Viruses In Foods Research Articles

Environmental virology linked to waterborne diseases and foodborne pathogens: Human and animal food viruses

Foodborne illnesses constitute a serious global one health issue. The transmission of foodborne viruses can occur due to inadequate hygiene of food, lack of basic sanitation, poor personal hygiene and consumption of raw or undercooked food. This study aims overview about environmental virology linked to waterborne diseases and foodborne pathogens in human and animal food viruses. Thus, epidemiological studies have proposed some emerging pathogens as possible transmitters of contaminated water ingestion based on biochemical properties and viral structures such as: Picornaviridae (polioviruses, enteroviruses, coxsakiviruses, hepatitis A viruses and echoviruses), Adenoviridae (adenoviruses), Caliciviridae (noroviruses, caliciviruses, astroviruses), and Reoviridae (reoviruses and rotaviruses). These viruses can cause gastrointestinal, liver and central nervous system disorders. In controlling foodborne infections, accurately identifying sources of contamination is a significant challenge, given the complexity of the food production chain. The lack of adequate monitoring and neglect of good health practices contribute to the spread of these infections. Large-scale immunization has contributed to the reduction of enteric outbreaks and eradication of cases in many regions of the world as a prevention of hepatitis A and poliomyelitis. However, challenges such as the lack of effectiveness in inspecting viruses in food increase the risk of outbreaks and epidemics, with emphasis on Noroviruses as the main causes of these infections today. Investment in vaccine research and development has been a fundamental strategy to reduce these statistics, especially in vulnerable groups. Moreover, the implementation molecular-based tools and wastewater-based epidemiology studies to high-throughput for aquatic biomonitoring should be up to date to avoid the risk and impact on public health and aquatic environments. Novel perspectives to control of waterborne viruses has been introduced in continuous surveillance programs for ecosystems monitoring.

Duplex Droplet Digital PCR Assay for Quantification of Hepatitis E Virus in Food.

Hepatitis E virus (HEV) represents an emerging risk in industrialized countries where the consumption of contaminated food plays a pivotal role. Quantitative real-time RT-PCR (RT-qPCR) is one of the most suitable methods for the detection and quantification of viruses in food. Nevertheless, quantification using RT-qPCR has limitations. Droplet digital PCR (ddPCR) provides the precise quantification of nucleic acids without the need for a standard curve and a reduction in the effect on virus quantification due to the presence of inhibitors. The objectives of the present work were (i) to develop a method for the absolute quantification of HEV in swine tissues based on ddPCR technology and provide internal process control for recovery assessment and (ii) to evaluate the performance of the method by analyzing a selection of naturally contaminated wild boar muscle samples previously tested using RT-qPCR. The method was optimized using a set of in vitro synthesized HEV RNA and quantified dsDNA. The limit of detection of the developed ddPCR assay was 0.34 genome copies/µL. The analysis of the wild boar samples confirmed the validity of the ddPCR assay. The duplex ddPCR method showed no reduction in efficiency compared to individual assays. The method developed in the present study could represent a sensitive assay for the detection and absolute quantification of HEV RNA in food samples with the advantage of presenting the co-amplification of internal process control.

Recent Advances in Biosensor Development for the Detection of Viral Particles in Foods: A Comprehensive Review.

Viral foodborne diseases cause serious harm to human health and the economy. Rapid, accurate, and convenient approaches for detecting foodborne viruses are crucial for preventing diseases. Biosensors integrating electrochemical and optical properties of nanomaterials have emerged as effective tools for the detection of viruses in foods. However, they still face several challenges, including substantial sample preparation and relatively poor sensitivity due to complex food matrices, which limit their field applications. Hence, the purpose of this review is to provide an overview of recent advances in biosensing techniques, including electrochemical, SERS-based, and colorimetric biosensors, for detecting viral particles in food samples, with emerging techniques for extraction/concentration of virus particles from food samples. Moreover, the principle, design, and advantages/disadvantages of each biosensing method are comprehensively described. This review covers the recent development of rapid and sensitive biosensors that can be used as new standards for monitoring food safety and food quality in the food industry.

