This study is intended to determine the antimicrobial resistance properties and class 1 integrons of 30 Citrobacter freundii strains isolated from pet turtles in order to determine their threat to human health. Citrobacter freundii isolates were isolated and identified by employing biochemical tests and 16SrRNA gene sequencing. Disc diffusion test and PCR amplification were employed to detect antimicrobial resistance patterns and genes, respectively. Ampicillin, amoxicillin, cephalothin and nalidixic acid resistance were observed among all isolates. Rifampicin and cefoxitin resistance was the second most prevalent and observed in 97% of the isolates, respectively. All isolates scored multiple antimicrobial resistance (MAR) indices ≥ 0.2. Aminoglycoside resistant genes were the most prevalent. aac(3')-IIa was detected in 80% of the isolates followed by aphAI-IAB and strA-strB genes in 33% and 50% of the isolates, respectively. β-lactamase encoding blaTEM, blaCTX and blaSHV genes were detected in 53%, 43% and 17% of the isolates, respectively. tetA and tetB genes were the only tetracycline resistance genes detected in 17% and 10% of the isolates, respectively. Class 1 integron integrase encoding intI1 gene (47%) and dfrA17-aadA5 gene cassette array (7%) were also detected. Pet turtle-borne multidrug-resistant C. freundii is a reservoir of antimicrobial resistance determinants in the domestic environment which poses a risk of infection.

Earthworms, especially Eisenia andrei, are cultivated for a variety of purposes including waste disposal and compost production. In this study, bacteria from earthworm gut were cultured and the dominant species identified. Subsequently, we isolated bacteriophages able to lyse the isolated gut bacteria. Two dominant genera of gut bacteria, Aeromonas sp. and Citrobacter sp. were identified by using MALDI-TOF MS analysis and a library was constructed to find lytic phages. Phage EF1 showed lytic activity for C. freundii and two Citrobacter isolates, C. braakii and C. murliniae. These 3 species have similar sensitivities to EF1. Several aspects of the life cycle of EF1 were investigated by using C. freundii under optimal growth conditions. EF1 infects C. freundii with a moderate latent period, approximately 25 min, and a large burst size averaging 5 × 109 per infected cell. Moreover, the antimicrobial activity of EF1 was well maintained under diverse conditions including a broad temperature range of 40°C to 50°C and a wide pH range of 4 to 11. In conclusion, the results indicate that earthworm casting contain a wide range of bacteria species, for which there are various corresponding bacteriophages.