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In this study, we report the results of two non-invasive optical methods, Raman microscopy (RM) and polarization-sensitive digital holographic imaging (PSDHI), for distinguishing prostate cancer cells from healthy ones. RM reveals cancer cells metabolize glucose faster, storing it as fatty acids and cholesteryl esters in lipid droplets (LDs). On the other hand, PSDHI shows significant morphological changes in LDs in glucose-incubated cancer cells, including number, volume, and refractive index. High birefringence in cancer LDs under perpendicular polarizations was observed, enabling fast discrimination with over 90% accuracy. PSDHI results align closely with Raman microscopy, suggesting its potential as a promising, high-speed technique for cancer screening purposes.

With continuously improving resolution of today’s (super-resolution) microscopes, a major technical limitation of light microscopy based image analysis is linkage error – a visualization error that is measured by the distance between the cellular target to be detected and the fluorescence emitter used for detection. The linkage error of standard labelled antibodies is caused by the size of the antibody and the random distribution of fluorescent emitters on the antibody surface. In this study, we describe a class of staining reagents that effectively reduce the linkage error by more than five-fold when compared to conventional staining techniques. We believe this class of reagents realize an unmet need in cell biological super resolution imaging studies where the precise localization of the target of interest is crucial for the understanding of complex biological phenomena

ABSTRACT: Staphylococcus aureus is a major pathogen in humans and animals. In cattle, it is one of the most important agents of mastitis, causing serious costs in the dairy industry. Early diagnosis and adequate therapy are therefore 2 key factors to deal with the problems caused by this bacterium, and benzylpenicillin (penicillin) is usually the first choice to treat these infections. Unfortunately, penicillin resistance testing in bovine S. aureus strains shows discrepant results depending on the test used; consequently, the best method for assessing penicillin resistance is still unknown. The aim of this study was therefore to find a method that assesses penicillin resistance in S. aureus and to elucidate the mechanisms leading to the observed discrepancies. A total of 146 methicillin-sensitive S. aureus strains isolated from bovine mastitis were tested for penicillin resistance using a broth microdilution [minimum inhibitory concentration (MIC)] and 2 different disk diffusion protocols. Furthermore, the strains were analyzed for the presence of the bla operon genes (blaI, blaR1, blaZ) by PCR, and a subset of 45 strains was also subjected to whole genome sequencing (WGS). Discrepant results were obtained when penicillin resistance of bovine S. aureus was evaluated by disk diffusion, MIC, and PCR methods. The discrepancies, however, could be fully explained by WGS analysis. In fact, it turned out that penicillin resistance is highly dependent on the completeness of the bla operon promotor: when the bla operon was complete based on WGS analysis, all strains showed MIC ≥1 µg/mL, whereas when the bla operon was mutated (31-nucleotide deletion), they were penicillin sensitive except in those strains where an additional, bla operon-independent resistance mechanism was observed. Further, WGS analyses showed that penicillin resistance is truly assessed by the MIC assay. In contrast, caution is required when interpreting disk diffusion and PCR results.

Nos últimos 15 anos o tratamento cirúrgico do câncer de mama foi revolucionado por alguns novos conceitos na história natural destes tumores. A impossibilidade de controlar a doença com uma cirurgia alargada, a identificação de tumores cada vez menores com o uso da mamografia, a possibilidade da "paciente atual" participar da escolha do tratamento, criaram as condições para um tratamento conservador. Os resultados retrospectivos e, especialmente, prospectivos, dos estudos sobre o tratamento conservador têm demonstrado que a terapia de combinação pode fornecer um adequado controle local do tumor. O Trial 1 de Milão, comparando a mastectomia de Halsted versus a QUART(quadrantectomia, dissecção axilare radioterapia), forneceu uma importante contribuição mostrando não existir diferenças na sobrevida global, período livre de doença e recidivas locais entre os dois tipos de tratamento.