Short Communication — Stability of “Free” Norovirus RNA on Fresh Produce

AbstractThe accuracy and dependability of results generated through molecular detection of specific target sequences of RNA, commonly used for detecting viruses in food, have been extensively debated within the scientific community. Such concerns have been raised by researchers, clients, and regulators alike, highlighting the need for further investigation and clarification. In particular, there has been debate about the possibility of molecular methods to detect enteric viruses, such as norovirus, in foods, producing false positive RT-PCR results due to the presence of “free” target RNA sequences in the food supply chain environment. This study aimed to investigate this issue by evaluating the recovery of “free” norovirus RNA from lettuce leaves. The “free” RNA was produced using heat treatment and chemical extraction methods. The study findings indicate that recovery of heat-extracted RNA from the lettuce leaves decreased markedly within 24 h, while recovery of chemically extracted RNA remained stable and even increased over time. The results of this study suggest that positive molecular detection of viruses in food is more likely to be associated with intact and potentially infectious virus particles, rather than “free” unprotected RNA. These findings have significant implications for the food industry and regulatory bodies in terms of the interpretation, accuracy and reliability of molecular detection methods for virus detection in food.

Development of enhanced primer sets for detection of Norovirus and Hepatitis A in food samples from Guayaquil (Ecuador) by reverse transcriptase-heminested PCR

Norovirus (NV) is an infectious biological agent that causes gastrointestinal problems of the original nonbacterial appearance of foodborne illnesses. The genotype of NV responsible for the most frequent NV disease outbreaks is GII, accounting for 60–80% of the cases. Moreover, original and new NV variants are continuously emerging, concurrent with the recent global increase in NV infections. Hepatitis A virus (HAV) is another foodborne pathogen frequently implicated in acute gastroenteritis cases around the world. The virus is transmitted among humans via the fecal-oral route, and infection by HAV causes the most severe form of viral illness acquired from foods. In this study, we implemented primer sets to detect NV genotypes I and II. We also developed primer sets for the detection of HAV. The primers were used in a heminested reverse transcriptase PCR (hnRT-PCR) protocol that was rapid and sensitive for detecting NVG1, NVGII and HAV virus in food. The hnRT-PCR was applied successfully to strawberries and spinach obtained from a local fresh-food market, where we could see NVGI, NVGII and HAV. Keywords: Norovirus1; Hepatitis A2; gastroenteritis3; genotypes4; NVG15; NVGII6; hnRT-PCR7.

Development of enhanced primer sets for detection of Norovirus and Hepatitis A in food samples from Guayaquil (Ecuador) by reverse transcriptase-heminested PCR

Norovirus (NV) is an infectious biological agent that causes gastrointestinal problems of the original nonbacterial appearance of foodborne illnesses. The genotype of NV responsible for the most frequent NV disease outbreaks is GII, accounting for 60–80% of the cases. Moreover, original and new NV variants are continuously emerging, concurrent with the recent global increase in NV infections. Hepatitis A virus (HAV) is another foodborne pathogen frequently implicated in acute gastroenteritis cases around the world. The virus is transmitted among humans via the fecal-oral route, and infection by HAV causes the most severe form of viral illness acquired from foods. In this study, we implemented primer sets to detect NV genotypes I and II. We also developed primer sets for the detection of HAV. The primers were used in a heminested reverse transcriptase PCR (hnRT-PCR) protocol that was rapid and sensitive for detecting NVG1, NVGII and HAV virus in food. The hnRT-PCR was applied successfully to strawberries and spinach obtained from a local fresh-food market, where we could see NVGI, NVGII and HAV. Keywords: Norovirus1; Hepatitis A2; gastroenteritis3; genotypes4; NVG15; NVGII6; hnRT-PCR7.

