Survival of Pathogens in Frozen Fruits

March 2013

Outbreaks of foodborne illness have been associated with consumption of contaminated berries. Contamination can occur pre- or post-harvest, or even during handling by consumers. Freezing is a common practice to preserve berries on a commercial level as well as in home settings but survival characteristics of foodborne pathogens on frozen fruits is not well understood.

We did a study to determine if foodborne pathogens can survive freezing in water and syrup, and after adding sugar and freezing. Red raspberries and sliced or whole strawberries were inoculated with Salmonella (mixture of five serotypes), Shiga toxin-producing Escherichia coli O157:H7 (mixture of five strains), Cryptosporidium parvum oocysts, Encephalitozoon intestinalis (microsporidia) spores, or murine norovirus (MNV-1), a surrogate for human norovirus.

Each fruit type (20 - 30 g) was separately inoculated with 100 µl of a suspension of test pathogen. Inocula were allowed to dry in a biosafety hood at 21 - 23°C for 1 h. Fruits were then immersed in water or syrup (40 g sugar/500 ml water), or combined with sugar (sucrose, 4 g/20-30 g of berries) before freezing or refrigerating.

Berries were stored for 16 - 18 h at 4°C or at -23°C followed by thawing at 20°C for 30 min. Berries inoculated with bacterial pathogens were combined with Dey Engley (DE) broth and washed. Samples serially diluted in 0.1% peptone were plated on McConkey agar to enumerate E. coli O157:H7 or XLD agar to enumerate Salmonella. Fruits on which microsporidia or Cryptosporidium were inoculated were resuspended in DE broth, repeatedly washed, resuspended in DMEM-F-12 medium, and inoculated onto monolayers of RK-13 or HCT-8 cell lines, respectively. MNV-1 was recovered in phosphate-buffered saline (1M NaCl) followed by 8% polyethylene glycol (PEG) precipitation. PEG pellets were resuspended in the infection medium and inoculated onto monolayers of the RAW 264-7 cell line. Live virus was quantified by plaque formation. Log reductions caused by freezing were calculated by subtracting the number of test organisms that survived at -23°C from the number surviving at 4°C.

Less than a 1-log reduction of Salmonella and E. coli O157:H7 was observed on raspberries and whole and sliced strawberries, with a non-significant but larger reduction in water than in syrup or sugar. A reduction of at least 4 log CFU/g occurred when Cryptosporidium was exposed to all treatments.

With microsporidia, reductions of 3, 0.8, and 1 log CFU/g were observed on raspberries frozen in water, syrup, and sugar, respectively. When inoculated onto whole strawberries, reductions of 1, 0.5 and 0.7 log CFU/g occurred in water, syrup, and sugar, respectively. Similar results were observed for sliced frozen strawberries.

The number of plaque-forming units (PFU) of MNV-1 was reduced by 0.4 log PFU/g in whole strawberries frozen in water and 0.3 log in whole strawberries frozen in syrup. Less than a 0.2-log PFU reduction of MNV-1 occurred on sliced strawberries and whole raspberries frozen in water, syrup, or sugar.

In summary, freezing strawberries or raspberries with water, syrup, or sugar minimally affects the viability of Salmonella, E. coli O157:H7, microsporidia, and MNV-1. In contrast, the viability of Cryptosporidium oocysts is substantially reduced but not eliminated by all of the freezing conditions tested.