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17 pages, 1712 KiB

Open AccessArticle

Human Complement Inhibits Myophages against Pseudomonas aeruginosa

by Julia E. Egido, Simon O. Dekker, Catherine Toner-Bartelds, Cédric Lood, Suzan H. M. Rooijakkers, Bart W. Bardoel and Pieter-Jan Haas

Viruses 2023, 15(11), 2211; https://doi.org/10.3390/v15112211 - 3 Nov 2023

Cited by 4 | Viewed by 1824

Abstract

Therapeutic bacteriophages (phages) are primarily chosen based on their in vitro bacteriolytic activity. Although anti-phage antibodies are known to inhibit phage infection, the influence of other immune system components is less well known. An important anti-bacterial and anti-viral innate immune system that may [...] Read more.

Therapeutic bacteriophages (phages) are primarily chosen based on their in vitro bacteriolytic activity. Although anti-phage antibodies are known to inhibit phage infection, the influence of other immune system components is less well known. An important anti-bacterial and anti-viral innate immune system that may interact with phages is the complement system, a cascade of proteases that recognizes and targets invading microorganisms. In this research, we aimed to study the effects of serum components such as complement on the infectivity of different phages targeting Pseudomonas aeruginosa. We used a fluorescence-based assay to monitor the killing of P. aeruginosa by phages of different morphotypes in the presence of human serum. Our results reveal that several myophages are inhibited by serum in a concentration-dependent way, while the activity of four podophages and one siphophage tested in this study is not affected by serum. By using specific nanobodies blocking different components of the complement cascade, we showed that activation of the classical complement pathway is a driver of phage inhibition. To determine the mechanism of inhibition, we produced bioorthogonally labeled fluorescent phages to study their binding by means of microscopy and flow cytometry. We show that phage adsorption is hampered in the presence of active complement. Our results indicate that interactions with complement may affect the in vivo activity of therapeutically administered phages. A better understanding of this phenomenon is essential to optimize the design and application of therapeutic phage cocktails. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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23 pages, 4140 KiB

Open AccessArticle

Compounding Achromobacter Phages for Therapeutic Applications

by Ana Georgina Cobián Güemes, Tram Le, Maria Isabel Rojas, Nicole E. Jacobson, Helena Villela, Katelyn McNair, Shr-Hau Hung, Lili Han, Lance Boling, Jessica Claire Octavio, Lorena Dominguez, Vito Adrian Cantú, Sinéad Archdeacon, Alejandro A. Vega, Michelle A. An, Hamza Hajama, Gregory Burkeen, Robert A. Edwards, Douglas J. Conrad, Forest Rohwer and Anca M. Segall Show full author list Hide full author list

Viruses 2023, 15(8), 1665; https://doi.org/10.3390/v15081665 - 30 Jul 2023

Cited by 3 | Viewed by 2961

Abstract

Achromobacter species colonization of Cystic Fibrosis respiratory airways is an increasing concern. Two adult patients with Cystic Fibrosis colonized by Achromobacter xylosoxidans CF418 or Achromobacter ruhlandii CF116 experienced fatal exacerbations. Achromobacter spp. are naturally resistant to several antibiotics. Therefore, phages could be valuable [...] Read more.

Achromobacter species colonization of Cystic Fibrosis respiratory airways is an increasing concern. Two adult patients with Cystic Fibrosis colonized by Achromobacter xylosoxidans CF418 or Achromobacter ruhlandii CF116 experienced fatal exacerbations. Achromobacter spp. are naturally resistant to several antibiotics. Therefore, phages could be valuable as therapeutics for the control of Achromobacter. In this study, thirteen lytic phages were isolated and characterized at the morphological and genomic levels for potential future use in phage therapy. They are presented here as the Achromobacter Kumeyaay phage collection. Six distinct Achromobacter phage genome clusters were identified based on a comprehensive phylogenetic analysis of the Kumeyaay collection as well as the publicly available Achromobacter phages. The infectivity of all phages in the Kumeyaay collection was tested in 23 Achromobacter clinical isolates; 78% of these isolates were lysed by at least one phage. A cryptic prophage was induced in Achromobacter xylosoxidans CF418 when infected with some of the lytic phages. This prophage genome was characterized and is presented as Achromobacter phage CF418-P1. Prophage induction during lytic phage preparation for therapy interventions require further exploration. Large-scale production of phages and removal of endotoxins using an octanol-based procedure resulted in a phage concentrate of 1 × 10⁹ plaque-forming units per milliliter with an endotoxin concentration of 65 endotoxin units per milliliter, which is below the Food and Drugs Administration recommended maximum threshold for human administration. This study provides a comprehensive framework for the isolation, bioinformatic characterization, and safe production of phages to kill Achromobacter spp. in order to potentially manage Cystic Fibrosis (CF) pulmonary infections. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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23 pages, 7809 KiB