Inactivation of foodborne viruses by the cold plasma technology

Cold plasma (CP) is an innovative non-thermal food processing method. CP refers to a partially or completely ionized gas containing reactive chemical species, which are active against microorganisms, including viruses or enzymes of foods. CP has a minimal effect on the quality attributes of foods and can also elongate the shelf life of foods. Foodborne outbreaks caused by viruses have been increased in various countries in recent years. The research works on the inactivation effect of CP against viruses including foodborne viruses have been also increased in recent years. The most important foodborne viruses are human norovirus (HuNoV) and hepatitis A virus (HAV), involved in several outbreaks worldwide. Human astrovirus (HAstV), human adenovirus (HuAdV), Aichi virus (AiV), sapovirus (SaV) and enterovirus (EV) are also notable foodborne viruses and were associated in sporadic cases. The CP treatment proved efficient for the inactivation of foodborne viruses such as HuNoV and HAV. The present work reviews the CP as a non-thermal food processing technology and present the published data on the effect of CP process on foodborne viruses in foods.

Los alimentos como transmisores de virus: Una revisión

Viruses have been present throughout human history, causing diseases due to infections and food poisoning; they have caused frequent public health problems worldwide. These illnesses are usually mild, moderate, or severe in nature. The personal hygiene of food handlers and processing processes should be checked periodically. Virus detection protocols and safety measures should be continually reviewed as viruses change their mode of infection. The objective of this review was to discuss the possible routes of virus transmission to humans through food. Important topics have been reviewed such as: definition of food viruses, presence, and types of viruses in food, enteric viruses, zoonotic viruses, water as a means of transmission, risks of infection, other non-conventional foods as potential transmitters of viruses and food safety, in addition to current and future challenges, research work on viruses more resistant to heat treatments in food should be sought. Also, future work on survival time of active viruses on food surfaces. In addition, studies that determine the mechanisms of virus mutation in relation to the conditions of food handling and processing.

Enteric viruses in food safety: New threats for an old problem.

Foodborne diseases are one of the most serious concerns in public health. It is estimated that around 600 million cases of gastroenteritis occur worldwide each year. At present, more than 200 food-borne diseases are known, which can cause from mild gastroenteritis to syndromes with a fatal outcome, with the added possibility of chronic complications. One of the major etiological agents in foodborne diseases are the food and waterborne viruses, which are attracting a great deal of attention to researchers, food hygienists and policy makers. Several aspects differentiate these pathogens from foodborne pathogenic bacteria: their high capacity for infection and preservation in food environments, and their difficulty for a correct and sensitive detection. In recent years, different initiatives have been carried out to prioritize research in the area of viruses in food, prioritizing different aspects of their detection, epidemiology and control. There is clear evidence that the existing data on their prevalence may be underestimated due to the lack of robust methods for their sensitive detection. It is also necessary to know exactly what the incidence is in the different stages of the food production chain, and particularly in that which is dedicated to the transformation of products of animal origin. Finally, it is also necessary to calibrate the current disinfection procedures in the food industry in order to reliably establish a quantitative evaluation of the viral risk in food.

Monitoring of SARS-CoV 2 on primary production and manually processed foods in Northern Italy

Abstract Background SARS CoV2 pandemic has generated interest in research of virus spread sources, apart from human contagion. In particular different studies are performed to understand surfaces involvement; during the epidemic event, different types of studies on surfaces have been performed. However, there is little information regarding the presence of the virus in food; wastewater contamination and poor hygiene practices could be the sources of viral contamination in handled food products. The aim of this work was to analyze different types of food for Sars CoV 2 presence. Methods 390 samples of molluscs, 353 vegetables, 10 meat, 72 water, 169 berries, 16 seafood products and ready-to-eat dishes were analysed (2020-2021). Samples were selected from those provided for routine analysis during the pandemic period. The preparation was done based on ISO 15216-2 indications, followed by extraction with Nuclisense®Minimag® system; real-time PCR was applied to detect the Sars CoV2 virus Open Reading Frame 1ab, with two different master mixes. Results All samples tested were negative for ORF1ab detection. Many hypothesis can be made about negative Results: it could be possible that contamination levels in water, or on surfaces in contact with food, are too low to generate a detection signal; again, handling practices may be sufficient to avoid the transmission of the virus to food. Otherwise, the selected genetic target and the Real-time PCR performed, could have been not so efficient in detecting virus as expected. Conclusions In this preliminary work, food does not seem to be a substrate for Sars CoV2 contamination; other food categories or transformation/production processes could be investigated to confirm the obtained results. Also, different targets and amplification systems would have to be evaluated to verify the rule of foodstuffs as Sars CoV2 vehicles. Key messages Primary production and manual processed food don't seem to be at risk for SARS CoV2 contamination. Food has not been identified as a source of Sars CoV2 spreading.

