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12 pages, 1808 KiB

Open AccessArticle

A Bioluminescence-Based Ex Vivo Burn Wound Model for Real-Time Assessment of Novel Phage-Inspired Enzybiotics

by Vincent De Maesschalck, Diana Gutiérrez, Jan Paeshuyse, Yves Briers, Greetje Vande Velde and Rob Lavigne

Pharmaceutics 2022, 14(12), 2553; https://0-doi-org.brum.beds.ac.uk/10.3390/pharmaceutics14122553 - 22 Nov 2022

Cited by 1 | Viewed by 1643

Abstract

The silent pandemic of antibiotic resistance is thriving, prompting the urgent need for the development of new antibacterial drugs. However, within the preclinical pipeline, in vitro screening conditions can differ significantly from the final in vivo settings. To bridge the gap between in [...] Read more.

The silent pandemic of antibiotic resistance is thriving, prompting the urgent need for the development of new antibacterial drugs. However, within the preclinical pipeline, in vitro screening conditions can differ significantly from the final in vivo settings. To bridge the gap between in vitro and in vivo assays, we developed a pig-skin-based bioluminescent ex vivo burn wound infection model, enabling real-time assessment of antibacterials in a longitudinal, non-destructive manner. We provide a proof-of-concept for A. baumannii NCTC13423, a multidrug-resistant clinical isolate, which was equipped with the luxCDABE operon as a reporter using a Tn7-based tagging system. This bioluminescence model provided a linear correlation between the number of bacteria and a broad dynamic range (10⁴ to 10⁹ CFU). This longitudinal model was subsequently validated using a fast-acting enzybiotic, 1D10. Since this model combines a realistic, clinically relevant yet strictly controlled environment with real-time measurement of bacterial burden, we put forward this ex vivo model as a valuable tool to assess the preclinical potential of novel phage-inspired enzybiotics. Full article

(This article belongs to the Special Issue Bacteriophages, Critical Biocatalysts of Bacterial Doom: The Dawn of a New Era in the Fight against Multidrug Resistant Bacteria)

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40 pages, 8937 KiB

Open AccessArticle

Isolation and Molecular Characterization of Two Novel Lytic Bacteriophages for the Biocontrol of Escherichia coli in Uterine Infections: In Vitro and Ex Vivo Preliminary Studies in Veterinary Medicine

by Victor M. Balcão, Bianca G. Belline, Erica C. Silva, Pablo F. F. B. Almeida, Denicezar Â. Baldo, Lara R. P. Amorim, José M. Oliveira Júnior, Marta M. D. C. Vila and Fernando S. Del Fiol

Pharmaceutics 2022, 14(11), 2344; https://0-doi-org.brum.beds.ac.uk/10.3390/pharmaceutics14112344 - 30 Oct 2022

Cited by 8 | Viewed by 2272

Abstract

E. coli is one of the etiological agents responsible for pyometra in female dogs, with conventional treatment involving ovariohysterectomy. Here, we report the isolation and full characterization of two novel lytic phages, viz. vB_EcoM_Uniso11 (ph0011) and vB_EcoM_Uniso21 (ph0021). Both phages belong to the [...] Read more.

E. coli is one of the etiological agents responsible for pyometra in female dogs, with conventional treatment involving ovariohysterectomy. Here, we report the isolation and full characterization of two novel lytic phages, viz. vB_EcoM_Uniso11 (ph0011) and vB_EcoM_Uniso21 (ph0021). Both phages belong to the order Caudovirales and present myovirus-like morphotypes, with phage ph0011 being classified as Myoviridae genus Asteriusvirus and phage ph0021 being classified as Myoviridae genus Tequatrovirus, based on their complete genome sequences. The 348,288 bp phage ph0011 and 165,222 bp phage ph0021 genomes do not encode toxins, integrases or antimicrobial resistance genes neither depolymerases related sequences. Both phages were shown to be effective against at least twelve E. coli clinical isolates in in vitro antibacterial activity assays. Based on their features, both phages have potential for controlling pyometra infections caused by E. coli. Phage ph0011 (reduction of 4.24 log CFU/mL) was more effective than phage ph0021 (reduction of 1.90 log CFU/mL) after 12 h of incubation at MOI 1000. As a cocktail, the two phages were highly effective in reducing the bacterial load (reduction of 5.57 log CFU/mL) at MOI 100, after 12 h of treatment. Both phages were structurally and functionally stabilized in vaginal egg formulations. Full article

