Comparative analysis of multiple inducible phages from Mannheimia haemolytica

Abstract

Background: Mannheimia haemolytica is a commensal bacterium that resides in the upper respiratory tract of cattle that can play a role in bovine respiratory disease. Prophages are common in the M. haemolytica genome and contribute significantly to host diversity. The objective of this research was to undertake comparative genomic analysis of phages induced from strains of M. haemolytica serotype A1 (535A and 2256A), A2 (587A and 1127A) and A6 (1152A and 3927A).

Results: Overall, four P2-like (535AP1, 587AP1, 1127AP1 and 2256AP1; genomes: 34.9-35.7 kb; G+C content: 41.5-42.1 %; genes: 51-53 coding sequences, CDSs), four λ-like (535AP2, 587AP2, 1152AP2 and 3927AP1; genomes: 48.6-52.1 kb; 41.1-41.4 % mol G+C; genes: 77-83 CDSs and 2 tRNAs) and one Mu-like (3927AP2; genome: 33.8 kb; 43.1 % mol G+C; encoding 50 CDSs) phages were identified. All P2-like phages are collinear with the temperate phage φMhaA1-PHL101 with 535AP1, 2256AP1 and 1152AP1 being most closely related, followed by 587AP1 and 1127AP1. Lambdoid phages are not collinear with any other known λ-type phages, with 587AP2 being distinct from 535AP2, 3927AP1 and 1152AP2. All λ-like phages contain genes encoding a toxin-antitoxin (TA) system and cell-associated haemolysin XhlA. The Mu-like phage induced from 3927A is closely related to the phage remnant φMhaMu2 from M. haemolytica PHL21, with similar Mu-like phages existing in the genomes of M. haemolytica 535A and 587A.

Conclusions: This is among the first reports of both λ- and Mu-type phages being induced from M. haemolytica. Compared to phages induced from commensal strains of M. haemolytica serotype A2, those induced from the more virulent A1 and A6 serotypes are more closely related. Moreover, when P2-, λ- and Mu-like phages co-existed in the M. haemolytica genome, only P2- and λ-like phages were detected upon induction, suggesting that Mu-type phages may be more resistant to induction. Toxin-antitoxin gene cassettes in λ-like phages may contribute to their genomic persistence or the establishment of persister subpopulations of M. haemolytica. Further work is required to determine if the cell-associated haemolysin XhlA encoded by λ-like phages contributes to the pathogenicity and ecological fitness of M. haemolytica.

Fig. 1

Absorbance of dual wavelength (450–600 nm) measures of log-phase Mannheimia haemolytica strains. Strains were induced at 0 h with mitomycin C (●) or untreated (○). Values are means with average SD error bar on the final point, n = 7

Fig. 2

Whole genome comparisons of φMhaA1-PHL101 and all-P2 like Mannheimia haemolytica phages as well as using a progressive MAUVE alignment. The degree of sequence similarity is indicated by the intensity of the red region. The contiguous black boxes under the red region represent the position of genes. a, PHL101; b, 1152AP1; c, 535AP1; d, 2256AP1; e, 587AP1; f, 1127AP1

Fig. 3

Genomic structure and nucleotide sequence comparison of λ-like Mannheimia haemolytica phages. Genetic map was constructed using Easyfig [67]. The four genomes were aligned using a progressive MAUVE alignment and the degree of sequence similarity is indicated by the intensity of the red region

Fig. 4

Genomic structure of Mannheimia haemolytica phage 3927AP2 and comparison with related Mu-like phages. Genetic map was constructed using Easyfig [67]. The four genomes were aligned using a progressive MAUVE alignment and the degree of sequence similarity is indicated by the intensity of the red region