No abstract

A multiplex PCR test based on the simultaneous amplification of two lipoprotein genes, oprI and oprL, was designed and evaluated for its ability to directly detect fluorescent pseudomonads (amplification of oprI open reading frame, 249 bp) and Pseudomonas aeruginosa (amplification of oprL open reading frame, 504 bp) in clinical material. A collection of reference strains including 20 different species of fluorescent pseudomonads was tested. Positive PCR results for both genes were observed only for P. aeruginosa isolates (n ‫؍‬ 150), including strains of clinical and environmental origin, while only one gene, oprI, was amplified from the other fluorescent pseudomonads. All other bacteria tested (n ‫؍‬ 15) were negative by the amplification test. The lower detection level for P. aeruginosa was estimated to be 10 2 cells/ml. Preliminary evaluation on testing skin biopsy specimens from patients with burns (n ‫؍‬ 14) and sputum samples from cystic fibrosis patients (n ‫؍‬ 49) and other patients (n ‫؍‬ 19) showed 100% sensitivity and 74% specificity in comparison with culture. This multiplex PCR assay appears promising for the rapid and sensitive detection of P. aeruginosa in clinical specimens. Further evaluation of its specificity in longitudinal clinical studies is warranted.

Like the sword of Damocles, the threat of a post-antibiotic era is hanging over humanity's head. The scientific and medical community is thus reconsidering bacteriophage therapy (BT) as a partial but realistic solution for treatment of difficult to eradicate bacterial infections. Here, we summarize the latest developments in clinical BT applications, with a focus on developments in the following areas: i) pharmacology of bacteriophages of major clinical importance and their synergy with antibiotics; ii) production of therapeutic phages; and iii) clinical trials, case studies, and case reports in the field. We address regulatory concerns, which are of paramount importance insofar as they dictate the conduct of clinical trials, which are needed for broader BT application. The increasing amount of new available data confirm the particularities of BT as being innovative and highly personalized. The current circumstances suggest that the immediate future of BT may be advanced within the framework of national BT centers in collaboration with competent authorities, which are urged to adopt incisive initiatives originally launched by some national regulatory authorities.

The emergence of multidrug resistant bacteria in both community-and hospitalacquired infections is recognized as a major public health threat. Phage therapy is increasingly mediatized and researched as an additional tool for combatting antibiotic resistant infections. However, phages exhibit a number of properties that differ from antibiotics and hamper their development as pharmaceutical products and their application in therapy. This paper advocates a paradigm shift in the development and application of infectious disease therapeutics to cater for personalized phage therapy, which could be realized by the year 2035. More specifically, it presents a sustainable and ethical supply chain of instant synthetic phages, based on a community effort, supported and steered by public health organizations, and managed by a platform combining Artificial Intelligence (AI) and Distributed Ledger (DL) Technology.

With rising antibiotic resistance, there has been increasing interest in treating pathogenic bacteria with bacteriophages (phage therapy). One limitation of phage therapy is the ease at which bacteria can evolve resistance. Negative effects of resistance may be mitigated when resistance results in reduced bacterial growth and virulence, or when phage coevolve to overcome resistance. Resistance evolution and its consequences are contingent on the bacteria-phage combination and their environmental context, making therapeutic outcomes hard to predict. One solution might be to conduct 'in vitro evolutionary simulations' using bacteria-phage combinations from the therapeutic context. Overall, our aim was to investigate parallels between in vitro experiments and in vivo dynamics in a human participant. Evolutionary dynamics were similar, with high levels of resistance evolving quickly with limited evidence of phage evolution. Resistant bacteria - evolved in vitro and in vivo - had lower virulence. In vivo, this was linked to lower growth rates of resistant isolates, whereas in vitro phage resistant isolates evolved greater biofilm production. Population sequencing suggests resistance resulted from selection on de novo mutations rather than sorting of existing variants. These results highlight the speed at which phage resistance can evolve in vivo, and how in vitro experiments may give useful insights for clinical evolutionary outcomes.