Quantifying the Dynamics of Bacterial Biofilm Formation on the Surface of Soft Contact Lens Materials Using Digital Holographic Tomography to Advance Biofilm Research.

The increase in bacterial resistance to antibiotics in recent years demands innovative strategies for the detection and combating of biofilms, which are notoriously resilient. Biofilms, particularly those on contact lenses, can lead to biofilm-related infections (e.g., conjunctivitis and keratitis), posing a significant risk to patients. Non-destructive and non-contact sensing techniques are essential in addressing this threat. Digital holographic tomography emerges as a promising solution. This allows for the 3D reconstruction of the refractive index distribution in biological samples, enabling label-free visualization and the quantitative analysis of biofilms. This tool provides insight into the dynamics of biofilm formation and maturation on the surface of transparent materials. Applying digital holographic tomography for biofilm examination has the potential to advance our ability to combat the antibiotic bacterial resistance crisis. A recent study focused on characterizing biofilm formation and maturation on six soft contact lens materials (three silicone hydrogels, three hydrogels), with a particular emphasis on Staphylococcus epidermis and Pseudomonas aeruginosa, both common culprits in ocular infections. The results revealed species- and time-dependent variations in the refractive indexes and volumes of biofilms, shedding light on cell dynamics, cell death, and contact lens material-related factors. The use of digital holographic tomography enables the quantitative analysis of biofilm dynamics, providing us with a better understanding and characterization of bacterial biofilms.

Isolation and characterization of multidrug resistant Gallibacterium anatis biovar haemolytica strains from Polish geese and hens.

Gallibacterium anatis biovar haemolytica is a bacterium that is frequently associated with infections of the reproductive tract and respiratory system in poultry. To assess the current prevalence and resistance profile of these bacteria in Poland, we collected and investigated 63 strains of Gallibacterium from diseased domestic poultry flocks including geese, laying hens, breeding hens and an ornamental hen. Detailed characterization of the isolates included the analysis of phenotypic antimicrobial resistance profiles and biofilm formation ability. Furthermore, the genetic background of 40 selected isolates regarding the presence of virulence and antimicrobial resistance genes and mobile genetic elements was determined. All investigated isolates were multidrug resistant, most prominently to ß-lactams, fluoroquinolones, sulfonamides and macrolides. A total of 48 different resistance profiles were detected. Of all isolates, 50.8% formed a strong biofilm, where strains isolated from geese appeared to be better at biofilm formation than strains isolated from laying and breeding hens. Single-nucleotide polymorphism genotyping revealed that G. anatis bv. haemolytica strains are restricted in host and geographical distribution, and the geese isolates showed greater phylogenetic similarity. Whole genome sequencing enabled identification of 25 different antimicrobial resistance determinants. The most common resistance genes were tetB, blaROB-1, and blaTEM-1 which may be located on mobile genetic elements. All isolates possessed the toxin gene gtxA, and the fimbrial gene flfA was identified in 95% of strains. Our results indicated that all G. anatis bv. haemolytica isolates showed multidrug resistant phenotypes. Strains isolated from geese were characterized by the highest percentage of isolates resistant to selected antimicrobials, probably reflecting host-related adaptations.

The Enhancement of Antimicrobial Photodynamic Therapy of Escherichia Coli by a Functionalized Combination of Photosensitizers: In Vitro Examination of Single Cells by Quantitative Phase Imaging.

The prevention of biofilm formation is crucial for the limitation of bacterial infections typically associated with postoperative infections, complications in bedridden patients, and a short-term prognosis in affected cancer patients or mechanically ventilated patients. Antimicrobial photodynamic therapy (aPDT) emerges as a promising alternative for the prevention of infections due to the inability of bacteria to become resistant to aPDT inactivation processes. The aim of this study was to demonstrate the use of a functionalized combination of Chlorin e6 and Pheophorbide as a new approach to more effective aPDT by increasing the accumulation of photosensitizers (PSs) within Escherichia coli cells. The accumulation of PSs and changes in the dry mass density of single-cell bacteria before and after aPDT treatment were investigated by digital holotomography (DHT) using the refractive index as an imaging contrast for 3D label-free live bacteria cell imaging. The results confirmed that DHT can be used in complex examination of the cell-photosensitizer interaction and characterization of the efficiency of aPDT. Furthermore, the use of Pheophorbide a as an efflux pomp inhibitor in combination with Chlorin e6 increases photosensitizers accumulation within E. coli and overcomes the limited penetration of Gram-negative cells by anionic and neutral photosensitizers.

