The genus Lactobacillus includes, among others, Lactobacillus casei, Lactobacillus paracasei and Lactobacillus rhamnosus, species that are collectively referred to as the Lactobacillus casei group. Many studies have shown that strains belonging to this group may decrease lactose intolerance, the effects of inflammatory bowel disease, diarrhea, constipation, food allergies and even colon cancer. Moreover, evidences exists of positive effects of these bacteria on mucosal immunity and blood cholesterol level. Because of their beneficial influence on human health, many of them are used as food additives and probiotic pharmaceuticals. It should be stressed that health-promoting properties are not attributed at the species level, but to specific strains. Therefore, procedures are necessary to allow specific identification at each phylogenetic level—genus, species and strain. In this paper we present a practical overview of molecular methods for the identification and differentiation of L. casei bacteria. The research included 30 bacterial strains belonging to three species: L.casei, L. paracasei and L. rhamnosus. Among the tested procedures were genus- and species-specific PCR, multiplex-PCR, Real-Time HRM analysis, RFLP-PCR, rep-PCR, RAPD-PCR, AFLP-PCR, and proteomic methods such as MALDI-TOF MS typing and SDS-PAGE fingerprinting. The obtained results showed that multiplex-PCR and MALDI-TOF MS turned out to be the most useful methods to identify the tested bacteria at the species level. At the strain level, the AFLP-PCR method showed the highest discriminatory power. We hope that the presented results will allow for the easy selection of an appropriate procedure, depending on the experiment conducted and the equipment capabilities of any given laboratory.

Oral health is a window to a patient’s general well-being. Balance in oral microbiome functions is crucial for health maintenance. A state of oral dysbiosis may lead to a variety of local and systemic pathological conditions. The presence of dental plaque is related to the majority of oral infections. Proper oral hygiene is crucial and the most economic practice contributing to oral health prophylaxis. Aside from prophylactic treatments provided by dental practitioners, mouth rinses, containing antimicrobial agents, are one of the possible tools used for oral care. Our study was to determine whether available mouth rinses and selected products dedicated for professional use are efficient to eradicate biofilm formed by reference and clinical strains of Streptococcus mutans, Streptococcus sanguinis, Streptococcus oralis, Streptococcus mitis, Staphylococcus aureus, Enterococcus faecalis, Lactobacillus rhamnosus and Candida albicans on the surface of hydroxyapatite – major mineral component of a tooth. Therefore, such antimicrobials as chlorhexidine, cetylpyridine chloride, polyhexanide, silver nanoparticles, sulphonated phenolics, and natural antiplaque essential oils and coconut oil were analyzed. Applied experimental settings in in vitro models were designed to reflect accurately the recommended use of the tested substances, therefore four types of eradication procedure were conducted. Sialorrhea simulation was also performed to evaluate antibiofilm potential of diluted mouth rinses. Biofilm was investigated with quantitative method where absorbance values were measured. Statistical differences were assessed using the Kruskal–Wallis test with a post-hoc Dunnett’s analysis. Results have shown that biofilms displayed a diversified sensitivity to the tested antimicrobials. The highest antibiofilm activity was detected for cetylpyridine chloride while the lowest for chlorhexidine. However the differences in E. faecalis biofilm reduction observed after the use of these two compounds were not statistically significant (p > 0.05), whereas all observed differences in S. aureus survival after exposure to the examined antimicrobial agents were statistically significant (p < 0.5). The PHMB, both in standard and in sialorrhea simulated conditions had the highest potential against streptococci. The coconut oil reduced C. albicans fungus biofilm by 65.48% but low eradication level was observed in case of bacterial biofilms. The dehydrating mechanism of action of sulfonated phenolics turned out to be ineffective against streptococcal biofilm which in turn was effectively eradicated by silver nanoparticles. The implementation of Antibiofilm Dressing's Activity Measurement method allowed to observe strain-related differences in terms of antimicrobial sensitivity. The obtained results may be introduced in everyday out-patient dental plaque prophylaxis as well as clinical environment.

The impact of the Gram-negative bacterium Escherichia coli (E. coli) on the microbiomic and pathogenic phenomena occurring in humans and other warm-blooded animals is relatively well-recognized. At the same time, there are scant data concerning the role of E. coli strains in the health and disease of cold-blooded animals. It is presently known that reptiles are common asymptomatic carriers of another human pathogen, Salmonella, which, when transferred to humans, may cause a disease referred to as reptile-associated salmonellosis (RAS). We therefore hypothesized that reptiles may also be carriers of specific E. coli strains (reptilian Escherichia coli, RepEC) which may differ in their genetic composition from the human uropathogenic strain (UPEC) and avian pathogenic E. coli (APEC). Therefore, we isolated RepECs (n = 24) from reptile feces and compared isolated strains’ pathogenic potentials and phylogenic relations with the aforementioned UPEC (n = 24) and APEC (n = 24) strains. To this end, we conducted an array of molecular analyses, including determination of the phylogenetic groups of E. coli, virulence genotyping, Pulsed-Field Gel Electrophoresis-Restriction Analysis (RA-PFGE) and genetic population structure analysis using Multi-Locus Sequence Typing (MLST). The majority of the tested RepEC strains belonged to nonpathogenic phylogroups, with an important exception of one strain, which belonged to the pathogenic group B2, typical of extraintestinal pathogenic E. coli. This strain was part of the globally disseminated ST131 lineage. Unlike RepEC strains and in line with previous studies, a high percentage of UPEC strains belonged to the phylogroup B2, and the percentage distribution of phylogroups among the tested APEC strains was relatively homogenous, with most coming from the following nonpathogenic groups: C, A and B1. The RA-PFGE displayed a high genetic diversity among all the tested E. coli groups. In the case of RepEC strains, the frequency of occurrence of virulence genes (VGs) was lower than in the UPEC and APEC strains. The presented study is one of the first attempting to compare the phylogenetic structures of E. coli populations isolated from three groups of vertebrates: reptiles, birds and mammals (humans).

