Gut-liver axis calibrates intestinal stem cell fitness.

The gut and liver are recognized to mutually communicate through the biliary tract, portal vein, and systemic circulation. However, it remains unclear how this gut-liver axis regulates intestinal physiology. Through hepatectomy and transcriptomic and proteomic profiling, we identified pigment epithelium-derived factor (PEDF), a liver-derived soluble Wnt inhibitor, which restrains intestinal stem cell (ISC) hyperproliferation to maintain gut homeostasis by suppressing the Wnt/ß-catenin signaling pathway. Furthermore, we found that microbial danger signals resulting from intestinal inflammation can be sensed by the liver, leading to the repression of PEDF production through peroxisome proliferator-activated receptor-α (PPARα). This repression liberates ISC proliferation to accelerate tissue repair in the gut. Additionally, treating mice with fenofibrate, a clinical PPARα agonist used for hypolipidemia, enhances colitis susceptibility due to PEDF activity. Therefore, we have identified a distinct role for PEDF in calibrating ISC expansion for intestinal homeostasis through reciprocal interactions between the gut and liver.

BRD4 directs mitotic cell division by inhibiting DNA damage.

BRD4 binds to acetylated histones to regulate transcription and drive cancer cell proliferation. However, the role of BRD4 in normal cell growth remains to be elucidated. Here we investigated the question by using mouse embryonic fibroblasts with conditional Brd4 knockout (KO). We found that Brd4KO cells grow more slowly than wild type cells: they do not complete replication, fail to achieve mitosis, and exhibit extensive DNA damage throughout all cell cycle stages. BRD4 was required for expression of more than 450 cell cycle genes including genes encoding core histones and centromere/kinetochore proteins that are critical for genome replication and chromosomal segregation. Moreover, we show that many genes controlling R-loop formation and DNA damage response (DDR) require BRD4 for expression. Finally, BRD4 constitutively occupied genes controlling R-loop, DDR and cell cycle progression. We suggest that BRD4 epigenetically marks those genes and serves as a master regulator of normal cell growth.

Ten Years of Animal Tuberculosis Monitoring in Free-Living European Bison (Bison bonasus) in Poland.

In the period 1996-2012, two outbreaks of animal tuberculosis were noted in the population of free-living European bison (Bison bonasus caucasicus) in the Bieszczady Mountains, Southern Poland. As the European bison is an endangered species and particularly susceptible to tuberculosis, not to mention a national icon, the decision was made to test all deceased bison for TB in Poland. The screened bison were obtained by elimination due to poor health or natural death. A total of 159 European bison have been examined over the last 10 years. The individuals came from four regions of Poland (Bialowieza Forest, Bieszczady Mountains, Borecka Forest, Knyszynska Forest), not only from the area where tuberculosis is still endemic. Mycobacterium bovis and Mycobacterium avium spp. hominisuis were identified in two different herds. The isolation of M. bovis from European bison was the first case described in Poland. So far, the only causative agent of tuberculosis identified in European bison in Poland, both in the wild and in captive herds, was Mycobacterium caprae. The isolated M. bovis spoligotype has not previously been registered in international spoligotype databases so far. The obtained results highlight the need to monitor TB in European bison in Poland.

The Influence of Storage Temperature and Packaging Technology on the Durability of Ready-to-Eat Preservative-Free Meat Bars with Dried Plasma.

Food business operators must include the results of shelf life testing in their HACCP plan. Ready-to-eat preservative-free meat products enriched with blood plasma are an unfathomable area of research in food safety. We tested modified atmosphere (80% N2 and 20% CO2) and vacuum packaged RTE preservative-free baked and smoked pork bars with dried blood plasma for Aerobic Plate Count, yeast and mould, lactic acid bacteria, Staphylococcus aureus, Enterobacteriaceae, Escherichia coli, and Campylobacter spp., and the presence of Listeria monocytogenes and Salmonella spp. during storage (temperatures from 4 to 34 °C) up to 35 days after production. The obtained data on the count of individual groups of microorganisms were subjected to analysis of variance (ANOVA) and statistically tested (Student's t-test with the Bonferroni correction); for temperatures at which there were statistically significant differences and high numerical variability, the trend of changes in bacterial counts were visualised using mathematical modelling. The results show that the optimal storage conditions are refrigerated temperatures (up to 8 °C) for two weeks. At higher temperatures, food spoilage occurred due to the growth of aerobic bacteria, lactic acid bacteria, yeast, and mould. The MAP packaging method was more conducive to spoilage of the bars, especially in temperatures over 8 °C.