Open AccessArticle

Characterization and Genomic Analysis of a Bacteriophage with Potential in Lysing Vibrio alginolyticus

by Jingyun Fu, Ying Li, Lihong Zhao, Chunguang Wu and Zengguo He

Viruses 2023, 15(1), 135; https://doi.org/10.3390/v15010135 - 31 Dec 2022

Cited by 7 | Viewed by 2658

Abstract

Vibrio alginolyticus is one of the major pathogens causing vibriosis to a variety of aquatic animals as well as bringing about severe food safety concerns. Nowadays, phage therapy has received increasing attention as an alternative to the antibiotics that have being limited for use [...] Read more.

Vibrio alginolyticus is one of the major pathogens causing vibriosis to a variety of aquatic animals as well as bringing about severe food safety concerns. Nowadays, phage therapy has received increasing attention as an alternative to the antibiotics that have being limited for use in aquaculture industries. In this work, a potent bacteriophage, vB_ValM_PVA23 (PVA23), which efficiently infects pathogenic strains of V. alginolyticus, was isolated from sewage water and characterized by microbiological and genomic analyses. Based on the transmission electronic observation, the phage was characterized to be the Myoviridae family. It has a latent period of 10 min and a burst size of 203 PFUs/infected bacterium, and was stable over a broad pH range (5.0–11.0) and a wide temperature span (−80 °C to 60 °C), respectively. Genome sequencing results show that PVA23 has a 246,962-bp double-stranded DNA with a G + C content of 41.25%. The lab and plant shrimp farming trials demonstrated that phage preparation derived from PVA23 out-performed the chemical disinfectant iodine treatment in the prevention of V. alginolyticus propagation, and the phage application could rapidly yet significantly reduce the level of V. alginolyticus in the pond within 12 h, with negligible rebound observed. These results suggests that phage PVA23 has the potential to be used as an anti-V. alginolyticus agent in aquaculture industries. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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21 pages, 4488 KiB

Open AccessArticle

In Vitro Activity, Stability and Molecular Characterization of Eight Potent Bacteriophages Infecting Carbapenem-Resistant Klebsiella pneumoniae

by Abeer Ameen Baqer, Kokxin Fang, Norfarhan Mohd-Assaad, Siti Noor Adnalizawati Adnan and Norefrina Shafinaz Md Nor

Viruses 2023, 15(1), 117; https://doi.org/10.3390/v15010117 - 30 Dec 2022

Cited by 5 | Viewed by 2504

Abstract

Background: Members of the genus Klebsiella are among the leading microbial pathogens associated with nosocomial infection. The increased incidence of antimicrobial resistance in these species has propelled the need for alternate/combination therapeutic regimens to aid clinical treatment, including bacteriophage therapy. Bacteriophages are considered [...] Read more.

Background: Members of the genus Klebsiella are among the leading microbial pathogens associated with nosocomial infection. The increased incidence of antimicrobial resistance in these species has propelled the need for alternate/combination therapeutic regimens to aid clinical treatment, including bacteriophage therapy. Bacteriophages are considered very safe and effective in treating bacterial infections. In this study, we characterize eight lytic bacteriophages that were previously isolated by our team against carbapenem-resistant Klebsiella pneumoniae. Methods: The one-step-growth curves, stability and lytic ability of eight bacteriophages were characterized. Restriction fragment length polymorphism (RFLP), random amplification of polymorphic DNA (RAPD) typing analysis and protein profiling were used to characterize the microbes at the molecular level. Phylogenetic trees of four important proteins were constructed for the two selected bacteriophages. Results and conclusions: All eight bacteriophages showed high efficiency for reducing bacterial concentration with high stability under different physical and chemical conditions. We found four major protein bands out of at least ten 15–190 KDa bands that were clearly separated by SDS-PAGE, which were assumed to be the major head and tail proteins. The genomes were found to be dsDNA, with sizes of approximately 36–87 Kb. All bacteriophages reduced the optical density of the planktonic K. pneumoniae abruptly, indicating great potential to reduce K. pneumoniae infection. In this study, we have found that tail fiber protein can further distinguished closely related bacteriophages. The characterised bacteriophages showed promising potential as candidates against carbapenem-resistant Klebsiella pneumoniae via bacteriophage therapy. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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26 pages, 4639 KiB

