An outbreak of severe Kawasaki-like disease at the Italian epicentre of the SARS-CoV-2 epidemic: an observational cohort study.

BACKGROUND: The Bergamo province, which is extensively affected by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) epidemic, is a natural observatory of virus manifestations in the general population. In the past month we recorded an outbreak of Kawasaki disease; we aimed to evaluate incidence and features of patients with Kawasaki-like disease diagnosed during the SARS-CoV-2 epidemic. METHODS: All patients diagnosed with a Kawasaki-like disease at our centre in the past 5 years were divided according to symptomatic presentation before (group 1) or after (group 2) the beginning of the SARS-CoV-2 epidemic. Kawasaki- like presentations were managed as Kawasaki disease according to the American Heart Association indications. Kawasaki disease shock syndrome (KDSS) was defined by presence of circulatory dysfunction, and macrophage activation syndrome (MAS) by the Paediatric Rheumatology International Trials Organisation criteria. Current or previous infection was sought by reverse-transcriptase quantitative PCR in nasopharyngeal and oropharyngeal swabs, and by serological qualitative test detecting SARS-CoV-2 IgM and IgG, respectively. FINDINGS: Group 1 comprised 19 patients (seven boys, 12 girls; aged 3·0 years [SD 2·5]) diagnosed between Jan 1, 2015, and Feb 17, 2020. Group 2 included ten patients (seven boys, three girls; aged 7·5 years [SD 3·5]) diagnosed between Feb 18 and April 20, 2020; eight of ten were positive for IgG or IgM, or both. The two groups differed in disease incidence (group 1 vs group 2, 0·3 vs ten per month), mean age (3·0 vs 7·5 years), cardiac involvement (two of 19 vs six of ten), KDSS (zero of 19 vs five of ten), MAS (zero of 19 vs five of ten), and need for adjunctive steroid treatment (three of 19 vs eight of ten; all p<0·01). INTERPRETATION: In the past month we found a 30-fold increased incidence of Kawasaki-like disease. Children diagnosed after the SARS-CoV-2 epidemic began showed evidence of immune response to the virus, were older, had a higher rate of cardiac involvement, and features of MAS. The SARS-CoV-2 epidemic was associated with high incidence of a severe form of Kawasaki disease. A similar outbreak of Kawasaki-like disease is expected in countries involved in the SARS-CoV-2 epidemic. FUNDING: None.

A Pediatric Emergency Department Protocol to Avoid Intrahospital Spread of SARS-CoV-2 during the Outbreak in Bergamo, Italy.

The pandemic of coronavirus SARS-CoV-2 disease affected Northern Italy, spreading from the Bergamo province to the entire country. During reorganization of our emergency department to support patients presenting with coronavirus SARS-CoV-2 disease, we aimed to evaluate whether children play a role in intrahospital spread of the infection.

Molecular signature detection of circulating tumor cells using a panel of selected genes.

Circulating tumor cell (CTC) detection in peripheral blood of colon and other epithelial cancer patients is becoming a scientifically recognised indicator for the presence of primary tumors and/or metastasis. The resulting need to further develop CTC detection-based systems for improved diagnosis, prognosis and assessment of therapy efficacy in tumour patients has prompted the application of different approaches, including expression analysis of tissue-specific and epithelial genes. In this context, lack of specificity of the analysed genes remains a fundamental problem for reliable CTC detection. In this study, we have selected a panel of highly specific epithelial genes: cytokeratin 20 (CK20), cytokeratin 19 (CK19), carcinoembryonic antigen (CEA) and guanylyl cyclase C (GCC), and performed RT-PCR analysis to assess their expression in total blood and in different cell fractions of peripheral blood (PBMC and CD45-negative population) of cancer patients and healthy controls. Our results demonstrate that analysis of a single gene in a CTC-enriched population (CD45(-) peripheral blood cells) of cancer patients allows detection of a CTC molecular signature in at most 63.3% of cases, while analysis of all four genes performed in all three sample types increases the detection of positive patient samples to 87.7%. Healthy controls did not show positivity for any combination of these genes, although positivity was observed for the CEA marker alone, which was detected in 3 (6.6%) out of 45 donors, and only in the CD45(-) fraction. Here, we demonstrate that combined analysis of the genes above, in multiple blood fractions, results in a highly specific and sensitive CTC detection system in patients with metastatic solid tumors. Therefore, we believe that validation on a large scale of this approach, which demonstrates higher specificity in patients compared to controls, could become a relevant CTC screening test in patients with established metastatic disease, and furthermore, may also be useful for evaluating the possible presence of CTCs before the onset of clinically manifested metastatic spreading.

Human GluR6c, a functional splicing variants of GluR6, is mainly expressed in non-nervous cells.

Kainate-type glutamate receptors (KARs) are receptor channels with a variety of distinct physiological functions in synaptic transmission, depending on their sub-cellular location in functional neuronal compartments. The kainate receptor subunit GluR6 presents different splice variants involving the C-terminal domain, namely GluR6a, GluR6b and GluR6c. In this study, we report the analysis of the three human splicing isoforms and in particular of the uncharacterized hGluR6c. When expressed in COS-7 cells, hGluR6a receptor subunit was highly present on the surface of the plasma membrane, whereas hGluR6b and hGluRc were poorly transported to the membrane. Electrophysiological studies of homomeric receptors showed that hGluR6c subunit can generate functional receptors with characteristics similar to the GluR6b variant. mRNA expression analysis demonstrated that hGluR6c variant is mainly expressed in non-neuronal cells and barely expressed in neuronal ones. Interestingly, undifferentiated NT2 cells expressing only the hGluR6c isoform, during neuronal differentiation induced by retinoic acid, increased the expression level of the neuronal form hGluR6a with a parallel decreased of hGluR6c. Overall, our data indicate that hGluR6c might have unique properties in non-nervous cells and in the first stages of CNS development.

Comparison of three distinct methods for the detection of circulating tumor cells in colorectal cancer patients.

The detection of circulating tumor cells (CTCs) has considerable utility in the clinical management of patients with solid cancers. However, the phenotypic heterogeneity of CTCs and their low numbers in the bloodstream of patients means that no standardized detection method currently exists for these cells. This, together with differences in pre-analytical sample processing, has led to the collection and accumulation of inconsistent data among independent studies. Here, we compare the ability of three methods to detect CTCs in the blood of colorectal cancer patients. Specifically, different aliquots of the same blood sample were screened for the presence of CTCs by a multimarker RT-PCR assay, the standardized CellSearch assay and dHPLC-based gene mutation analysis. In the population tested, none of the blood samples analysed appeared to be positive by all three methods. Of the samples, 75% were positive for the presence of CTCs by the RT-PCR method. Only 20% were positive by the CellSearch assay, while 14.3% of samples displayed gene mutations consistent with the presence of CTCs when the dHPLC method was applied. The samples which were positive for CTCs by the CellSearch assay did not overlap with those that were positive by dHPLC. Interestingly, however, all of these samples were positive when assessed by RT-PCR. Conversely, of the samples that resulted negative by RT-PCR analysis, none appeared to be positive by either of the other methods. These data, therefore, indicate that of the three methods tested, the multimarker RT-PCR assay provides maximal probability of CTC detection. Here, we present the preliminary results of an ongoing clinical study. Future follow-up involving detection of CTCs in the blood of colorectal cancer patients using these three distinct methods will allow us to verify whether either a single method, or a combination of different assays, is necessary to uncover further prognostic significance of circulating tumor cells.