Oncogenic RAS commandeers amino acid sensing machinery to aberrantly activate mTORC1 in multiple myeloma.

Oncogenic RAS mutations are common in multiple myeloma (MM), an incurable malignancy of plasma cells. However, the mechanisms of pathogenic RAS signaling in this disease remain enigmatic and difficult to inhibit therapeutically. We employ an unbiased proteogenomic approach to dissect RAS signaling in MM. We discover that mutant isoforms of RAS organize a signaling complex with the amino acid transporter, SLC3A2, and MTOR on endolysosomes, which directly activates mTORC1 by co-opting amino acid sensing pathways. MM tumors with high expression of mTORC1-dependent genes are more aggressive and enriched in RAS mutations, and we detect interactions between RAS and MTOR in MM patient tumors harboring mutant RAS isoforms. Inhibition of RAS-dependent mTORC1 activity synergizes with MEK and ERK inhibitors to quench pathogenic RAS signaling in MM cells. This study redefines the RAS pathway in MM and provides a mechanistic and rational basis to target this mode of RAS signaling.

Kinetic principles underlying pioneer function of GAGA transcription factor in live cells.

How pioneer factors interface with chromatin to promote accessibility for transcription control is poorly understood in vivo. Here, we directly visualize chromatin association by the prototypical GAGA pioneer factor (GAF) in live Drosophila hemocytes. Single-particle tracking reveals that most GAF is chromatin bound, with a stable-binding fraction showing nucleosome-like confinement residing on chromatin for more than 2 min, far longer than the dynamic range of most transcription factors. These kinetic properties require the full complement of GAF's DNA-binding, multimerization and intrinsically disordered domains, and are autonomous from recruited chromatin remodelers NURF and PBAP, whose activities primarily benefit GAF's neighbors such as Heat Shock Factor. Evaluation of GAF kinetics together with its endogenous abundance indicates that, despite on-off dynamics, GAF constitutively and fully occupies major chromatin targets, thereby providing a temporal mechanism that sustains open chromatin for transcriptional responses to homeostatic, environmental and developmental signals.

Wild Duck (Anas platyrhynchos) as a Source of Antibiotic-Resistant Salmonella enterica subsp. diarizonae O58-The First Report in Poland.

The "One Health" approach increasingly demonstrates the global spread of pathogenic microorganisms and their antimicrobial resistance in the environment, both in animals and humans. Salmonella enterica subsp. diarizonae is nowadays very often isolated from cold-blooded reptiles to a lesser extent from sheep, but unfortunately more and more often from humans. However, there are a few studies describing the isolation of Salmonella enterica subsp. diarizonae from migratory wild birds. The mallard duck (Anas platyrhynchos), a wild animal that traverses the continent of Eurasia, can be an excellent indicator of the spread of intestinal microbes as well as their resistance to antibiotics. This is the first report of the Salmonella enterica subsp. diarizonae detection in Poland in a migrating mallard duck. This research presented the identification difficulties associated with the isolation of Salmonella enterica subsp. diarizonae using three different biochemical tests and advanced serology tests. At the same time, we detected very high antimicrobial resistance in the isolated strain. By using the minimum inhibitory concentration (MIC) method, it was found that the isolated strain of S. enterica subsp. diarizonae has high antibiotic resistance against 14 of the 33 tested antimicrobials agents. The resistance genes that have been identified in S. enterica subsp. diarizonae include aadA, strA/strB, and blaTEM.

Predicting the risk of Alaria alata infestation in wild boar on the basis of environmental factors.

Alaria alata is an emerging parasite that poses a potential risk for those consuming game, pork, snails and frogs. One paratenic host of A. alata that is known to play an important role in its spread through its feeding habitats is the wild boar. However, no statistical analysis of the influence of aquatic environments and carnivores on the occurrence of A. alata in wild boars has yet been performed. The present study combines a small-scale analysis based on hunting districts in the Mazowieckie province with a large-scale analysis based on data for all provinces in Poland. We applied various modeling approaches, including logistic regression and a generalized linear model in order to determine the presence, intensity and prevalence of A. alata. We used the Alaria mesocercariae migration technique (AMT) to estimate the risk of A. alata among wild boar in a given hunting district or province. The small-scale analysis found that mesopredators (red fox (Vulpes vulpes)) and racoon dog (Nyctereutes procyinoides) were likely to influence A. alata infestation of wild boar; however, the effect was weak, probably as a result of the large home range size of these animals. The large-scale analysis found that wetlands influence the prevalence of A. alata in wild boar, with the estimated risk increasing in the north of the country; this finding is consistent with other studies. Our findings indicate that the occurrence of A. alata in wild boar requires analysis on many levels, and environmental factors play a key role in risk assessment.