Molecular detection of avian hepatitis E virus (Orthohepevirus B) in chickens, ducks, geese, and western capercaillies in Poland.

Orthohepevirus B, commonly known as avian hepatitis E virus (aHEV), causes big liver and spleen disease (BLS) or hepatitis-splenomegaly syndrome (HSS) in chickens. BLS is an emerging disease among chicken flocks in several countries around the world. In our previous studies, serology and molecular biology screening revealed that chicken flocks are widely affected by aHEV in Poland. The present study, which was conducted between 2019 and 2020, aimed to investigate the prevalence of aHEV in chicken flocks and other poultry, including ducks, geese, and turkeys. A total of 307 flocks were examined. In addition, 29 samples from captive wild birds (western capercaillies, Tetrao urogallus) were analyzed. In all the investigated poultry species, except turkeys, the nucleic acid sequence covering part of the ORF1 gene of the aHEV genome was detected (34/336 samples, 10.1%). The infection rate was found to be the highest in broiler breeder chicken flocks (14/40 samples; 35%). Phylogenetic analysis of partial ORF1 gene, which encodes helicase, revealed that the obtained sequences belonged to genotypes 2 and 4, while one belonged to genotype 3. Genotype 2 was detected for the first time in domestic geese and ducks, and genotype 4 was detected for the first time in Poland. The study demonstrated the presence of aHEV among the investigated western capercaillies, suggesting that this species is susceptible to aHEV infections and biosecurity is therefore required in western capercaillie breeding facilities.

Genomic and functional characterization of five novel Salmonella-targeting bacteriophages.

BACKGROUND: The host-unrestricted, non-typhoidal Salmonella enterica serovar Enteritidis (S. Enteritidis) and the serovar Typhimurium (S. Typhimurium) are major causative agents of food-borne gastroenteritis, and the host-restricted Salmonella enterica serovar Gallinarum (S. Gallinarum) is responsible for fowl typhoid. Increasing drug resistance in Salmonella contributes to the reduction of effective therapeutic and/or preventive options. Bacteriophages appear to be promising antibacterial tools, able to combat infectious diseases caused by a wide range of Salmonella strains belonging to both host-unrestricted and host-restricted Salmonella serovars. METHODS: In this study, five novel lytic Salmonella phages, named UPWr_S1-5, were isolated and characterized, including host range determination by plaque formation, morphology visualization with transmission electron microscopy, and establishment of physiological parameters. Moreover, phage genomes were sequenced, annotated and analyzed, and their genomes were compared with reference Salmonella phages by use of average nucleotide identity, phylogeny, dot plot, single nucleotide variation and protein function analysis. RESULTS: It was found that UPWr_S1-5 phages belong to the genus Jerseyvirus within the Siphoviridae family. All UPWr_S phages were found to efficiently infect various Salmonella serovars. Host range determination revealed differences in host infection profiles and exhibited ability to infect Salmonella enterica serovars such as Enteritidis, Gallinarum, Senftenberg, Stanley and Chester. The lytic life cycle of UPWr_S phages was confirmed using the mitomycin C test assay. Genomic analysis revealed that genomes of UPWr_S phages are composed of 51 core and 19 accessory genes, with 33 of all predicted genes having assigned functions. UPWr_S genome organization comparison revealed 3 kinds of genomes and mosaic structure. UPWr_S phages showed very high sequence similarity to each other, with more than 95% average nucleotide identity. CONCLUSIONS: Five novel UPWr_S1-5 bacteriophages were isolated and characterized. They exhibit host lysis range within 5 different serovars and are efficient in lysis of both host-unrestricted and host-restricted Salmonella serovars. Therefore, because of their ability to infect various Salmonella serovars and lytic life cycle, UPWr_S1-5 phages can be considered as useful tools in biological control of salmonellosis.