Impact of COVID-19 Pandemic Crisis and Food Safety System: A Literature Review

The new outbreak as unknown pneumonia that occurred in Wuhan province of China in December 2019, is a new coronavirus from Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), and has been termed Coronavirus Disease 2019 (COVID-19). Coronaviruses are a family of viruses that usually cause respiratory illness. Since food plays an essential role in human health as an integral part of human life, food safety is critical in such cases. It is essential to adopt practical strategies in controlling the COVID-19 crisis. Also, extreme economic consequences and threats to human health were imposed in the COVID-19 pandemic in 2019-2021. Some studies have been published by World Health Organization (WHO), Environmental Protection Agency (EPA), and Food and Drug Administration (FDA); however, there is little information about food safety and COVID-19. Although COVID-19 transmission routes through food are not currently known, contamination through contaminated food and environmental surfaces cannot be ignored, especially in manufacturing companies, restaurants, and communities that are unsanitary. Nevertheless, World Health Organization precautions on good hygiene when preparing food, and recommends effective management of food safety during COVID-19. In food safety, COVID-19 is known as a potential risk through food especially in restaurants and industrial areas. There have been very few studies on the relationship between food safety and COVID- 19. Studies have shown that COVID-19 can survive longer than MERS-CoV in food stored at 4 °C. In the current situation, several methods are used to disinfect and control the spread of COVID-19 disease, some of which are not effective and can also have problems and limitations. Heat treatment, pasteurization, UV light-based, and chemical disinfectants can be ways to prevent COVID-19 probable transmission. The temperatures (30 or 40 °C) reduced the survival time of HCoVs on the inanimate surfaces. The cooking processes (70 °C) are effective in inactivating the virus in food. SARS-CoVs were inactivated by exposure to ultraviolet (254 nm) for 1 to 6 min with an increase of up to 400-fold. Also, usage of disinfectants can be effective for inanimate surfaces. This review summarizes the available data related to some topics and methods to inactivate COVID-19 and the role and importance of the food industry and food supply chain during the pandemic.

Inactivation of viruses using nonthermal plasma in viral suspensions and foodstuff: A short review of recent studies

AbstractThis review tries to introduce the novel and promising technology of the nonthermal plasma against different viruses and virus surrogates, which can trigger food infections and gastroenteritis. It has been reported that the noroviruses are responsible for 21 million cases of gastrointestinal illness annually just in the United States. The article is divided into the introduction for nonthermal plasma, its antiviral mechanism, and the most effective parameters on its antiviral functionality. This technology offers many advantages, including simple in designing, leaving no toxic residues, fast processing, and no thermal degradation of samples. According to the accomplished studies, the exposure distance, processing time, and generated amount of reactive species, particularly singlet oxygen are the main parameters affecting the performance of nonthermal plasma against food viruses. Especially, the capsid and genome of viruses seem to be under influence of nonthermal plasma. Deformation of the capsid envelope through oxidation and damage to the sugar phosphate backbone of viral RNA and DNA are considered the most important factors of reactive species generated by nonthermal plasma on viruses.

