Biology subjects

Aracil, B.

Publications and source records attributed to Aracil, B..

4 recordsLinked to original sources

bioRxiv2025

Identification and Functional Insights into New Phage Tail-Like Bacteriocins (PTLBs) Targeting Pseudomonas aeruginosa as new antimicrobials

The current health crisis caused by multidrug resistant (MDR) pathogens is one of the health problems of most concern globally. Infections caused by these pathogens, such as Pseudomonas aeruginosa, lead to high rates of complications, particularly in compromised patients such as cystic fibrosis (CF) patients. The need to counteract and minimize the forecast future impact has led to the rescue of phage therapy. The use of bacteriophages has important advantages, including highly specific targeting, self-amplification at the infection site, minimal disruption of the microbiome, safety and biocompatibility. However, the capacity of bacteria to escape these entities results in a form of resistance that compromises the effectiveness of the therapy. This involves the search for potential alternatives, such as the phage tail-like bacteriocins (PTLBs), also named as tailocins. These high molecular weight particles resemble the tail structure of bacteriophages and are characterized by the absence of genetic material, avoiding the development of resistance, one of the major handicaps associated with phage therapy. In this study, we detected 34 different PTLBs in 75 P. aeruginosa genomes, with different serotypes and sequence types, 11 of which were characterized as novel F-type PTLBs subtypes (F13-F24). Furthermore, we report that four selected PTLBs (R1, F15, F19, R3-F24) can deal with bacterial infection, with the R1 and the F15 PTLBs being the most efficient in clearing infection in vitro, yielding a survival rate of more than 75% in the Galleria mellonella larvae in vivo model. This reaffirms the potential of PTLBs to control P. aeruginosa infections, which can cause chronic infections in some patients, such as people with CF, due to its strong impact as a MDR bacterium. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=194 HEIGHT=200 SRC="FIGDIR/small/671207v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@76b60borg.highwire.dtl.DTLVardef@1a9cd0aorg.highwire.dtl.DTLVardef@d45dd0org.highwire.dtl.DTLVardef@5b345a_HPS_FORMAT_FIGEXP M_FIG C_FIG HIGHLIGHTS-The 75 Pseudomonas aeruginosa genomes from people with cystic fibrosis in the study collection included at least one Phage tail-like bacteriocins (PTLB) cluster. -From the 34 different PTLBs detected in the study collection, 7 were R-type, 10 were complex (R and F-type encoded) and 14 were F-type PTLBs. -11 new F-type PTLBs were described in the Pseudomonas aeruginosa collection under study. -An association between the O-antigen present on the surface of the Pseudomonas aeruginosa isolate and the encoded PTLB subtype was detected. -The R1 and F15 PTLBs subtypes display high antimicrobial activity both in vitro and in vivo (Galleria mellonella).

microbiology↗

bioRxiv2024

The LAMP-CRISPR-Cas13a technique for detecting CBASS-mechanism of phage resistance in bacteria

Antimicrobial resistance (AMR) is an important threat to public health that has led to the development of innovative alternative treatments for bacterial infections, such as phage therapy. However, one of the greatest disadvantages of phage therapy is the generation of phage-resistant bacterial mutants via bacterial defence mechanisms, which are mainly contained in genomic islands (GIs) and controlled by the quorum sensing (QS) network. In this study, 309 pathogenic islands (PAIs) harbouring a total of 22.1 % of proteins related to anti-phage defence (APD) were detected in the genome of 48 K. pneumoniae strains. Several type I and type II CBASS systems were also detected in the genome of the 48 K. pneumoniae strains, but only 2 type II CBASS systems were located in PAIs. We constructed a knockout K. pneumoniae strain, not expressing the cyclase gene from the type II CBASS system present in PAIs, to study the regulatory role of QS in expressing the gene. As the anti-phage CBASS system is an abortive infection (Abi) system, the role of the type II CBASS system in regulating cell viability was assessed. The knockout strain was confirmed by targeting the LAMP-CRISPR-Cas13a technique specifically to the cyclase gene, and the same protocol was also used to detect the gene of the main cyclase in these type I CBASS systems, i.e. APECO1. The study findings demonstrate the regulatory role of the QS network in anti-phage defence systems. Finally, this is the first work which development an innovative biotechnological application for the LAMP-CRISPR-Cas13a rapid-technique (<2 hours) in optimizing phage therapy by detecting bacterial resistance mechanisms, by predicting the potential inefficacy of a therapeutic phage and thus improving patient prognosis.

