Modifying the surface charge of pathogen-sized microspheres for studying pathogen transport in groundwater.

Consuming pathogen-contaminated groundwater has caused many waterborne disease worldwide. Microspheres are often used as pathogen surrogates because they can be made similar to pathogens in terms of their sizes, buoyant densities, and shapes. Laboratory studies have, however, shown that the surface charges of microspheres are very different from those of pathogens of comparable sizes, and that their attenuation and transport behaviors differ significantly to those of the pathogens mimicked. Thus, for microspheres to be better surrogates, their surface charges need to be modified. We have demonstrated that the surface charge of a microorganism can be closely mimicked by microspheres covalently coated with a protein that has a similar pHPZC to the microorganism. Using MS2 bacteriophage to test our concept, 20 nm carboxylated microspheres were covalently coated with casein. Zeta potentials as a function of pH were determined for purified MS2, casein, and uncoated and coated microspheres. The uncoated microspheres were significantly more negatively charged than MS2. The coated microspheres displayed zeta potentials and a pHPZC value similar to MS2. The modified surface charge on the microspheres was stable for at least 4 mo. Using the concept developed from this study, surrogates for many specific pathogens of concern can be developed, and the results can be corrected with pathogen die-off determined independently in the laboratory. Protein-coated microspheres could provide a new and alternative approach to investigate pathogen transport in groundwater. Future research is required to validate the surrogates' resemblances to pathogens in terms of their attenuation and transport behaviors in groundwater.

Comparison of particle size spectrum determination from images made using manual and automated image analysis.

A digital processing method is described for determining the size spectra of sub-micron particles in natural water from transmission electron microscopy images of particles collected by ultracentrifugation, and is compared with traditional manual counting and size measurement methods. The processing method is based on the use of the MatLab Image Processing toolbox. The manual method was found to underestimate the population of particles smaller than 40 nm equivalent radius, primarily because of a "fatigue factor" in counting the very large numbers of particles in this size range. By contrast, the manual method produced higher particle counts of particles >50 nm radius, primarily because manual counters tend to group together particles as aggregates that digital processing indicates are not contiguous. The digitally-produced size spectrum for a river water sample was found to closely follow a power-series law down to the smallest particle size.