Carbapenem-resistant Acinetobacter baumannii is responsible for frequent, hard-to-treat and often fatal healthcare-associated infections. Phage therapy, the use of viruses that infect and kill bacteria, is an approach gaining significant clinical interest to combat antibiotic-resistant infections. However, a major limitation is that bacteria can develop resistance against phages. Here, we isolated phages with activity against a panel of A.baumannii strains and focused on clinical isolates AB900 and A9844 and their phages for detailed characterization. As expected, coincubation of the phages with their hosts in vitro resulted in the emergence of phage-resistant bacterial mutants. Genome sequence analysis revealed that phage-resistant mutants harbored loss-of-function mutations in genes from the K locus, responsible for the biosynthesis of the bacterial capsule.Using molecular biology techniques, phage adsorption assays, and quantitative evaluation of capsule production, we established that the bacterial capsule serves as the primary receptor for these phages. As a collateral phenotype of impaired capsule production, the phage-resistant strains could not form biofilms, became fully sensitized to the human complement system, showed increased susceptibility to beta-lactam antibiotics, and became vulnerable to additional phages. Finally, in a murine model of bacteremia, the phage-resistant A.baumannii demonstrated a diminished capacity to colonize blood and solid tissues. This study demonstrates that phages can be used not only for their lytic activity but, if combined with a posteriori knowledge of their receptors and the mechanism of bacterial resistance, for their potential synergy with other antimicrobial agents, thus providing even broader clinical options for phage therapy.

The large and complex genome of Pseudomonas aeruginosa, which consists of significant portions (up to 20%) of transferable genetic elements contributes to the rapid development of antibiotic resistance. The whole genome sequences of 22 strains isolated from eye and cystic fibrosis patients in Australia and India between 1992 and 2007 were used to compare genomic divergence and phylogenetic relationships as well as genes for antibiotic resistance and virulence factors. Analysis of the pangenome indicated a large variation in the size of accessory genome amongst 22 stains and the size of the accessory genome correlated with number of genomic islands, insertion sequences and prophages. The strains were diverse in terms of sequence type and dissimilar to that of global epidemic P. aeruginosa clones. Of the eye isolates, 62% clustered together within a single lineage. Indian eye isolates possessed genes associated with resistance to aminoglycoside, beta-lactams, sulphonamide, quaternary ammonium compounds, tetracycline, trimethoprims and chloramphenicols. These genes were, however, absent in Australian isolates regardless of source. Overall, our results provide valuable information for understanding the genomic diversity of P. aeruginosa isolated from two different infection types and countries.

Treatment of Pseudomonas aeruginosa eye infections often becomes a challenge due to the ability of this bacterium to be resistant to antibiotics via intrinsic and acquired mechanisms. Transfer of resistance due to interchangeable genetic elements is an important mechanism for the rapid transfer of antibiotic resistance in this pathogen. As a result, drug-resistant strains are becoming increasingly prevalent worldwide. This review systematically analyses data from recent publications to describe the global prevalence and antibiotic sensitivity of ocular P. aeruginosa. Thirty-seven studies were selected for review from PubMed-based searches using the criteria 'microbial keratitis OR eye infection AND Pseudomonas aeruginosa AND antibiotic resistance' and limiting to papers from 2011 onward, to demonstrate the antibiotic resistance from isolates from around the world. Subsequently, we reviewed the ways in which P. aeruginosa can become resistant to antibiotics. Both the rate of isolation of bacteria in general (79 per cent of cases), and prevalence of P. aeruginosa (68 per cent of all isolates) were highest in contact lens-related microbial keratitis. The average resistance rate to common ocular antibiotics such as ciprofloxacin (9 per cent), gentamicin (22 per cent) and ceftazidime (13 per cent) remained relatively low. However, there were large variations in resistance rates reported in studies from different countries, for example resistance to ciprofloxacin reached up to 33 per cent. We next reviewed the types of mobile genetic elements (MGEs) such as plasmids, integrons and transposons that are frequently associated with drug resistance in P. aeruginosa. MGEs are important for the transmission of resistance to beta-lactams and aminoglycosides and recently have been shown to be potential factors for the transmission of fluoroquinolone resistance. Studies on the molecular mechanisms of resistance transfer in ocular P. aeruginosa have begun to be reported and will provide valuable information on the emergence of new antibiotic resistance and potential to treat resistant strains.

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Acanthamoeba, an opportunistic pathogen is known to cause an infection of the cornea, central nervous system, and skin. Acanthamoeba feeds different microorganisms, including potentially pathogenic prokaryotes; some of microbes have developed ways of surviving intracellularly and this may mean that Acanthamoeba acts as incubator of important pathogens. A systematic review of the literature was performed in order to capture a comprehensive picture of the variety of microbial species identified within Acanthamoeba following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. Forty-three studies met the inclusion criteria, 26 studies (60.5%) examined environmental samples, eight (18.6%) studies examined clinical specimens, and another nine (20.9%) studies analysed both types of samples. Polymerase chain reaction (PCR) followed by gene sequencing was the most common technique used to identify the intracellular microorganisms. Important pathogenic bacteria, such as E. coli, Mycobacterium spp. and P. aeruginosa, were observed in clinical isolates of Acanthamoeba, whereas Legionella, adenovirus, mimivirus, and unidentified bacteria (Candidatus) were often identified in environmental Acanthamoeba. Increasing resistance of Acanthamoeba associated intracellular pathogens to antimicrobials is an increased risk to public health. Molecular-based future studies are needed in order to assess the microbiome residing in Acanthamoeba, as a research on the hypotheses that intracellular microbes can affect the pathogenicity of Acanthamoeba infections.