Bacteria Single-Cell and Photosensitizer Interaction Revealed by Quantitative Phase Imaging.

Quantifying changes in bacteria cells in the presence of antibacterial treatment is one of the main challenges facing contemporary medicine; it is a challenge that is relevant for tackling issues pertaining to bacterial biofilm formation that substantially decreases susceptibility to biocidal agents. Three-dimensional label-free imaging and quantitative analysis of bacteria-photosensitizer interactions, crucial for antimicrobial photodynamic therapy, is still limited due to the use of conventional imaging techniques. We present a new method for investigating the alterations in living cells and quantitatively analyzing the process of bacteria photodynamic inactivation. Digital holographic tomography (DHT) was used for in situ examination of the response of Escherichia coli and Staphylococcus aureus to the accumulation of the photosensitizers immobilized in the copolymer revealed by the changes in the 3D refractive index distributions of single cells. Obtained results were confirmed by confocal microscopy and statistical analysis. We demonstrated that DHT enables real-time characterization of the subcellular structures, the biophysical processes, and the induced local changes of the intracellular density in a label-free manner and at sub-micrometer spatial resolution.

Phenotypic and genotypic characterization of mcr-1-positive multidrug-resistant Escherichia coli ST93, ST117, ST156, ST10, and ST744 isolated from poultry in Poland.

BACKGROUND: A plasmid-mediated mechanism of bacterial resistance to polymyxin is a serious threat to public health worldwide. The present study aimed to determine the occurrence of plasmid-mediated colistin resistance genes and to conduct the molecular characterization of mcr-positive Escherichia coli strains isolated from Polish poultry. METHODS: In this study, 318 E. coli strains were characterized by the prevalence of mcr1-mcr5 genes, antimicrobial susceptibility testing by minimal inhibitory concentration method, the presence of antimicrobial resistance genes was screened by PCR, and the biofilm formation ability was tested using the crystal violet staining method. Genetic relatedness of mcr-1-positive E. coli strains was evaluated by multilocus sequence typing method. RESULTS: Among the 318 E. coli isolates, 17 (5.35%) harbored the mcr-1 gene. High antimicrobial resistance rates were observed for ampicillin (100%), tetracycline (88.24%), and chloramphenicol (82.35%). All mcr-1-positive E. coli strains were multidrug-resistant, and as many as 88.24% of the isolates contained the blaTEM gene, tetracycline (tetA and tetB), and sulfonamide (sul1, sul2, and sul3) resistance genes. Additionally, 41.18% of multidrug-resistant, mcr-1-positive E. coli isolates were moderate biofilm producers, while the rest of the strains showed weak biofilm production. Nine different sequence types were identified, and the dominant ST was ST93 (29.41%), followed by ST117 (17.65%), ST156 (11.76%), ST 8979 (11.76%), ST744 (5.88%), and ST10 (5.88%). Moreover, the new ST was identified in this study. CONCLUSIONS: Our results showed a low occurrence of mcr-1-positive E. coli strains isolated from Polish poultry; however, all the isolated strains were resistant to multiple antimicrobial agents and were able to form biofilms at low or medium level.

Myricetin as an Antivirulence Compound Interfering with a Morphological Transformation into Coccoid Forms and Potentiating Activity of Antibiotics against Helicobacter pylori.

