Detection of circoviruses and porcine adenoviruses in water samples collected from swine manure treatment systems

Padilha

Schissi

et al. 2015

Environ Sci Pollut Res

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This study was designed to assess the presence of human adenovirus (HAdV), rotavirus-A (RVA), hepatitis A virus (HAV), and porcine circovirus-2 (PCV2) in groundwater from deep wells, and recreational and network waters. The water samples were collected and concentrated and the virus genomes were assessed and quantified by quantitative PCR (qPCR). Infectious HAdV was evaluated in groundwater and network water samples by integrated cell culture using transcribed messenger RNA (mRNA) (ICC-RT-qPCR). In recreational water samples, HAdV was detected in 100 % (6/6), HAV in 66.6 % (4/6), and RVA in 66.6 % (4/6). In network water, HAdV was detected in 100 % (6/6) of the samples (these 83 % contained infectious HAdV), although HAV and RVA were not detected and PCV2 was not evaluated. In groundwater from deep wells, during rainy period, HAdV and RVA were detected in 80 % (4/5) of the samples, and HAV and PCV2 were not detected; however, during dry period, HAdV and RVA were detected in 60 % (3/5), HAV in only one sample, and PCV2 in 60 % (4/5). In groundwater, all samples contained infectious HAdV. PCV2 presence in groundwater is indicative of contamination caused by swine manure in Concórdia, Santa Catarina, Brazil. The disinfection of human and animal wastes is urgent, since they can contaminate surface and groundwater, being a potential threat for public and animal health.

Section: Real-time Qpcrmentioning

confidence: 99%

Human and animal enteric virus in groundwater from deep wells, and recreational and network water

Padilha

Schissi

et al. 2015

Environ Sci Pollut Res

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“…During the experiment, 20 mL of each fraction (liquid and solid) was collected at t 0h , t 3h , and t 24h and concentrated using the glycine buffer method (Viancelli et al 2012). The DNA was extracted from each sample using the QIAmp MinElute Virus Spin Kit (Qiagen, Germany) according to the manufacturer's instructions.…”

Section: Pcv2 Analysesmentioning

confidence: 99%

“…The DNA was extracted from each sample using the QIAmp MinElute Virus Spin Kit (Qiagen, Germany) according to the manufacturer's instructions. For PCV2 detection, DNA was submitted to quantitative PCR (qPCR) following the protocols described by Viancelli et al (2012). The results were expressed as genome copies (gc).…”

Section: Pcv2 Analysesmentioning

confidence: 99%

Pathogen Inactivation and the Chemical Removal of Phosphorus from Swine Wastewater

Viancelli

Kunz

et al. 2015

Water Air Soil Pollut

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Inactivation of pathogens present in animal manure prior to land application has justified the use of advanced technologies. However, some alternatives are expensive or not effective due to the organic material and suspended solids present in the effluent (e.g., ozone, UV light). The use of hydrated lime (calcium hydroxide, Ca(OH) 2 ) is an attractive wastewater treatment option due to the ability of lime to kill pathogens and to extract phosphorus from manure at an alkaline pH. The present study aimed to evaluate the soluble phosphorus removal and pathogen inactivation (Escherichia coli, Salmonella enterica serovar typhymurium and Porcine circovirus type 2), in the liquid fraction and in the solid generated after Ca(OH) 2 addition in swine wastewater, exposed for 3 and 24 h at different pH conditions: 9.0, 9.5, and 10.0. The results showed the efficiency of pH elevation with Ca(OH) 2 in the removal of soluble P at pH 9.0 and the total inactivation of E. coli, Salmonella, and P. circovirus type 2 at pH 10.0. The liquid fraction (reuse water) could be safely used for cleaning the swine production facilities, and the solid fraction (precipitated P) could be used as a secondary product and fertilizer.

“…To infer the presence of undamaged viral particles, HAdV-positive samples, as detected by qPCR, were treated with DNase I, as described by Viancelli et al (2011) [42]. To verify potential inhibitors of DNAse I present in the sample matrix, a known amount of previously inactivated HAdV-2 (1 h at 99°C and 30 min under UV irradiation) was added in concentrated samples (previously HAdV-2 negative) of all sites and in nuclease-free water (NFW), as a control.…”

Section: Enzymatic Assaymentioning

confidence: 99%

Evaluation and molecular characterization of human adenovirus in drinking water supplies: viral integrity and viability assays

Nascimento

Rigotto

et al. 2013

Virol J

BackgroundHuman adenoviruses (HAdVs) are the second-leading cause of childhood gastroenteritis worldwide. This virus is commonly found in environmental waters and is very resistant to water disinfection and environmental stressors, especially UV light inactivation. Molecular techniques, such as PCR-based methods (Polymerase Chain Reaction), are commonly used to detect and identify viral contamination in water, although PCR alone does not allow the discrimination between infectious and non-infectious viral particles. A combination of cell culture and PCR has allowed detection of infectious viruses that grow slowly or fail to produce cytopathic effects (CPE) in cell culture. This study aimed to assess the integrity and viability of human adenovirus (HAdV) in environmental water and evaluate circulating strains by molecular characterization in three sites of the water supply in Florianópolis, Santa Catarina Island, Brazil: Peri Lagoon water, spring source water, and water from the public water supply system.MethodsWater samples were collected, concentrated and HAdV quantified by real-time PCR. Viral integrity was evaluated by enzymatic assay (DNase I) and infectivity by plaque assay (PA) and integrated cell culture using transcribed mRNA (ICC-RT-qPCR). Samples containing particles of infectious HAdV were selected for sequencing and molecular characterization.ResultsThe analyzed sites contained 83, 66 and 58% undamaged HAdV particles (defined as those in which the genetic material is protected by the viral capsid) at Peri Lagoon, spring source water and public supply system water, respectively. Of these, 66% of the particles (by PA) and 75% (by ICC-RT-qPCR) HAdV were shown to be infectious, due to being undamaged in Peri Lagoon, 33% (by PA) and 58% (by ICC-RT-qPCR) in spring source water and 8% (by PA) and 25% (by ICC-RT-qPCR) in the public water supply system. ICC-RT-qPCR, a very sensitive and rapid technique, was able to detect as low as 1 × 102 HAdV genome copies per milliliter of infectious viral particles in the environmental water samples. The molecular characterization studies indicated that HAdV-2 was the prevalent serotype.ConclusionsThese results indicate a lack of proper public health measures. We suggest that HAdV can be efficiently used as a marker of environmental and drinking water contamination and ICC-RT-qPCR demonstrated greater sensitivity and speed of detection of infectious viral particles compared to PA.