Searching for a Better Animal Model for Chronic Tympanic Membrane Perforation.

Chronic tympanic membrane perforation represents a prevalent otological condition, necessitating a reliable animal model for the validation and safety assessment of surgical techniques and materials employed in myringoplasty. This prospective study involved the establishment of chronic tympanic membrane perforation animal models in 16 chinchillas. A thermic myringotomy was conducted on the right ear (study group), followed by cold instrument myringotomy, coupled with the topical application of mitomycin C and dexamethasone solution on the left ear (control group). Results revealed that tympanic membrane perforations in the study group persisted for a minimum of 4 weeks in 93.7% of cases and extended to 12 weeks in 62.5% of the cases. In contrast, all tympanic membrane perforations in the control group were present at 4 weeks, with only 37.5% persisting after 12 weeks, although statistical tests did not find significant differences between the two groups (chi-square: p-value = 0.157, Kruskal-Wallis: p-value = 0.093, Mann-Whitney: p-value = 0.121). The thermic myringotomy employed to induce chronic tympanic membrane perforation in animals demonstrated efficiency and sustainability. This model, characterized by stability and reproducibility, holds promise for future experimental applications in the field.

Antibacterial and Antifungal Silver Nanoparticles with Tunable Size Embedded in Various Cellulose-Based Matrices.

The aim of this study was to synthesize silver nanoparticles (AgNPs) using cellulose derivatives and to evaluate their antimicrobial potential. As effective reducing and stabilizing agents for AgNPs, cellulose derivatives, such as hydroxypropyl cellulose (HPC), methylcellulose (MC), ethylcellulose (EC), and cellulose acetate (CA), were used. Their ability to reduce silver ions as well as the size of the resulting AgNPs were compared. The formation and stability of the reduced AgNPs in the solution were monitored using UV-Vis analysis. The size, morphology, and charge of the AgNPs were evaluated. We found that, when using cellulosic derivatives, AgNPs with sizes ranging from 17 to 89 nm and different stabilities were obtained. The parameters, such as size and ζ potential indicate the stability of AgNPs, with AgNPs-CA and AgNPs-HPC being considered more stable than AgNPs-EC and AgNPs-MC since they show higher ζ potential values. In addition, the AgNPs showed antimicrobial activity against all reference strains and clinical isolates. MIC values between 0.0312 and 0.125 mM had a bactericidal effect on both Gram-positive and Gram-negative bacteria. The fungicidal effect was obtained at a MIC value of 0.125 mM. These results may provide rational support in the design of medical gauze products, including gauze pads, rolls, and sponges.

Trichinella spiralis and T. britovi in North-Eastern Romania: A Six-Year Retrospective Multicentric Survey.

The genus Trichinella includes species with a wide geographical spread that cause pathology in humans and animals. In this context, an epidemiological study of Trichinella infection was carried out in the northeastern part of Romania to investigate for the first time the prevalence of this infection in pigs, horses, wild boars and bears, the geographical distribution of Trichinella species and the natural reservoir of Trichinella infection. Between 2010 and 2015, a total of 166,270 animals were examined by the method of artificial digestion, in order to calculate the annual and general prevalence of Trichinella infection, according to the host and the Trichinella species involved, the Pearson correlation coefficient (r), trendline and geographical distribution of species of the genus Trichinella. Taxonomic framing was performed by the multiplex PCR method. The overall prevalence of Trichinella infection in animals was 0.188%. Within the host species, the prevalence varied as follows: in pigs 0.096%, horses 0.021%, wild boar 1.46% and bears 36.76%. The geographical distribution showed that T. spiralis was dominant, occupying the entire northeastern part of Romania, being identified in pigs, horses, wild boars and bears. T britovi occupied five mountain counties, being identified only in wild boars and bears. These results validate the presence of T. spiralis and T. britovi in domestic and game animals in the northeastern part of Romania.

Chitosan-Based Therapeutic Systems for Superficial Candidiasis Treatment. Synergetic Activity of Nystatin and Propolis.

