Microbiological quality and safety of fresh cultivated and wild mushrooms commercialized in Spain

Abstract

402 samples of 22 species of cultivated and wild fresh mushrooms sold in retail markets and supermarkets in Zaragoza (Spain) were studied to quantify their microbial load (mesophilic aerobic microorganisms, Pseudomonas genus, Enterobacteriaceae, lactic acid bacteria, total and thermotolerant coliform bacteria, Escherichia coli, yeasts and moulds) and to investigate the presence of E. coli O157:H7, Listeria monocytogenes, Salmonella spp., Staphylococcus aureus and Yersinia enterocolitica. The total microbial counts ranged from 4.4 to 9.4 log cfu/g, the genus Pseudomonas being the most prevalent with counts from 3.7 to 9.3 log cfu/g and Auricularia auricula-judae the species with the highest microbial load (9.4 log cfu/g). No significant differences (p > 0.05) were detected between mean counts of wild and cultivated species in all the microbial groups studied. The microbiological safety level of the cultivated mushrooms was excellent since no pathogens were isolated, and the microbial counts of indicator microorganisms were low, being detected in only half of the species. Salmonella spp, E. coli O157:H7 and S. aureus were not isolated from any sample, Y. enterocolitica was detected in only four samples of wild mushrooms whereas twenty-six (6.5%) were positive for L. monocytogenes, their occurrence being relatively high in Calocybe gambosa (40%), Hygrophorus limacinus (40%) and Tuber indicum (100%). These results suggest that a strategy to reduce bacterial populations, and to improve the microbiological safety of some species of fresh mushrooms, should be investigated.

Introduction

In recent years there has been an increase in the worldwide consumption of fresh mushrooms. According to the Food and Agriculture Organization (2010) data, the world production of mushrooms and truffles in 2008 exceeded 3.4 million t. In Spain, according to data compiled by the Ministry of the Environment and Rural and Marine Affairs (2009), the production of mushrooms doubled in the period 1998–2008, reaching 120,000 t. The greater demand for fresh mushrooms has led to an increase in the quantity and variety of produce available to the consumer in the Spanish market. Traditionally used in cuisine as a vegetable, they are appreciated not only for their organoleptic characteristics but also for their nutritional value (Manzi et al., 2001, Díez and Alvarez, 2001) and functional properties (Wasser and Weis, 1999, Zhang et al., 2001). Historically, the fresh mushroom market has been dominated by the species Agaricus bisporus (white button mushroom), widely cultivated in Spain. However, during recent years the production of other edible mushrooms such as Pleurotus ostreatus (oyster mushroom), Lentinula edodes (shiitake) and A. bisporus var. Portobello (brown button mushroom) has increased considerably. Also, various species of wild mushrooms (e.g. Lactarius deliciosus, Boletus edulis, Cantharellus spp., Hygrophorus spp., and Tricholoma spp.) are seasonally available in their fresh form, and species of fresh exotic cultivated mushrooms not grown in Spain (such as Flammulina velutipes and Auricularia auricula-judae) or fresh wild mushroom (during off-season periods) are imported from China, France, Poland or South Africa to supply the increasing demand. Thus, a wide range of edible fresh mushrooms are now available all the year round from retail markets and supermarkets in Spain.

Fresh mushrooms are an ideal medium for microbial growth because they have a high moisture content, a water activity of 0.98 or higher and a neutral pH (Martínez-Carrera et al., 1998). Physiologically they are classified as high respiring products (Kader, 2002) so the loss of water and the consequent wilting occur very fast. These characteristics coupled with their high initial microbial load limit their post-harvest shelf life to a few days. Mushrooms destined for the fresh market are hand-harvested, transported to a processing facility where they are packed into retail containers always without being washed, and commercialized in many cases at ambient temperature. However, in contrast to other fresh products, information on the microbiological profile of fresh mushrooms is very limited. What information there is focuses on cultivated A. bisporus (Doores et al., 1987, González-Fandos et al., 2001).Mushroom quality is defined by a combination of parameters, including whiteness, texture and microbial counts (Gormley, 1975). Beelman et al. (1989) determined that the presence of high bacterial populations in fresh mushrooms is a major factor that significantly diminishes quality by causing a brown, blotchy appearance and that the rate of post-harvest deterioration of fresh mushrooms has been directly related to the initial microbial load. Doores et al. (1987) found that bacterial populations during post-harvest storage at 13 °C increased from an initial load of 7 log cfu/g to almost 11 log cfu/g over a 10-day storage period. These authors also reported that the deterioration of mushroom quality, as indicated by maturity and colour measurement, appeared to be concomitant with increase in bacterial numbers. The genus Pseudomonas, the most prevalent in mushrooms, has been associated with brown stains when its counts exceed 6 log cfu/cm² (Wong et al., 1982, Soler-Rivas et al., 1999). Just as for many years research has been carried out into the potential of many treatments to reduce the microbial populations in fruit and vegetables, whether whole or peeled and cut, so in recent years studies have also appeared in the literature evaluating such treatments on mushrooms. Sapers et al. (2001) investigated how the treatment of mushrooms with hydrogen peroxide and a sodium erythorbarte-based browning inhibitor made the product less subject to brown blotching. Recently, Simón et al. (2010) determined that the reduction of pseudomonas by 2.5 logarithmic units in A. bisporus washed with citric acid before being sliced avoided bacterial blotch caused by Pseudomonas species during 17 days of storage at 5 °C. In A. bisporus, Gautam et al. (1998) reported that a dose of gamma radiation of 2 kGy was necessary to reduce microbial growth and to extend the shelf life from 2 to 10 days. In recent studies, Rivera et al. showed that for Tuber aestivum and Tuber melanosporum a decontaminating strategy with ethanol and ultrasound (2011a) or with ionizing radiation (Rivera et al., 2011b, Rivera et al., 2011c) significantly extended the commercial life of fresh truffles as a result of the reduction in Pseudomonas populations and the elimination of moulds.

