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Background Preliminary data suggested that fat embolism could explain the importance of visceral obesity as a critical determinant of coronavirus disease-2019 (COVID-19). Methods We performed a comprehensive histomorphologic analysis of autoptic visceral adipose tissue (VAT), lungs and livers of 19 subjects with COVID-19 (COVID-19+), and 23 people without COVID-19 (controls). Human adipocytes (hMADS) infected with SARS-CoV-2 were also studied. Results Although there were no between-group differences in body-mass-index and adipocytes size, a higher prevalence of CD68+ macrophages among COVID-19+ VAT was detected (p = 0.005) and accompanied by crown-like structures presence, signs of adipocytes stress and death. Consistently, human adipocytes were successfully infected by SARS-CoV-2 in vitro and displayed lower cell viability. Being VAT inflammation associated with lipids spill-over from dead adipocytes, we studied lipids distribution by ORO. Lipids were observed within lungs and livers interstitial spaces, macrophages, endothelial cells, and vessels lumen, features suggestive of fat embolism syndrome, more prevalent among COVID-19+ (p < 0.001). Notably, signs of fat embolism were more prevalent among people with obesity (p = 0.03) independently of COVID-19 diagnosis, suggesting that such condition may be an obesity complication exacerbated by SARS-CoV-2 infection. Importantly, all infected subjects’ lungs presented lipids-rich (ORO+) hyaline membranes, formations associated with COVID-19-related pneumonia, present only in one control patient with non-COVID-19-related pneumonia. Importantly, transition aspects between embolic fat and hyaline membranes were also observed. Conclusions This study confirms the lung fat embolism in COVID-19+ patients and describes for the first time novel COVID-19-related features possibly underlying the unfavorable prognosis in people with COVID-19 and obesity.

Mycobacterium chimaera is a non-tuberculous mycobacterium, member of the Mycobacterium avium complex (MAC), which has become a global public health concern due to infection following cardiac surgery performed with contaminated heater-cooler units. M. chimaera infection is characterized by a long latency, non-specific signs and symptoms and high mortality rates. Thus, the diagnosis is still challenging both for forensic pathologists and for clinicians. Clinical manifestations of M. chimaera infection include endocarditis, hepatitis, nephritis, encephalitis and chorioretinitis. A constant histopathologic finding is the presence of non-caseating granulomas, with multinucleated giant cells and histiocytes. Hereby, we present two cases of fatal disseminated M. chimaera infection following aortic valve surgery reporting clinical history and post-mortem findings. Further, we provide a brief overview of the literature with a special focus on histopathological characteristics of M. chimaera infection. The aim of this article is to provide a complete synopsis of histopathological characteristics useful for forensic pathologists.

Introduction Defining extreme temperatures as the cause of death remains challenging. It is mostly based on circumstantial, macroscopic and microscopic features. Methods We retrospectively compared groups of cases of fatal hypothermia, fatal hyperthermia and non-extreme temperature-related deaths. We analysed specific histological findings, focusing on samples from the liver, pancreas and kidney. Results Between 1 January 2013 and 31 December 2016, 15 autopsies were performed for deaths related to extreme temperatures. They included 11 cases of fatal hypothermia (group A), four cases of fatal hyperthermia (group B) and eight controls (group C). Perinuclear hepatocyte vacuolisation was observed in seven cases of hypothermia, one case of hyperthermia and four controls. Pancreatic cytoarchitecture was well preserved in two cases of hypothermia, one case of hyperthermia and two controls. No particular microscopic feature was found in pancreatic samples. Renal epithelial tubular cell vacuolisation was observed in seven cases of hypothermia and one case of hyperthermia, while it was absent in all controls. Chromogranin A (CgA) was markedly positive in the pancreatic tissue of five cases of fatal hypothermia and one control, and mildly positive in one case of fatal hyperthermia. No significant p-values were observed for any comparisons ( p > 0.05), except when hypothermia cases group were compared to the control group for the Armanni–Ebstein phenomenon test ( p = 0.0078). Conclusions Although our study did not find a specific microscopic marker, hepatocyte vacuolisation, the Armanni–Ebstein phenomenon and pancreatic CgA positivity, taken together, may be useful tools to confirm hypo- and hyperthermia-related deaths, in addition to circumstantial and macroscopic findings.