Open AccessArticle

Isolation and Characterization of a Phapecoctavirus Infecting Multidrug-Resistant Acinetobacter baumannii in A549 Alveolar Epithelial Cells

by Phitchayapak Wintachai, Komwit Surachat, Ganyalak Chaimaha, Abdi Wira Septama and Duncan R. Smith

Viruses 2022, 14(11), 2561; https://doi.org/10.3390/v14112561 - 19 Nov 2022

Cited by 7 | Viewed by 2735

Abstract

Multidrug-resistant Acinetobacter baumannii (MDR A. baumannii) is an emerging pathogen in the ESKAPE group. The global burden of antimicrobial resistance has led to renewed interest in alternative antimicrobial treatment strategies, including phage therapy. This study isolated and characterized a phage vB_AbaM_ ABPW7 [...] Read more.

Multidrug-resistant Acinetobacter baumannii (MDR A. baumannii) is an emerging pathogen in the ESKAPE group. The global burden of antimicrobial resistance has led to renewed interest in alternative antimicrobial treatment strategies, including phage therapy. This study isolated and characterized a phage vB_AbaM_ ABPW7 (vABPW7) specific to MDR A. baumannii. Morphological analysis showed that phage vABPW7 belongs to the Myoviridae family. Genome analysis showed that the phage DNA genome consists of 148,647 bp and that the phage is a member of the Phapecoctavirus genus of the order Caudovirales. A short latent period and a large burst size indicated that phage vABPW7 was a lytic phage that could potentially be used in phage therapy. Phage vABPW7 is a high-stability phage that has high lytic activity. Phage vABPW7 could effectively reduce biofilm formation and remove preformed biofilm. The utility of phage vABPW7 was investigated in a human A549 alveolar epithelial cell culture model. Phage vABPW7 was not cytotoxic to A549 cells, and the phage could significantly reduce planktonic MDR A. baumannii and MDR A. baumannii adhesion on A549 cells without cytotoxicity. This study suggests that phage vABPW7 has the potential to be developed further as a new antimicrobial agent against MDR A. baumannii. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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23 pages, 5865 KiB

Open AccessArticle

Isolation and Characterization of Lytic Bacteriophages Active against Clinical Strains of E. coli and Development of a Phage Antimicrobial Cocktail

by Pavel Alexyuk, Andrey Bogoyavlenskiy, Madina Alexyuk, Kuralay Akanova, Yergali Moldakhanov and Vladimir Berezin

Viruses 2022, 14(11), 2381; https://doi.org/10.3390/v14112381 - 28 Oct 2022

Cited by 16 | Viewed by 3665

Abstract

Pathogenic E. coli cause urinary tract, soft tissue and central nervous system infections, sepsis, etc. Lytic bacteriophages can be used to combat such infections. We investigated six lytic E. coli bacteriophages isolated from wastewater. Transmission electron microscopy and whole genome sequencing showed that [...] Read more.

Pathogenic E. coli cause urinary tract, soft tissue and central nervous system infections, sepsis, etc. Lytic bacteriophages can be used to combat such infections. We investigated six lytic E. coli bacteriophages isolated from wastewater. Transmission electron microscopy and whole genome sequencing showed that the isolated bacteriophages are tailed phages of the Caudoviricetes class. One-step growth curves revealed that their latent period of reproduction is 20–30 min, and the average value of the burst size is 117–155. During co-cultivation with various E. coli strains, the phages completely suppressed bacterial host culture growth within the first 4 h at MOIs 10⁻⁷ to 10⁻³. The host range lysed by each bacteriophage varied from six to two bacterial strains out of nine used in the study. The cocktail formed from the isolated bacteriophages possessed the ability to completely suppress the growth of all the E. coli strains used in the study within 6 h and maintain its lytic activity for 8 months of storage. All the isolated bacteriophages may be useful in fighting pathogenic E. coli strains and in the development of phage cocktails with a long storage period and high efficiency in the treatment of bacterial infections. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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12 pages, 2571 KiB

Open AccessArticle

Isolation and Characterization of the Lytic Pseudoxanthomonas kaohsiungensi Phage PW916

by Chang Wen, Chaofan Ai, Shiyun Lu, Qiue Yang, Hanpeng Liao and Shungui Zhou

Viruses 2022, 14(8), 1709; https://doi.org/10.3390/v14081709 - 2 Aug 2022

Cited by 5 | Viewed by 3049

Abstract

The emergence of multidrug-resistant bacterial pathogens poses a serious global health threat. While patient infections by the opportunistic human pathogen Pseudoxanthomonas spp. have been increasingly reported worldwide, no phage associated with this bacterial genus has yet been isolated and reported. In this study, [...] Read more.

