Analyzing the safety of the parasiticide fungus Mucor circinelloides: first insights on its virulence profile and interactions with the avian gut microbial community.

Parasiticide fungi are considered an accurate, sustainable, and safe solution for the biocontrol of animal gastrointestinal (GI) parasites. This research provides an initial characterization of the virulence of the native parasiticide fungus Mucor circinelloides (FMV-FR1) and an assessment of its impact on birds' gut microbes. The genome of this fungus was sequenced to identify the genes coding for virulence factors. Also, this fungus was checked for the phenotypic expression of proteinase, lecithinase, DNase, gelatinase, hemolysin, and biofilm production. Finally, an in vivo trial was developed based on feeding M. circinelloides spores to laying hens and peacocks three times a week. Bird feces were collected for 3 months, with total genomic DNA being extracted and subjected to long-read 16S and 25S-28S sequencing. Genes coding for an iron permease (FTR1), iron receptors (FOB1 and FOB2), ADP-ribosylation factors (ARFs) (ARF2 and ARF6), and a GTPase (CDC42) were identified in this M. circinelloides genome. Also, this fungus was positive only for lecithinase activity. The field trial revealed a fecal microbiome dominated by Firmicutes and Proteobacteria in laying hens, and Firmicutes and Bacteroidetes in peacocks, whereas the fecal mycobiome of both bird species was mainly composed of Ascomycetes and Basidiomycetes fungi. Bacterial and fungal alpha-diversities did not differ between sampling time points after M. circinelloides administrations (P = 0.62 and P = 0.15, respectively). Although findings from this research suggest the lack of virulence of this M. circinelloides parasiticide isolate, more complementary in vitro and in vivo research is needed to conclude about the safety of its administration to birds, aiming at controlling their GI parasites.IMPORTANCEA previous study revealed that the native Mucor circinelloides isolate (FMV-FR1) can develop parasiticide activity toward coccidia oocysts, one of the most pathogenic GI parasites in birds. However, ensuring its safety for birds is of utmost importance, namely by studying its virulence profile and potential effect on commensal gut microbes. This initial study revealed that although this M. circinelloides isolate had genes coding for four types of virulence factors-iron permease, iron receptors, ADP-ribosylation factors, and GTPase-and only expressed phenotypically the enzyme lecithinase, the administration of its spores to laying hens and peacocks did not interfere with the abundances and diversities of their gut commensal bacteria and fungi. Although overall results suggest the lack of virulence of this M. circinelloides isolate, more complementary research is needed to conclude about the safety of its administration to birds in the scope of parasite biocontrol programs.

Nisin Z Potential for the Control of Diabetic Foot Infections Promoted by Pseudomonas aeruginosa Persisters.

Diabetic foot ulcers (DFU) are a major complication of diabetes mellitus and a public health concern worldwide. The ability of P. aeruginosa to form biofilms is a key factor responsible for the chronicity of diabetic foot infections (DFIs) and frequently associated with the presence of persister cells. These are a subpopulation of phenotypic variants highly tolerant to antibiotics for which new therapeutic alternatives are urgently needed, such as those based on antimicrobial peptides. This study aimed to evaluate the inhibitory effect of nisin Z on P. aeruginosa DFI persisters. To induce the development of a persister state in both planktonic suspensions and biofilms, P. aeruginosa DFI isolates were exposed to carbonyl cyanide m-chlorophenylhydrazone (CCCP) and ciprofloxacin, respectively. After RNA extraction from CCCP-induced persisters, transcriptome analysis was performed to evaluate the differential gene expression between the control, persisters, and persister cells exposed to nisin Z. Nisin Z presented a high inhibitory effect against P. aeruginosa persister cells but was unable to eradicate them when present in established biofilms. Transcriptome analysis revealed that persistence was associated with downregulation of genes related to metabolic processes, cell wall synthesis, and dysregulation of stress response and biofilm formation. After nisin Z treatment, some of the transcriptomic changes induced by persistence were reversed. In conclusion, nisin Z could be considered as a potential complementary therapy for treating P. aeruginosa DFI, but it should be applied as an early treatment or after wound debridement.

Potential of two delivery systems for nisin topical application to dental plaque biofilms in dogs.

BACKGROUND: Periodontal disease (PD) is caused by the development of a microbial biofilm (dental plaque) in the periodontium, affecting approximately 80% of dogs. Several bacterial species present in the canine oral cavity can be implicated in the development of this disease, including Enterococcus spp. To decrease antibiotic administration, a possible control strategy for dog's enterococcal PD may involve the use of the antimicrobial peptide (AMP) nisin. Nisin's inhibitory activity was evaluated against a collection of previously characterized enterococci obtained from the oral cavity of dogs with PD (n = 20), as well as the potential of a guar-gum gel and a veterinary toothpaste as topical delivery systems for this AMP. The Minimum Inhibitory (MIC) and Bactericidal Concentrations (MBC) and the Minimum Biofilm Eradication (MBEC) and Inhibitory Concentrations (MBIC) were determined for nisin and for the supplemented guar-gum gel. For the supplemented veterinary toothpaste an agar-well diffusion assay was used to evaluate its inhibitory potential. RESULTS: Nisin was effective against all isolates. Independently of being or not incorporated in the guar-gum gel, its inhibitory activity on biofilms was higher, with MBIC (12.46 ± 5.16 and 13.60 ± 4.31 µg/mL, respectively) and MBEC values (21.87 ± 11.33 and 42.34 ± 16.61 µg/mL) being lower than MIC (24.61 ± 4.64 and 14.90 ± 4.10 µg/mL) and MBC (63.09 ± 13.22 and 66.63 ± 19.55 µg/mL) values. The supplemented toothpaste was also effective, showing inhibitory activity against 95% of the isolates. CONCLUSIONS: The inhibitory ability of nisin when incorporated in the two delivery systems was maintained or increased, demonstrating the potential of these supplemented vehicles to be applied to PD control in dogs.