The One Health approach emphasizes the importance of zoonoses due to their pandemic potential, highlighting the need to characterize emerging bacterial pathogens across animal reservoirs. Non-typhoidal Salmonella (NTS) species are among the most common zoonotic agents and can be transmitted by various reservoirs, including reptiles. Both direct and indirect contact with reptiles may result in Reptile-Associated Salmonellosis (RAS), which mainly affects children, immunocompromised individuals, pregnant women, and the elderly. This study aimed to isolate and characterize the Gram-negative intestinal microbiota from free-living snakes in Poland (Natrix natrix, Natrix tessellata, Coronella austriaca, Zamenis longissimus, and Elaphe dione) and to determine the prevalence and virulence potential of Salmonella. Using MALDI-TOF Mass Spectrometry, 432 isolates were identified. Serological analysis of 62 Salmonella isolates revealed 10 distinct O-antigen groups, and rare serovars O:38, O:48, O:57 and others were confirmed. Salmonella isolates were tested for antibiotic susceptibility and resistance to Human Serum; most isolates survived exposure to serum while remaining susceptible to antibiotics. One isolate was classified as multidrug-resistant (MDR), showing resistance to amoxicillin/clavulanic acid, ampicillin, cefuroxime, cephalexin, tigecycline, and fosfomycin. These findings demonstrate that wild snakes in Poland can act as reservoirs of pathogenic and zoonotic Salmonella, emphasizing their epidemiological significance in natural ecosystems.

Fungi belonging to the Cryptococcus neoformans/C. gattii species complex (CNGSC) are etiological agents of serious and not infrequently fatal infections in both humans and animals. Trees are the main ecological niche and source of potential exposition concerning these pathogens. With regard to epidemiology of cryptococcosis, various surveys were performed worldwide, enabling the establishment of a map of distribution and genetic structure of the arboreal population of the CNGSC. However, there are regions, among them Central and Eastern Europe, in which the data are lacking. The present study shows the results of such an environmental study performed in Wrocław, Poland. The CNGSC strains were detected in 2.2% of the tested trees belonging to four genera. The obtained pathogen population consisted exclusively of C. neoformans, represented by both the major molecular type VNI and VNIV. Within the tested group of isolates, resistance to commonly used antimycotics was not found, except for 5-fluorocytosine, in which about 5% of the strains were classified as a non-wild type.

(1) Background: Meyerozyma guilliermondii is a yeast species widely distributed in the natural environment and one of the rare emerging pathogens capable of causing difficult to treat, severe infections. The species’ susceptibility profile is not fully defined; however, the species could be more prone to develop resistance than other Candida species. The objective of this research was to determine the susceptibility of a local collection of Meyerozyma guilliermondii clinical isolates to classical antifungal drugs as well as a new one—manogepix. (2) Methods: The study included 20 clinical isolates identified using the MALDI–TOF method followed with sequencing of ITS1-2 region of DNA. Overall, the susceptibility to 12 antifungal drugs was tested. Nine drugs (amphotericin B, flucytosine, fluconazole, itraconazole, posaconazole, voriconazole, anidulafungin, caspofungin, and micafungin) were assessed using the MICRONAUT–AT test. The susceptibility to the new drug, manogepix, as well as isavuconazole, clotrimazole and anidulafungin, was determined using the microdilution method recommended by EUCAST. Additionally, anidulafungin and voriconazole MIC was also examined with commercial gradient tests. (3) Results: Overall, the isolates showed low MIC values for amphotericin B (0.125 to 1 mg/L) and for flucytosine (≤0.06 to 32 mg/L), with the exception of one isolate with a high MIC value. The MIC ranges for azoles were 2–64 mg/L (fluconazole), 0.008–0.5 mg/L (voriconazole), ≤0.03–≥4 mg/L (itraconazole) and 0.008–0.5 mg/L (posaconazole). One isolate showed non-WT phenotype to all tested azoles. For anidulafungin, the MIC values ranged from ≤0.06 to 0.25 mg/L; however, in the reference method, higher values were observed, but they did not exceed 2 mg/L (ECOFF value). For manogepix, the MIC values ranged from 0.002 to 0.125 mg/L. Finally, the comparison of the obtained and published susceptibility data was conducted. (4) Conclusions: The data obtained in this study are consistent with reports by other authors and indicate that resistance to azoles or 5-fluorocytosine among clinical isolates of Meyerozyma guilliermondii should be considered. The low MIC values of manogepix suggest the potentially good efficacy of this drug against Meyerozyma guilliermondii species.