Molecular characterization of SARS-CoV-2 Omicron clade and clinical presentation in children.

Since its emergence, SARS-CoV-2 Omicron clade has shown a marked degree of variability and different clinical presentation compared with previous clades. Here we demonstrate that at least four Omicron lineages circulated in children since December 2021, and studied until November 2022: BA.1 (33.6%), BA.2 (40.6%), BA.5 (23.7%) and BQ.1 (2.1%). At least 70% of infections concerned children under 1 year, most of them being infected with BA.2 lineages (n = 201, 75.6%). Looking at SARS-CoV-2 genetic variability, 69 SNPs were found to be significantly associated in pairs, (phi < - 0.3 or > 0.3 and p-value < 0.001). 16 SNPs were involved in 4 distinct clusters (bootstrap > 0.75). One of these clusters (A23040G, A27259C, T23617G, T23620G) was also positively associated with moderate/severe COVID-19 presentation (AOR [95% CI] 2.49 [1.26-4.89] p-value: 0.008) together with comorbidities (AOR [95% CI] 2.67 [1.36-5.24] p-value: 0.004). Overall, these results highlight the extensive SARS-CoV-2 Omicron circulation in children, mostly aged < 1 year, and provide insights on viral diversification even considering low-abundant SNPs, finally suggesting the potential contribution of viral diversification in affecting disease severity.

Identification of the blaOXA-23 Gene in the First Mucoid XDR Acinetobacter baumannii Isolated from a Patient with Cystic Fibrosis.

Acinetobacter baumannii is one of the pathogens most involved in health care-associated infections in recent decades. Known for its ability to accumulate several antimicrobial resistance mechanisms, it possesses the oxacillinase blaoxa-23, a carbapenemase now endemic in Italy. Acinetobacter species are not frequently observed in patients with cystic fibrosis, and multidrug-resistant A. baumannii is a rare event in these patients. Non-mucoid A. baumannii carrying the blaoxa-23 gene has been sporadically detected. Here, we describe the methods used to detect blaoxa-23 in the first established case of pulmonary infection via a mucoid strain of A. baumannii producing carbapenemase in a 24-year-old cystic fibrosis patient admitted to Bambino Gesù Children's Hospital in Rome, Italy. This strain, which exhibited an extensively drug-resistant antibiotype, also showed a great ability to further increase its resistance in a short time.

A case of SARS-CoV-2 Omicron reinfection resulting in a significant immunity boost in a paediatric patient affected by B-cell acute lymphoblastic leukemia.

BACKGROUND: Since its emergence in November 2021, SARS-CoV-2 Omicron clade has quickly become dominant, due to its increased transmissibility and immune evasion. Different sublineages are currently circulating, which differ in mutations and deletions in regions of the SARS-CoV-2 genome implicated in the immune response. In May 2022, BA.1 and BA.2 were the most prevalent sublineages in Europe, both characterized by ability of evading natural acquired and vaccine-induced immunity and of escaping monoclonal antibodies neutralization. CASE PRESENTATION: A 5-years old male affected by B-cell acute lymphoblastic leukemia in reinduction was tested positive for SARS-CoV-2 by RT-PCR at the Bambino Gesù Children Hospital in Rome in December 2021. He experienced a mild COVID-19 manifestation, and a peak of nasopharyngeal viral load corresponding to 15.5 Ct. Whole genome sequencing identified the clade 21 K (Omicron), sublineage BA.1.1. The patient was monitored over time and tested negative for SARS-CoV-2 after 30 days. Anti-S antibodies were detected positive with modest titre (3.86 BAU/mL), while anti-N antibodies were negative. 74 days after the onset of the first infection and 23 days after the last negative test, the patient was readmitted to hospital with fever, and tested positive for SARS-CoV-2 by RT-PCR (peak of viral load corresponding to 23.3 Ct). Again, he experienced a mild COVID-19. Whole genome sequencing revealed an infection with the Omicron lineage BA.2 (21L clade). Sotrovimab administration was started at the fifth day of positivity, and RT-PCR negativity occurred 10 days later. Surveillance SARS-CoV-2 RT-PCR were persistently negative, and in May 2022, anti-N antibodies were found positive and anti-S antibodies reached titres > 5000 BAU/mL. CONCLUSIONS: By this clinical case, we showed that SARS-CoV-2 reinfection within the Omicron clade can occur and can be correlated to inadequate immune responses to primary infection. We also showed that the infection's length was shorter in the second respect to first episode, suggesting that pre-existing T cell-mediated immunity, though not preventing re-infection, might have limited the SARS-CoV-2 replication capacity. Lastly, Sotrovimab treatment retained activity against BA.2, probably accelerating the viral clearance in the second infectious episode, after which seroconversion and increase of anti-S antibodies titres were observed.