The paper deals with new approaches to chitosan (CS)-based antifungal therapeutic formulations designed to fulfill the requirements of specific applications. Gel-like formulations were prepared by mixing CS dissolved in aqueous lactic acid (LA) solution with nystatin (NYS) powder and/or propolis (PRO) aqueous solution dispersed in glycerin, followed by water evaporation to yield flexible mesoporous (pore widths of 2-4 nm) films of high specific surfaces between 1 × 103 and 1.7 × 103 m2/g. Morphological evaluation of the antifungal films showed uniform dispersion and downsizing of NYS crystallites (with initial sizes up to 50 µm). Their mechanical properties were found to be close to those of soft tissues (Young's modulus values between 0.044-0.025 MPa). The films presented hydration capacities in physiological condition depending on their composition, i.e., higher for NYS-charged (628%), as compared with PRO loaded films (118-129%). All NYS charged films presented a quick release for the first 10 min followed by a progressive increase of the release efficiency at 48.6%, for the samples containing NYS alone and decreasing values with increasing amount of PRO to 45.9% and 42.8% after 5 h. By in vitro analysis, the hydrogels with acidic pH values around 3.8 were proven to be active against Candida albicans and Candida glabrata species. The time-killing assay performed during 24 h on Candida albicans in synthetic vagina-simulative medium showed that the hydrogel formulations containing both NYS and PRO presented the faster slowing down of the fungal growth, from colony-forming unit (CFU)/mL of 1.24 × 107 to CFU/mL < 10 (starting from the first 6 h).

Apiaceae Essential Oils: Boosters of Terbinafine Activity against Dermatophytes and Potent Anti-Inflammatory Effectors.

Dermatophyte infections represent an important public health concern, affecting up to 25% of the world's population. Trichophyton rubrum and T. mentagrophytes are the predominant dermatophytes in cutaneous infections, with a prevalence accounting for 70% of dermatophytoses. Although terbinafine represents the preferred treatment, its clinical use is hampered by side effects, drug-drug interactions, and the emergence of resistant clinical isolates. Combination therapy, associating terbinafine and essential oils (EOs), represents a promising strategy in the treatment of dermatophytosis. In this study, we screened the potential of selected Apiaceae EOs (ajowan, coriander, caraway, and anise) to improve the antifungal activity of terbinafine against T. rubrum ATCC 28188 and T. mentagrophytes ATCC 9533. The chemical profile of EOs was analyzed by gas chromatography. The minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) of EOs/main compounds were determined according to EUCAST-AFST guidelines, with minor modifications. The checkerboard microtiter method was used to identify putative synergistic combinations of EOs/main constituents with terbinafine. The influence of EOs on the viability and pro-inflammatory cytokine production (IL-1ß, IL-8 and TNF-α) was determined using an ex vivo human neutrophils model. The binary associations of tested EOs with terbinafine were found to be synergistic against T. rubrum, with FICI values of 0.26-0.31. At the tested concentrations (6.25-25 mg/L), EOs did not exert cytotoxic effects towards human neutrophils. Anise EO was the most potent inhibitor of IL-1ß release (46.49% inhibition at 25 mg/L), while coriander EO displayed the highest inhibition towards IL-8 and TNF-α production (54.15% and 54.91%, respectively). In conclusion, the synergistic combinations of terbinafine and investigated Apiaceae EOs could be a starting point in the development of novel topical therapies against T. rubrum-related dermatophytosis.

Honokiol and Magnolol: Insights into Their Antidermatophytic Effects.

Dermatophyte infections represent a significant public health concern, with an alarming negative impact caused by unsuccessful therapeutic regimens. Natural products have been highlighted as a promising alternative, due to their long-standing traditional use and increasing scientific recognition. In this study, honokiol and magnolol, the main bioactives from Magnolia spp. bark, were investigated for their antidermatophytic activity. The antifungal screening was performed using dermatophyte standard strains and clinical isolates. The minimal inhibitory concentration (MIC) and the minimal fungicidal concentration (MFC) were determined in accordance with EUCAST-AFST guidelines, with minor modifications. The effects on ergosterol biosynthesis were assessed in Trichophyton rubrum cells by HPLC-DAD. Putative interactions with terbinafine against T. rubrum were evaluated by the checkerboard method. Their impact on cells' viability and pro-inflammatory cytokines (IL-1ß, IL-8 and TNF-α) was shown using an ex vivo human neutrophils model. Honokiol and magnolol were highly active against tested dermatophytes, with MIC and MFC values of 8 and 16 mg/L, respectively. The mechanism of action involved the inhibition of ergosterol biosynthesis, with accumulation of squalene in T. rubrum cells. Synergy was assessed for binary mixtures of magnolol with terbinafine (FICI = 0.50), while honokiol-terbinafine combinations displayed only additive effects (FICI = 0.56). In addition, magnolol displayed inhibitory effects towards IL-1ß, IL-8 and TNF-α released from lipopolysaccharide (LPS)-stimulated human neutrophils, while honokiol only decreased IL-1ß secretion, compared to the untreated control. Overall, honokiol and magnolol acted as fungicidal agents against dermatophytes, with impairment of ergosterol biosynthesis.

