Gut Microbiota Composition of Insectivorous Synanthropic and Fructivorous Zoo Bats: A Direct Metagenomic Comparison.

Bats are natural reservoirs for many emerging viral diseases. That is why their virome is widely studied. But at the same time, studies of their bacterial gut microbiota are limited, creating a degree of uncertainty about the role of bats in global microbial ecology. In this study, we analyzed gut microbiota of insectivorous Nyctalus noctula and Vespertilio murinus from rehabilitation centers from Rostov-on-Don and Moscow, respectively, and fructivorous Carollia perspicillata from the Moscow Zoo based on V3-V4 16S rRNA metagenomic sequencing. We revealed that microbial diversity significantly differs between the insectivorous and fructivorous species studied, while the differences between N. noctula and V. murinus are less pronounced, which shows that bats' gut microbiota is not strictly species-specific and depends more on diet type. In the gut microbiota of synanthropic bats, we observed bacteria that are important for public health and animal welfare such as Bacteroides, Enterobacter, Clostridiaceae, Enterococcus, Ureaplasma, Faecalibacterium, and Helicobacter, as well as some lactic acid bacteria such as Pediococcus, Lactobacillus, Lactococcus, and Weisella. All these bacteria, except for Bacteroides and Weisella, were significantly less abundant in C. perspicillata. This study provides a direct metagenomic comparison of synanthropic insectivorous and zoo fructivorous bats, suggesting future directions for studying these animals' role in microbial ecology.

Confocal Laser and Electron Microscopic Investigation of Gap Junctions in Anaplastic Astrocytomas.

Connexin 43 (Cx43) is a multifunction protein that forms gap junction channels and hemichannels and is suggested to play an essential role in oxygen-glucose deprivation, induced via neuroinflammation during astrocytoma expansion into healthy tissue. To prove this assumption we studied connexin 43 localisation and ultrastructure of gap junctions in samples of malignant brain tumour (anaplastic astrocytomas grade III). For confocal laser microscopy, vibratome sections of tumour fragments were incubated in a mixture of primary antibodies to connexin 43 and glial fibrillary acidic protein (GFAP), then in a mixture of secondary antibodies conjugated with a fluorescent label. After the immunofluorescence study, sections were washed in phosphate buffer, additionally postfixed with 1% OsO4 solution, dehydrated and embedded in epoxy resin by a plane-parallel method. Ultra-thin sections obtained from these samples were contrasted with uranyl acetate and lead citrate and viewed under a Jem 1011 electron microscope. Confocal laser examination detected a positive reaction to Cx43 in the form of point fluorescence. These points were of various sizes. Most of them were localised around or at the intersection of small processes containing GFAP. Electron microscopy of the tumour samples containing the most significant number of Cx43 revealed single and closely spaced gap junctions with a typical ultrastructure on the processes and bodies of tumour cells. Sequential analysis in the fields of view revealed 62 gap junctions in the area of 100 µm2. Numerous gap junctions in anaplastic astrocytomas revealed in our study may indicate electrotonic and metabolic transmission between glioma cells, possibly promoting its progression.

Visualization of different anatomical parts of the enucleated human eye using X-ray micro-CT imaging.

Micro-CT visualization allows reconstruction of eye structures with the resolution of light microscopy and estimation of tissue densities. Moreover, this method excludes damaging procedures and allows further histological staining due to the similar steps in the beginning. We have shown the feasibility of the lab-based micro-CT machine usage for visualization of clinically important compartments of human eye such as trabecular outflow pathway, retina, iris and ciliary body after pre-treatment with iodine in ethanol. We also identified the challenges of applying this contrasting technique to lens, cornea, and retina and proposed alternative staining methods for these tissues. Thereby this work provides a starting point for other studies for imaging of human eyes in normal and pathological conditions using lab-based micro-CT systems.

Modulation of Swine Gut Microbiota by Phytogenic Blends and High Concentrations of Casein in a Validated Swine Large Intestinal In Vitro Model.