Fresh mushrooms have also the potential to carry food-borne bacteria derived from different sources. In the particular case of cultivated mushrooms, contamination with pathogenic bacteria may take place during all growth, harvest and sorting stages, the substrate where they grow (casing soil) being an important source of contamination. Chikthimmah (2006) demonstrated that Listeria monocytogenes and Salmonella spp. died rapidly in sterile treated soil but grow in sterile soil. In this case the thermal pasteurization eliminate the natural microflora that supress the pathogen growth. The potential of wild mushrooms to become contaminated with food-borne bacterial pathogens is relatively greater than that of cultivated varieties. Wild mushrooms are hand-harvested from forest and woodlands ecosystems which are silvopastoral systems on which livestock (bovine, ovine, or equine) and wild animals (wild boar, roe deer, red deer, etc.) usually graze. These ecosystems also support a great diversity of invertebrate and vertebrate organisms such as insects, molluscs (snails), birds, reptiles, amphibians and small mammals (principally rodents), some of which are mycophagous or fungivorous. Consequently, the contamination of wild mushrooms with food-borne bacteria can occur directly or indirectly via animals or insects.

However, food-borne illness outbreaks attributed to fresh mushrooms contaminated with pathogenic bacteria have not been reported (Strapp et al., 2003). Although they are considered a safe product, occurrences of pathogenic bacteria in commercial fresh mushroom samples have been reported and some studies to determine their ability to grow and survive during commercialization have also been carried out. Low prevalence of Campylobacter jejuni, 1.5% and 0.9%, was detected in fresh mushrooms by Doyle and Schoeni (1986) and by Whyte et al. (2004), respectively. Staphylococcus spp. was isolated from 22.4% of the samples of mushrooms analyzed by Johannessen et al. (2002), but only four isolates were identified as Staphylococcus aureus. These researchers also isolated L. monocytogenes from one sample, Samadpour et al. (1999) detected this bacterium in 5% of the mushroom samples and Rivera et al. (2010) in 10% of the fresh ascocarps of T. aestivum analyzed. In 2001, a health alert was issued in Ireland by government authorities after Salmonella Kedougou was detected on mushrooms (Food Safety Authority of Ireland, 2001). The pathogen was found in the mushroom compost, the mushroom casing soil and in several mushroom samples. It was concluded that the most likely source of the pathogen was the dried sugar beet lime used as a mushroom casing soil amendment (Meikle, 2001). The ability of L. monocytogenes and S. aureus to survive on fresh mushrooms has also been described. Martin and Beelman (1996) noted that the gas composition (low oxygen and high carbon dioxide) inside unventilated polyvinyl chloride (PVC) packages would favour the growth of facultative anaerobic microorganisms such as S. aureus. González-Fandos et al. (2001) reported that the modified atmosphere packaging followed by storage at 4 °C and 10 °C allowed the growth and survival of L. monocytogenes.

This study was undertaken to determine and compare the microbial quality and safety of 22 species of fresh cultivated and wild mushrooms obtained in retail outlets in Zaragoza (Spain). For these purposes, we quantified the number of aerobic mesophilic microorganisms, Pseudomonas spp., Enterobacteriaceae, lactic acid bacteria, yeasts and moulds; total and thermotolerant coliform bacteria and Escherichia coli were also analyzed as fecal contamination indicator microorganisms and the microbiological safety was evaluated by the prevalence of selected food-borne pathogens, E. coli O157:H7, L. monocytogenes, Salmonella spp., S. aureus and Yersinia enterocolitica. These data should contribute to evaluating the microbiological risk associated with fresh mushrooms as well as establishing international criteria for the microbial quality of these highly appreciated foods.