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We report an index patient with complete insensitivity to pain and a history of painless fractures, joint hypermobility, and behavioral problems. The index patient descends from a family with notable cases among his maternal relatives, including his aunt and his mother’s first cousin, both of whom suffer from congenital insensitivity to pain. The patient had normal results for prior genetic testing: fragile-X syndrome testing, chromosomal microarray analysis, and exome sequencing. Optical genome mapping detected a homozygous deletion affecting the noncoding 5′ untranslated region (UTR) and the first non-coding exon of the SCN9A gene in all affected family members, compatible with recessive disease transmission. Pathogenic homozygous loss-of-function variants in the SCN9A gene are associated with impaired pain sensation in humans. Optical genome mapping can thus detect pathogenic structural variants in patients without molecular etiology by standard diagnostic procedures and is a more accessible diagnostic tool than short-read or long-read whole-genome sequencing.

AbstractPneumocystis jirovecii is an unculturable fungus and the causative agent of Pneumocystis pneumonia, a life-threatening opportunistic infection. Although molecular diagnosis is often based on the mtLSU rRNA mitochondrial gene due to its greater sensitivity, physiology and the dynamics of the mitochondria in this fungus remains largely unknown. We developed and optimized six real-time PCR assays in order to determine the copy number of four mitochondrial genes (mtSSU rRNA, mtLSU rRNA, NAD1 and CYTB) in comparison to nuclear genome (DHPS and HSP70) and tested 84 bronchoalveolar fluids of patients at different stages of the infection. Unexpectedly, we found that copy number of mitochondrial genes varied from gene to gene with mtSSU rRNA gene being more represented (37 copies) than NAD1 (23 copies), mtLSU rRNA (15 copies) and CYTB (6 copies) genes compared to nuclear genome. Hierarchical clustering analysis (HCA) allowed us to define five major clusters, significantly associated with fungal load (p=0.029), in which copy number of mitochondrial genes was significantly different among them. More importantly, copy number of mtLSU rRNA, NAD1 and CYTB but not mtSSU rRNA differed according to P. jirovecii physiological state with a decreased number of copies when the fungal load is low. This suggests the existence of a mixture of various subspecies of mtDNA that can harbor different amplification rates. Overall, we revealed here an unexpected plasticity and dynamics of P. jirovecii mitochondrial DNA that vary according to P. jirovecii’s physiological state.