged as serious global health problems, highlighting the immediate need for novel therapeutics. Consequently, we aimed to isolate and characterize endophytic <i>Streptomyces</i> strains from the rhizospheres of the Himalayan region of Nepal and identify specialized metabolites with antibacterial, antifungal, and cytotoxic potential. <b>Methods:</b> To isolate <i>Streptomyces</i> sp., we collected two soil samples and cultured them on an ISP4 medium after pretreatment. We isolated and identified the strains PY108 and PY109 using a combination of morphological observations and 16S rRNA gene sequencing. <b>Results:</b> The BLAST results showed that PY108 and PY109 resembled <i>Streptomyces hundungensis</i> PSB170 and <i>Streptomyces</i> sp. Ed-065 with 99.28% and 99.36% nucleotide similarity, respectively. Antibacterial assays of ethyl acetate (EA) extracts from both isolates PY108 and PY109 in a tryptic soy broth (TSB) medium were conducted against four pathogenic bacteria. They showed significant antibacterial potential against <i>Staphylococcus aureus</i> and <i>Klebsiella pneumoniae</i>. Similarly, these extracts exhibited moderate antifungal activities against <i>Saccharomyces cerevisiae</i> and <i>Aspergillus niger</i>. Cytotoxicity assays on cervical cancer cells (HeLa) and breast cancer cells (MCF-7) revealed significant potential for both extracts. LC-MS/MS profiling of the EA extracts identified 27 specialized metabolites, including diketopiperazine derivatives, aureolic acid derivatives such as chromomycin A, and lipopeptide derivatives. In comparison, GC-MS analysis detected 34 metabolites, including actinomycin D and γ-sitosterol. Furthermore, a global natural product social molecular networking (GNPS)-based molecular networking analysis dereplicated 24 metabolites in both extracts. <b>Conclusions:</b> These findings underscore the potential of endophytic <i>Streptomyces</i> sp. PY108 and PY109 to develop new therapeutics in the future.