A proof-of-concept study on the genomic evolution of Sars-Cov-2 in molnupiravir-treated, paxlovid-treated and drug-naïve patients.

Little is known about SARS-CoV-2 evolution under Molnupiravir and Paxlovid, the only antivirals approved for COVID-19 treatment. By investigating SARS-CoV-2 variability in 8 Molnupiravir-treated, 7 Paxlovid-treated and 5 drug-naïve individuals at 4 time-points (Days 0-2-5-7), a higher genetic distance is found under Molnupiravir pressure compared to Paxlovid and no-drug pressure (nucleotide-substitutions/site mean±Standard error: 18.7 × 10-4 ± 2.1 × 10-4 vs. 3.3 × 10-4 ± 0.8 × 10-4 vs. 3.1 × 10-4 ± 0.8 × 10-4, P = 0.0003), peaking between Day 2 and 5. Molnupiravir drives the emergence of more G-A and C-T transitions than other mutations (P = 0.031). SARS-CoV-2 selective evolution under Molnupiravir pressure does not differ from that under Paxlovid or no-drug pressure, except for orf8 (dN > dS, P = 0.001); few amino acid mutations are enriched at specific sites. No RNA-dependent RNA polymerase (RdRp) or main proteases (Mpro) mutations conferring resistance to Molnupiravir or Paxlovid are found. This proof-of-concept study defines the SARS-CoV-2 within-host evolution during antiviral treatment, confirming higher in vivo variability induced by Molnupiravir compared to Paxlovid and drug-naive, albeit not resulting in apparent mutation selection.

Pathology of triple negative breast cancer.

Triple negative breast cancer (TNBC) is a subtype of breast tumor lacking hormone receptors expression and HER2 gene amplification and represents 24 % of newly diagnosed breast neoplasms. In this review, pathological aspects of triple-negative breast cancer are illustrated, with particular attention to the seminal studies that defined this subtype of breast cancer by a molecular point of view. This paper also focuses on practical issues raised in clinical routine by the introduction of genetic expression breast cancer profiling and the innovative prognostic and predictive impact on triple-negative breast cancer pathology. Moreover, histopathological aspects of triple-negative neoplasms are also mentioned, underlying the importance of histologic diagnosis of particular cancer subtypes with decisive impact on clinical outcome. Importantly, focus on new therapeutic frontier represented by immunotherapy is illustrated, with particular mention of immune checkpoint inhibitors introduction in TNBC therapy and their impact on future treatments.

Molecular Aspects and Prognostic Significance of Microcalcifications in Human Pathology: A Narrative Review.

The presence of calcium deposits in human lesions is largely used as imaging biomarkers of human diseases such as breast cancer. Indeed, the presence of micro- or macrocalcifications is frequently associated with the development of both benign and malignant lesions. Nevertheless, the molecular mechanisms involved in the formation of these calcium deposits, as well as the prognostic significance of their presence in human tissues, have not been completely elucidated. Therefore, a better characterization of the biological process related to the formation of calcifications in different tissues and organs, as well as the understanding of the prognostic significance of the presence of these calcium deposits into human tissues could significantly improve the management of patients characterized by microcalcifications associated lesions. Starting from these considerations, this narrative review highlights the most recent histopathological and molecular data concerning the formation of calcifications in breast, thyroid, lung, and ovarian diseases. Evidence reported here could deeply change the current point of view concerning the role of ectopic calcifications in the progression of human diseases and also in the patients' management. In fact, the presence of calcifications can suggest an unfavorable prognosis due to dysregulation of normal tissues homeostasis.