The emergence of multidrug-resistant bacterial pathogens poses a serious global health threat. While patient infections by the opportunistic human pathogen Pseudoxanthomonas spp. have been increasingly reported worldwide, no phage associated with this bacterial genus has yet been isolated and reported. In this study, we isolated and characterized the novel phage PW916 to subsequently be used to lyse the multidrug-resistant Pseudoxanthomonas kaohsiungensi which was isolated from soil samples obtained from Chongqing, China. We studied the morphological features, thermal stability, pH stability, optimal multiplicity of infection, and genomic sequence of phage PW916. Transmission electron microscopy revealed the morphology of PW916 and indicated it to belong to the Siphoviridae family, with the morphological characteristics of a rounded head and a long noncontractile tail. The optimal multiplicity of infection of PW916 was 0.1. Moreover, PW916 was found to be stable under a wide range of temperatures (4–60 °C), pH (4–11) as well as treatment with 1% (v/w) chloroform. The genome of PW916 was determined to be a circular double-stranded structure with a length of 47,760 bp, containing 64 open reading frames that encoded functional and structural proteins, while no antibiotic resistance nor virulence factor genes were detected. The genomic sequencing and phylogenetic tree analysis showed that PW916 was a novel phage belonging to the Siphoviridae family that was closely related to the Stenotrophomonas phage. This is the first study to identify a novel phage infecting the multidrug-resistant P. kaohsiungensi and the findings provide insight into the potential application of PW916 in future phage therapies. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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13 pages, 2415 KiB

Open AccessArticle

Isolation and Characterization of a Novel Siphoviridae Phage, vB_AbaS_TCUP2199, Infecting Multidrug-Resistant Acinetobacter baumannii

by Meity Mardiana, Soon-Hian Teh, Ling-Chun Lin and Nien-Tsung Lin

Viruses 2022, 14(6), 1240; https://doi.org/10.3390/v14061240 - 7 Jun 2022

Cited by 17 | Viewed by 3430

Abstract

Multidrug-resistant Acinetobacter baumannii (MDRAB) is a pathogen recognized as antimicrobial-resistant bacteria involved in healthcare-associated infections. Resistance to antibiotics has made alternative therapies necessary. Bacteriophage therapy is considered a potential solution to treat MDRAB. In this study, we isolated and characterized the phage vB_AbaS_TCUP2199 [...] Read more.

Multidrug-resistant Acinetobacter baumannii (MDRAB) is a pathogen recognized as antimicrobial-resistant bacteria involved in healthcare-associated infections. Resistance to antibiotics has made alternative therapies necessary. Bacteriophage therapy is considered a potential solution to treat MDRAB. In this study, we isolated and characterized the phage vB_AbaS_TCUP2199 (TCUP2199), which can infect MDRAB. Morphological analysis revealed that TCUP2199 belongs to the Siphoviridae family. TCUP2199 has a wide host range, can adsorb rapidly (68.28% in 2 min), and has a burst size of 196 PFU/cell. At least 16 distinct structural proteins were visualized by SDS polyacrylamide gel electrophoresis. A stability test showed that TCUP2199 was stable at 37 °C and pH 7. Genome analysis of TCUP2199 showed that it consists of a double-stranded DNA genome of 79,572 bp with a G+C content of 40.39%, which contains 98 putative open reading frames, none of which is closely related to the bacteriophage genome sequence that was found in the public database. TCUP2199 shows similarity in genomic organization and putative packaging mechanism with Achromobacter phage JWF and Pseudoalteromonas phage KB12-38 based on protein BLAST and phylogenetic analysis. Because of those unique characteristics, we consider TCUP2199 to be a novel phage that is suitable for inclusion in a phage cocktail to treat A. baumannii infection. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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11 pages, 1709 KiB

Open AccessArticle

Global Transcriptomic Response of Staphylococcus aureus to Virulent Bacteriophage Infection

by Nikita Kuptsov, Maria Kornienko, Dmitry Bespiatykh, Roman Gorodnichev, Ksenia Klimina, Vladimir Veselovsky and Egor Shitikov

Viruses 2022, 14(3), 567; https://doi.org/10.3390/v14030567 - 9 Mar 2022

Cited by 6 | Viewed by 3375

Abstract

In light of the ever-increasing number of multidrug-resistant bacteria worldwide, bacteriophages are becoming a valid alternative to antibiotics; therefore, their interactions with host bacteria must be thoroughly investigated. Here, we report genome-wide transcriptional changes in a clinical Staphylococcus aureus SA515 strain for three [...] Read more.