Phytogenic feed additives are gaining popularity in livestock as a replacement for antibiotic growth promotors. Some phytogenic blends (PB) positively affect the production performance, inhibit pathogens within the gut microbiota, and improve the overall health of farm animals. In this study, a swine large intestine in vitro model was used to evaluate the effect of two PBs, alone or in combination with casein, on swine gut microbiota. As a result, the combination of casein with PB1 had the most beneficial effects on swine gut microbiota, as it increased the relative abundance of some commensal bacteria and two genera (Lactobacillus and Oscillospiraceae UCG-002), which are associated with greater production performance in pigs. At the same time, supplementation with PBs did not lead to an increase in opportunistic pathogens, indicating their safety for pigs. Both PBs showed fewer changes in swine gut microbiota compared to interventions with added casein. In contrast, casein supplementation significantly increased beta diversity and the relative abundance of commensal as well as potentially beneficial bacteria. In conclusion, the combination of casein with PBs, in particular PB1, had the most beneficial effects among the studied supplements in vitro, with respect to microbiota modulation and metabolite production, although this data should be proven in further in vivo studies.

Cultivable Gut Microbiota in Synanthropic Bats: Shifts of Its Composition and Diversity Associated with Hibernation.

The role of bats in the global microbial ecology no doubt is significant due to their unique immune responses, ability to fly, and long lifespan, all contributing to pathogen spread. Some of these animals hibernate during winter, which results in the altering of their physiology. However, gut microbiota shifts during hibernation is little studied. In this research, we studied cultivable gut microbiota composition and diversity of Nyctalus noctula before, during, and after hibernation in a bat rehabilitation center. Gut microorganisms were isolated on a broad spectrum of culture media, counted, and identified with mass spectrometry. Linear modeling was used to investigate associations between microorganism abundance and N. noctula physiological status, and alpha- and beta-diversity indexes were used to explore diversity changes. As a result, most notable changes were observed in Serratia liquefaciens, Hafnia alvei, Staphylococcus sciuri, and Staphylococcus xylosus, which were significantly more highly abundant in hibernating bats, while Citrobacter freundii, Klebsiella oxytoca, Providencia rettgeri, Citrobacter braakii, and Pedicoccus pentosaceus were more abundant in active bats before hibernation. The alpha-diversity was the lowest in hibernating bats, while the beta-diversity differed significantly among all studied periods. Overall, this study shows that hibernation contributes to changes in bat cultivable gut microbiota composition and diversity.

Detection of coronaviruses in insectivorous bats of Fore-Caucasus, 2021.

Coronaviruses (CoVs) pose a huge threat to public health as emerging viruses. Bat-borne CoVs are especially unpredictable in their evolution due to some unique features of bat physiology boosting the rate of mutations in CoVs, which is already high by itself compared to other viruses. Among bats, a meta-analysis of overall CoVs epizootiology identified a nucleic acid observed prevalence of 9.8% (95% CI 8.7-10.9%). The main objectives of our study were to conduct a qPCR screening of CoVs' prevalence in the insectivorous bat population of Fore-Caucasus and perform their characterization based on the metagenomic NGS of samples with detected CoV RNA. According to the qPCR screening, CoV RNA was detected in 5 samples, resulting in a 3.33% (95% CI 1.1-7.6%) prevalence of CoVs in bats from these studied locations. BetaCoVs reads were identified in raw metagenomic NGS data, however, detailed characterization was not possible due to relatively low RNA concentration in samples. Our results correspond to other studies, although a lower prevalence in qPCR studies was observed compared to other regions and countries. Further studies should require deeper metagenomic NGS investigation, as a supplementary method, which will allow detailed CoV characterization.

Anti-obesity effects of Chenpi: an artificial gastrointestinal system study.

