Immunoprevalence to Six Waterborne Pathogens in Beachgoers at Boquerón Beach, Puerto Rico: Application of a Microsphere-Based Salivary Antibody Multiplex Immunoassay.

Waterborne infectious diseases are a major public health concern worldwide. Few methods have been established that are capable of measuring human exposure to multiple waterborne pathogens simultaneously using non-invasive samples such as saliva. Most current methods measure exposure to only one pathogen at a time, require large volumes of individual samples collected using invasive procedures, and are very labor intensive.

A Spike Cocktail Approach to Improve Microbial Performance Monitoring for Water Reuse.

  Water reuse, via either centralized treatment of traditional wastewater or decentralized treatment and on-site reuse, is becoming an increasingly important element of sustainable water management. Despite advances in waterborne pathogen detection methods, low and highly variable pathogen levels limit their utility for routine evaluation of health risks in water reuse systems. Therefore, there is a need to improve our understanding of the linkage between pathogens and more readily measured process indicators during treatment.

The influence of incubation time on adenovirus quantitation in A549 cells by most probable number.

Cell culture based assays used to detect waterborne viruses typically call for incubating the sample for at least two weeks in order to ensure that all the culturable virus present is detected. Historically, this estimate was based, at least in part, on the length of time used for detecting poliovirus. In this study, we have examined A549 cells infected with human adenovirus type 2, and have found that a three week incubation of virus infected cells results in a higher number of detected viruses by quantal assay than what is seen after two weeks of incubation, with an average 955% increase in Most Probable Number (MPN) from 2 weeks to 3 weeks.

Optimization and evaluation of a method to detect adenoviruses in river water.

Adenoviruses are often implicated in recreational water disease outbreaks but existing methods for their detection perform poorly within these matrices. In this study, small volume (100mL) concentration was used to identify processes that promoted recovery of adenovirus from river water. Several alternative secondary concentration techniques were investigated and compared to the baseline method consisting of primary concentration via filtration, followed by celite mediated secondary concentration.

Statistical approaches to developing a multiplex immunoassay for determining human exposure to environmental pathogens.

There are numerous pathogens that can be transmitted through water. Identifying and understanding the routes and magnitude of exposure or infection to these microbial contaminants are critical to assessing and mitigating risk. Conventional approaches of studying immunological responses to exposure or infection such as Enzyme-Linked Immunosorbent Assays (ELISAs) and other monoplex antibody-based immunoassays can be very costly, laborious, and consume large quantities of patient sample.

EPA Method 1615. Measurement of enterovirus and norovirus occurrence in water by culture and RT-qPCR. I. Collection of virus samples.

EPA Method 1615 was developed with a goal of providing a standard method for measuring enteroviruses and noroviruses in environmental and drinking waters. The standardized sampling component of the method concentrates viruses that may be present in water by passage of a minimum specified volume of water through an electropositive cartridge filter. The minimum specified volumes for surface and finished/ground water are 300 L and 1,500 L, respectively.

Development and evaluation of EPA method 1615 for detection of enterovirus and norovirus in water.

The U.S. EPA developed a sample concentration and preparation assay in conjunction with the total culturable virus assay for concentrating and measuring culturable viruses in source and drinking waters as part of the Information Collection Rule (ICR) promulgated in 1996.

Evaluation of the celite secondary concentration procedure and an alternate elution buffer for the recovery of enteric adenoviruses 40 and 41.

The effective recovery of adenovirus from water is a critical first step in developing a virus occurrence method able to provide accurate data for risk assessments and other applications. During virus concentration, electropositive filters are typically eluted with beef extract, undergo secondary concentration using either an organic flocculation or polyethylene glycol (PEG) precipitation technique and are ultimately resuspended in sodium phosphate buffer. In this study, an alternative secondary concentration procedure using celite was optimized by identifying the optimal celite and elution buffer to use.

Development of an astrovirus RT-PCR detection assay for use with conventional, real-time, and integrated cell culture/RT-PCR.

Astrovirus is a common cause of gastroenteritis in humans that has been determined to be responsible for outbreaks of illness in several countries. Since astrovirus can be waterborne, it is important to be able to identify this virus in environmental water. We have developed and optimized a reverse transcription - polymerase chain reaction (RT-PCR) method that was able to amplify all eight astrovirus serotypes in a single reaction.

Development of a molecular method to identify hepatitis E virus in water.

Hepatitis E virus (HEV) causes an infectious form of hepatitis associated with contaminated water. By analyzing the sequence of several HEV isolates, a reverse transcription-polymerase chain reaction method was developed and optimized that should be able to identify all of the known HEV strains. When tested under laboratory conditions, this method was able to detect low levels of five diverse HEV variants.