MOL-PCR and xMAP Technology: A Multiplex System for Fast Detection of Food- and Waterborne Viruses

Viruses are common causes of food- and waterborne diseases worldwide. Conventional identification of these agents is based on cultivation, antigen detection, electron microscopy, or real-time PCR. Because recent technological advancements in detection methods are focused on fast and robust analysis, a rapid multiplexing technology, which can detect a broad spectrum of pathogenic viruses connected to food or water contamination, was utilized. A new semiquantitative magnetic bead-based multiplex system has been designed for simultaneous detection of several targets in one reaction. The system includes adenoviruses 40/41 (AdV), rotavirus A (RVA), norovirus (NoV), hepatitis E virus (HEV), hepatitis A virus (HAV), and a target for external control of the system. To evaluate the detection system, interlaboratory ring tests were performed in four independent laboratories. Analytical specificity of the tool was tested on a cohort of pathogenic agents and biological samples with quantitative PCR as a reference method. Limit of detection (analytical sensitivity) of 5×100 (AdV, HEV, and RVA) and 5×101 (HAV and NoV) genome equivalents per reaction was reached. This robust, senstivie, and rapid multiplexing technology may be used to routinely monitor and manage viruses in food and water to prevent food and waterborne diseases.

Respiratory viruses in foods and their potential transmission through the diet: A review of the literature

Respiratory viruses are the main agents causing respiratory tract diseases. Nowadays, coronaviruses – and specifically, SARS-CoV-1, MERS-CoV and SARS-CoV-2 – are the principal responsible for the major epidemic outbreaks of the 21st century. The major routes of transmission for respiratory viruses – including coronaviruses – are via direct and indirect contacts. However, transmission through contaminated foods has not been extensively assessed. The present paper was aimed at reviewing scientific data on the transmission of respiratory viruses through potentially contaminated foods. While the current data seem to suggest that this route of transmission is not likely to occur, in order to increase the knowledge on this issue further investigations are still clearly necessary for a more complete prevention of the risks. Studies should include fresh produce and cooked foods. Anyway, prevention measures and good hygienic practices for both consumers and workers are mandatory when handling and cooking foods.

Inactivation of Foodborne Viruses by High-Pressure Processing (HPP).

High-pressure processing (HPP) is an innovative non-thermal food preservation method. HPP can inactivate microorganisms, including viruses, with minimal influence on the physicochemical and sensory properties of foods. The most significant foodborne viruses are human norovirus (HuNoV), hepatitis A virus (HAV), human rotavirus (HRV), hepatitis E virus (HEV), human astrovirus (HAstV), human adenovirus (HuAdV), Aichi virus (AiV), sapovirus (SaV), and enterovirus (EV), which have also been implicated in foodborne outbreaks in various countries. The HPP inactivation of foodborne viruses in foods depends on high-pressure processing parameters (pressure, temperature, and duration time) or non-processing parameters such as virus type, food matrix, water activity (aw), and the pH of foods. HPP was found to be effective for the inactivation of foodborne viruses such as HuNoV, HAV, HAstV, and HuAdV in foods. HPP treatments have been found to be effective at eliminating foodborne viruses in high-risk foods such as shellfish and vegetables. The present work reviews the published data on the effect of HPP processing on foodborne viruses in laboratory media and foods.

Foodborne Viruses and Innovative Non-Thermal Food-Processing Technologies.

In recent years, several foodborne viruses’ outbreaks have been recorded worldwide. Μost of the foodborne viruses have a low infection dose, are stable and can persist and survive in foods for a long time without loss of infectivity. The most important foodborne viruses are: human norovirus (HuNoV), human rotavirus (HRV), hepatitis A virus (HAV), hepatitis E virus (HEV), human astrovirus (HAstV), Aichi virus (AiV), sapovirus (SaV), human adenovirus (HAdV) and enterovirus (EV). In recent years, innovative non-thermal food-processing technologies including high-pressure processing (HPP), cold plasma (CP), ultraviolet light (UV), irradiation and pulsed electric field (PEF) for improving the quality and safety of foods, including foods of animal origin, have been under research. This review presents the recent data on foodborne viruses and reviews the innovative non-thermal technologies for the control of the foodborne viruses in foods.