(This article belongs to the Special Issue Bacteriophages, Critical Biocatalysts of Bacterial Doom: The Dawn of a New Era in the Fight against Multidrug Resistant Bacteria)

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16 pages, 4861 KiB

Open AccessArticle

Novel Stenotrophomonas maltophilia Bacteriophage as Potential Therapeutic Agent

by Rima Fanaei Pirlar, Jeroen Wagemans, Fabian Kunisch, Rob Lavigne, Andrej Trampuz and Mercedes Gonzalez Moreno

Pharmaceutics 2022, 14(10), 2216; https://0-doi-org.brum.beds.ac.uk/10.3390/pharmaceutics14102216 - 18 Oct 2022

Cited by 5 | Viewed by 2055

Abstract

A novel bacteriophage CUB19 specific to the bacterial species Stenotrophomonas maltophilia was isolated from hospital sewage and characterized as a new species belonging to a proposed new phage genus ‘Cubvirus’ (Caudoviricetes). Its genome contains a total of 48,301 bp and 79 [...] Read more.

A novel bacteriophage CUB19 specific to the bacterial species Stenotrophomonas maltophilia was isolated from hospital sewage and characterized as a new species belonging to a proposed new phage genus ‘Cubvirus’ (Caudoviricetes). Its genome contains a total of 48,301 bp and 79 predicted genes, among which some have been associated with packaging and lysis-associated proteins, structural proteins, or DNA- and metabolism-associated proteins. No lysogeny-associated proteins or known virulence proteins were identified on the phage genome. CUB19 showed stability over a wide range of temperatures (−20 °C–60 °C) and pH values (pH 3–pH 13). Despite its narrow host range, this phage has potent observed antimicrobial and antibiofilm activity. A time-killing curve assay showed significant biofilm reduction after 24 h exposure to CUP19. Isothermal microcalorimetry assays investigating phage-antibiotic combinations revealed the effectiveness of CUB19 during co-administration with increasing antibiotic doses, regardless of the administration approach (simultaneous or staggered). These are encouraging indications for its application as a targeted therapeutic agent against resilient biofilm-associated Stenotrophomonas infections. Full article

(This article belongs to the Special Issue Bacteriophages, Critical Biocatalysts of Bacterial Doom: The Dawn of a New Era in the Fight against Multidrug Resistant Bacteria)

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20 pages, 4594 KiB

Open AccessArticle

Characterization of Novel Bacteriophage vB_KpnP_ZX1 and Its Depolymerases with Therapeutic Potential for K57 Klebsiella pneumoniae Infection

by Ping Li, Wenjie Ma, Jiayin Shen and Xin Zhou

Pharmaceutics 2022, 14(9), 1916; https://0-doi-org.brum.beds.ac.uk/10.3390/pharmaceutics14091916 - 10 Sep 2022

Cited by 9 | Viewed by 1929

Abstract

A novel temperate phage vB_KpnP_ZX1 was isolated from hospital sewage samples using the clinically derived K57-type Klebsiella pneumoniae as a host. Phage vB_KpnP_ZX1, encoding three lysogen genes, the repressor, anti-repressor, and integrase, is the fourth phage of the genus Uetakevirus, family Podoviridae [...] Read more.

A novel temperate phage vB_KpnP_ZX1 was isolated from hospital sewage samples using the clinically derived K57-type Klebsiella pneumoniae as a host. Phage vB_KpnP_ZX1, encoding three lysogen genes, the repressor, anti-repressor, and integrase, is the fourth phage of the genus Uetakevirus, family Podoviridae, ever discovered. Phage vB_KpnP_ZX1 did not show ideal bactericidal effect on K. pneumoniae 111-2, but TEM showed that the depolymerase Dep_ZX1 encoded on the short tail fiber protein has efficient capsule degradation activity. In vitro antibacterial results show that purified recombinant Dep_ZX1 can significantly prevent the formation of biofilm, degrade the formed biofilm, and improve the sensitivity of the bacteria in the biofilm to the antibiotics kanamycin, gentamicin, and streptomycin. Furthermore, the results of animal experiments show that 50 µg Dep_ZX1 can protect all K. pneumoniae 111-2-infected mice from death, whereas the control mice infected with the same dose of K. pneumoniae 111-2 all died. The degradation activity of Dep_ZX1 on capsular polysaccharide makes the bacteria weaken their resistance to immune cells, such as complement-mediated serum killing and phagocytosis, which are the key factors for its therapeutic action. In conclusion, Dep_ZX1 is a promising anti-virulence agent for the K57-type K. pneumoniae infection or biofilm diseases. Full article