Intra-Palpebral Tuberculin Skin Test and Interferon Gamma Release Assay in Diagnosing Tuberculosis Due to Mycobacterium caprae in European Bison (Bison bonasus).

Despite the threat posed by tuberculosis (TB) to the protected European bison (Bison bonasus), no validated TB tests exist for this species. This pilot study evaluates two tests based on detecting cellular immunity for this purpose: interferon gamma release assay (IGRA) and tuberculin skin test (TST). Ten animals were subjected to ante-mortem and post-mortem examinations. IGRA was performed using a commercial test, and the comparative TST was performed in the eyelids. The lesions were assessed post-mortem and material was collected for mycobacterial culture. The isolated strains were subjected to genotyping. At post-mortem examination, five out of ten individuals demonstrated both tuberculous lesions and positive culture results (Mycobacterium caprae). Compared to the palpebral TST, the findings of the IGRA are easier to interpret when diagnosing tuberculosis in European bison.

A serological survey of Toxoplasma gondii in polish pigs from organic farms, other housing systems and in pigs of different age groups.

BACKGROUND: The consumption of raw or undercooked meat, especially pork, and offal containing infective tissue cysts is suspected to be a significant route of infection with Toxoplasma gondii. Although the use of "animal-friendly pig production systems" ensuring direct contact with the natural environment offers ethical benefits, it limits the ability to ensure animal health; it may also increase the probability of infections by pathogens such as T. gondii, and thus their entry into the food chain. This study determines the seroprevalence of T. gondii in pigs from different housing systems and farms with different hygiene standards in Poland, as well as among pigs of different age groups from farms with high hygiene standards. In total 760 pig serum samples were examined for the presence of specific antibodies using the PrioCHECK® Toxoplasma Ab porcine commercial ELISA test (Prionics, Switzerland). RESULTS: Test results with PP ≥ 20% were regarded as positive, as indicated by the manufacturer. Antibodies to T. gondii were found in 193 of 760 (25.4%) tested sera. Regarding different housing systems, antibodies were found in 117 pigs: of these, 52.6% (61/116) were from organic farms, 40.9% (47/115) from farms with low hygiene standards, 5.4% (9/167) from farms with high hygiene standards and 0% (0/40) from a farm with a high level of biosecurity. Regarding age groups, antibodies were found in 76 animals on farms with high hygiene standards: 11.1% (7/63) were pigs younger than 3 months, 0% (0/60) aged 3-4 months, 12.3% (7/57) aged 5-6 months (final fattening stage) and 43.7% (62/142) were sows aged 9 months and older. CONCLUSIONS: Antibodies to T. gondii were most often found in pigs from organic and low-hygiene farms, as well as in pigs aged 9 months and older. Meat derived from seropositive animals can pose a potential source of infection for humans. As maternal antibodies to T. gondii can be present in the blood of piglets aged up to 3-4 months, serological examination is unjustified in piglets up to this age.

MYC assembles and stimulates topoisomerases 1 and 2 in a "topoisome".

High-intensity transcription and replication supercoil DNA to levels that can impede or halt these processes. As a potent transcription amplifier and replication accelerator, the proto-oncogene MYC must manage this interfering torsional stress. By comparing gene expression with the recruitment of topoisomerases and MYC to promoters, we surmised a direct association of MYC with topoisomerase 1 (TOP1) and TOP2 that was confirmed in vitro and in cells. Beyond recruiting topoisomerases, MYC directly stimulates their activities. We identify a MYC-nucleated "topoisome" complex that unites TOP1 and TOP2 and increases their levels and activities at promoters, gene bodies, and enhancers. Whether TOP2A or TOP2B is included in the topoisome is dictated by the presence of MYC versus MYCN, respectively. Thus, in vitro and in cells, MYC assembles tools that simplify DNA topology and promote genome function under high output conditions.

Microbiological and molecular monitoring for bovine tuberculosis in the Polish population of European bison (Bison bonasus).