BMP-2 Variants in Breast Epithelial to Mesenchymal Transition and Microcalcifications Origin.

This study aims to investigate the possible different roles of the BMP-2 variants, cytoplasmic and nuclear variant, in both epithelial to mesenchymal transition and in microcalcifications origin in human breast cancers. To this end, the in situ expression of cytoplasmic and nuclear BMP-2 was associated with the expression of the main epithelial to mesenchymal transition biomarkers (e-cadherin and vimentin) and molecules involved in bone metabolisms (RUNX2, RANKL, SDF-1) by immunohistochemistry. In addition, the expression of cytoplasmic and nuclear BMP-2 was associated with the presence of microcalcifications. Our data showed a significant association among the number of cytoplasmic BMP-2-positive cells and the number of both vimentin (positive association) and e-cadherin (negative association) positive breast cells. Conversely, no associations were found concerning the nuclear BMP-2-positive breast cells. Surprisingly, the opposite result was obtained by analyzing the variants of BMP-2 and both the expression of RANKL and SDF-1 and the presence of microcalcifications. Specifically, the presence of microcalcifications was related to the expression of nuclear BMP-2 variant rather than the cytoplasmic one, as well as a strong association between the number of nuclear BMP-2 and the expression of the main breast osteoblast-like cells (BOLCs) biomarkers. To further corroborate these data, an in vitro experiment for demonstrating the co-expression of nBMP-2 and RANKL or vimentin or SDF-1 in breast cancer cells that acquire the capability to produce microcalcifications was developed. These investigations confirmed the association between the nBMP-2 expression and both RANKL and SDF-1. The data supports the idea that whilst cytoplasmic BMP-2 can be involved in epithelial to mesenchymal transition phenomenon, the nuclear variant is related to the essential mechanisms for the formation of breast microcalcifications. In conclusion, from these experimental and translational perspectives, the complexity of BMP-2 signaling will require a detailed understanding of the involvement of specific BMP-2 variants in breast cancers.

Plaque calcification is driven by different mechanisms of mineralization associated with specific cardiovascular risk factors.

BACKGROUND AND AIMS: The aim of this study was to investigate possible associations among markers of mineralization, plaque instability and the main risk factors of atherosclerosis. METHODS AND RESULTS: A Tissue MicroArray containing 52 samples of calcified carotid plaques from 52 symptomatic and asymptomatic patients were built. TMA serial sections were used to study the expression of inflammatory and mineralization markers (BMP-2, BMP-4, VDR, RANKL, Osteopontin, Sclerostin, ß-catenin and calmodulin) by immunohistochemistry. Our data clearly demonstrated the expression of mineralization markers in atheromatic plaques. Indeed, with the exception of RANKL, all investigated markers were expressed in at least 60% of cases. Specifically, multivariate analysis displayed significant associations between both the expression of BMP-2 and the presence of unstable plaques as well as between the expression of ß-catenin and the presence of stable plaques. We also found a significant inverse association between both a) the presence of hypertension and VDR and b) smoking habits and calmodulin expression. Finally, we noted a higher density of RANKL positive cells in plaques from diabetic patients as compared to non-diabetic ones and a significant positive association between hypertriglyceridemia and BMP-4 expression. CONCLUSION: Our results support the hypothesis that the process of atherosclerotic plaque calcification presents a number of similarities with the physiological processes that occur in bone, involving both osteoblasts- and osteoclasts-like arterial cells. Finally, the present study suggests that risk factors, such as hypertension, cigarette smoke and diabetes, can cause the destabilization of the atheromatic plaque acting on calcification process as well as inflammation.