(This article belongs to the Special Issue Bacteriophages, Critical Biocatalysts of Bacterial Doom: The Dawn of a New Era in the Fight against Multidrug Resistant Bacteria)

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14 pages, 1827 KiB

Open AccessArticle

In Vitro Evaluation of Five Newly Isolated Bacteriophages against E. faecalis Biofilm for Their Potential Use against Post-Treatment Apical Periodontitis

by Marie Voit, Andrej Trampuz and Mercedes Gonzalez Moreno

Pharmaceutics 2022, 14(9), 1779; https://0-doi-org.brum.beds.ac.uk/10.3390/pharmaceutics14091779 - 25 Aug 2022

Cited by 5 | Viewed by 1831

Abstract

State-of-the-art treatment of root canal infection includes the use of mechanical debridement and chemical agents. This disinfection method is limited, and microorganisms can remain in the canal system. Enterococcus faecalis appears with a high prevalence in secondary and persistent root canal infections and [...] Read more.

State-of-the-art treatment of root canal infection includes the use of mechanical debridement and chemical agents. This disinfection method is limited, and microorganisms can remain in the canal system. Enterococcus faecalis appears with a high prevalence in secondary and persistent root canal infections and can be linked to endodontic treatment failure due to its various resistance mechanisms. Here, we evaluated the activity of newly isolated bacteriophages against clinical isolates of E. faecalis (including one vancomycin- and gentamicin-resistant strain) as a single treatment or in combination with gentamicin and vancomycin. For the resistant strain, daptomycin and fosfomycin were tested. Sixteen E. faecalis strains were used to screen for the presence of bacteriophages in sewage. Five different bacteriophages were characterized in terms of virion morphology, host range and killing-kinetics against each E. faecalis host strain. To investigate the antibiofilm effect of antibiotic and phages, E. faecalis biofilm was grown on porous glass beads and treated with different antibiotic concentrations and with isolated bacteriophages alone or in staggered combinations. A strong biofilm reduction was observed when phages were combined with antibiotic, where combinations with gentamicin showed a better outcome compared to vancomycin. Regarding the resistant strain, daptomycin had a superior antibiofilm effect than fosfomycin. Full article

(This article belongs to the Special Issue Bacteriophages, Critical Biocatalysts of Bacterial Doom: The Dawn of a New Era in the Fight against Multidrug Resistant Bacteria)

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11 pages, 1995 KiB

Open AccessArticle

Genomic and Functional Characterization of Vancomycin-Resistant Enterococci-Specific Bacteriophages in the Galleria mellonella Wax Moth Larvae Model

by Lynn El Haddad, Georgios Angelidakis, Justin R. Clark, Jesus F. Mendoza, Austen L. Terwilliger, Christopher P. Chaftari, Mark Duna, Serena T. Yusuf, Cynthia P. Harb, Mark Stibich, Anthony Maresso and Roy F. Chemaly

Pharmaceutics 2022, 14(8), 1591; https://0-doi-org.brum.beds.ac.uk/10.3390/pharmaceutics14081591 - 30 Jul 2022

Cited by 7 | Viewed by 2192

Abstract

Phages are naturally occurring viruses that selectively kill bacterial species without disturbing the individual’s normal flora, averting the collateral damage of antimicrobial usage. The safety and the effectiveness of phages have been mainly confirmed in the food industry as well as in animal [...] Read more.

Phages are naturally occurring viruses that selectively kill bacterial species without disturbing the individual’s normal flora, averting the collateral damage of antimicrobial usage. The safety and the effectiveness of phages have been mainly confirmed in the food industry as well as in animal models. In this study, we report on the successful isolation of phages specific to Vancomycin-resistant Enterococci, including Enterococcus faecium (VREfm) and Enterococcus faecalis from sewage samples, and demonstrate their efficacy and safety for VREfm infection in the greater wax moth Galleria mellonella model. No virulence-associated genes, antibiotic resistance genes or integrases were detected in the phages’ genomes, rendering them safe to be used in an in vivo model. Phages may be considered as potential agents for therapy for bacterial infections secondary to multidrug-resistant organisms such as VREfm. Full article

(This article belongs to the Special Issue Bacteriophages, Critical Biocatalysts of Bacterial Doom: The Dawn of a New Era in the Fight against Multidrug Resistant Bacteria)