INTRODUCTION AND OBJECTIVE: In recent years, bovine tuberculosis (BTB) has become one of the major health hazards facing the European bison (EB, Bison bonasus), a vulnerable species that requires active protection, including regular and effective health monitoring. Monitoring of zoonotic disease in wildlife is also an important part of public health protection. The aim of the study was to determine whether BTB still influences the EB population in Poland. MATERIAL AND METHODS: During 2017-2019, mandibular, retropharyngeal and mediastinal lymph nodes were collected from 90 EB during post-mortem examination, and then cultivated on Lowenstein-Jensen and Stonebrink media. Isolated strains were subjected to molecular analysis to determine the species, spoligotype and MIRU-VNTR pattern. RESULTS: Lesions were found in lymph nodes originating from eight EB (8.89%). Positive microbiological cultures for mycobacteria were obtained in samples from six (6.67%) EB. The isolated strains were identified as Mycobacterium caprae (material from four EB) and atypical mycobacteria (material from two EB). For M. caprae strains spoligotype M. bovis 4_CA 1600 was identified and the MIRU-VNTR pattern was identified as 345751355413232. CONCLUSIONS: It is recommended that this potentially dangerous disease should be monitored in EB via a comprehensive strategy based on a combination of microbiological and molecular methods. Such monitoring will protect the health of both animals and humans.

Spatiotemporal coordination of transcription preinitiation complex assembly in live cells.

Transcription initiation by RNA polymerase II (RNA Pol II) requires preinitiation complex (PIC) assembly at gene promoters. In the dynamic nucleus, where thousands of promoters are broadly distributed in chromatin, it is unclear how multiple individual components converge on any target to establish the PIC. Here we use live-cell, single-molecule tracking in S. cerevisiae to visualize constrained exploration of the nucleoplasm by PIC components and Mediator's key role in guiding this process. On chromatin, TFIID/TATA-binding protein (TBP), Mediator, and RNA Pol II instruct assembly of a short-lived PIC, which occurs infrequently but efficiently within a few seconds on average. Moreover, PIC exclusion by nucleosome encroachment underscores regulated promoter accessibility by chromatin remodeling. Thus, coordinated nuclear exploration and recruitment to accessible targets underlies dynamic PIC establishment in yeast. Our study provides a global spatiotemporal model for transcription initiation in live cells.

A comparison of detection methods of Alaria alata mesocercariae in wild boar (Sus scrofa) meat.

Distomum musculorum suis (DMS), the mesocercariae of Alaria alata, is typically found accidently during examination of wild boar meat for Trichinella spp. The aim of the study was to compare DMS detection methods. Briefly, 232 wild boar meat samples were tested by mesocercariae migration technique (AMT) as a reference method; of these, 104 were found to be positive. Selected positive samples were tested again with the three other methods: compressorium method (Compressor), digestion with magnetic stirrer (Digestion) and by modified digestion with Pancreatin® bile and pancreatic enzymes (D + P). The results were analyzed by logistic regression, the non-parametric Kruskal-Wallis test and the Mann-Whitney U test. Of the 43 samples found positive by the AMT, 20 were found positive by Digestion and 25 by D + P. The Compressor identified DMS in seven of the 19 tested samples. The Digestion and D + P methods gave similar intensities (P = 0.506), i.e. 1.4 and 1.3 DMS respectively, but the AMT detected seven times higher number of parasites. The probability of detection of DMS in the meat sample by the Digestion or by D + P was higher than 0.5 when at least seven (Digestion) or five (D + P) DMS were present in the sample (AMT). The Compressor was the least sensitive method: at least 14 DMS must be present in the meat sample for detection. AMT should be considered the most accurate method of DMS detection.

Prevalence of Toxoplasma gondii antibodies in wild boar (Sus scrofa) from Strzalowo Forest Division, Warmia and Mazury Region, Poland.

INTRODUCTION AND OBJECTIVE: Toxoplasmosis is an important zoonosis caused by a protozoan, Toxoplasma gondii. Raw or undercooked venison may be a source of infection in humans. The aim of the study was to determine the prevalence of T. gondii antibodies in wild boar from the Strzalowo Forest Division of the Warmia and Mazury Region of Poland. MATERIAL AND METHODS: A total of 90 samples were collected from 50 wild boar: 40 from both tongue and diaphragm muscles, 4 from diaphragm muscles and six from tongue muscles. Samples were analyzed using the commercial PrioCHECK® Toxoplasma Ab porcine ELISA, according to the manufacturer's instructions. RESULTS: T. gondii antibodies were detected in 24 of 50 (48%) tested animals. T. gondii antibodies were detected in 40 of 90 (44.4%) tested samples (21 of tongue muscles and 19 of diaphragm muscles). In the 40 wild boar that provided samples of meat juice from the tongue and diaphragm muscles, specific antibodies were more prevalent in the tongue (20 of 40 animals - 50%) than in the diaphragm muscles (17 of 40 animals - 42.5%). CONCLUSIONS: The study revealed a high percentage of wild boar seropositive to T. gondii. Muscle samples to obtain meat juice are easily available and simple to collect, even on the hunting grounds, which makes them suitable material for detecting T. gondii antibodies in wild boar. Wild boar are essential to T. gondii circulation in the environment, and raw or undercooked venison may be a source of human infections with this parasite.