Helicobacter pylori, a gastric pathogen associated with a broad range of stomach diseases, has a high tendency to become resistant to antibiotics. One of the most important factors related to therapeutic failures is its ability to change from a spiral to a coccoid form. Therefore, the main aim of our original article was to determine the influence of myricetin, a natural compound with an antivirulence action, on the morphological transformation of H. pylori and check the potential of myricetin to increase the activity of antibiotics against this pathogen. We observed that sub-minimal inhibitory concentrations (sub-MICs) of this compound have the ability to slow down the process of transformation into coccoid forms and reduce biofilm formation of this bacterium. Using checkerboard assays, we noticed that the exposure of H. pylori to sub-MICs of myricetin enabled a 4-16-fold reduction in MICs of all classically used antibiotics (amoxicillin, clarithromycin, tetracycline, metronidazole, and levofloxacin). Additionally, RT-qPCR studies of genes related to the H. pylori morphogenesis showed a decrease in their expression during exposure to myricetin. This inhibitory effect was more strongly seen for genes involved in the muropeptide monomers shortening (csd3, csd6, csd4, and amiA), suggesting their significant participation in the spiral-to-coccoid transition. To our knowledge, this is the first research showing the ability of any compound to synergistically interact with all five antibiotics against H. pylori and the first one showing the capacity of a natural substance to interfere with the morphological transition of H. pylori from spiral to coccoid forms.

The Phylogenetic Structure of Reptile, Avian and Uropathogenic Escherichia coli with Particular Reference to Extraintestinal Pathotypes.

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).

Genetic Diversity and Distribution of Virulence-Associated Genes in Y. enterocolitica and Y. enterocolitica-Like Isolates from Humans and Animals in Poland.

Yersinia enterocolitica, widespread within domestic and wild-living animals, is a foodborne pathogen causing yersiniosis. The goal of this study was to assess a genetic similarity of Y. enterocolitica and Y. enterocolitica-like strains isolated from different hosts using Multiple Locus Variable-Number Tandem Repeat Analysis (MLVA) and Pulsed-Field Gel Electrophoresis (PFGE) methods, and analyze the prevalence of virulence genes using multiplex-Polymerase Chain Reaction (PCR) assays. Among 51 Yersinia sp. strains 20 virulotypes were determined. The most common virulence genes were ymoA, ureC, inv, myfA, and yst. Yersinia sp. strains had genes which may contribute to the bacterial invasion and colonization of the intestines as well as survival in serum. One wild boar Y. enterocolitica 1A strain possessed ail gene implying the possible pathogenicity of 1A biotype. Wild boar strains, represented mainly by 1A biotype, were not classified into the predominant Variable-Number Tandem Repeats (VNTR)/PFGE profile and virulotype. There was a clustering tendency among VNTR/PFGE profiles of pig origin, 4/O:3, and virulence profile. Pig and human strains formed the most related group, characterized by ~80% of genetic similarity what suggest the role of pigs as a potential source of infection for the pork consumers.

Photolon Nanoporous Photoactive Material with Antibacterial Activity and Label-Free Noncontact Method for Free Radical Detection.

The worldwide increase in bacterial resistance and healthcare-associated bacterial infections pose a serious threat to human health. The antimicrobial photodynamic method reveals the opportunity for a new therapeutic approach that is based on the limited delivery of photosensitizer from the material surface. Nanoporous inorganic-organic composites were obtained by entrapment of photosensitizer Photolon in polysiloxanes that was prepared by the sol-gel method. The material was characterized by its porosity, optical properties (fluorescence and absorbance), and laser-induced antimicrobial activity against Staphylococcus epidermidis, Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli. The permanent encapsulation of Photolon in the silica coating and the antimicrobial efficiency was confirmed by confocal microscope and digital holotomography. The generation of free radicals from nanoporous surfaces was proved by scanning Kelvin probe microscopy. For the first time, it was confirmed that Kelvin probe microscopy can be a label-free, noncontact alternative to other conventional methods based on fluorescence or chemiluminescence probes, etc. It was confirmed that the proposed photoactive coating enables the antibacterial photodynamic effect based on free radicals released from the surface of the coating. The highest bactericidal efficiency of the proposed coating was 87.16%. This coating can selectively limit the multiplication of bacterial cells, while protecting the environment and reducing the risk of surface contamination.

On the application of multi-parametric optical phenotyping of bacterial colonies for multipurpose microbiological diagnostics.

