Chitosan-based matrix as a carrier for bacteriophages.

Wound healing is a dynamic and complex process where infection prevention is essential. Chitosan, thanks to its bactericidal activity against gram-positive and gram-negative bacteria, as well as anti-inflammatory and hemostatic properties, is an excellent candidate to design dressings for difficult-to-heal wound treatment. The great advantage of this biopolymer is its capacity to be chemically modified, which allows for the production of various functional forms, depending on the needs and subsequent use. Moreover, chitosan can be an excellent polymer matrix for bacteriophage (phage) packing as a novel alternative/supportive antibacterial therapy approach. This study is focused on the preparation and characteristics of chitosan-based material in the form of a film with the addition of Pseudomonas lytic phages (KTN4, KT28, and LUZ19), which would exhibit antibacterial activity as a potential dressing that accelerates the wound healing. We investigated the method of producing a polymer based on microcrystalline chitosan (MKCh) to serve as the matrix for phage deposition. We described some important parameters such as average molar mass, swelling capacity, surface morphology, phage release profile, and antibacterial activity tested in the Pseudomonas aeruginosa bacterial model. The chitosan polysaccharide turned out to interact with phage particles immobilizing them within a material matrix. Nevertheless, with the high hydrophilicity and swelling features of the prepared material, the external solution of bacterial culture was absorbed and phages went in direct contact with bacteria causing their lysis in the polymer matrix. KEY POINTS: • A novel chitosan-based matrix with the addition of active phages was prepared • Phage interactions with the chitosan matrix were determined as electrostatic • Phages in the matrix work through direct contact with the bacterial cells.

Chitin and Chitosan as Polymers of the Future-Obtaining, Modification, Life Cycle Assessment and Main Directions of Application.

Natural polymers are very widespread in the world, which is why it is so important to know about the possibilities of their use. Chitin is the second most abundant reproducible natural polymer in nature; however, it is insoluble in water and basic solvents. Chitin is an unused waste of the food industry, for which there are possibilities of secondary management. The research led to obtaining a soluble, environmentally friendly form of chitin, which has found potential applications in the many fields, e.g., medicine, cosmetics, food and textile industries, agriculture, etc. The deacetylated form of chitin, which is chitosan, has a number of beneficial properties and wide possibilities of modification. Modification possibilities mean that we can obtain chitosan with the desired functional properties, facilitating, for example, the processing of this polymer and expanding the possibilities of its application, also as biomimetic materials. The review contains a rich description of the possibilities of modifying chitin and chitosan and the main directions of their application, and life cycle assessment (LCA)-from the source of the polymer through production materials to various applications with the reduction of waste.

Reliability of histopathological examination and immunohistochemistry of a single biopsy for evaluation of endometrial health in Icelandic mares.

INTRODUCTION: Endometritis and endometrosis have been and still are the major reasons for infertility in mares. The diagnosis of endometritis can be based on cytology and microbiology, but endometrial biopsy is still the only way to diagnose endometrosis in the mare. Our study attempted to determine if a single biopsy using his-topathology and immunohistochemistry is sufficient to ascertain reasons for infertility in Icelandic mares. The objectives of this study were to examine the relationship between deviations in endometrial biopsies in terms of prostaglandin-endoperoxide synthase-2 (PTGS-2) and fibronectin expression and polymorphonuclear cells (PMNs) infiltration, as well as scoring degeneration in two endometrial biopsies. MATERIAL AND METHODS: Materials were collected from 53 Icelandic breed mares, from whom two endometrial biopsies were collected and they were used for histopathology and for immunohistochemistry for PTGS-2 and fibronectin. RESULTS: In our study, twenty-six of 53 mares (49%) showed differences in the biopsy score between the left and the right uterine horns (p = 0.002). There were statistically significant differences in fibronectin expression (p = 0.001), as well as in PTGS-2 expression in the superficial epithelium (p = 0.017). CONCLUSIONS: Significant differences in the biopsy score, and fibronectin and PTGS-2 expression, between two endometrial biopsies obtained from individual mares demonstrated that a single biopsy could be insufficient for diagnosing uterine health status in Icelandic mares.

The usefulness of uterine lavage and acute phase protein levels as a diagnostic tool for subclinical endometritis in Icelandic mares.