Detection of Human polyomavirus 2 (HPyV2) in oyster samples in northern Brazil

BackgroundHuman polyomavirus 2 (HPyV2 or JCPyV) is persistent in the environment due to its excretion in urine and feces; it is detected in samples of wastewater, surface water and drinking water. A lack of basic sanitation and sewage collection results in the presence of this virus in food, especially in oysters, since they are bioaccumulators and are consumed in their natural form, thus posing a risk to human health.MethodsThis study investigated the frequency of HPyV2 in samples of oysters marketed in northeastern Pará State, Brazil, and optimized a real-time PCR (qPCR) protocol for the detection of an endogenous oyster control. A total of 217 oysters in 22 pools from five municipalities in the state of Pará were analyzed. Samples underwent dissection and total maceration of oyster tissue using a viral concentration technique, followed by DNA extraction with phenol-chloroform and amplification of the VP1 region for molecular detection via qPCR.ResultsHPyV2 was detected in 18.2% (4/22) of the pooled samples, with frequencies of 25, 20, 20 and 16% in the municipalities of Salinópolis, Augusto Corrêa, São Caetano de Odivelas and Curuçá, respectively. Notably, the sample pool from the municipality of Bragança did not have detectable HPyV2 and this was the only sampled location with a water treatment station. In this study, Crassostrea genus-specific primers (AFL52 ribosomal RNA gene) of oyster were developed for use as an endogenous control in the qPCR analysis, which will be useful for future studies.ConclusionsThe detection of HPyV2 in oyster samples commercialized in the state of Pará shows the circulation of this virus in the studied municipalities. Thus, it is necessary to implement measures for improving sewage collection and basic sanitation to avoid contamination of water and food with HPyV2.

Metagenomic to evaluate norovirus genomic diversity in oysters: Impact on hexamer selection and targeted capture-based enrichment

Human virus transmission through food consumption has been identified since many years and the international trade increases the risk of dissemination of viral pathogens. The development of metagenomic approach holds many promises for the surveillance of viruses in food and water. This work aimed to analyze norovirus diversity and to evaluate strain-dependent accumulation patterns in three oyster types by using a metagenomic approach. Different hexamer sets to prime cDNA were evaluated before capture-based approach to enhance virus reads recovery during deep sequencing. The study includes the use of technical replicates of artificially contaminated oysters and the analysis of multiple negatives controls. Results showed a clear impact of the hexamer set used for cDNA synthesis. A set of In-house designed (I-HD) hexamers, selected to lower mollusk amplification, gave promising results in terms of viral reads abundancy. However, the best correlation between CT values, thus concentrations, and number of reads was observed using random hexamers. Random hexamers also provided the highest numbers of reads and allowed the identification of sequence of different human enteric viruses. Regarding human norovirus, different genogroups and genotypes were identified among contigs longer than 500 bp. Two full genomes and six sequences longer than 3600 bases were obtained allowing a precise strain identification. The use of technical triplicates was found valuable to increase the chances to sequence viral strains present at low concentrations. Analyzing viral contamination in shellfish samples is quite challenging, however this work demonstrates that the recovery of full genome or long contigs, allowing clear identification of viral strains is possible.

Foodborne toxinfections caused by virus: characteristics of the main viruses, prevention, treatment and clinical method of laboratory diagnosis by RT-qPCR.

