Smart water disinfection project (2014-18)

Primary Investigator Benito J. Mariñas reports several advances in 2016-17. Technically speaking:

 

• Fundamental Advances: Viral pathogen inactivation mechanisms using free chlorine and ultraviolet (UV) light
  1. “We identified that free chlorine transforms the capsid in a manner that alters its integrity,” he said. “We are now using confocal microscopy to examine individual steps of the virus replication cycle. These findings will elucidate what are the possible proteins that are being mutagenized by free chlorine and how this disinfectant inactivates adenovirus.”
  2. In addition, the team found that exposure to UV irradiation that would produce a 99.99% virus inactivation does not seem to inhibit the association of HAdV-2 to the host cells. “It is quite remarkable that different UV wavelengths (UV220, UV254, and UV280), have different mechanisms of inactivation,” Mariñas said. “We now know that UV220 minimally impacted genome integrity but seemed to produce a structural transformation of the viral capsid that may inhibit the delivery of viral genome into the host cell nucleus. Quite surprisingly, it appears that UV254 and UV280 have at least two mechanisms of inactivation, which includes both mutagenesis of the viral genome and (we suspect) viral entry into the host.”
  3. Team members have also found that viruses are more susceptible to disinfection if treated sequentially with UV and monochloramine. “This is unexpected and may provide a new strategy for treating drinking water,” he said.
  4. As a second means to identify inactivation mechanisms of free chlorine, the team will perform in vitro evolution experiments in which they will isolate and propagate adenoviruses that are resistant to free chlorine. They will use deep sequencing of the genomes of these resistant adenoviruses, comparing them to the genomes of wild-type adenoviruses, to identify potential mutations responsible for resistance. “We will continue to use molecular techniques to determine how sequential application of UV light irradiation and free or combined chlorine inactivates HAdV-2,” Mariñas said.

• Innovative Sensing: Real-time detection of infective viruses
  1. The team has finished SELEX of aptamers for infectious adenovirus using graphene oxide to separate the DNA-adenovirus complex from non-binding DNA and obtained sequences of aptamers from the SELEX. Team members have identified and characterized DNA sequencing that are responsible for the selective binding.
  2. “We will continue to characterize the DNA aptamers for the infectious adenovirus and transform them into fluorescent sensors for quantitative detection, by labeling the DNA aptamers with a fluorophore and quencher,” Mariñas said. “Upon binding of the target, the fluorescent signal will increase due to changes of melting temperatures of the DNA aptamers. We will use a potable fluorimeter for such quantification.”
  3. In addition, to lower the costs for on-site and real-time detection, the team will label the DNA aptamer with an enzyme called invertase so that, upon binding of adenovirus, the sensing system will generate glucose, which can then be quantified by a glucose meter. For rapid semi-quantitative detection, the team will conjugate the DNA aptamer with gold nanoparticles and immobilize them onto a lateral flow device so that the binding of the DNA aptamer will result in change of color from blue to red.
  4. Sensitivity in terms of limit of detection and selectivity against other viruses, including non-infectious viruses, will be tested for sensing performance in water.

The team reported significant progress in some major areas of its research. Technically speaking:

 

• Viral inactivation: Team members determined that UV irradiation emitted by a medium pressure (MP) source inactivates human adenovirus (HAdV) through different mechanisms depending on specific wavelengths. The fast inactivation of HAdV by MPUV may be a combination of direct and indirect viral capsid protein damage. The knowledge of which viral component gets affected during UV light inactivation will allow the monitoring of this pathogen in drinking water, and effectiveness inspection of water disinfection by UV light.
• Viral inactivation: In the case of free chlorine and monochloramine disinfection, the team has shown that mRNA production is being disrupted at the same level as the kinetic rates of virus inactivation. This means that inhibition of an event occurring at or before early gene transcription must be the cause of adenovirus inactivation. Our findings will be the foundation to develop novel effective disinfectants and rapid methods for detection of infectious viruses in drinking water.
• Detection: The mechanism for adenovirus inactivation by free chlorine remains unknown, and is most likely caused by modifications to the viral capsid resulting in host cell entry inhibition. To gain an understanding of how the major capsid proteins of adenovirus are damaged by free chlorine, the team developed a method to express the proteins in Escherichia coli. The recombinant proteins were then subjected to increasing free chlorine exposure. Several modified residues in fiber, penton base, and hexon major capsid proteins were detected that are important for viral entry. These modifications may play a role in inhibition of viral entry, thus leading to adenovirus inactivation by free chlorine.
• Detection: The team has reported a simple and highly sensitive amplified aptamer-based fluorescent sensor for AFB₁, which relies on the ability of nano-graphene oxide (GO) to protect aptamers from nuclease cleavage for amplified detection and on the nanometer size effect of GO to tune the dynamic range and sensitivity. The sensor was highly selective against other aflatoxins and common molecules in foods, and its performance was verified in corn samples spiked with known concentration of AFB₁.
• Local entrepreneur business implementation: The team engaged an interdisciplinary group of students in a yearlong project with field research that involved developing sensors for water tanks. Working with an NGO partner in Tanzania, team members have begun marketplace and sustainability literacy education, using trainers. The groundwork of designing materials and translating them has been completed, and co-PI Madhu Viswanathan will be traveling to Tanzania to guide training of trainers.

SPRING 2015 —  In February, Professor Benito Mariñas and Illinois students from the CEE 449 Civil and Environmental Engineering Class visited Kenya and Uganda under the umbrella of the Safe Global Water Institute (SGWI) to learn about the needs, available resources and challenges in rural communities that need safe water and proper sanitation. The 12 undergraduate students who traveled, among other things, researched fluorosis (too much fluoride in water) and anaerobic digestion of waste (turning it into energy or fertilizer). To read more, visit the students’ blog site.

 

 

Pictured left:

CEE undergraduate student Gabrielle Levato collects a water sample from a borewell while fellow Illinois undergrads perform additional water quality analyses with the help of Makerere University graduate students during a February visit to the Oruchinga Refugee Settlement in Isingiro district, Uganda.