New aspects of the esophageal histology of the domestic goat (Capra hircus) and European roe deer (Capreolus capreolus).

The present study examines the esophageal wall of animals from two distinct families of the Ruminantia: domestic goats and European roe deer. Five fragments were collected from the entire length of the esophageal wall in five goats and four roe deer and subjected to microscopic and morphometric analyses. All layers of the esophageal wall except the tela submucosa were found to be thicker in the goats. In both species, the esophagus was lined by parakeratinized stratified squamous epithelium, and the tela submucosa was deprived of glands along its entire length. However, the structure of the lamina muscularis mucosae was better developed in goats: it was found to be discontinuous in the proximal part, and then became fused in the cervical part, that is around the most proximal quarter of its length. In contrast, in roe deer, the lamina muscularis mucosae began as sparse, thin muscle bundles at the pharyngeal-esophageal junction, which thickened and clustered further down the esophagus, but did not fuse. Our findings regarding the microscopic structure of the ruminant esophagus are not fully consistent with the widely-accepted view and suggest that the histological structure of the esophagus demonstrates interspecies variation within this large suborder. More precisely, species-specific differences can be seen regarding the presence of esophageal glands and parakeratinized epithelium, and in the organization of the lamina muscularis mucosae.

Dual-view light-sheet imaging through a tilted glass interface using a deformable mirror.

Light-sheet microscopy has become indispensable for imaging developing organisms, and imaging from multiple directions (views) is essential to improve its spatial resolution. We combine multi-view light-sheet microscopy with microfluidics using adaptive optics (deformable mirror) which corrects aberrations introduced by the 45o-tilted glass coverslip. The optimal shape of the deformable mirror is computed by an iterative algorithm that optimizes the point-spread function in two orthogonal views. Simultaneous correction in two optical arms is achieved via a knife-edge mirror that splits the excitation path and combines the detection paths. Our design allows multi-view light-sheet microscopy with microfluidic devices for precisely controlled experiments and high-content screening.

The first visually-guided bronchoscopy in European bison (Bison bonasus) - An additional tool in the diagnosis of bovine tuberculosis?

The European bison (Bison bonasus, EB) is an endangered species, and as about 1/3 of its global population is found in Poland, it is particularly important that Polish herds should be monitored. One particular concern is tuberculosis, which is not a marginal problem in wildlife in Poland, and has been microbiologically confirmed in EB, wolves (Canis lupus) and wild boar (Sus scrofa). However, ante mortem diagnosis of tuberculosis in EB is troublesome. Therefore, the present paper evaluates the potential of bronchoscopy as a diagnostic tool. Seven EB were studied, four of which were found to be naturally infected with M. caprae; in two of these, endoscopy identified abnormalities in the respiratory tract ante mortem. Therefore, despite some limitations, endoscopy can be an additional tool for diagnosing tuberculosis in EB, especially in highly valuable animals, and to assess the stage of the disease.

The occurrence of Alaria alata mesocercariae in wild boars (Sus scrofa) in north-eastern Poland.

Various species of mammals, including humans and wild boars, can serve as paratenic hosts of Alaria alata mesocercariae - Distomum musculorum suis (DMS). Cases of DMS can be reliably detected by the recent introduction of the A. alata mesocercariae migration technique (AMT). The aim of this study is to present current data on the occurrence of DMS in wild boars in north-eastern Poland, and to compare the findings with those obtained in other European countries. A. alata was identified in 98 of 221 (44.3%) tissue samples of wild boar taken from five provinces in north-eastern Poland during the hunting seasons 2015-2016 and 2016-2017 based on AMT analysis. Positive results were found in all studied regions, but the percentage of infected individuals ranged from 26.3% in the Pomorskie province to 65.5% in the Warminsko-Mazurskie province. The mean number of larvae exceeded seven larvae per 30 g sample for three provinces: Pomorskie, Mazowieckie and Lubelskie. In turn, lower values were found in the Warminsko-Mazurskie province (3.3 larvae per 30 g), and the lowest in the Kujawsko-Pomorskie province (1.8 larvae per 30 g). The occurrence and intensity of A. alata infestation in wild boars was found to depend on the environment in which they live. Neither the sex or the age of the wild boar appeared to influence the occurrence nor the intensity of infestation.