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21 pages, 3835 KiB

Open AccessArticle

Isolation and Molecular Characterization of a Novel Lytic Bacteriophage That Inactivates MDR Klebsiella pneumoniae Strains

by Victor M. Balcão, Fernanda C. Moreli, Erica C. Silva, Bianca G. Belline, Layla F. Martins, Fernando P. N. Rossi, Carla Pereira, Marta M. D. C. Vila and Aline M. da Silva

Pharmaceutics 2022, 14(7), 1421; https://0-doi-org.brum.beds.ac.uk/10.3390/pharmaceutics14071421 - 06 Jul 2022

Cited by 13 | Viewed by 2610

Abstract

The worldwide increase in serious infections caused by multidrug-resistant (MDR) K. pneumoniae emphasizes the urgent need of new therapeutic strategies for the control of this pathogen. There is growing interest in the use of bacteriophages (or phages) to treat K. pneumoniae infections, and [...] Read more.

The worldwide increase in serious infections caused by multidrug-resistant (MDR) K. pneumoniae emphasizes the urgent need of new therapeutic strategies for the control of this pathogen. There is growing interest in the use of bacteriophages (or phages) to treat K. pneumoniae infections, and newly isolated phages are needed. Here, we report the isolation and physical/biological/molecular characterization of a novel lytic phage and its efficacy in the control of MDR K. pneumoniae. The phage vB_KpnS_Uniso31, referred to hereafter as phage Kpn31, was isolated from hospital wastewater using K. pneumoniae CCCD-K001 as the host. Phage Kpn31 presents a siphovirus-like morphotype and was classified as Demerecviridae; Sugarlandvirus based on its complete genome sequence. The 113,444 bp Kpn31 genome does not encode known toxins or antimicrobial resistance genes, nor does it encode depolymerases related sequences. Phage Kpn31 showed an eclipse time of 15 min and a burst size of 9.12 PFU/host cell, allowing us to conclude it replicates well in K. pneumoniae CCCD-K001 with a latency period of 30 min. Phage Kpn31 was shown to be effective against at least six MDR K. pneumoniae clinical isolates in in vitro antibacterial activity assays. Based on its features, phage Kpn31 has potential for controlling infections caused by MDR K. pneumoniae. Full article

(This article belongs to the Special Issue Bacteriophages, Critical Biocatalysts of Bacterial Doom: The Dawn of a New Era in the Fight against Multidrug Resistant Bacteria)

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Review

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17 pages, 1456 KiB

Open AccessReview

Bacteriophage and Bacterial Susceptibility, Resistance, and Tolerance to Antibiotics

by Qingquan Chen, Tejas Dharmaraj, Pamela C. Cai, Elizabeth B. Burgener, Naomi L. Haddock, Andy J. Spakowitz and Paul L. Bollyky

Pharmaceutics 2022, 14(7), 1425; https://0-doi-org.brum.beds.ac.uk/10.3390/pharmaceutics14071425 - 07 Jul 2022

Cited by 13 | Viewed by 4150

Abstract

Bacteriophages, viruses that infect and replicate within bacteria, impact bacterial responses to antibiotics in complex ways. Recent studies using lytic bacteriophages to treat bacterial infections (phage therapy) demonstrate that phages can promote susceptibility to chemical antibiotics and that phage/antibiotic synergy is possible. However, [...] Read more.

Bacteriophages, viruses that infect and replicate within bacteria, impact bacterial responses to antibiotics in complex ways. Recent studies using lytic bacteriophages to treat bacterial infections (phage therapy) demonstrate that phages can promote susceptibility to chemical antibiotics and that phage/antibiotic synergy is possible. However, both lytic and lysogenic bacteriophages can contribute to antimicrobial resistance. In particular, some phages mediate the horizontal transfer of antibiotic resistance genes between bacteria via transduction and other mechanisms. In addition, chronic infection filamentous phages can promote antimicrobial tolerance, the ability of bacteria to persist in the face of antibiotics. In particular, filamentous phages serve as structural elements in bacterial biofilms and prevent the penetration of antibiotics. Over time, these contributions to antibiotic tolerance favor the selection of resistance clones. Here, we review recent insights into bacteriophage contributions to antibiotic susceptibility, resistance, and tolerance. We discuss the mechanisms involved in these effects and address their impact on bacterial fitness. Full article

(This article belongs to the Special Issue Bacteriophages, Critical Biocatalysts of Bacterial Doom: The Dawn of a New Era in the Fight against Multidrug Resistant Bacteria)

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