The gut microbiota plays a significant role in human health; however, the complex relationship between gut microbial communities and host health is still to be thoroughly studied and understood. Microbes in the distal gut contribute to host health through the biosynthesis of vitamins and essential amino acids and the generation of important metabolic by-products from dietary components that are left undigested by the small intestine. Aged citrus peel (Chenpi) is used in traditional Chinese medicine to lower cholesterol, promote weight loss and treat various gastrointestinal symptoms. This study investigated how the microbial community changes during treatment with Chenpi using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME). Two preparations of Chenpi extract were tested: Chenpi suspended in oil only and Chenpi in a viscoelastic emulsion. Short-chain fatty acids (SCFAs) were measured during treatment to monitor changes in the microbial community of the colon presenting a decrease in production for acetic, propionic and butyric acid (ANOVA (P < 0.001) during the 15 days of treatment. 16S rRNA sequencing of microbial samples showed a clear difference between the two treatments at the different sampling times (ANOSIM P < 0.003; ADOSIM P < 0.002 [R2 = 69%]). Beta diversity analysis by PcoA showed differences between the two Chenpi formulations for treatment day 6. These differences were no longer detectable as soon as the Chenpi treatment was stopped, showing a reversible effect of Chenpi on the human microbiome. 16S rRNA sequencing of microbial samples from the descending colon showed an increase in Firmicutes for the treatment with the viscoelastic emulsion. At the genus level, Roseburia, Blautia, Subdoligranulum and Eubacterium increased in numbers during the viscoelastic emulsion treatment. This study sheds light on the anti-obesity effect of a polymethoxyflavone (PMFs)-enriched Chenpi extract and creates a foundation for the identification of 'obesity-prevention' biomarkers in the gut microbiota.

Beneficial Effects of Spore-Forming Bacillus Probiotic Bacteria Isolated From Poultry Microbiota on Broilers' Health, Growth Performance, and Immune System.

Probiotics are known for their beneficial effects on poultry health and wellbeing. One promising strategy for discovering Bacillus probiotics is selecting strains from the microbiota of healthy chickens and subsequent screening for potential biological activity. In this study, we focused on three probiotic strains isolated from the gastrointestinal tract of chickens bred in different housing types. In addition to the previously reported poultry probiotic Bacillus subtilis KATMIRA1933, three strains with antimutagenic and antioxidant properties Bacillus subtilis KB16, Bacillus subtilis KB41, and Bacillus amyloliquefaciens KB54, were investigated. Their potential effects on broiler health, growth performance, and the immune system were evaluated in vivo. Two hundred newly hatched Cobb500 broiler chickens were randomly divided into five groups (n = 40). Four groups received a standard diet supplemented with the studied bacilli for 42 days, and one group with no supplements was used as a control. Our data showed that all probiotics except Bacillus subtilis KATMIRA1933 colonized the intestines. Treatment with Bacillus subtilis KB54 showed a significant improvement in growth performance compared to other treated groups. When Bacillus subtilis KB41 and Bacillus amyloliquefaciens KB54 were applied, the most significant immune modulation was noticed through the promotion of IL-6 and IL-10. We concluded that Bacillus subtilis KB54 supplementation had the largest positive impact on broilers' health and growth performance.

Factors Associated With Highest Symptoms of Anxiety During COVID-19: Cross-Cultural Study of 23 Countries.