In light of the ever-increasing number of multidrug-resistant bacteria worldwide, bacteriophages are becoming a valid alternative to antibiotics; therefore, their interactions with host bacteria must be thoroughly investigated. Here, we report genome-wide transcriptional changes in a clinical Staphylococcus aureus SA515 strain for three time points after infection with the vB_SauM-515A1 kayvirus. Using an RNA sequencing approach, we identify 263 genes that were differentially expressed (DEGs) between phage-infected and uninfected host samples. Most of the DEGs were identified at an early stage of phage infection and were mainly involved in nucleotide and amino acid metabolism, as well as in cell death prevention. At the subsequent infection stages, the vast majority of DEGs were upregulated. Interestingly, 39 upregulated DEGs were common between the 15th and 30th minutes post-infection, and a substantial number of them belonged to the prophages. Furthermore, some virulence factors were overexpressed at the late infection stage, which necessitates more stringent host strain selection requirements for further use of bacteriophages for therapeutic purposes. Thus, this work allows us to better understand the influence of kayviruses on the metabolic systems of S. aureus and contributes to a better comprehension of phage therapy. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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Review

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17 pages, 2538 KiB

Open AccessReview

Genetic Engineering and Biosynthesis Technology: Keys to Unlocking the Chains of Phage Therapy

by Sixuan Lv, Yuhan Wang, Kaixin Jiang, Xinge Guo, Jing Zhang, Fang Zhou, Qiming Li, Yuan Jiang, Changyong Yang and Tieshan Teng

Viruses 2023, 15(8), 1736; https://doi.org/10.3390/v15081736 - 14 Aug 2023

Cited by 4 | Viewed by 4442

Abstract

Phages possess the ability to selectively eliminate pathogenic bacteria by recognizing bacterial surface receptors. Since their discovery, phages have been recognized for their potent bactericidal properties, making them a promising alternative to antibiotics in the context of rising antibiotic resistance. However, the rapid [...] Read more.

Phages possess the ability to selectively eliminate pathogenic bacteria by recognizing bacterial surface receptors. Since their discovery, phages have been recognized for their potent bactericidal properties, making them a promising alternative to antibiotics in the context of rising antibiotic resistance. However, the rapid emergence of phage-resistant strains (generally involving temperature phage) and the limited host range of most phage strains have hindered their antibacterial efficacy, impeding their full potential. In recent years, advancements in genetic engineering and biosynthesis technology have facilitated the precise engineering of phages, thereby unleashing their potential as a novel source of antibacterial agents. In this review, we present a comprehensive overview of the diverse strategies employed for phage genetic engineering, as well as discuss their benefits and drawbacks in terms of bactericidal effect. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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27 pages, 1877 KiB

Open AccessReview

Acinetobacter Baumannii Phages: Past, Present and Future

by Qihang Tu, Mingfang Pu, Yahao Li, Yuer Wang, Maochen Li, Lihua Song, Mengzhe Li, Xiaoping An, Huahao Fan and Yigang Tong

Viruses 2023, 15(3), 673; https://doi.org/10.3390/v15030673 - 3 Mar 2023

Cited by 15 | Viewed by 5962

Abstract

Acinetobacter baumannii (A. baumannii) is one of the most common clinical pathogens and a typical multi-drug resistant (MDR) bacterium. With the increase of drug-resistant A. baumannii infections, it is urgent to find some new treatment strategies, such as phage therapy. In [...] Read more.

Acinetobacter baumannii (A. baumannii) is one of the most common clinical pathogens and a typical multi-drug resistant (MDR) bacterium. With the increase of drug-resistant A. baumannii infections, it is urgent to find some new treatment strategies, such as phage therapy. In this paper, we described the different drug resistances of A. baumannii and some basic properties of A. baumannii phages, analyzed the interaction between phages and their hosts, and focused on A. baumannii phage therapies. Finally, we discussed the chance and challenge of phage therapy. This paper aims to provide a more comprehensive understanding of A. baumannii phages and theoretical support for the clinical application of A. baumannii phages. Full article

(This article belongs to the Special Issue Viruses versus Bacteria—Novel Approaches to Phage Therapy as a Tool against Multidrug-Resistant Pathogens)

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