Foodborne viruses were recognized among the top-rated food safety priorities and have become a greater concern to the food industry over the past few years. Food safety experts agreed that control measures for viruses throughout the food chain are required. This manuscript provides a description of foodborne viruses (Rotavirus, Adenovirus, Norovirus, Astrovirus, Hepatitis A, Hepatitis E, and Poliovirus) and their characteristics, and technologies developed for viral detection and control. A bibliographical research was carried out to collect data and information on viral diseases of the last nineteen years. The research sites accessed were: Books, theses and the database of Scielo, Google Scholar, Medline, Pubmed, Science Direct, CAPES periodical portal and virtual health library. This study was carried to clarify to the health professionals the importance of the need for a greater notification and update of these viral food toxinfections, in order to reduce the cases number of these diseases. Effective prevention and biological technologies to ensure control of viruses in the food chain can significantly reduce foodborne toxinfections caused by virus. Key words: Enteric viruses, virus in food, quantitative reverse transcription polymerase chain reaction (RT-qPCR) in virus detection.

Human Adenovirus, Mesophilic Bacteria and Fungi in Puppies’ Food Marketed in Bulk in Southern Brazil

Background: The Brazilian domestic canine population are the second largest in the world and their feeding means 0.4% of the Brazilian gross domestic product. For maintaining the quality of the food, the companies use worldwide standards for technical prevention and control of contaminants and biological conservation. The packaging is part of this process, since it provides a barrier between food and environment. However, in Brazil, packagings are often opened in retail stores for bulk marketing. The objectives of this work were to develop a methodology to detect viruses in foods and to analyze the bacterial and fungal contamination in puppies’ food sold in bulk in Ivoti and Estância Velha, cities in Southern Brazil. Materials & Methods & Results: Twenty samples collected between September and October 2016 were analyzed for most probable number of coliforms, Salmonella sp., mesophilic aerobic bacteria and yeast/mold following the regulation of the Brazilian Ministry of Agriculture, Livestock and Food Supply guidelines. They were also tested for Human Mastadenovirus C (HAdV), Canine Mastadenovirus A (CAdV), and Carnivore Protoparvovirus 1 (CPV) genomes. Viral analysis were performed by polymerase chain reaction (PCR) detection. During the collection of the samples hygienic-sanitary conditions, storage of feeds, animals’ access, dog grooming, and veterinary care were considered to evaluate the conditions of each store. A pilot study was carried out using one food sample marketed in bulk and one sample from the original package (closed package) and testing them for bacterial and fungal contamination for standardizing viral detection. Ten grams of food from the original package were mixed with 90 mL of Eagles’ Minimal Essential Medium (E-MEM) in 100 mL sterile bottles. These bottles were kept in room temperature and shaken for 60 min. Subsequently, aliquots were obtained by sequentially diluting the sample (10-2 to 10-4). All final specimens contained 10 mL and each diluted sample was spiked with HAdV-C prototype viral strain (AdV5). A standard solution of HAdV-C was diluted from 3.6x106TCID50mL, (50% tissue culture infective dose) to 3.6x103 TCID50mL, and DNA extraction was performed. Nested-PCR targeting AdV DNApol was performed to detect adenoviruses from different hosts. AdV-positive samples were submitted to nucleotide sequencing and phylogenetic analysis. Specific PCRs were also carried out for CAdV and CPV. Mesophilic aerobic bacteria were detected in all samples and Aspergillus sp. was found in five samples, among which one sample was co-infected with Penicillium sp. One sample was positive for AdV, which was identified as HAdV by sequencing; while coliforms and Salmonella sp. were not detected.Discussion: The presence of fungi with mycotoxigenic potential, such as Aspergillus sp. and Penicillium sp. represents a threat for canines, due to toxins that may persist for a long period even after the fungus is not viable. Moreover, dogs seem to be more susceptible to the effects of the toxins, which is probably because of low glutathione s-transferase activity. Some species of Penicillium sp. genus may produce ocrathoxin A, which nephrotoxic and immunosuppressive effects in dogs are widely reported. Mesophilic bacteria were detected in all samples (at 104 CFU/g) and considered harmless. The detection of human viruses points to the presence of anthropic contamination; on the other hand, ingestion of contaminated feed, even if it is by a heterologous species, turns the dogs into carriers of the virus. In addition, manipulation of those feeds by children who share the same space with dogs can result in gastroenteritis episodes.