The COVID-19 restrictions have impacted people's lifestyles in all spheres (social, psychological, political, economic, and others). This study explored which factors affected the level of anxiety during the time of the first wave of COVID-19 and subsequent quarantine in a substantial proportion of 23 countries, included in this study. The data was collected from May to August 2020 (5 June 2020). The sample included 15,375 participants from 23 countries: (seven from Europe: Belarus, Bulgaria, Croatia, Hungary, Italy, Romania, Russia; 11 from West, South and Southeast Asia: Armenia, India, Indonesia, Iran, Iraq, Jordan, Malaysia, Pakistan, Saudi Arabia, Thailand, Turkey; two African: Nigeria and Tanzania; and three from North, South, and Central America: Brazil, Canada, United States). Level of anxiety was measured by means of the 7-item Generalized Anxiety Disorder Scale (GAD-7) and the 20-item first part of The State-Trait Anxiety Inventory (STAI)-State Anxiety Inventory (SAI). Respondents were also asked about their personal experiences with COVID-19, attitudes toward measures introduced by governments, changes in attitudes toward migrants during a pandemic, family income, isolation conditions, etc. The factor analysis revealed that four factors explained 45.08% of variance in increase of anxiety, and these components were interpreted as follows: (1) personal awareness of the threat of COVID-19, (2) personal reaction toward officially undertaken measures and attitudes to foreigners, (3) personal trust in official sources, (4) personal experience with COVID-19. Three out of four factors demonstrated strong associations with both scales of anxiety: high level of anxiety was significantly correlated with high level of personal awareness of the threat of COVID-19, low level of personal reaction toward officially undertaken measures and attitudes to foreigners, and high level of presence of personal experience with COVID-19. Our study revealed significant main effects of sex, country, and all four factors on the level of anxiety. It was demonstrated that countries with higher levels of anxiety assessed the real danger of a pandemic as higher, and had more personal experience with COVID-19. Respondents who trusted the government demonstrated lower levels of anxiety. Finally, foreigners were perceived as the cause of epidemic spread.

Antimicrobial and Anti-Biofilm Activity of Polymyxin E Alone and in Combination with Probiotic Strains of Bacillus subtilis KATMIRA1933 and Bacillus amyloliquefaciens B-1895 against Clinical Isolates of Selected Acinetobacter spp.: A Preliminary Study.

Acinetobacter spp., the nosocomial pathogen, forms strong biofilms and is resistant to numerous antibiotics, causing persistent infections. This study investigates the antibacterial and anti-biofilm activity of polymyxin E alone and in combination with the cell-free supernatants (CFS) of the tested probiotic bacilli, Bacillus subtilis KATMIRA1933 and Bacillus amyloliquefaciens B-1895 against the selected Acinetobacter spp. starins. Three isolates of Acinetobacter spp., designated as Acinetobacter spp. isolate 1; Acinetobacter spp. isolate 2, and Acinetobacter spp. isolate 3, were collected from patients with burns, wounds, and blood infections, respectively. Bacterial identification and antibiotic susceptibility testing were conducted using the VITEK2 system. Auto-aggregation and coaggregation of the tested bacilli strains with the selected Acinetobacter spp. isolates were evaluated. A disk diffusion assay was used to identify the microorganism's susceptibility to the selected antibiotics, alone and in combination with the CFS of the bacilli. The MIC and MBIC (minimum inhibitory and minimum biofilm inhibitory concentrations) of polymyxin E combined with bacilli CFS were determined. Acinetobacter spp. isolates were (i) sensitive to polymyxin E, (ii) able to form a strong biofilm, and (iii) resistant to the tested antibiotics and the CFS of tested bacilli. Significant inhibition of biofilm formation was noticed when CFS of the tested bacilli were combined with polymyxin E. The bacilli CFS showed synergy with polymyxin E against planktonic cells and biofilms of the isolated pathogens.

Halocins, natural antimicrobials of Archaea: Exotic or special or both?

Haloarchaea are adapted to survive under extreme saline conditions by accumulating osmolytes and salts to counteract the high osmotic pressure in their habitats. As a consequence, their proteins have evolved to remain active, or even most active, at very high ionic strength. Halocins are proteinaceous antimicrobial substances that are ribosomally-synthesized by haloarchaea and they provide the producers an advantage in the competition for nutrients and ecological niches. These antimicrobials are stable at high temperature, elevated salt concentrations, and alkaline pH conditions. These properties have endowed them with great potential in diverse biotechnological applications, which involve extreme processing conditions (such as high salt concentrations, high pressure, or high temperatures). They kill target cells by inhibition of Na+/H+ antiporter in the membrane or modification/disruption of the cell membrane leading to cell lysis. In general, the taxonomy of haloarchaea and their typical phenotypic and genotypic characteristics are well studied; however, information regarding their halocins, especially aspects related to genetics, biosynthetic pathways, mechanism of action, and structure-function relationship is very limited. A few studies have demonstrated the potential applications of halocins in the preservation of salted food products and brine-cured hides in leather industries, protecting the myocardium from ischemia and reperfusion injury, as well as from life-threatening diseases such as cardiac arrest and cancers. In recent years, genome mining has been an essential tool to decipher the genetic basis of halocin biosynthesis. Nevertheless, this is likely the tip of the iceberg as genome analyses have revealed many putative halocins in databases waiting for further investigation. Identification and characterization of this source of halocins may lead to antimicrobials for future therapeutics and/or food preservation. Hence, the present review analyzes different aspects of halocins such as biosynthesis, mechanism of action against target cells, and potential biotechnological applications.

