# Biparental Inheritance and Instability of kDNA in Experimental Hybrids of Trypanosoma cruzi: A Proposal for a Mechanism

## TL;DR

This study explores how genetic material is inherited in hybrids of the Chagas disease parasite, revealing that small DNA rings come from both parents while large ones come from one.

## Contribution

The study proposes a new mechanism called 'Replicative Mixing' to explain biparental inheritance of minicircles and uniparental inheritance of maxicircles in T. cruzi hybrids.

## Key findings

- Minicircles from both parents persist in hybrid offspring for hundreds of generations.
- Maxicircles are inherited exclusively from one parent in hybrids.
- Hybrids show increased and unstable levels of kinetoplast DNA compared to their parents.

## Abstract

Chagas disease is caused by the parasite Trypanosoma cruzi, which carries a dense bundle of thousands of interlocked DNA rings called the kinetoplast in its mitochondrion. How this genetic material is passed on when two parasites form a hybrid has remained unclear. Here, we analyzed laboratory hybrids of T. cruzi and tracked the parental origin and quantity of the small DNA rings (“minicircles”) and the large DNA rings (“maxicircles”) across many generations. We found that minicircles from both parents persist together in the same offspring lines for hundreds of generations, while maxicircles persist only from one parent. We also observed that hybrids often carry increased amounts of this kinetoplast DNA compared with their parents, and that these levels vary over time. To explain these patterns, we propose a “replicative mixing” mechanism: after mating, the two parental kinetoplasts replicate, minicircles intermingle across them, and later the networks separate so that each daughter cell retains maxicircles from one parent and minicircles from both.

The mitochondrial DNA of trypanosomatid parasites consists of thousands of catenated minicircles and dozens of maxicircles that form a complex network structure, the kinetoplast (kDNA). Although kDNA replication and segregation during mitotic division are well studied, its inheritance during genetic exchange events remains unclear. In Trypanosoma brucei, hybrids inherit minicircles biparentally but retain maxicircles from a single parent. Although biparental inheritance of minicircles has been described in natural Trypanosoma cruzi hybrids, this process has not been explored in laboratory-generated hybrids of this parasite. In the present study, we analyzed kDNA inheritance in T. cruzi experimental hybrids using a comprehensive minicircle hypervariable region (mHVR) database and genome sequencing data. Our findings revealed biparental inheritance of minicircles, with hybrid lines retaining mHVRs from both parents for over 800 generations. In contrast, maxicircles were exclusively inherited from one parent. Unexpectedly, we observed an increase in kDNA content in hybrids, affecting both minicircles and maxicircles, and exhibiting instability over time. To explain these findings, we propose a Replicative Mixing (REMIX) model, where the hybrid inherits one kinetoplast from each parent and they are replicated allowing minicircle mixing. Instead maxicircle networks remain physically separated, leading to uniparental fixation after segregation in the first cell division of the hybrid. This model challenges previous assumptions regarding kDNA inheritance and provides a new framework for understanding kinetoplast dynamics in hybrid trypanosomes.