Peanuts (Arachis hypogaea) are a vital and affordable source of nutrients, providing essential proteins, lipids, vitamins, and minerals. However, their susceptibility to microbial contamination poses significant food safety and shelf-life concerns. This study aimed to assess the microbiological quality and diversity of freshly harvested and stored peanuts, with implications for food safety and shelf life. Microbial analyses revealed that freshly harvested peanut seeds sampled from local markets in Obio-Akpor, Rivers State, Nigeria exhibited higher bacterial and fungal loads compared to stored samples, with viable bacterial counts of 5.4×10⁴ and 7.2×10⁴ cfu/g for freshly harvested seeds, and 3.2×10³ and 3.35×10³ cfu/g for stored seeds. Fungal counts followed a similar trend, with higher loads observed in freshly harvested samples (3.6×10³ and 4.0×10³ cfu/g) compared to stored seeds (1.2×10³ and 1.8×10³ cfu/g). The higher microbial load in freshly harvested peanuts was attributed to the lack of preprocessing, while the lower counts in stored samples were linked to industrial processing steps, including drying and salting, which reduce moisture content and inhibit microbial proliferation. Microbial diversity analysis identified Bacillus sp., Micrococcus sp., Staphylococcus sp., and Proteus sp. in both fresh and stored samples, with Serratia sp. isolated exclusively from freshly harvested peanuts and Enterobacter sp. found only in stored samples. Among fungal species, Aspergillus niger and Aspergillus flavus were predominant in both sample types, while Fusarium sp., Alternaria sp., and Trichoderma sp. were isolated solely from freshly harvested peanuts. Shelf-life studies demonstrated a progressive increase in microbial load over time, with higher rates observed in freshly harvested peanuts. The antibiotic susceptibility profile indicated resistance to Nalidixic acid, cephalexin, and penicillin among Gram-positive isolates, with high sensitivity to Ciprofloxacin, Ofloxacin, and Clindamycin. Gram-negative isolates were generally susceptible to the tested antibiotics, except for Norfloxacin, which was ineffective against Proteus sp. and Serratia sp. from freshly harvested samples. This study underscores the importance of preprocessing, such as salting and drying, to extend the shelf life of peanuts and mitigate microbial spoilage. The findings highlight the critical role of moisture control in reducing microbial contamination, thereby enhancing food safety and extending the shelf life of peanuts. Recommendations include the adoption of preprocessing techniques for subsistence farmers and the need for timely consumption of freshly harvested peanuts to minimize microbial deterioration.