Novel bi-allelic variants in DNAH2 cause severe asthenoteratozoospermia with multiple morphological abnormalities of the flagella

Abstract Research Question Multiple morphological abnormalities of the flagella (MMAF) is a severe subtype of asthenoteratozoospermia characterised by excessive immotile spermatozoa with severe flagellar abnormalities in the ejaculate. Previous studies reported a heterogeneous genetic profile associated with MMAF. However, the identified genetic variants cannot fully explain the aetiology of the MMAF-affected asthenoteratozoospermia. Design Whole-exome sequencing in a cohort of 90 Chinese MMAF patients was performed and three unrelated men with bi-allelic DNAH2 variants were identified. Results Sanger PCR verified that the six novel variants originated from the parents of the MMAF-affected men. All these variants were located at the conserved domains of DNAH2, and predicted to be deleterious by bioinformatic tools. H&E staining and scanning electron microscopy revealed that spermatozoa harbouring DNAH2 variants displayed severely aberrant morphology mainly with absent and short flagella (>78%). Moreover, transmission electron microscopy revealed the obvious absence of a central pair of microtubules and inner dynein arms in the spermatozoa with mutated DNAH2. Immunofluorescence data further validated these findings, showing reduced DNAH2 protein expression in the spermatozoa with DNAH2 variants, compared with normal sperm. Intracytoplasmic sperm injection was used to achieve good quality blastocysts with the spermatozoa from all the three men with mutated DNAH2, ultimately resulting in clinical pregnancy. Conclusions These experimental and clinical data suggest that bi-allelic DNAH2 variants might induce MMAF-associated asthenoteratozoospermia, which can be overcome through intracytoplasmic sperm injection. These findings contribute to the knowledge of the genetic landscape of asthenoteratozoospermia and clinical counselling of male infertility.