Silver Nanoparticles Biosynthesized with Spruce Bark Extract-A Molecular Aggregate with Antifungal Activity against Candida Species.

Due to their high content of biomolecules, combined with silver's well known antimicrobial potential, silver nanoparticles biosynthesized using spruce bark (AgNP SBEs) demonstrate antibacterial and antioxidant activity, making them a versatile option for developing new antimicrobial agents that might be used for medical treatment or as adjuvants for the classical agents. This study aims to analyze if silver nanoparticles (AgNPs) mediated by spruce bark extract (SBE) and silver salts (AgNP SBE Acetate, AgNP SBE Nitrate) presents antifungal activity against five different Candida spp., synergistic activity with fluconazole, and if they interact with some virulence factors of C. albicans. AgNP SBEs presented MICs (minimum inhibitory concentrations) for all the five tested Candida spp. AgNP SBEs inhibited the growth of C. parapsilosis, C. krusei, and C. guilliermondii, exerted synergistic activity with fluconazole for C. parapsilosis and C. guilliermondii, and inhibited biofilm production for C. albicans, C. auris, and C. guilliermondii. MICs of AgNP SBE Acetate significantly inhibited the production of germ tubes of C. albicans. The expression of C. albicans SAP2 gene was down-regulated by the short-time treatment with MICs of AgNP SBE Acetate, while ALS3 and HSP70 genes were up-regulated by the AgNPs MICs. These results emphasize the potential of using the AgNP SBEs as treatments/adjuvants options, not only against the redundant C. albicans but also for the non-albicans Candida species (which are not as frequently involved in human pathologies, but, sometimes, can be more aggressive).

In Vitro Antifungal Activity of Silver Nanoparticles Biosynthesized with Beech Bark Extract.

Biosynthesis is a green method for the synthesis of silver nanoparticles (AgNPs). This study aimed to assess the antifungal activity of two silver nanoparticle solutions, synthesized using beech bark extract (BBE) and acetate and nitrate silver salts (AgNP Acetate BBE and AgNP Nitrate BBE), their influence on biofilm production, their potential synergistic effects with fluconazole, on different Candida spp., and their influence on virulence factors of C. albicans (germ tube production, gene expression for ALS3, SAP2, HSP70). Both the AgNP BBEs presented different minimum inhibitory concentrations for all the studied Candida spp., but biofilm production was inhibited only for C. albicans and C. guilliermondii. The growth rates of all the studied Candida spp. were inhibited in the presence of both AgNP BBEs, except for C. auris. Synergistic activity was observed for C. parapsilosis and C. guilliermondii, for different combinations of fluconazole with both the AgNP BBEs. The germ tube production of C. albicans was slightly inhibited by the AgNP BBEs. Only AgNP Acetate BBE was able to down-regulate the expression of SAP2. Overall, we can conclude that, even if more studies are necessary, AgNPs synthesized with beech bark extract might be an interesting alternative to classic antifungal treatments.

Antibiotic Susceptibility Profile of Pseudomonas aeruginosa Canine Isolates from a Multicentric Study in Romania.

Treating infections caused by Pseudomonas aeruginosa is increasingly difficult due to high antibiotic resistance, materialized through the presence of multiple resistance strains, as well as due to rapid development of resistance throughout treatment. The present survey was conducted to investigate the antibiotic susceptibility profile of Pseudomonas aeruginosa pathogens in two University Veterinary hospitals from different geographical regions of Romania (i.e., Southwest Timișoara county and Northeast Iași county) involved in superficial canine infections. A total of 142 swab specimens were collected from dogs with superficial infections (superficial skin infections, otitis externa, and perianal abscess) to assess the presence of Pseudomonas aeruginosa, based on phenotypic and molecular characterization. According to their confirmed morphological and molecular features, 58 samples (40.84%; 58/142) were positive for Pseudomonas aeruginosa (according to their confirmed morphological and molecular features). Antibiotic susceptibility testing for 12 antibiotics was conducted using the Kirby-Bauer disc diffusion method. Drug resistance was observed in the case of all tested antibiotics. The susceptibility rate of P. aeruginosa strains that were tested in this study was in the following order: ceftazidime (53.44%; 31/58), followed by aztreonam (51.72%; 30/58), amikacin (44.82%; 26/58), azithromycin (41.37%; 24/58), gentamicin (37.93%; 22/58), cefepime (36.20%; 21/58), meropenem (25.86%; 13/58), piperacillin-tazobactam (25.86%; 13/58), imipenem (22.41%; 13/158), ciprofloxacin (17.24%; 10/58), tobramycin (8.62; 5/58), and polymyxin B (1.72; 1/58). The results highlight the importance of antibiotic susceptibility testing in Pseudomonas aeruginosa isolates from dogs with superficial infections to use an adequate treatment plan to manage the skin condition and other pathologies (otitis externa and perianal abscesses).

