Background: Garlic is an oldest agricultural harvest that has historical dating since 800 BC and today being used worldwide as food and medicine. Garlic has many different uses like antimicrobial activities of garlic volatile oil, garlic powder, and garlic juice. The main garlic species is Allium sativum, which is accepted as an ethnopharmaceutical drug and proved to have therapeutic effects. The Citrus limon fruit has well-known nutritional properties, but its valuable biological activities are underestimated in modern phytotherapy and cosmetology. Garlic and lemon possess several valuable properties, in particular anti-inflammatory, antioxidant, anticancer, antidiabetic, antiviral, antifungal and antimicrobial activities. Objective: To determine the antibacterial effectiveness of Garlic (Allium sativum) and Lemon (Citrus limon) on gram positive and negative bacterial isolates from clinical specimen Methods: Fresh garlic and lemon were bought from local market in Hawassa city and were subjected to solvent extraction. The fresh juice, aqueous and ethanol extracts of garlic and lemon were tested for susceptibility pattern with isolates of gram positive and negative bacteria. Data were be sorted manually, computed for descriptive statistics, summarized and presented with tables, frequencies, percentage, mean and standard deviation. Results: A total of 27 (21.3%) clinical bacteria were isolated from 127 different clinical samples. From these 13 (48.1%) were gram positive bacteria and 14 (51.9%) isolates were gram negative. Fresh juice of garlic showed antibacterial effects on 26 (96.3%) isolates of the study that is 100% effect on gram positives and 92.8% effect on gram negative isolates. This fresh garlic juice showed no zone of inhibition (ineffective) to one Pseudomonas aeruginosa. Zone of inhibition for gram positive bacteria ranges from 9.7±0.6mm (Enterococcus faecalis) to 35.3±0.6mm (MRSA) and for gram negative bacteria it ranges from 10.7±0.6mm (Pseudomonas aeruginosa) to 33.7±0.6mm (Escherichia coli). Heat treatment of garlic revealed decreasing effect with increasing temperature. But at 100 0C for 20 minutes showed no antibacterial effect on all (100%) study isolates. Aqueous extract was showed lower antibacterial effect than fresh garlic juice. All (100%) isolates showed resistance for 500gm/ml, 200mg/ml and 100mg/ml ethanolic extract of garlic. Fresh lemon juice shown antibacterial effect to all 27 (100%) of isolates with zone of inhibition ranging from 8.3±0.58mm to 23.7±0.58mm but the lemon peel ethanolic extract showed no antibacterial effect to all (100%) isolates.