The development of new diagnostics techniques and modalities is critical for early detection of microbial contamination. In this study, the novel integrated system for multi-parametric optical phenotyping and characterization of bacterial colonies, is presented. The system combines Mach-Zehnder interferometer with a spectral imaging system for capturing multispectral diffraction patterns and multispectral two-dimensional transmission maps of bacterial colonies, along with the simultaneous interferometric profilometry. The herein presented investigation was carried out on five representative bacteria species and nearly 3000 registered multispectral optical signatures. The interferograms were analyzed by four-step phase shift algorithm to reconstruct the colony profile to enable the obtaining of the comparable optical signatures. The dedicated image processing algorithms were used for extraction of quantitative features of these signatures. The random forest algorithm was applied for selection of the most predictive set of features, which were used in classification model based on Support-Vector Machine. Obtained results have shown that the use of multiple multispectral optical signatures provide a multi-parametric bacteria identification at an exceptionally high accuracy (99.4-100%), significantly better than in case of classification based on each of these signatures (multispectral diffraction patterns, two-dimensional transmission coefficient maps), separately. Obtained results revealed that analysis of multispectral signatures can also be applied for characterisation of physical, physicochemical and chemical properties of the bacterial colonies in the presence of the antimicrobial factors. Therefore, the proposed label-free, non-destructive optical technique has perspectives to be exploited in the multipurpose diagnostics and it can be used as a pre-screening tool in microbiological laboratories.

Short Beak and Dwarfism Syndrome in Ducks in Poland Caused by Novel Goose Parvovirus.

Short beak and dwarfism syndrome (SBDS), which was previously identified only in mule ducks, is now an emerging disease of Pekin ducks in China and Egypt. The disease is caused by the infection of ducks with a genetic variant of goose parvovirus-novel goose parvovirus (nGPV). In 2019, SBDS was observed for the first time in Poland in eight farms of Pekin ducks. Birds in the affected flock were found to show growth retardation and beak atrophy with tongue protrusions. Morbidity ranged between 15% and 40% (in one flock), while the mortality rate was 4-6%. Co-infection with duck circovirus, a known immunosuppressive agent, was observed in 85.7% of ducks. The complete coding regions of four isolates were sequenced and submitted to GenBank. The phylogenetic analysis revealed a close relationship of Polish viral sequences with the Chinese nGPV. Genomic sequence alignments showed 98.57-99.28% identity with the nGPV sequences obtained in China, and 96.42% identity with the classical GPV (cGPV; Derzsy's disease). The rate of amino acid mutations in comparison to cGPV and Chinese nGPV was higher in the Rep protein than in the Vp1 protein. To our knowledge, this is the first report of nGPV infection in Pekin ducks in Poland and Europe. It should be emphasized that monitoring and sequencing of waterfowl parvoviruses is important for tracking the viral genetic changes that enable adaptation to new species of waterbirds.

Development of the Correction Algorithm to Limit the Deformation of Bacterial Colonies Diffraction Patterns Caused by Misalignment and Its Impact on the Bacteria Identification in the Proposed Optical Biosensor.

Recently proposed methods of bacteria identification in optical biosensors based on the phenomenon of light diffraction on macro-colonies offer over 98% classification accuracy. However, such high accuracy relies on the comparable and repeatable spatial intensity distribution of diffraction patterns. Therefore, it is essential to eliminate all non-species/strain-dependent factors affecting the diffraction patterns. In this study, the impact of the bacterial colony and illuminating beam misalignment on the variation of classification features extracted from diffraction patterns was examined. It was demonstrated that misalignment introduced by the scanning module significantly affected diffraction patterns and extracted classification features used for bacteria identification. Therefore, it is a crucial system-dependent factor limiting the identification accuracy. The acceptable misalignment level, when the accuracy and quality of the classification features are not affected, was determined as no greater than 50 µm. Obtained results led to development of image-processing algorithms for determination of the direction of misalignment and concurrent alignment of the bacterial colonies' diffraction patterns. The proposed algorithms enable the rigorous monitoring and controlling of the measurement's conditions in order to preserve the high accuracy of bacteria identification.

Antimicrobial Resistance and Biofilm Formation Capacity of Salmonella enterica Serovar Enteritidis Strains Isolated from Poultry and Humans in Poland.

