Interactions between Human Norovirus Surrogates and Acanthamoeba spp

Abstract

Human noroviruses (HuNoVs) are the most common cause of food-borne disease outbreaks, as well as virus-related waterborne disease outbreaks in the United States. Here, we hypothesize that common free-living amoebae (FLA)-ubiquitous in the environment, known to interact with pathogens, and frequently isolated from water and fresh produce-could potentially act as reservoirs of HuNoV and facilitate the environmental transmission of HuNoVs. To investigate FLA as reservoirs for HuNoV, the interactions between two Acanthamoeba species, A. castellanii and A. polyphaga, as well as two HuNoV surrogates, murine norovirus type 1 (MNV-1) and feline calicivirus (FCV), were evaluated. The results showed that after 1 h of amoeba-virus incubation at 25°C, 490 and 337 PFU of MNV-1/ml were recovered from A. castellanii and A. polyphaga, respectively, while only few or no FCVs were detected. In addition, prolonged interaction of MNV-1 with amoebae was investigated for a period of 8 days, and MNV-1 was demonstrated to remain stable at around 200 PFU/ml from day 2 to day 8 after virus inoculation in A. castellanii. Moreover, after a complete amoeba life cycle (i.e., encystment and excystment), infectious viruses could still be detected. To determine the location of virus associated with amoebae, immunofluorescence experiments were performed and showed MNV-1 transitioning from the amoeba surface to inside the amoeba over a 24-h period. These results are significant to the understanding of how HuNoVs may interact with other microorganisms in the environment in order to aid in its persistence and survival, as well as potential transmission in water and to vulnerable food products such as fresh produce.N oroviruses belong to a highly genetically and antigenically diverse genus Norovirus of the Caliciviridae family. Noroviruses are divided into five genogroups (GI, GII, GIII, GIV, and GV) according to the genetic similarity of highly conserved areas of their genomes, such as the RNA-dependent RNA polymerase and the VP1 protein (1), with GI, GII, and GIV associated with human diseases. They are important enteric pathogens responsible for at least 50% of all gastroenteritis outbreaks worldwide (2) and, partially due to their high infectivity and low infectious dose (18 to 1,000 virus particles) (3), human noroviruses (HuNoVs) are the most common cause of food-borne disease outbreaks in the United States (4).Transmission of HuNoV occurs primarily through exposure to contaminated food or water, person-to-person contact, aerosolized particles from vomitus, and contact with contaminated fomites (e.g., door handles, toilets, tables, and elevator buttons) (5). Numerous waterborne disease outbreaks have been linked to HuNoV (6-9), and HuNoVs are often detected in surface water (10), wastewater (11), and even finished drinking water (12). With respect to food-borne disease outbreaks, the most common food commodity implicated in HuNoV outbreaks is fresh produce, specifically leafy greens (13); however, the route of transmission and point of contami...

Transfer ofAcanthamoebaspp. to fresh produce from water and environmental surfaces

“…Based on previous studies by Hsueh and Gibson (2015), after virus inoculation, the number of MNV-1 associated with 6Á5 log 10 Acanthamoeba was approx. 2Á5 log 10 following a 1-h incubation at room temperature.…”

“…2Á5 log 10 following a 1-h incubation at room temperature. In addition, the virus inoculation did not affect morphology under microscopy nor the normal growth of either Acanthamoeba species (Hsueh and Gibson 2015).…”

“…Moreover, a recent study conducted by the current manuscript authors shows that that Acanthamoeba associates with infectious murine norovirus type 1(MNV-1)-one of the surrogates for the study of human noroviruses (HuNoV)-for a prolonged period of time (i.e. at least 8 days) as well as through a complete life cycle and can even become internalized by Acanthamoeba (Hsueh and Gibson 2015). This is of particular importance as HuNoV are the primary cause of food-borne illnesses in the United States as indicated below.…”

Survival of Viruses in Water

“…Likewise, amoebas have been shown to carry viruses such as adenovirus, mimivirus [47], murine norovirus, and others [48]. Although the viral population will decrease by a small amount once internalized, viruses are able to survive inside amoebas, even as cysts, for several days [48].…”

АКАНТАМЕБИ ЯК РЕЗЕРВУАР ПАТОГЕННИХ БАКТЕРІЙ ТА ВІРУСІВ (Огляд Літератури)

“…У результаті проведеного дослідження було встановлено, що у вірусу MNV-1 процес переходу від зовнішньої поверхні амеби до її внутрішньої частини завершується вже через 24 години, тоді як FCL не виділявся вже через годину. Крім того, було досліджено тривалу взаємодію вірусу (протягом 8 діб) MNV-1 і показано, що вірус виживає після проходження амебою повного життєвого циклу (інцистування та ексцистування) [37]. Отже, вільноживучі амеби можуть сприяти підвищенню виживання норовірусів, як і ряду інших мікробних чинників.…”