Brandon C. Iker

Relative abundance and treatment reduction of viruses during wastewater treatment processes--identification of potential viral indicators.

Waterborne pathogenic viruses discharged from wastewater treatment plants (WWTPs) pose potential public health risks. In the present study, we investigated the relative abundance, occurrence, and reduction of eleven different viruses at two WWTPs in southern Arizona over a 12-month period, from August 2011 to July 2012. Influent and effluent samples from the two WWTPs were collected monthly. Viruses were concentrated using an electronegative filter method and quantified using TaqMan-based quantitative PCR (qPCR) assays for each of the virus types (i.e., genogroup I, II and IV noroviruses, sapovirus, enterovirus, group A rotavirus, Aichi virus, pepper mild mottle virus, adenovirus, and JC and BK polyomaviruses), with murine norovirus internal control for the monitoring of extraction-RT-qPCR efficiencies. The pepper mild mottle virus, a plant virus, was found to be the most prevalent virus in both influent and effluent wastewater (annual mean concentration of 3.7-4.4×10(6) copies/L and 4.6-6.3×10(5) copies/L in influent and effluent wastewater, respectively), showing a low reduction by the treatment processes (0.76-0.99 annual mean log10 reduction), and no significant seasonal change in concentration. Aichi virus, a human enteric virus, was also found in greater abundance, and showed lower reduction during wastewater treatment than other human enteric viruses. Our results suggest that these viruses could be used as potential indicators of wastewater reclamation system performance, with respect to virus occurrence and removal.

Occurrence of Cryptosporidium, Giardia, and Cyclospora in influent and effluent water at wastewater treatment plants in Arizona

We investigated the occurrence of Cryptosporidium, Giardia, and Cyclospora at two wastewater treatment plants (WWTPs) in Arizona over a 12-month period, from August 2011 to July 2012. Influent and effluent wastewater samples were collected monthly, and protozoan (oo)cysts were concentrated using an electronegative filter, followed by the detection of protozoa using fluorescent microscopy (Cryptosporidium oocysts and Giardia cysts) and PCR-based methods (Cryptosporidium spp., Giardia intestinalis, and Cyclospora cayetanensis). The concentration of Giardia cysts in the influent was always higher than that of Cryptosporidium oocysts (mean concentration of 4.8-6.4×10(3) versus 7.4×10(1)-1.0×10(2)(oo)cysts/l) with no clear seasonality, and log10 reduction of Giardia cysts was significantly higher than that of Cryptosporidium oocysts for both WWTPs (P<0.05). Log10 reduction of Giardia cysts at the WWTP utilizing activated sludge was significantly higher than the other WWTP using trickling filter (P=0.014), while no statistically significant difference between the two WWTPs was observed for the log10 reduction of Cryptosporidium oocysts (P=0.207). Phylogenetic analysis revealed that G. intestinalis strains identified in wastewater belonged to two assemblages, AII and B, which are potentially infectious to humans. C. cayetanensis was also detected from both influent and effluent using a newly developed quantitative PCR, with the highest influent concentration of 1.2×10(4)copies/l. Our results demonstrated that these protozoan pathogens are prevalent in the study area and that efficacy of the conventional wastewater treatment processes at physically removing (oo)cysts is limited.

Evaluation of commercial kits for the extraction and purification of viral nucleic acids from environmental and fecal samples.

The extraction and purification of nucleic acids is a critical step in the molecular detection of enteric viruses from environmental or fecal samples. In the present study, the performance of three commercially available kits was assessed: the MO BIO PowerViral Environmental DNA/RNA Isolation kit, the Qiagen QIAamp Viral RNA Mini kit, and the Zymo ZR Virus DNA/RNA Extraction kit. Viral particles of adenovirus 2 (AdV), murine norovirus (MNV), and poliovirus type 1 (PV1) were spiked in molecular grade water and three different types of sample matrices (i.e., biosolids, feces, and surface water concentrates), extracted with the kits, and the yields of the nucleic acids were determined by quantitative PCR (qPCR). The MO BIO kit performed the best with the biosolids, which were considered to contain the highest level of inhibitors and provided the most consistent detection of spiked virus from all of the samples. A qPCR inhibition test using an internal control plasmid DNA and a nucleic acid purity test using an absorbance at 230 nm for the nucleic acid extracts demonstrated that the MO BIO kit was able to remove qPCR inhibitors more effectively than the Qiagen and Zymo kits. These results suggest that the MO BIO kit is appropriate for the extraction and purification of viral nucleic acids from environmental and clinical samples that contain high levels of inhibitors.

Bacterial populations within copper mine tailings: long-term effects of amendment with Class A biosolids.

This study evaluates the effect of surface application of dried Class A biosolids on microbial populations within copper mine tailings.

Viral presence in waste water and sewage and control methods.

Abstract: This chapter discusses the occurrence of different virus types in wastewaters (sewage) and their removal by various treatment processes. Factors that influence the occurrence and concentration of viruses in wastewater are reviewed and the mechanisms involved in their removal discussed.

Norovirus Outbreaks Among Colorado River Rafters in the Grand Canyon, Summer 2012

OBJECTIVE To investigate the incidence and causative agent of the recurrent outbreaks of acute gastrointestinal illness (AGI) among different rafting groups on the Colorado River in the Grand Canyon National Park during the 2012 summer season. METHODS Confidential illness reports were completed by all individuals with symptoms of AGI, and samples of fecal matter and vomitus, surface swabs of rafting equipment, and environmental swabs at stops along the hiking corridor were collected and tested for the presence of norovirus using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). RESULTS During the active outbreak period between May 9 and July 9, 2012, 97 rafters (1.4%) from 10 trips (2.9% of all trips) declared AGI symptoms. AGI incidence within the 10 infected trips varied from 6% to 88%. Outbreaks occurred in 3 distinct temporal clusters that involved 2 different genogroups of norovirus. All available toilet fecal samples (5 samples) were positive for norovirus RNA: 1 with genogroup I (GI) and 4 with GII. The vomitus sample tested positive for GI. None of the fomite samples from rafting equipment or from the hiking corridors were confirmed for norovirus. CONCLUSIONS The results suggest that norovirus may have been introduced by ill or asymptomatic individuals actively shedding the virus in their vomitus or feces, and spread within, or between, river trips by different modes of transmission. This study reinforces the importance of appropriate guidance and practice regarding norovirus prevention and the necessity of postoutbreak containment in relatively isolated groups of individuals.

Temporal variations in genotype distribution of human sapoviruses and Aichi virus 1 in wastewater in Southern Arizona, United States

To investigate the molecular epidemiology, especially temporal variations in genotype distribution, of sapoviruses and Aichi virus 1 (AiV‐1) in Arizona, United States, by examining wastewater.