Biology subjects

Chaverra-Rodriguez, D.

Publications and source records attributed to Chaverra-Rodriguez, D..

4 recordsLinked to original sources

bioRxiv2025

Virome profiling of Culex tarsalis through small RNA-seq: A challenge of suboptimal samples

Viral infections in mosquitoes trigger the RNA interference (RNAi) pathway, a key antiviral defense mechanism that generates virus-derived small RNAs (vsRNAs). Given the natural enrichment of vsRNAs during infection and their stability, small RNA sequencing (sRNA-seq) has emerged as a powerful tool for virome characterization. Culex tarsalis is a widely distributed mosquito species in North America and is an important vector of West Nile virus (WNV). Previous studies have shown that co-infection with insect-specific viruses (ISVs) can modulate WNV replication in Cx. tarsalis, highlighting the importance of characterizing the virome of this species. Here, we investigated the virome of Cx. tarsalis populations across 5 states of the Midwestern United States using sRNA-seq. We analyzed samples from 17 geographic locations which were collected under suboptimal field conditions during the COVID-19 pandemic, presenting challenges related to sample integrity. Despite these challenges, sRNA-seq proved to be a reliable method for virome analysis. We identified seven viruses associated with Cx. tarsalis, along with their respective sRNA (siRNA and piRNA) profiles. These findings not only deepen our understanding of ISVs, but also demonstrate the utility of sRNA-seq in non-ideal situations, enabling the collection and analysis of samples under real-world surveillance scenarios.

microbiology↗

bioRxiv2023

CRISPR/Cas9-mediated mutagenesis of the Asian Citrus Psyllid, Diaphorina citri

The most devastating disease affecting the global citrus industry is Huanglongbing (HLB), caused by the pathogen Candidatus Liberibacter asiaticus. HLB is primarily spread by the insect vector Diaphorina citri (Asian Citrus Psyllid). To counteract the rapid spread of HLB by D. citri, traditional vector control strategies such as insecticide sprays, the release of natural predators, and mass introductions of natural parasitoids are used. However, these methods alone have not managed to contain the spread of disease. To further expand the available tools for D. citri control via generating specific modifications of the D. citri genome, we have developed protocols for CRISPR/Cas9-based genetic modification. Until now, genome editing in D. citri has been challenging due to the general fragility and size of D.citri eggs. Here we present optimized methods for collecting and preparing eggs to introduce the Cas9 ribonucleoprotein (RNP) into early embryos and alternative methods (ReMOT Control) for injecting RNP into the hemocoel of adult females for ovarian transduction. Using these methods, we have generated visible somatic mutations, indicating their suitability for gene editing in D. citri. These methods represent the first steps towards advancing D. citri research in preparation for future genetic-based systems for controlling HLB.

bioengineering↗

bioRxiv2020

Germline mutagenesis of Nasonia vitripennis through ovarian delivery of CRISPR-Cas9 ribonucleoprotein

CRISPR/Cas9 gene editing is a powerful technology to study the genetics of rising model organisms, such as the jewel wasp Nasonia vitripennis. However, current methods involving embryonic microinjection of CRISPR reagents are challenging. Delivery of Cas9 ribonucleoprotein into female ovaries is an alternative that has only been explored in a small handful of insects, such as mosquitoes and whiteflies. Here, we developed a simple protocol for germline gene editing by injecting Cas9 ribonucleoprotein in adult N. vitripennis females using either ReMOT control (Receptor-Mediated Ovary Transduction of Cargo) or BAPC (Branched Amphiphilic Peptide Capsules) as ovary delivery methods. We demonstrate efficient delivery of protein cargo such as EGFP and Cas9 into developing oocytes via P2C peptide and BAPC. Additionally, somatic and germline gene editing have been demonstrated. This approach will greatly facilitate CRISPR-applied genetic manipulation in this and other rising model organisms.

molecular biology↗

bioRxiv2019

Cas9-mediated gene-editing in the malaria mosquito Anopheles stephensi by ReMOT Control

Innovative tool development is essential for continued advancement in malaria control and depends on a deeper understanding of the molecular mechanisms that govern transmission of malaria parasites by Anopheles mosquitoes. Targeted disruption of genes in mosquito vectors is a powerful method to uncover the underlying biology of vector-pathogen interactions, and genome manipulation technologies can themselves form the basis of mosquito and pathogen control strategies. However, the embryo injection methods used to genetically manipulate mosquitoes, and in particular Anopheles species, are difficult and inefficient, particularly for non-specialist laboratories. We have adapted a strategy called ReMOT Control (Receptor-mediated Ovary Transduction of Cargo) to deliver the Cas9 ribonucleoprotein complex to adult mosquito ovaries and generate targeted and heritable mutations in the malaria vector Anopheles stephensi. We found that gene editing by ReMOT Control in Anopheles mosquitoes was comparable to the technique in Ae. aegypti and as efficient in editing as standard embryo injections. The adaptation of this technology to Anopheles mosquitoes opens up the power of reverse genetics to malaria vector labs that do not have the equipment or technical expertise to perform embryo injections and establishes the flexibility of ReMOT Control for gene-editing in non-Aedes species.

genetics↗