Salmonella enterica ser. Enteritidis (S. enterica ser. Enteritidis) is the most frequently detected serovar in human salmonellosis, and its ability to produce a biofilm and the risk of transmission from animals and food of animal origin to humans are significant. The main aim of the present work was to compare S. enterica ser. Enteritidis strains isolated from poultry and human feces in terms of resistance profiles, prevalence of selected resistance genes, and their potential for biofilm formation, by assessing their biofilm growth intensity, the prevalence and expression of selected genes associated with this phenomenon, and the correlation between increased antimicrobial resistance and biofilm formation ability of the two tested groups of S. enterica ser. Enteritidis. This study showed a difference in antimicrobial resistance (minimal inhibitory concentration value) between S. enterica ser. Enteritidis groups; however, the majority of multidrug-resistant (MDR) strains were isolated from poultry (environmental samples from chicken broilers, turkey broilers, and laying hens). Differences in the prevalence of resistance genes were observed; the most common gene among poultry strains was floR, and that among strains from humans was blaTEM. S. enterica ser. Enteritidis strains isolated from poultry under the tested incubation conditions exhibited better biofilm growth than strains isolated from humans. A higher level of gene expression associated with the production of cellulose was only detected in the S48 strain isolated from poultry. On the other hand, increased expression of genes associated with quorum sensing was observed in two strains isolated from poultry farms and one strain isolated from human feces.

Comparison of the phylogenetic analysis of PFGE profiles and the characteristic of virulence genes in clinical and reptile associated Salmonella strains.

BACKGROUND: Salmonella is generally considered as a human pathogen causing typhoid fever and gastrointestinal infections called salmonellosis, with S. Enteritidis and S. Typhimurium strains as the main causative agents. Salmonella enterica strains have a wide host array including humans, birds, pigs, horses, dogs, cats, reptiles, amphibians and insects. Up to 90% of reptiles are the carriers of one or more serovars of Salmonella. Extraintestinal bacterial infections associated with reptiles pose serious health threat to humans. The import of exotic species of reptiles as pet animals to Europe correlates with the emergence of Salmonella serotypes, which not found previously in European countries. The presented study is a new report about Salmonella serotypes associated with exotic reptiles in Poland. The goal of this research was to examine the zoonotic potential of Salmonella strains isolated from reptiles by comparative analysis with S. Enteritidis strains occurring in human population and causing salmonellosis. RESULTS: The main findings of our work show that exotic reptiles are asymptomatic carriers of Salmonella serovars other than correlated with salmonellosis in humans (S. Enteritidis, S. Typhimurium). Among the isolated Salmonella strains we identified serovars that have not been reported earlier in Poland, for example belonging to subspecies diarizonae and salamae. Restriction analysis with Pulsed-field Gel Electrophoresis (PFGE), showed a great diversity among Salmonella strains isolated from reptiles. Almost all tested strains had distinct restriction patterns. While S. Enteritidis strains were quite homogeneous in term of phylogenetic relations. Most of the tested VGs were common for the two tested groups of Salmonella strains. CONCLUSIONS: The obtained results show that Salmonella strains isolated from reptiles share most of virulence genes with the S. Enteritidis strains and exhibit a greater phylogenetic diversity than the tested S. Enteritidis population.

Integrated multi-channel optical system for bacteria characterization and its potential use for monitoring of environmental bacteria.

The potential use of a novel multichannel optical system towards fast and non-destructive bacteria identification and its application for environmental bacteria characterisation on the strain level is presented. It is the first attempt to use the proposed optical method to study various bacteria species (Gram-negative, Gram-positive) commonly present in the environment. The novel configuration of the optical system enables multichannel examination of bacterial colonies and provides additional functionality such as registration of two-dimensional (2D) distribution of monochromatic transmission coefficient of examined colonies, what can be used as a novel optical signature for bacteria characterization. Performed statistical analysis indicates that it is possible to identify representatives of environmental soil bacteria on the species level with the 98.51% accuracy and in case of two strains of Rahnella aquatilis bacteria on the strain level with the 98.8% accuracy. The proposed method is an alternative to the currently used preliminary bacteria examination in environment safety control with the advantage of being fast, reliable, non-destructive and requiring minimal sample preparation.