Venous thromboembolism (VTE) is a multifactorial disease determined by a combination of inherited and acquired factors. Inherited factors include mutations in the genes coding for coagulation factors, some of which seem to exert a differential influence on the risk of developing deep vein thrombosis (DVT) and pulmonary embolism (PE). In post-mortem studies of subjects who have died from pulmonary embolism (PE), the analysis of the factors that may have augmented the VTE risk is often limited to acquired factors. This is due to the complexity-and sometimes the unfeasibility-of analyzing genetic factors and to insufficient knowledge of their individual roles in PE development. The present study used formalin-fixed paraffin-embedded (FFPE) tissue to investigate a panel of 12 polymorphisms-the largest ever studied-that affect the VTE risk. Tissue samples came from post-mortem examinations performed by the specialists of the Section of Legal Medicine of the Department of Pathology of Marche's Polytechnic University, and by the specialists of Health Care District Hospital of Imola, on 44 subjects who died from PE in the period 1997-2014. All individuals were found to have at least one mutation affecting the VTE risk. The present study demonstrates that genetic analysis can be performed post-mortem and the results are useful for forensic investigations, especially from MTHFR C677T and PAI-1 4G/5G polymorphisms. Broader studies using the techniques described herein are needed to determine the relative influence of the individual polymorphisms and their interaction in PE deaths.

To define the Y-chromosome genetic structure in Apennine populations, 17 Y-chromosome short tandem repeats (Y-STRs) and 37 Y-single nucleotide polymorphisms (Y-SNPs) were typed in 162 subjects living in the upland area of the Marches (Central Italy). A total number of 155 haplotypes (haplotype diversity was 0.9994) and 14 SNP haplogroups were observed. Testing high-resolution Y-chromosome data sets, e.g. using Yfiler and SNPs, increases the discriminatory capacity in individual identification for forensic purposes. It is also useful in autochthonous population and micro-population studies to highlight the most informative loci for evolutionary aims.

This work describes an efficient and rapid test for typing 37 single nucleotide polymorphisms (SNPs) of the non-recombining region of Y chromosome (NRY) from a minimal amount of DNA using six PCR multiplexes. Markers were drawn following a hierarchical strategy based on the phylogenetic tree of Y chromosome proposed by the Y Chromosome Consortium [The Y Chromosome Consortium, A nomenclature system for the tree of human Y-chromosomal binary haplogroups, Genome Res. 12 (2002) 339-348]. Two multiplexes--arbitrarily named MY1 and MY2--were developed to explore the basal branches of the tree encompassing all the major clades A-R: MY1 for markers M35, M89, M172, M170, M9, M173, M45 and MY2 for markers M52, M216, M174, M181, M201, M91, M96, M214. Four multiplexes able of typing the more superficial branches typical of most frequent European haplogroups E3b, J2, R1 and I, were also developed and named MY-E3b (M78, M107, M224, M165, M148, M81), MY-J2 (M158, M68, M47, M102, M137, M67), MY-R1 (M17, M269, M18, P25, SRY10831.2) and MY-I (M72, M223, M26, M21, M161). SNP genotyping was carried out by hot-start PCR amplification with primers yielding fragments between 63 and 210 nucleotides, followed by minisequencing reaction based on dideoxy single-base extension and capillary electrophoresis of extension products. The sequential application of these multiplexes is a robust and effective resource for typing the most frequent European Y-SNP haplogroups, and appears to be suitable for forensic purposes and evolutionary studies.