Anti-biofilm Potential of Elletaria cardamomum Essential Oil Against Escherichia coli O157:H7 and Salmonella Typhimurium JSG 1748.

Foodborne pathogens, microbial recurrent infections, and antibiotic resistance have driven researchers to explore natural compounds as safe alternative antimicrobials. In this study, the chemical profile, antimicrobial, and mutagenic activities of the Elletaria cardamomum essential oil were investigated. GC-MS analysis identified the major bioactive components as α-terpinyl acetate, 1,8-cineole, linalool acetate, and sabinene, at concentrations of 34.95, 25.30, 8.13, and 5.48% respectively, of the essential oil's content. Regarding antimicrobial activity, the minimum inhibitory concentration of green cardamom essential oil was 1% against Escherichia coli O157:H7 and Pseudomonas aeruginosa ATCC 14213. Green cardamom essential oil, when used at concentrations of 0.015, 0.031, 0.062, and 0.125% (v/v) prevented biofilm formation of Escherichia coli O157:H7 by 64.29, 65.98, 70.41, and 85.59%, respectively. Furthermore, these concentrations inhibited 6.13, 45.50, 49.45, and 100%, respectively, of the Salmonella Typhimurium JSG 1748 biofilm. A mutagenicity assay confirmed that green cardamom essential oil has no demonstrable mutagenic activity against the tested strains. The study's findings suggest that green cardamom derived bioactive compounds are safe organic antimicrobials, effective in controlling biofilm formation by Gram-negative pathogens. Moreover, such compounds could possibly be used in the food industry (e.g., bakery, dairy, meat, and other food products) as a safe alternative to chemical preservatives (antimicrobials) to enhance shelf life by improving the antimicrobial status while at the same time imparting a pleasant and appealing aroma for consumers.

Probiotic Bacilli Inhibit Salmonella Biofilm Formation Without Killing Planktonic Cells.

Salmonellosis is a foodborne infection caused by Salmonella. Domestic poultry species are one of the main reservoirs of Salmonella, which causes the foodborne infection salmonellosis, and are responsible for many cases of animal-to-human transmission. Keeping backyard chickens is now a growing trend, increasing the frequency of direct contact with the flock and, by consequence, the incidence of Salmonella infections. Bacillus subtilis KATMIRA1933 and Bacillus amyloliquefaciens B-1895 are probiotic bacilli that produce the bacteriocins subtilosin A and subtilin, respectively. The antimicrobial activity of the two strains was determined against the reference strain Micrococcus luteus ATCC 10420. The cell-free supernatant of B. subtilis KATMIRA1933 inhibited biofilm formation by Salmonella enterica subsp. enterica serovar Hadar, Salmonella enterica subsp. enterica serovar Enteritidis phage type 4, and Salmonella enterica subsp. enterica serovar Thompson by 51.1, 48.3, and 56.9%, respectively. The cell-free supernatant of B. amyloliquefaciens B-1895 inhibited the biofilm formation of these Salmonella strains by 30.4, 28.6, and 35.5%, respectively. These findings suggest that the bacillus strains may have the potential to be used as probiotics and antibiotic alternatives for the control of Salmonella in poultry. The number of planktonic cells was unaffected by treatment with the cell-free supernatant. A co-culture of the Salmonella strains with either bacilli showed no signs of growth inhibition, suggesting that it might have been quorum sensing that is affected by the two Bacillus strains.

