Bacteriophage ϕMAM1, a Viunalikevirus, Is a Broad-Host-Range, High-Efficiency Generalized Transducer That Infects Environmental and Clinical Isolates of the Enterobacterial Genera Serratia and Kluyvera

Abstract

Members of the enterobacterial genus Serratia are ecologically widespread, and some strains are opportunistic human pathogens. Bacteriophage MAM1 was isolated on Serratia plymuthica A153, a biocontrol rhizosphere strain that produces the potently bioactive antifungal and anticancer haterumalide oocydin A. The MAM1 phage is a generalized transducing phage that infects multiple environmental and clinical isolates of Serratia spp. and a rhizosphere strain of Kluyvera cryocrescens. Electron microscopy allowed classification of MAM1 in the family Myoviridae. Bacteriophage MAM1 is virulent, uses capsular polysaccharides as a receptor, and can transduce chromosomal markers at frequencies of up to 7 ؋ 10 ؊6 transductants per PFU. We also demonstrated transduction of the complete 77-kb oocydin A gene cluster and heterogeneric transduction of a plasmid carrying a type III toxin-antitoxin system. These results support the notion of the potential ecological importance of transducing phages in the acquisition of genes by horizontal gene transfer. Phylogenetic analyses grouped MAM1 within the ViI-like bacteriophages, and genomic analyses revealed that the major differences between MAM1 and other ViI-like phages arise in a region encoding the host recognition determinants. Our results predict that the wider genus of ViI-like phages could be efficient transducing phages, and this possibility has obvious implications for the ecology of horizontal gene transfer, bacterial functional genomics, and synthetic biology.

Biosynthesis of the acetyl‐CoA carboxylase‐inhibiting antibiotic, andrimid in Serratia is regulated by Hfq and the LysR‐type transcriptional regulator, AdmX

“…To identify the genes responsible for this bioactivity, a random transposon insertion strain library was constructed and screened for mutants defective in antibacterial activity against Bacillus subtilis . Several transposon insertion mutants showing loss of antibacterial properties were isolated and all the insertions were transduced back into the wild type genetic background using the transducing phage ϕMAM1 (Matilla and Salmond, ) to confirm the link between transposon insertions and mutant phenotype. Random primed PCR confirmed that most of the transposons were located in a hybrid PKS/NRPS gene cluster described previously as responsible for the biosynthesis of the broad‐spectrum antibiotic, andrimid (Figs.…”

“…In total, three thousand kanamycin‐resistant insertion mutants were screened for their antibacterial activity against Bacillus subtilis using dual drop culture bioassays. Auxotrophic mutants were discarded and insertion mutations were transduced into the wild type strain A153 using phage ϕMAM1 (Matilla and Salmond, ). The insertion site of transposon Tn‐KRCPN1 in mutants of interest was determined using random primed PCR following the method described previously (Matilla et al ., ) and using primers described in Supporting Information Table S3.…”

“…The generalized transducing viunalikevirus, ϕMAM1, was used for transduction of chromosomal mutations, as described previously (Matilla and Salmond, ).…”

Horizontal transfer of antibiotic resistance genes in clinical environments

“…Bacteriophages were able to transduce a 5667 bp plasmid containing tetracycline and aminoglycoside resistance genes between S. aureus strains in the laboratory (Groisman and Ochman 1993;Zeman et al 2017). Bacteriophages were also able to transduce a 5620 bp plasmid containing a kanamycin resistance gene between Serratia and Kluyvera species, albeit at a lower rate than transduction of chromosomal resistance genes (Matilla and Salmond 2014). Chloramphenicol and tetracycline resistance genes on 27 kb plasmids were also transduced by bacteriophage between Actinobacillus (Willi et al 1997).…”

<i>In vivo</i> cloning of large chromosomal segments into a BAC derivative by generalized transduction and recombineering in <i>Salmonella enterica</i>

“…Likewise, a similar method could be applied to many other bacteria, such as Vibrio cholera (Guidolin and Manning, 1987), Pseudomonas aeruginosa (Budzik et al, 2004), and Serratia spp. (Matilla and Salmond, 2014), where generalized transducing phage are available. Alternatively, or in addition, this could be extended for the subcloning of BAC clones into pAK1002 or other plasmids containing an FRT site in a recA -mutant strain expressing Flp.…”