Genomic wastewater monitoring for surveillance of viral threats
Background
Wastewater sequencing is a powerful, cost-effective, and non-invasive tool for virus detection and public health monitoring. It has been successfully used to detect SARS-CoV-2 variants and can be expanded to monitor drug resistance and vaccine escape in viruses. Wastewater sequencing can also facilitate community-level surveillance by monitoring the prevalence of different viruses in a community. Additionally, it has the potential to enable the early detection of outbreaks before clinical cases are reported, including the detection of emerging and re-emerging viruses. Importantly, it allows for the monitoring of large populations without extensive individual testing.
Aim
Metagenomic approaches using next-generation sequencing are crucial for detecting a wide range of viruses. However, they are costly and often not optimized for the fragmented, low-quality nucleic acid in this type of samples. This project aims to design a hyper-multiplexed primer set targeting specific regions of a comprehensive set of viruses, enabling rapid and cost-effective detection, and quantification.
Project outline
To achieve this aim, you will begin with a literature review to understand the existing work in this area, focusing particularly on the strategies employed. Next, you will collect genome sequences from databases. Given the large volume of data, you will apply methods to retain representative sequences. Following this, you will perform sequence alignments and simultaneously design the primer strategy. This involves considering various factors, such as including amplicons for classifying viruses at the genus/family level and others for species/variant identification. Finally, you will conduct a mock community simulation. This step involves testing the designed primers to assess their in silico coverage of the target viral sequences and ensuring primer specificity to avoid cross-reacting with non-target sequences. You will use alignment tools (MEGA, Clustal, MAFFT), primer design software (Primer3, NCBI Primer-BLAST), and tools for mock community simulation (In silico PCR, FastPCR), among others. The final output of this rotation will be a database of curated primers with annotations for their intended use (genus or species identification) and a comprehensive report.
Future work
Following the rotation, this project will extend to lab validation. This phase will assess the sensitivity and specificity against key viruses using simulated wastewater samples spiked with synthetic viral genomes in known proportions. The designed primers will include unique molecular barcodes to facilitate quantification of the viruses. After this stage of wet lab validation, the project will advance to processing and analysing real wastewater samples.