Gut microbiota of bats: pro-mutagenic properties and possible frontiers in preventing emerging disease.

Bats are potential natural reservoirs for emerging viruses, causing deadly human diseases, such as COVID-19, MERS, SARS, Nipah, Hendra, and Ebola infections. The fundamental mechanisms by which bats are considered "living bioreactors" for emerging viruses are not fully understood. Some studies suggest that tolerance to viruses is linked to suppressing antiviral immune and inflammatory responses due to DNA damage by energy generated to fly. Our study reveals that bats' gut bacteria could also be involved in the host and its microbiota's DNA damage. We performed screening of lactic acid bacteria and bacilli isolated from bats' feces for mutagenic and oxidative activity by lux-biosensors. The pro-mutagenic activity was determined when expression of recA increased with the appearance of double-strand breaks in the cell DNA, while an increase of katG expression in the presence of hydroxyl radicals indicated antioxidant activity. We identified that most of the isolated bacteria have pro-mutagenic and antioxidant properties at the same time. This study reveals new insights into bat gut microbiota's potential involvement in antiviral response and opens new frontiers in preventing emerging diseases originating from bats.

Probiotic biomarkers and models upside down: From humans to animals.

Probiotics development for animal farming implies thorough testing of a vast variety of properties, including adhesion, toxicity, host cells signaling modulation, and immune effects. Being diverse, these properties are often tested individually and using separate biological models, with great emphasis on the host organism. Although being precise, this approach is cost-ineffective, limits the probiotics screening throughput and lacks informativeness due to the 'one model - one test - one property' principle. There is а solution coming from human-derived cells and in vitro systems, an extraordinary example of human models serving animal research. In the present review, we focus on the current outlooks of employing human-derived in vitro biological models in probiotics development for animal applications, examples of such studies and the analysis of concordance between these models and host-derived in vivo data. In our opinion, human-cells derived screening systems allow to test several probiotic properties at once with reasonable precision, great informativeness and less expenses and labor effort.

Fermented Duckweed as a Potential Feed Additive with Poultry Beneficial Bacilli Probiotics.

In this study, the duckweed varieties Lemna minor, Spirodela polyrhiza, and a commercially processed duckweed food supplement were investigated as potential substrates for the propagation of two probiotic Bacillus strains, B. subtilis KATMIRA1933 and B. amyloliquefaciens B-1895. Both L. minor and S. polyrhiza were found to be suitable substrates for the propagation of both bacilli, with 8.47-9.48 Log CFU/g and 10.17-11.31 Log CFU/g after 24 and 48 h growth on the substrates, respectively. The commercial duckweed product was a less favorable substrate, with growth reaching a maximum of 7.89-8.91 CFU/g after 24 h with no further growth after 48 h. Growth and adherence of the bacilli to the three products were confirmed via electron microscopy. These strains have demonstrated health-promoting benefits for poultry and thereby have the potential to enhance duckweed as an animal feed through the process of fermentation. Duckweed has been shown to be a promising alternative resource for protein and has the opportunity to become a valuable resource in multiple industries as a potential means to increase sustainability, food security, and reduce environmental impact.

The Effect of Immunobiotic/Psychobiotic Lactobacillus acidophilus Strain INMIA 9602 Er 317/402 Narine on Gut Prevotella in Familial Mediterranean Fever: Gender-Associated Effects.

Possible mechanisms involved in sex-dependent differences in the gut microbiota have a growing interest worldwide, but the effects of probiotics dependence on the gender of the host have remained outside of researchers' attention until now. Previously, our research data described gender-specific differences in the gut microbiota of Armenian Familial Mediterranean fever (FMF) patients. Taking into account the possible association of Prevotella spp. with depressive disorders, the aim of the current investigations was an evaluation of changes in the abundance of gut Prevotella of FMF patients in association with the patient's depression and gender. The differences between healthy and FMF diseased gut microbiota in terms of Prevotella abundance were revealed. In addition, the gender-dependent effects of immunobiotic/psychobiotic Narine on the abundance of gut Prevotella of FMF patients and patients' depression scores were shown by us in this study.

