With ever increasing threats from emergent pathogens and multiple resistant bacteria, hospitals are cracking down with more rigorous monitoring and decontamination procedures to prevent pathogen spread.
Emergency Medical Service (EMS) vehicles and workers are often the first point of patient care, however, little is known about the importance of EMS vehicles for the transmission of pathogens, and detection methods lag behind those of hospitals.
We are Nanoresponder! A team dedicated to the advancement of pathogen detection and monitoring in EMS vehicles. We have taken a dual approach to pathogen detection: (1) Nanopore Next Generation Sequencing (NGS) of DNA samples from EMS vehicles; and (2) A rapid single-domain antibody detection system to generate antibodies that may be implemented in simple and reliable testing kits for fast, on-site pathogen detection.
EMS Vehicle Microbiome Investigation
Upon conducting an in-depth review of literature surrounding EMS vehicle pathogen detection and interviews with EMS officials, we discovered that no published study utilizes a Next Generation Sequencing approach to pathogen detection in EMS vehicles and instead solely cell culturing techniques were used. This is surprising considering that many microbes are unculturable in a laboratory setting and would otherwise escape detection. Moreover, cell culturing techniques may require many days before test results are available.
Recognizing the need for high-throughput and inclusive pathogen detection methods, we developed a detection platform, utilizing the MinION - a novel NGS sequencing technology from Oxford Nanopore Technologies.
The MinION Next Geneneration Sequencing Platform
In collaboration with our local EMS department, we devised a sampling strategy, DNA extraction procedure and amplification procedure that is compatible with the MinION sequencing technology.
Sampling locations for the EMS Microbiome Study
Using polymerase chair reaction, we amplified ribosomal small subunit gene sequences from extracted sample DNA and subjected them to MinION deep sequencing. We found marked differences in microbial community structure between sample and control DNAs, with many EMS vehicles samples containing a large proportion of opportunistic pathogens.
Sampling locations for the EMS Microbiome Study
Single Domain Antibody Development System
Recognizing that EMS vehicles may be reservoirs of pathogens, we next wished to meet the need for a rapid and on-site pathogen detection technology that uses specific antibodies for detection. Towards this we engineered a library of synthetic single domain antibodies (Nanobodies) as well as a bacterial two hybrid system for antibody selection.
Engineered Bacterial-two-hybrid Antibody Selection System
We successfully demonstrate an integrated DNA sequencing plus antibody detection platform for monitoring the presence of pathogens in EMS vehicles that may be ported to function in other portions of health, environment and other sectors.