Introduction: Legionnaires' disease is a severe pneumonia predominantly caused by Legionella pneumophila (Lp), whose major reservoirs are artificial water systems. As most human infections are caused by L. pneumophila serogroup 1 (Lp1), a reliable method for Lp distinction can be crucial for bacterial spread prevention. As the ability to withstand in environments and to cause the waterborne disease is strongly related to specific genes, the identification of virulent strains can be of great relevance to implement water environmental monitoring and to contain harmful outbreaks to public health. We aimed to test an assay for Lp identification among different Legionella species, and to determine the serogroups. Additionally, we investigated the carriage of virulence and antimicrobial resistance genes. Methods: A total of 90 Legionella spp. isolates identified by phenotypic tests were subjected to the designed quantitative PCR assay targeting specific mip for Lp, wzm for Lp1, pvcA and ahpD for biofilm production. Eleven serogroups were investigated in all our isolates tested positive for mip gene, subsequently analyzed for 12 virulence and 8 antimicrobial resistance genes. Results: Only the 70 Lp isolates were positive for mip. Out of 27 Lp isolates belonging to serogroup 1 based on agglutination test, 23 (85.2%) carried wzm. The presence of ahpD and pvcA was found in 94.3 and 98.6% of Lp isolates, respectively. By multiplex PCR, all 23 wzm-positive strains were confirmed as serogroup 1 that was the most predominant (33%). At least one virulence gene was detected in all Lp isolates. The most frequent gene was ispE (100%), followed by issD (96%), icmK and enhC (93%), cpxA (91%), rtxA2 (74%), lvhB8-B9 (61%), and prpA (54%). The other genes were less diffused in Lp strains (rtxA1, 44%; lvhB3-B4, 47%; pvcB, 27%; lvrE, 24%). Of the macrolide resistance genes, the ereA was found in 84% of Lp strains, while only 14 (20%) harbored the lpeAB among the efflux pump genes. Conclusion: The assays validated in this study enable the simultaneous Lp and Lp1 detection. The differentiation of Lp strains according to their virulence properties could be useful to predict the bacterial ability to survive and to cause the disease. [ABSTRACT FROM AUTHOR]

Aim: We aimed to evaluate the impact of Coronavirus Disease‐19 (COVID‐19) pandemic on the incidence and clinical presentation of celiac disease (CD) in children. Methods: The diagnoses of CD were compared between the COVID‐19 pandemic (from April 2020 to March 2022) and the pre‐pandemic period (from April 2018 to March 2020) in three Italian Paediatric Gastroenterology centres (Varese, Como, Catanzaro). Electronic patient records were reviewed and additional information were collected through parental interview. The diagnosis of CD was made according to ESPGHAN criteria. SARS‐CoV‐2 infection was diagnosed based on pre‐vaccination positive serum antibodies or nasopharyngeal swabs. Z test and chi‐square were used for statistical analysis. Results: The overall number of paediatric diagnosis of CD did not differ between the two years pre‐pandemic and pandemic periods (177 and 172 cases) in the three Italian participating centres. Clinical presentation of CD was also similar throughout the study periods. SARS‐CoV‐2 infection has been documented in 10.6% of children but only in 5.8% of these occurred before CD diagnosis. Conclusion: Different to what reported for other autoimmune diseases, the incidence and presenting symptoms of CD in our paediatric population did not change during the COVID‐19 pandemic compared to the previous 2 years. [ABSTRACT FROM AUTHOR]

Cannabidiol (CBD), the main non-psychoactive component of Cannabis sativa L., is widely used in therapy for the treatment of different diseases and as an adjuvant drug. Our aim was to assess the effects of CBD on proinflammatory cytokine production and cell proliferation in human peripheral blood mononuclear cells (PBMCs) and on CD4+ T lymphocyte differentiation, and, furthermore, to test CBD’s ability to affect the functional properties of regulatory T cells (Treg). Experiments were performed on isolated PBMCs and purified CD4+ T lymphocytes obtained from the buffy coats of healthy subjects. Cytokines produced by CD4+ T cells were evaluated by flow cytometry and intracellular cytokine staining techniques. PBMC cytokine production was measured by an ELISA assay. Real-time PCR was used to assess the mRNA expression of cytokines and the key transcription factors (TFs) of CD4+ T cells. Finally, the proliferation of PBMC and CD4+ T effector cells (Teff), alone and in the presence of Treg, was assessed by flow cytometry. Results showed that CBD affects both the frequency of IL-4-producing CD4+ and of IFN-γ/IL-17-producing cells and dramatically decreases the mRNA levels of all TFs. Stimuli-induced cytokine mRNA expression was decreased while protein production was unaffected. CBD was unable to affect the ability of Treg to prevent Teff cell proliferation while it slightly increased PBMC proliferation. In conclusion, CBD may inhibit the expression of proinflammatory cytokines; however, the effect of CBD on cell proliferation suggests that this cannabinoid exerts a complex activity on human PBMCs and CD4+ T cells which deserves further investigation.

