Clostridium difficile infection (CDI) is a common cause of morbidity and mortality in hospitalized patients worldwide. The major problem facing current treatment is multiple recurrences, prompting the need for alternative therapies. In this study we isolated bacterial species, from Egyptian individuals’ stool, with antimicrobial activity against clinical isolates of C. difficile and tried to examine the nature of the produced antimicrobials. In vitro antibacterial activity against C. difficile was initially screened in 123 fecal samples cultures using an agar overlay method. The isolates with antimicrobial activity against C. difficile in addition to Clostridium isolates were identified using partial 16S rDNA gene sequencing analysis. The isolates acting against C. difficile belonged to Lactobacillus, Enterococcus and Clostridium genera. The concentrated cell-free supernatants (CFSs) from these bacterial isolates were examined for antimicrobial activity against C. difficile growth by broth dilution method. 10 x concentrated CFSs of five isolates showed inhibition for C. difficile growth which was significantly different (p < 0.001) from control. Lactobacillus agilis T99A and Clostridium butyricum T58A isolates were selected for further evaluation of the produced antimicrobials. The antimicrobial activity of 10x CFSs of the two isolates was stable after enzymatic treatment with proteinase K or heating treatments up to 90 °C or neutralizing pH. The spectrum of activity of the two isolates was evaluated using different gram-positive and gram-negative bacterial species and did not show antimicrobial activity against these species. Our results showed two unconventional bacterial isolates: L. agilis T99A and C. butyricum T58A producing extracellular thermo stable antimicrobial agents against C. difficile clinical isolates.

艰难梭菌感染（CDI）是全球住院患者发病和死亡的常见原因。当前治疗所面临的主要问题是多次复发，提示需要替代疗法。在这项研究中，我们从埃及人的粪便中分离出了对艰难梭菌临床分离株具有抗菌活性的细菌，并试图检验所产生抗菌剂的性质。首先使用琼脂覆盖法在123个粪便样品培养物中筛选了对艰难梭菌的体外抗菌活性。除梭状芽胞杆菌外还对艰难梭菌具有抗菌活性的分离株使用部分16S rDNA基因测序分析鉴定了分离株。对抗艰难梭菌的分离株属于乳杆菌，肠球菌和梭状芽胞杆菌属。通过肉汤稀释法检查了来自这些细菌分离物的浓缩的无细胞上清液（CFS）对艰难梭菌的抗微生物活性。5个分离物的10 x浓缩CFS显示出对艰难梭菌生长的抑制作用，与对照相比有显着差异（p <0.001）。敏捷乳酸杆菌T99A和丁酸梭菌选择T58A分离株以进一步评估产生的抗菌素。在用蛋白酶K进行酶处理或加热至90°C或中和pH值后，两个分离株的10x CFS的抗菌活性稳定。使用不同的革兰氏阳性和革兰氏阴性细菌物种评估了两种分离物的活性谱，并且没有显示出针对这些物种的抗菌活性。我们的研究结果显示有两个非传统细菌分离：L. AGILIS T99A和丁酸梭菌T58A生产外热稳定对抗菌药物艰难梭菌临床分离株。