BACKGROUND: Endometritis is a common problem in a broodmare practice, often leading to infertility. The diagnosis is based on several methods such as cytology, bacteriology and histopathology; however the outcome of these methods may be inconclusive even when used together. The objectives of this study were: (1) to investigate the usefulness of acute phase proteins as an additional diagnostic tool for diagnosis of subclinical endometritis in mares and (2) to evaluate the association between macroscopic changes in uterine flushes and inflammation of the uterus. Materials were collected from 53 Icelandic mares with subclinical endometritis. Endometrial swabs and uterine lavage for cytological and bacteriological examinations and two endometrial biopsies were taken. Blood samples were collected 12-24 h after ovulation to determine the concentrations of serum amyloid A and haptoglobin in the 53 subfertile mares and, for comparison, from 20 non-pregnant mares that later conceived. RESULTS: Twenty-five mares were classified as positive for endometritis based on endometrial biopsy, which was used as the 'gold standard'. We observed a correlation between cloudy efflux in the lavage and (1) polymorphonuclear cell (PMN) infiltration of the endometrium (P = 0.031), (2) positive cytology in samples obtained by cotton swabs (P = 0.019) and uterine lavage (P = 0.011), and (3) positive microbiology from samples obtained by cotton swabs (P = 0.001) and uterine lavage (P = 0.047). The degree of agreement between PMN infiltration and positive cytology from samples taken by cotton swabs and uterine lavage was fair to moderate. We found no association between the concentration of acute phase proteins and infiltration of the endometrium by PMNs, or with positive results of cytological and microbiological examinations. CONCLUSIONS: Measurement of serum amyloid A and haptoglobin was not proven useful for diagnosis of subclinical endometritis in Icelandic mares. Macroscopic changes in the fluid collected by lavage were not consistently indicative of infection, but when present they indicate inflammation in the uterus with a high probability.

Relationship between uterine biopsy score, endometrial infection and inflammation in the mare.

OBJECTIVE: Endometrial biopsy score is an accepted marker of uterine health and predicted fertility, and it has been suggested that endometrial alternations are correlated with susceptibility to persistent infectious endometritis. The objective of this study was to investigate associations of endometrial biopsy score with: 1) presence of polymorphonuclear cells (PMNs) in the epithelium and stratum compactum in histopathology; 2) presence of PMNs in cytology and 3) presence of infection in microbiology. MATERIALS AND METHODS: The material for examination was collected from 69 mares suspected for subclinical endometritis (bred three or more times unsuccessfully in the same breeding season) and from 15 maiden mares. Samples were collected by endometrial biopsy and cytobrush technique. RESULTS: Endometrial alterations (biopsy score IIA, IIB, III) were found in 64 of 82 mares (78%). There was an increase in PMN occurrence for grades IIA, IIB and III. When comparing grades and PMNs infiltration, we observed statistically significant differences between grades I and IIA (p  = 0.222) and grades I and IIB (p = 0.042) in samples collected by endometrial biopsy. Statistically significant differences were found in microbiological examination (biopsy p = 0.036; cytobrush p = 0.189), cytological examination (biopsy p = 0.040; cytobrush p = 0.079) and PMN infiltration (p    =    0.042) between mares with biopsy scores I and IIB. Furthermore, the highest percentage of infected mares was in grade IIA and IIB, and we found statistically significant differences between grades I and IIA (p = 0.043), and grades I and IIB (p = 0.036) in biopsy samples. We observed a tendency to higher prevalence of endometrial infection in mares with biopsy score IIA, IIB and III than with biopsy score I in samples collected using cytobrush technique. However, there were no statistical significant differences. CONCLUSION: Degenerative endometrial changes can predispose to uterine infection and inflammation. Our study shows that mares with endometrial score I are less predisposed to infection than mares with category IIA, IIB and III. Endometrial biopsy is a reliable diagnostic tool.

Effects of cycle stage and sampling procedure on interpretation of endometrial cytology in mares.

The aim of this study was to ascertain if (1) the stage of reproductive cycle influences cytological results, (2) cytology obtained from an endometrial biopsy is more accurate than cytology derived using the cytobrush, and (3) different methods used for evaluation of cytological samples produce similar diagnostic results. Material was collected from 46 mares in estrus, 48 in diestrus and from 33 mares in anestrus. Smears were evaluated using two criteria. In criterion I, a total of 300cells were counted and the percentage of polymorphonuclear cells was recorded. In criterion II, the number of polymorphonuclear cells/high power field was determined. Endometrial biopsy samples were also examined histologically, and comparisons were made between cytological and histological results. Regardless of the method used for sample collection, endometritis was diagnosed significantly more frequently if smears were evaluated according to criterion I. The agreement between the number of polymorphonuclear cells infiltrated into the endometrial luminal epithelium and stratum compactum and the cytology results for smears evaluated according to criterion I was fair for anestrous mares, poor for diestrous mares, and moderate for mares in estrus. When cytological smears were evaluated according to criterion II the agreement between the histological and cytological and results was always poor. In summary, cytological evaluation of the mare endometrium should be based on counting the numbers of polymorphonuclear cells in relation to epithelial cells rather than counting the number of polymorphonuclear cells in relation to high power field.