Circulating tumor cells (CTCs) are tumor cells that have penetrated the circulatory system preserving tumor properties and heterogeneity. Detection and characterization of CTCs has high potential clinical values and many technologies have been developed for CTC identification. These approaches remain challenged by the extraordinary rarity of CTCs and the difficulty of efficiently distinguishing cancer from the much larger number of white blood cells in the bloodstream. Consequently, there is still a need for efficient and rapid methods to capture the broad spectrum of tumor cells circulating in the blood. Herein, we exploit the peculiarities of cancer metabolism for discriminating cancer from WBCs. Using deuterated glucose and Raman microscopy we show that a) the known ability of cancer cells to take up glucose at greatly increased rates compared to non-cancer cells results in the lipid generation and accumulation into lipid droplets and, b) by contrast, leukocytes do not appear to generate visible LDs. The difference in LD abundance is such that it provides a reliable parameter for distinguishing cancer from blood cells. For LD sensitive detections in a cell at rates suitable for screening purposes, we test a polarization-sensitive digital holographic imaging (PSDHI) technique that detects the birefringent properties of the LDs. By using polarization-sensitive digital holographic imaging, cancer cells (prostate cancer, PC3 and hepatocarcinoma cells, HepG2) can be rapidly discriminated from leukocytes with reliability close to 100%. The combined Raman and PSDHI microscopy platform lays the foundations for the future development of a new label-free, simple and universally applicable cancer cells’ isolation method.

Forty‑one tumors were detected in a population of 1,649,003 cattle slaughtered in 4 abattoirs in Lombardy over a 5‑year period, for an overall prevalence of 2.5 tumors per 100,000 cattle. Tumors were classified according to the WHO histological classification of tumors of domestic animals. Alimentary and hemopoietic systems were commonly affected with 9 cases each. Other affected sites were the respiratory (n = 3), urinary (n = 2), endocrine (n = 2), musculoskeletal (n = 2), nervous (n = 1), and cardiovascular (n = 1) systems. The peritoneum was affected by 6 cases, while the primary location of 3 tumors of the connective tissues and 3 metastatic carcinomas was unidentified. Liver tumors and mesotheliomas, for which environmental risk factors are well‑known in humans, were common, as well as tumors typically encountered in pediatric human patients (tumors of mesenchymal tissues, pulmonary blastomas and nephroblastomas). These findings suggest the useful role of bovines as sentinel and model for human carcinogenesis. Our study indicates that the establishment of a bovine cancer registry in Lombardy is feasible considering its potential contribution to understanding the role of environmental risk factors in the genesis of tumors in animals and humans.

En este artículo se presenta la automatización implementada a través de una serie de proyectos orientados hacia el continuo mejoramiento y desarrollo de los sistemas didácticos presentes en celdas de manufactura flexibles HAS200 de la Facultad Tecnológica de la Universidad Distrital Francisco José de Caldas. Por medio de dichos proyectos se ha logrado implementar una serie de mejoras automatizadas como por ejemplo: i) un entorno virtual de la celda de manufactura flexible, modelado geométricamente para la simulación remota de los procesos didácticos; ii) una novedosa estación automatizada para el reciclaje de pellets que se integra con las demás estaciones para clasificar la materia prima en tres colores separados; iii) una nueva interfaz de usuario-máquina para un brazo manipulador Melfa Mitsubishi RV-M1, que permite integrar la estación de reciclaje con el resto de la celda de manufactura flexible; iv) un diseño de gripper automatizado, para la manipulación y posterior apertura de recipientes dentro de la celda; v) un sistema de visión artificial implementado al transporte autónomo alterno de producto entre estaciones mediante vehículos terrestres no tripulados; vi) un curso virtual para el manejo y operación de la celda de manufactura flexible. La automatización propuesta en la HAS200 puede ser extrapolable hacia entornos productivos reales bajo los mismos conceptos de automatización, lo cual hace que cada resultado obtenido sea doblemente beneficioso.

Ground-based microwave radiometer (MWR) observations of downwelling brightness temperature (TB) are commonly used to estimate atmospheric attenuation at relative transparent channels for radio propagation and telecommunication purposes. The atmospheric attenuation is derived from TB by inverting the radiative transfer equation with a priori knowledge of the mean radiating temperature (TMR). TMR is usually estimated by either time-variant site climatology (e.g., monthly average computed from atmospheric thermodynamical profiles) or condition-variant estimation from surface meteorological sensors. However, information on TMR may also be extracted directly from MWR measurements at channels other than those used to estimate atmospheric attenuation. This paper proposes a novel approach to estimate TMR in clear and cloudy sky from independent MWR profiler measurements. A linear regression algorithm is trained with a simulated dataset obtained by processing 1 year of radiosonde observations of atmospheric thermodynamic profiles. The algorithm is trained to estimate TMR at K- and V–W-band frequencies (22–31 and 72–82 GHz, respectively) from independent MWR observations at the V band (54–58 GHz). The retrieval coefficients are then applied to a 1-year dataset of real V-band observations, and the estimated TMR at the K and V–W band is compared with estimates from nearly colocated and simultaneous radiosondes. The proposed method provides TMR estimates in better agreement with radiosondes than a traditional method, with 32 %–38 % improvement depending on frequency. This maps into an expected improvement in atmospheric attenuation of 10 %–20 % for K-band channels and ∼30 % for V–W-band channels.