Evaluation of Microbial Contamination in Bivalve Mollusks: Epidemiology and Diagnosis

Shellfish are filter-feeding organisms that can accumulate many bacteria and viruses. Considering that depuration procedures are not effective in removal of certain microorganisms, shellfish-borne diseases are frequent in many parts of the world, and their control must rely primarily on investigation of prevalence of human pathogens in shellfish and water environment. However, the diffusion of enteric viruses and Vibrio bacteria is not known in many geographical areas, for example in Sardinia, Italy. A survey aimed at investigating the prevalence of Norovirus (NoV), hepatitis A virus (HAV), V. parahaemolyticus, V. cholerae and V. vulnificus was carried out, analyzing both local and imported purified, non-purified and retail shellfish from North Italy and Sardinia. Shellfish from both areas were found contaminated by NoVs, HAV and Vibrio, including retail and purified animals. Molecular analysis evidenced different NoV genogroups and genotypes, including bovine NoVs, as well as pathogenic Vibrio strains, underlining the risk for shellfish consumers. However, also other approaches are needed to control the diffusion of shellfish-borne diseases. It was originally thought that enteric viruses are passively accumulated by shellfish. Recently, it was proven that NoVs bind to specific carbohydrate ligands in oysters, and various NoV strains are characterized by a different bioaccumulation pattern. To deepen the knowledge on this argument, a study was carried out, analyzing bioaccumulation of up to 8 different NoV strains in four different species of shellfish. Different bioaccumulation patterns were observed for each shellfish species and NoV strain used, potentially important in setting up effective shellfish purification protocols. Finally, a novel study of evaluation of viral contamination in shellfish from the French Atlantic coast was carried out following the passage of Xynthia tempest over Western Europe which caused massive destruction. Different enteric viruses were found over a one month period, evidencing the potential of these events of contaminating shellfish.

Epidemiology and population genetics of Podosphaera fusca and Golovinomyces orontii, causal agents of cucurbit powdery mildew

In 2010, 2011 and 2012 growing seasons, the occurrence of the ascomycetes Podosphaera fusca and Golovinomyces orontii, causal agents of powdery mildew disease, was monitored on cultivated cucurbits located in Bologna and Mantua provinces to determine the epidemiology of the species. To identify the pathogens, both morphological and molecular identifications were performed on infected leaf samples and a Multiplex-PCR was performed to identify the mating type genes of P. fusca isolates. The investigations indicated a temporal succession of the two species with the earlier infections caused by G. orontii, that seems to be the predominant species till the middle of July when it progressively disappears and P. fusca becomes the main species infecting cucurbits till the end of October. The temporal variation is likely due to the different overwintering strategies of the two species instead of climatic conditions. Only chasmothecia of P. fusca were recorded and mating type alleles ratio tended to be 1:1. Considering that only chasmothecia of P. fusca were found, molecular-genetic analysis were carried out to find some evidence of recombination within this species by MLST and AFLP methods. Surprisingly, no variations were observed within isolates for the 8 MLST markers used. According to this result, AFLP analysis showed a high similarity within isolates, with SM similarity coefficient ranging between 0.91-1.00 and also, sequencing of 12 polymorphic bands revealed identity to some gene involved in mutation and selection. The results suggest that populations of P. fusca are likely to be a clonal population with some differences among isolates probably due to agricultural practices such as fungicides treatments and cultivated hosts. Therefore, asexual reproduction, producing a lot of fungal biomass that can be easily transported by wind, is the most common and useful way to the spread and colonization of the pathogen.