Phenotypic Characterization and Whole-Genome Analysis of a Novel Bacteriophage HCF1 Infecting Citrobacter amalonaticus and C. freundii

Abstract

Citrobacter species often occur in sewage, food, soil, wastewater, and in the intestinal tract of animals and humans. Citrobacter spp. cause urinary tract infections (UTIs) and infantile meningitis in humans. Due to the presence of plasmid-encoded resistance genes, Citrobacter spp. are often resistant to many antibiotics. In this study, Citrobacter virus HCF1, a novel virulent bacteriophage capable of killing Citrobacter amalonaticus and Citrobacter freundii, was isolated from the sewage water. The isolated bacteriophage was characterized with respect to transmission electron microscopy, one-step growth curve, host range, in vitro efficacy, storage stability, and environmental stress tolerance. The one-step growth curve analysis revealed that the latent period of HCF1 was 30 min and the estimated burst size was 121 plaque-forming units (PFU) per bacterial cell. Host range testing indicated that the HCF1 was specific to the Citrobacter genus. In vitro efficacy assay in the effluent of an anaerobic biodigester showed that the HCF1 completely eliminated the host within 4 and 5 h at MOI:100 and MOI:10, respectively, thereby indicating its potential for combating C. amalonaticus infections. The isolated bacteriophage is considerably stable and tolerant to environmental stress. Furthermore, the complete genome of HCF1 was sequenced using Oxford Nanopore sequencing and the data were subjected to detailed bioinformatic analyses. NCBI-BLASTn analysis revealed that the HCF1 genome had a query coverage of 15–21% and a maximum similarity of 77.27–78.49% with 11 bacteriophages of the Drexlerviridae family. Detailed bioinformatic analysis of the genome profile suggests that HCF1 is a novel T1svirus belonging to the Tempevirinae subfamily of the Drexlerviridae family.

A Novel Disease (Water Bubble Disease) of the Giant Freshwater Prawn Macrobrachium rosenbergii Caused by Citrobacter freundii: Antibiotic Treatment and Effects on the Antioxidant Enzyme Activity and Immune Responses

“…Phylogenetic analysis revealed that our isolated strain GDZQ201912 showed a very close association with other strains of C. freundii submitted from China and UK and formed separate clades with other Citrobacter sp., such as C. portucalensis (AP022399.1, AP022486.1, and AP022513.1). The physiological and biochemical profiles of strain GDZQ201912 were similar to those of other isolates reported by earlier works [ 49 , 50 ]. Variations in the biochemical test, such as those of tyrosine, putrescine [ 38 ], indole, citrate, H 2 S, malonate, raffinose, and dulcitol, were also described by earlier studies [ 17 , 49 ].…”

“…In the case of pathogenic bacteria, such as C. freundii , multidrug resistance is one of the most prominent problems [ 50 ]. In this study, the isolated strain was resistant to four different β-lactam antibiotics, including ampicillin, clindamycin, lincomycin, and penicillin.…”

Isolation, phenotypic characterization and comparative genomic analysis of 2019SD1, a polyvalent enterobacteria phage

“…This was confirmed in another study which reported that pSf-2 phage produced clear plaques only in S. flexneri cultures (ATCC 12022, 11 836 and 29903) 56 . Similarly, a more recently described phage (HCF1) 58 belonging to Drexlerviridae family displayed lytic activity against two Citrobacter species ( C. freundii and C. amalanoticus ) but not against S. dysenteriae. In contrast, Hamdi et al 55 reported that S. flexneri and E. coli can be lysed by Drexlerviridae phage SH6 and Sfin-1.…”

The Bacteriophage vB_CbrM_HP1 Protects Crucian Carp Against Citrobacter braakii Infection

“…For example, from 2015 to 2019, the Kuty Everett GF team uploaded 9 complete genome sequences of C. freundii phage (Moogle, Merlin, Maleficent, Maroon, Mijalis, Moon, Michonne, Mordin, and Miller) (46)(47)(48)(49)(50)(51)(52)(53)(54), which were similar to vB_CbrM_HP1, but they did not further study their biological characteristics or antibacterial activities in vivo or in vitro. Only five Citrobacter phages (C. freundii phage IME-JL8, LK1, phiCFP-1, CfP1, and HCF1) have been characterized thus far (19)(20)(21)(22)(23). Research suggested that they were Siphoviridae phages (IME-JL8 and HCF1), Podoviridae phages (LK1 and phiCFP-1), or a Myovirus phage (CfP1).…”

“…There are approximately 40 Citrobacter phages published in PubMed. Only five Citrobacter phages (C. freundii phage IME-JL8, LK1, phiCFP-1, CfP1, and HCF1) have been characterized thus far (19)(20)(21)(22)(23). C. freundii phage IME-JL8 was used to study the antimicrobial activity in carp both in vivo and in vitro (19).…”