The postmortem diagnosis of anaphylactic death may be frustrating when victims are not hospital patients, even more so when they are recovered dead. The frequent lack of specific morphological findings in such cases means that diagnosis by the forensic pathologist must rely solely on exclusion criteria or circumstantial evidence. However, a diagnostic approach based on case history, analysis of circumstances, available clinical and necropsy findings, as well as toxicology, histopathology and biohumoral data, often allows demonstration of the cause of death. Some useful reflections on microscopic morphological data have come from two recent cases, where thorough data collection provided a reasonably certain diagnosis of anaphylactic death and systemic inflammatory response syndrome-related cardiac arrest, respectively. In both cases tissue histopathology proved crucial, since histochemical (GIEMSA) and immunohistochemical analysis (CD117 and tryptase) documented a large number of mast cells in tissues, particularly the laryngeal wall, and a discrepancy between cells positive for GIEMSA and tryptase and those positive for CD117. Staining for CD117 was also detected in cells with dendrite morphology and in a subpopulation of small lymphocytes with incised nuclei. The morphological findings of these cases are discussed, especially those obtained with immunohistochemistry, and the need for the latter data to be interpreted by experienced medical staff in the framework of a thorough analysis of all the data collected is highlighted.

Material recovered from 374 fingerprints left by eleven laboratory workers on three different substrates (glass, wood, metal) at a standard pressure time of 30 s, with and without preliminary handwashing, was submitted to morphological, quantitative, and type analysis. Morphological and agarose-gel electrophoresis analysis showed that a non-negligible amount of epidermal corneal cells presented apoptotic alterations. The quantity of DNA recovered from fingerprints ranged between 0.04 to 0.2 ng, and in a significant number of experiments no DNA was detected. Handwashing reduced the amount of DNA recovered from fingerprints. The "shedder status" of the donor was a very important factor, causing inter-individual variations in the amount of DNA left by fingerprints. Spurious alleles from laboratory-based and secondary transfer contamination, stutters, and other artifacts described when analyzing low-copy-number DNA and capable of affecting correct profiles were observed.

This study reports a paternity case analyzed by the AmpFlSTR Identifiler Kit (AB) in which father and daughter shared three rare alleles for D19S433, D18S51 and TH01 microsatellites. The case also showed an apparent exclusion, due to a mutation at the D3S 1358 microsatellite. Sequencing analysis was performed to assess the size of the rare alleles and to establish their structure, which revealed some molecular variations in regions flanking the motif repeats.

Single-nucleotide polymorphisms of Y chromosome (Y-SNPs) are a class of markers of interest in forensic investigations, because many of them show regional specificity, providing useful information about the geographic origin of a subject or evidence under investigation. A first multiplex with 7 SNPs (M35, M89, M9, M170, M172, M45, M173), which occur in the basal branches of the phylogenetic tree and are able to assign a subject to known most frequent European haplogroups, was designed. SNP genotyping was accomplished by hot-start PCR with primers amplifying fragments between 96 and 136 nucleotides, minisequencing, and capillary electrophoresis of extension products. Ninety seven subjects of known geographic provenance were studied, of which 68 from Europe. Of these, 57 had mutations found more frequently in European haplogroups and 11 more frequent in Asian populations. Subjects from non-European countries were also examined and had haplogroups common in their regions of provenance. Experiments with low molecular weight DNA gave positive amplification from 1 ng of DNA for all seven SNPs.

Background Sudden cardiac death (SCD) is the clinical outcome of a lethal arrhythmia that can develop on the background of unrecognized channelopathies or cardiomyopathies. Several susceptibility genes have been identified for the congenital forms of these cardiac diseases, including caveolin-3 (Cav-3) gene. In the heart Cav-3 is the main component of caveolae, plasma membrane domains that regulate multiple cellular processes highly relevant for cardiac excitability, such as trafficking, calcium homeostasis, signal transduction and cellular response to injury. Here we characterized a new putative Cav-3 variant, Cav-3 V82I, found in a patient with SCD. Results In heterologous systems Cav-3 V82I was expressed at significantly higher level than Cav-3 WT and accumulated within the cells. Cells expressing Cav-3 V82I exhibited a decreased activation of extracellular-signal-regulated kinases (ERKs) and were more vulnerable to sub-lethal osmotic stress. Conclusion Considering that abnormal loss of myocytes can play a mechanistic role in lethal cardiac diseases, we suggest that the detrimental effect of Cav-3 V82I variant on cell viability may participate in determining the susceptibility to cardiac death.