microbiology↗

bioRxiv2023

Molecular Studies of Phages-Klebsiella pneumoniae in a Mucoid Environment: Innovative use of mucolytic agents prior to the administration of lytic phages

Mucins are important glycoproteins that form a protective layer throughout the gastrointestinal and respiratory tracts. There is scientific evidence of increase in phage-resistance in the presence of mucin for some bacterial pathogens. Manipulation in mucin composition may ultimately influence the effectiveness of phage therapy. In this work, two clinical strains of K. pneumoniae (K3574 and K3325), were exposed to the lytic bacteriophage vB_KpnS-VAC35 in the presence and absence of mucin on a long-term co-evolution assay, in an attempt to mimic in vitro the exposure to mucins that bacteria and their phages face in vivo. Enumerations of the bacterial and phage counts at regular time intervals were conducted, and extraction of the genomic DNA of co- evolved bacteria to the phage, the mucin and both was performed. We determined the frequency of phage-resistant mutants in the presence and absence of mucin and including a mucolytic agent (N-acetyl L-cysteine, NAC), and sequenced these conditions using Nanopore. We phenotypically demonstrated that the presence of mucin induces the emergence of bacterial resistance against lytic phages, effectively decreased in the presence of NAC. In addition, the genomic analysis revealed some of the genes relevant to the development of phage resistance in long-term co- evolution, with a special focus on the mucoid environment. Genes involved in the metabolism of carbohydrates were mutated in the presence of mucin. In conclusion, the use of mucolytic agents prior to the administration of lytic phages could be an interesting therapeutic option when addressing K. pneumoniae infections in environments where mucin is overproduced.

microbiology↗

bioRxiv2023

In vitro and in vivo combination of lytic phages and octapeptin OPX10053 against B-lactamase-producing clinical isolates of Klebsiella pneumoniae

Backgroundnovel approaches to treat Klebsiella pneumoniae infections are desperately needed, such as the use of rationally designed combination therapies. Objectivesto evaluate the in vitro and in vivo therapeutic potential of lytic phages against K. pneumoniae in combination with octapeptin, a promising class of lipopeptides with broad spectrum Gram-negative activity. Methodswe determined the MICs to twenty-two lipopeptide compounds and chose one octapeptin (OPX10053) for evaluation of potential synergism in combination with lytic phages using checkerboard assays, optical density growth curves and time-kill (CFU enumeration). Toxicity and efficacy in vivo assays were conducted on Galleria mellonella larvae. Resultsthis study reports the synergy found in vitro between the octapeptin OPX10053 and two lytic phages previously characterized by our research group (vB_KpnM-VAC13 and vB_KpnM-VAC66) against clinical isolates of K. pneumoniae. This synergy was validated by the FIC index, OD growth curves and time-kill assay when OPX10053 was added following 4 hours of phage exposure. Preliminary evaluation of toxicity revealed that OPX10053, even at subinhibitory concentrations and in phage combinations, exerts a toxic effect on larvae, which requires further investigation. ConclusionsThe in vitro application of lytic phages in combination with octapeptin OPX10053 showed synergistic activity. Exposure of G. mellonella to the lytic phages was well tolerated, whereas combination treatment with subinhibitory concentrations of OPX10053 did not attenuate toxicity. Even so, this innovative approach of combining lytic phages could open the door to some interesting associations between chemically synthesized drugs and biological entities. Sequential or simultaneous application alongside time, dosing and stewardship warrants further research.

microbiology↗