Antimicrobial Resistance and Biofilm Formation in Enterococcus spp. Isolated from Humans and Turkeys in Poland.

Enterococci are a natural component of the intestinal flora of many organisms, including humans and birds. As opportunistic pathogens, they can cause fatal infections of the urinary tract and endocarditis in humans, whereas in poultry symptoms are joint disease, sepsis, and falls in the first week of life. The study covered 107 Enterococcus strains-56 isolated from humans and 51 from turkeys. Among the isolates investigated Enterococcus faecalis was detected in 80.36% of human and 80.39% of turkey samples. Enterococcus faecium was identified in 8.93% of human and 17.65% of turkey strains. The highest percentage of the strains was resistant to tetracycline as follows: 48 (85.71%) and 48 (94.12%) of human and turkey strains, respectively. Resistance to erythromycin occurred in 37.50% of the human and in 76.47% of turkey strains, otherwise 27.10% of all strains showed resistance to ciprofloxacin. Our study revealed that 25% of human and 15.69% of turkey strains were resistant to vancomycin. Multidrug resistance showed in 32.14% and 43.14% of human and turkey strains, respectively. The tetracycline resistance gene, tetM, was detected in 82.24% of all strains analyzed, whereas the tetO gene was found in 53.57% of human but only in 7.84% of turkey strains. The vancomycin resistance gene (vanA) was detected in seven Enterococcus strains (six isolated from turkeys and one from humans). The ermB gene (resistance to macrolide) was detected in 55.14% of all isolates (42.86% of human and 68.63% of turkey strains), whereas the ermA gene was detected in 17.65% of turkey but only in 3.57% of human isolates. All the strains had the ability to form biofilms. A stronger biofilm was formed after 24-hour incubation by strains isolated from turkeys, whereas after 48 hours of incubation all examined strains produced strong biofilm.

Avian hepatitis E virus is widespread among chickens in Poland and belongs to genotype 2.

Big liver and spleen disease, caused by avian hepatitis E virus, has been reported in Poland, but the prevalence of the virus has not yet been investigated. In this study, 1034 serum samples from 57 breeder broiler and laying hen flocks were screened for the presence of anti-aHEV antibodies. In a random serology study, 56.1% of flocks were positive. Seroprevalence was higher in laying hen flocks than in broiler breeder flocks. Phylogenetic analysis of partial ORF1 and ORF2 sequences revealed that all Polish isolates belonged to genotype 2. This is the first time this genotype has been detected in Central Europe.

High Prevalence of Resistance to Fluoroquinolones and Tetracycline Campylobacter Spp. Isolated from Poultry in Poland.

Campylobacter spp. is a major cause of foodborne diseases in humans, particularly when transmitted by the handling or consumption of undercooked poultry meat. Most Campylobacter infections are self-limiting, but antimicrobial treatment (e.g., fluoroquinolones and macrolides) is necessary in severe or prolonged cases. The indiscriminate use of these drugs, both in clinical medicine and animal production, has a major impact on public health. The aim of the present study was to identify Campylobacter strains, isolated from turkey and broilers, using both PCR and the matrix-assisted laser desorption-ionization time-of-flight (MALDI-TOF) methods to reveal the accuracy of identification, as well to evaluate the antimicrobial and genetic resistance of the investigated strains. MALDI-TOF and PCR methods were used to show differences, if any, in the specificity of that test. In this study, MALDI-TOF mass spectrometry gave the same results as multiplex PCR, in all cases. The highest rate of resistance (i.e., 100% of turkey and broiler strains) was detected against ciprofloxacin, whereas 58.1% of turkey and 78.6% of broiler strains were resistant to tetracycline. Multidrug-resistant isolates were not found in the study. All ciprofloxacin-resistant strains had a mutation in the gyrA gene, at the Thr-86 position. The presence of the tetO gene was found in 71% of turkey and in 100% of broiler strains. All resistant to tetracycline strains included tetO gene. Additionally, in five turkey and three broiler strains, susceptible to tetracycline, tetO gene was present. These results indicate the high prevalence of Campylobacter strains, which are phenotypically and genetically resistant to fluoroquinolones and tetracycline.