Needles in a haystack: Extremely rare invasive fungal infections reported in FungiScopeⓇ-Global Registry for Emerging Fungal Infections.

OBJECTIVES: Emerging invasive fungal infections (IFI) have become a notable challenge. Apart from the more frequently described fusariosis, lomentosporiosis, mucormycosis, scedosporiosis, and certain dematiaceae or yeasts, little is known about extremely rare IFI. METHODS: Extremely rare IFI collected in the FungiScopeⓇ registry were grouped as Dematiaceae, Hypocreales, Saccharomycetales, Eurotiales, Dermatomycetes, Agaricales, and Mucorales. RESULTS: Between 2003 and June 2019, 186 extremely rare IFI were documented in FungiScopeⓇ. Dematiaceae (35.5%), Hypocreales (23.1%), Mucorales (11.8%), and Saccharomycetales (11.3%) caused most IFI. Most patients had an underlying malignancy (38.7%) with acute leukemia accounting for 50% of cancers. Dissemination was observed in 26.9% of the patients. Complete or partial clinical response rate was 68.3%, being highest in Eurotiales (82.4%) and in Agaricales (80.0%). Overall mortality rate was 29.3%, ranging from 11.8% in Eurotiales to 50.0% in Mucorales. CONCLUSIONS: Physicians are confronted with a complex variety of fungal pathogens, for which treatment recommendations are lacking and successful outcome might be incidental. Through an international consortium of physicians and scientists, these cases of extremely rare IFI can be collected to further investigate their epidemiology and eventually identify effective treatment regimens.

Invasive pulmonary infection due to Thermoascus crustaceus in a kidney transplant recipient.

The first case of invasive pulmonary infection due to the thermophilic fungus Thermoascus crustaceus in a kidney transplant recipient is described. For the identification of the fungal isolate, morphological aspects and molecular analysis have been used. The case report emphasizes this fungal species as an opportunistic human pathogen and underlines the importance of an accurate laboratory diagnosis for the correct management of the patient.

The Burden of Fungal Diseases in Romania.

OBJECTIVE: To estimate for the first time the burden of fungal infections in Romania. METHODS: Data derived from the World Health Organization (WHO), National Institute of Statistics, Romanian public health agencies and non-profit health organizations, and published annual reports on local epidemiology were used in the present study. When no data were available, specific at-risk populations were used to calculate frequencies of serious fungal diseases, using previously published epidemiological parameters. All data refer to the year 2016. RESULTS: The estimated number of serious fungal infections in Romanian population was 436,230 in 2016. Recurrent vulvovaginal candidiasis accounted for up to 80% of total cases (more than 350,000 women annually). Concerning HIV-related infections, among 14,349 infected persons, Pneumocystis pneumonia occurred in about 10% of late presenters (30 cases in 2016), while cryptococcal meningitis was rarely diagnosed (less than 20 cases). Annually, the total number of oesophageal candidiasis and oral thrush cases in HIV-positive patients may have been as high as 1229 and 3066, respectively. In immunocompromised and cancer patient populations, the annual incidence of candidaemia was 295, and at least 458 invasive aspergillosis cases and 4 mucormycosis cases occurred yearly. With 4966 critical care beds and approximately 200,000 abdominal surgeries performed, the estimated annual incidence of candidaemia and Candida peritonitis was 689 and 344, respectively. The annual incidence of pulmonary tuberculosis is still high in Romania (12,747 cases). Thus, the prevalence of post-TB chronic pulmonary aspergillosis is estimated to be 8.98/100,000 (1768 cases). The prevalence of chronic obstructive pulmonary disease (COPD) and asthma in adults is 6% and 6.5%, respectively. Therefore, allergic bronchopulmonary aspergillosis prevalence is estimated at 29,387 and severe asthma with fungal sensitisation at 38,731 cases annually. CONCLUSIONS: Not being on the list of reportable diseases, the number of patients presenting with severe mycoses in Romania can only be roughly estimated. Based on local reports and prevalence estimation, we consider that at least 2.23% of Romanians suffer from a serious form of fungal disease.