genes G C A T T A C G G C A T Article Complex Modes of Inheritance in Hereditary Red Blood Cell Disorders: A Case Series Study of 155 Patients Immacolata Andolfo 1,2 , Stefania Martone 1,2 , Barbara Eleni Rosato 1,2 , Roberta Marra 1,2 , Antonella Gambale 2,3 , Gian Luca Forni 4 , Valeria Pinto 4 , Magnus Göransson 5 , Vasiliki Papadopoulou 6 , Mathilde Gavillet 6 , Mohsen Elalfy 7 , Antonella Panarelli 2 , Giovanna Tomaiuolo 2,8 , Achille Iolascon 1,2, * and Roberta Russo 1   Citation: Andolfo, I.; Martone, S.; Rosato, B.E.; Marra, R.; Gambale, A.; Forni, G.L.; Pinto, V.; Göransson, M.; Papadopoulou, V.; Gavillet, M.; et al. Complex Modes of Inheritance in Hereditary Red Blood Cell Disorders: A Case Series Study of 155 Patients. Genes 2021, 12, 958. https://doi.org/ 10.3390/genes12070958 Academic Editor: Julia Horsfield Received: 27 May 2021 Accepted: 19 June 2021 Published: 23 June 2021 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). 1 Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Università degli Studi di Napoli Federico II, 80131 Napoli, Italy; andolfo@ceinge.unina.it (I.A.); stefaniamartone85@libero.it (S.M.); rosato.barbara@gmail.com (B.E.R.); robertamarra.r@gmail.com (R.M.); roberta.russo@unina.it (R.R.) 2 CEINGE Biotecnologie Avanzate, 80145 Naples, Italy; antonellagambale@gmail.com (A.G.); panarelli@ceinge.unina.it (A.P.); g.tomaiuolo@unina.it (G.T.) 3 Department of Laboratory Medicine (DAIMedLab), UOC Medical Genetics, ‘Federico II’ University Hospital, 80131 Naples, Italy 4 Centro della Microcitemia e delle Anemie Congenite, Ospedale Galliera, 16128 Genoa, Italy; gianluca.forni@galliera.it (G.L.F.); valeria.pinto@galliera.it (V.P.) 5 Department of Paediatrics, The Queen Silvia Children’s Hospital, Sahlgrenska University Hospital, 41345 Gothenburg, Sweden; magnus.l.goransson@vgregion.se 6 Service and Central Laboratory of Haematology, Department of Oncology and Department of Laboratory Medicine and Pathology, Lausanne University Hospital (CHUV), 1011 Lausanne, Switzerland; vasiliki.papadopoulou@chuv.ch (V.P.); mathilde.gavillet@chuv.ch (M.G.) 7 Thalassemia Centre, Faculty of Medicine, Ain Shams University, Cairo 11566, Egypt; elalfym@hotmail.com 8 Department of Chemical Engineering, Materials and Industrial Production, ‘Federico II’ University of Naples, 80125 Naples, Italy * Correspondence: achille.iolascon@unina.it Abstract: Hereditary erythrocytes disorders include a large group of conditions with heteroge- neous molecular bases and phenotypes. We analyzed here a case series of 155 consecutive patients with clinical suspicion of hereditary erythrocyte defects referred to the Medical Genetics Unit from 2018 to 2020. All of the cases followed a diagnostic workflow based on a targeted next-generation sequencing panel of 86 genes causative of hereditary red blood cell defects. We obtained an overall diagnostic yield of 84% of the tested patients. Monogenic inheritance was seen for 69% (107/155), and multi-locus inheritance for 15% (23/155). PIEZO1 and SPTA1 were the most mutated loci. Accordingly, 16/23 patients with multi-locus inheritance showed dual molecular diagnosis of dehy- drated hereditary stomatocytosis/xerocytosis and hereditary spherocytosis. These dual inheritance cases were fully characterized and were clinically indistinguishable from patients with hereditary spherocytosis. Additionally, their ektacytometry curves highlighted alterations of dual inheritance patients compared to both dehydrated hereditary stomatocytosis and hereditary spherocytosis. Our findings expand the genotypic spectrum of red blood cell disorders and indicate that multi-locus inheritance should be considered for analysis and counseling of these patients. Of note, the genetic testing was crucial for diagnosis of patients with a complex mode of inheritance. Keywords: red blood cell defects; targeted next-generation sequencing; multi-locus inheritance; PIEZO1; SPTA1 1. Introduction Hereditary anemias are a heterogeneous group of conditions that are characterized by complex genotype–phenotype correlations. Based on clinical manifestations and mor- phological red blood cell (RBC) alterations, hereditary anemias can be broadly classified into four different subtypes: (i) disorders of hemoglobin synthesis, such as thalassemia Genes 2021, 12, 958. https://doi.org/10.3390/genes12070958 https://www.mdpi.com/journal/genes