Pierre-Louis, E. | Search bioRxiv

bioRxiv2023

Massive invasion of organellar DNA drives nuclear genome evolution in Toxoplasma

Toxoplasma gondii is a zoonotic protist pathogen that infects up to 1/3 of the human population. This apicomplexan parasite contains three genome sequences: nuclear (63 Mb); plastid organellar, ptDNA (35 kb); and mitochondrial organellar, mtDNA (5.9 kb of non-repetitive sequence). We find that the nuclear genome contains a significant amount of NUMTs (nuclear DNA of mitochondrial origin) and NUPTs (nuclear DNA of plastid origin) that are continuously acquired and represent a significant source of intraspecific genetic variation. NUOT (nuclear DNA of organellar origin) accretion has generated 1.6% of the extant T. gondii ME49 nuclear genome; the highest fraction ever reported in any organism. NUOTs are primarily found in organisms that retain the non-homologous end-joining repair pathway. Significant movement of organellar DNA was experimentally captured via amplicon sequencing of a CRISPR-induced double-strand break in non-homologous end-joining repair competent, but not ku80 mutant, Toxoplasma parasites. Comparisons with Neospora caninum, a species that diverged from Toxoplasma [~]28 MY ago, revealed that the movement and fixation of 5 NUMTs predates the split of the two genera. This unexpected level of NUMT conservation suggests evolutionary constraint for cellular function. Most NUMT insertions reside within (60%) or nearby genes (23% within 1.5 kb) and reporter assays indicate that some NUMTs have the ability to function as cis-regulatory elements modulating gene expression. Together these findings portray a role for organellar sequence insertion in dynamically shaping the genomic architecture and likely contributing to adaptation and phenotypic changes in this important human pathogen. Significance StatementThis study reveals how DNA located in cellular compartments called organelles can be transferred to the nucleus of the cell and inserted into the nuclear genome of apicomplexan parasite Toxoplasma. Insertions alter the DNA sequence and may lead to significant changes in how genes function. Unexpectedly, we found that the human protist pathogen, Toxoplasma gondii and closely-related species have the largest observed organellar genome fragment content (>11,000 insertion comprising over 1 Mb of DNA) inserted into their nuclear genome sequence despite their compact 65 Mb nuclear genome. Insertions are occurring at a rate that makes them a significant mutational force that deserves further investigation when examining causes of adaptation and virulence of these parasites.

evolutionary biology↗

bioRxiv2020

The need to harmonize insecticide resistance testing: methodology, intensity concentrations and molecular mechanisms evaluated in Aedes aegypti populations in Central America and Hispaniola

BackgroundThe Zika AIRS Project, a USAID-funded initiative worked across the Latin America and Caribbean regions from 2016 to 2019, as an emergency to contain the spread of the Zika virus. All entomological records in the target countries showed wide distribution and high abundance of Aedes aegypti populations, however the susceptibility profiles of these insects to insecticides commonly employed by vector control campaigns were in most cases incomplete or inexistent. In close collaboration with the Ministries of Health of individual countries, Zika-AIRS teams conducted insecticide susceptibility testing of an array of insecticides in A. aegypti populations of each country. Procedures applied met the standard international protocols instructed by the World Health Organization and Centers for Disease Control and Prevention. Methodology and main findingsThe insecticides tested were selected under categories such as pyrethroids, organophosphates and carbamate. Results showed A. aegypti populations displaying high and widely distributed resistance to all pyrethroids across countries, tolerance to organophosphates and full susceptibility to a carbamate. Key inconsistencies between testing methods are presented and discussed. Additionally, four kdr mutations were analyzed to detect molecular mechanisms of insecticide resistance. The screening for kdr mutations suggested the widespread nature of V1016I mutation, linked to pyrethroid resistance in A. aegypti populations distributed and sampled in the above mentioned regions. Conclusions and perspectivesThis multi-country study contributes with updated information to the public health decision-makers across Central America and the Caribbean. This study provided training and established technical networks for more effective and sustainable insecticide surveillance programs. Most but not all records of insecticide resistance in A. aegypti were consistent between methodologies, thus inconsistent issues are discussed here to call for further improvement in procedures and convey more practical guidelines for surveillance teams in countries where Aedes-borne diseases are endemic. Author summaryAt the forefront of the fight against arboviruses transmission is the insecticide-based vector control. All countries in the Latin American and Caribbean region invest valuable resources from their limited budget to acquire and implement insecticide-based tools, with non-existent or weak insecticide resistance monitoring programs. Hence, the USAID-funded Zika AIRS Project (ZAP) collaborated with the Ministries of Health of multiple countries to update the profile of susceptibility to insecticides in Aedes aegypti populations. We found widespread resistance to pyrethroid and organophosphate insecticides, which account to almost 100% of the products available to control adult mosquitoes. As we used both of World Health Organization and Centers for Disease Control and Prevention standard methods, we found many similarities and some inconsistencies in the susceptibility profiles obtained for the very same vector populations. Additionally, we obtained insight on potential molecular mechanisms of resistance across the countries, finding the kdr mutation V1016I possibly involved in loss of susceptibility. This study is the biggest cross-country update of insecticide resistance for Aedes aegypti in years, and it should be used as evidence for improving the selection of insecticides in these countries and a call for further support to maintain insecticide resistance monitoring programs.

ecology↗