The first research on the insect Galleria mellonella was published 85 years ago, and the larva is now widely used as a model to study infections caused by bacterial and fungal pathogens, for screening new antimicrobials, to study the adjacent immune response in co-infections or in host-pathogen interaction, as well as in a toxicity model. The immune system of the G. mellonella model shows remarkable similarities with mammals. Furthermore, results from G. mellonella correlate positively with mammalian models and with other invertebrate models. Unlike other invertebrate models, G. mellonella can withstand temperatures of 37 °C, and its handling and experimental procedures are simpler. Despite having some disadvantages, G. mellonella is a virtuous in vivo model to be used in preclinical studies, as an intermediate model between in vitro and mammalian in vivo studies, and is a great example on how to apply the bioethics principle of the 3Rs (Replacement, Reduction, and Refinement) in animal experimentation. This review aims to discuss the progress of the G. mellonella model, highlighting the key aspects of its use, including experimental design considerations and the necessity to standardize them. A different score in the “cocoon” category included in the G. mellonella Health Index Scoring System is also proposed.

Hospital-acquired infections (HAIs) are a rising problem worldwide, and the best way of coping with them is through infection tracking and surveillance systems, combined with prevention strategies, namely efficient disinfection protocols, that employ various biocides. However, increasing reports about reductions in biocide susceptibility and the development of cross-resistance to antimicrobials emphasize the need for identifying the factors influencing biocide efficiency. In this study, 29 bacterial isolates (n = 3 E. coli, n = 2 Pseudomonas spp., n = 23 Enterococcus spp., and n = 1 Staphylococcus pseudintermedius), obtained from environmental samples collected from the Biological Isolation and Containment Unit (BICU), of the Veterinary Teaching Hospital of the Faculty of Veterinary Medicine, University of Lisbon, were tested in order to determine their antimicrobial susceptibility to various antibiotics. Thirteen of these isolates were further selected in order to determine their antimicrobial susceptibility to Virkon™ S, with and without the presence of organic matter. Afterward, seven of these isolates were incubated in the presence of sub-lethal concentrations of this formulation and, subsequently, new susceptibility profiles were determined. Fourteen of the 29 isolates (48.3%) were classified as multidrug resistant, all previously identified as enterococci. Concerning Virkon™ S’s susceptibility, the Minimal Bactericidal Concentration (MBC) of this biocide regarding all isolates was at least eight times lower than the concentration regularly used, when no organic matter was present. However, when organic matter was added, MBC values rose up to 23 times. After exposure to sub-lethal concentrations of Virkon™ S, four enterococci presented a phenotypical change regarding antimicrobial susceptibility towards gentamicin. Virkon™ S also resulted in higher MBC values, up to 1.5 times, in the presence of low concentrations of organic matter, but no rise in these values was observed in assays without interfering substance. Virkon™ S seemed to be an efficient formulation in eliminating all bacteria isolates isolated from the BICU. However, organic matter could represent a hindrance to this ability, which emphasizes the importance of sanitization before disinfection procedures. The changes seen in antimicrobial susceptibility could be explained by a general stress-induced response promoted by the sub-lethal levels of Virkon™ S. Additionally, when no organic matter was present, a decrease in susceptibility to this biocide seemed to be non-existent.

Background The Biological Isolation and Containment Unit (BICU) is a subunit of the Teaching Hospital of the Faculty of Veterinary Medicine of the University of Lisbon, Portugal, for the admission of animals with confirmed infectious diseases or under clinical suspicion and waiting for a diagnosis. As a high-risk environment for the transmission of infectious agents, it is extremely important to implement programs for the surveillance of nosocomial microorganisms in these facilities. The purpose of this study was to evaluate the level of bacterial contamination of the BICU environmental surfaces and to implement corrective actions on disinfection protocols. Swab samples were collected from selected environmental surfaces in 3 different areas of the BICU (isolation, work, and preparatory rooms) to evaluate the total aerobic bacterial load and investigate the presence of 4 nosocomial microorganisms: vancomycin-resistant Enterococcus spp., methicillin-resistant Staphylococcus aureus, 3rd-generation cephalosporin-resistant Escherichia coli, and carbapenem-resistant Pseudomonas aeruginosa. Bacterial quantification was performed by using non-selective media, while specific selective media were used for the isolation of the target microorganisms. Isolates were identified based on their macro and microscopic characteristics and their biochemical profile. Subsequently, new disinfection protocols were implemented, and their effectiveness evaluated. Results The surfaces with the highest bacterial load in the isolation, preparatory, and worker’s rooms were the cages, hand-held sponge, and telephone, respectively. Regarding the 4 pathogens investigated, Enterococcus spp. were the most frequently isolated (11.3%), followed by E. coli (1.5%) and P. aeruginosa (1.5%). One of the P. aeruginosa isolates obtained was resistant to imipenem. In the end, new disinfection protocols were implemented, which proved to be effective in reducing bacterial counts by 99.99% in cages and the sponge, and by 90 to 99% on the telephone. Conclusions This study allows to conclude that the cages and the human contact surfaces were the most contaminated in the isolation rooms. Nevertheless, the new disinfection strategies seemed to be effective in reducing environmental contamination, including by some potentially nosocomial agents, although more samples must be analyzed for definitive conclusions. These results may contribute to highlight the importance of infection prevention and control measures, as fundamental tools to reduce the spread of infectious agents in the hospital environment.

Pseudomonas aeruginosa is an opportunistic bacterial pathogen responsible for multiple hospital- and community-acquired infections, both in human and veterinary medicine. P. aeruginosa persistence in clinical settings is worrisome and is a result of its remarkable flexibility and adaptability. This species exhibits several characteristics that allow it to thrive under different environmental conditions, including the ability to colonize inert materials such as medical equipment and hospital surfaces. P. aeruginosa presents several intrinsic mechanisms of defense that allow it to survive external aggressions, but it is also able to develop strategies and evolve into multiple phenotypes to persevere, which include antimicrobial-tolerant strains, persister cells, and biofilms. Currently, these emergent pathogenic strains are a worldwide problem and a major concern. Biocides are frequently used as a complementary/combination strategy to control the dissemination of P. aeruginosa-resistant strains; however, tolerance to commonly used biocides has also already been reported, representing an impediment to the effective elimination of this important pathogen from clinical settings. This review focuses on the characteristics of P. aeruginosa responsible for its persistence in hospital environments, including those associated with its antibiotic and biocide resistance ability.