A Review of the Effects and Production of Spore-Forming Probiotics for Poultry.

One of the main problems in the poultry industry is the search for a viable replacement for antibiotic growth promoters. This issue requires a "one health" approach because the uncontrolled use of antibiotics in poultry can lead to the development of antimicrobial resistance, which is a concern not only in animals, but for humans as well. One of the promising ways to overcome this challenge is found in probiotics due to their wide range of features and mechanisms of action for health promotion. Moreover, spore-forming probiotics are suitable for use in the poultry industry because of their unique ability, encapsulation, granting them protection from the harshest conditions and resulting in improved availability for hosts' organisms. This review summarizes the information on gastrointestinal tract microbiota of poultry and their interaction with commensal and probiotic spore-forming bacteria. One of the most important topics of this review is the absence of uniformity in spore-forming probiotic trials in poultry. In our opinion, this problem can be solved by the creation of standards and checklists for these kinds of trials such as those used for pre-clinical and clinical trials in human medicine. Last but not least, this review covers problems and challenges related to spore-forming probiotic manufacturing.

Mechanisms of Candida Resistance to Antimycotics and Promising Ways to Overcome It: The Role of Probiotics.

Pathogenic Candida and infections caused by those species are now considered as a serious threat to public health. The treatment of candidiasis is significantly complicated by the increasing resistance of pathogenic strains to current treatments and the stagnant development of new antimycotic drugs. Many species, such as Candida auris, have a wide range of resistance mechanisms. Among the currently used synthetic and semi-synthetic antifungal drugs, the most effective are azoles, echinocandins, polyenes, nucleotide analogs, and their combinations. However, the use of probiotic microorganisms and/or the compounds they produce is quite promising, although underestimated by modern pharmacology, to control the spread of pathogenic Candida species.

X-ray micro-computed tomography in the assessment of penile cavernous fibrosis in a rabbit castration model.

BACKGROUND: Current assessment methods of penile cavernous fibrosis in animal models have limitations due to the inability to provide complex and volume analysis of fibrotic alterations. OBJECTIVE: The aim was to evaluate micro-computed tomography for assessment of cavernous fibrosis and compare it with histological, histochemical, immunohistochemical, and RT-PCR analysis. MATERIALS AND METHODS: A controlled trial was performed involving 25 New Zealand male rabbits with induced testosterone deficiency by orchidectomy. Penile samples were obtained before and after 7, 14, 21, and 84 days from orchidectomy. We consistently performed (a) gray value analysis of corpora cavernosa 3D models reconstructed after micro-computed tomography, (b) morphometry of smooth muscles/connective tissue ratio, collagen type I/III ratio, and area of TGF-beta-1 expression in corpora cavernosa, and (c) RT-PCR of TGF-beta-1 expression. RESULTS: Micro-computed tomography allowed visualization of penile structures at a resolution comparable to light microscopy. Gray values of corpora cavernosa decreased from 1673 (1512-1773) on the initial day to 1184 (1089-1232) on the 21st day (p < 0.005). However, on the 84th day, it increased to 1610 (1551-1768). On 21st and 84th days, there was observed a significant decrease in smooth muscle/connective tissue ratio and a significant increase in collagen type I/III ratio (p < 0.05). TGF-beta1 expression increased on the 84th day according to immunohistochemistry (p < 0.005). RT-PCR was impossible to conduct due to the absence of RNA in obtained samples after micro-CT. DISCUSSION AND CONCLUSIONS: Micro-computed tomography provided 3D visualization of entire corpora cavernosa and assessment of radiodensity alterations by gray value analysis in fibrosis progression. We speculate that gray value changes at early and late fibrosis stages could be related to tissue reorganization. RT-PCR is impossible to conduct on tissue samples studied by micro-CT due to RNA destruction. We also suggest that micro-computed tomography could negatively affect the immunohistochemical outcome, as a significant increase of TGF-beta-1 expression occurs later than histological fibrotic signs.