Survival of a serotype 4b strain and a serotype 1/2a strain of Listeria monocytogenes, isolated from a stone fruit outbreak investigation, on whole stone fruit at 4 °C

Highlights

• Survival of natural isolates of Listeria monocytogenes on stone fruit was analyzed.

• Effects of strain and fruit type on survival were investigated.

• A whole fruit rinsing method was evaluated for enumerating Listeria monocytogenes.

• Two chromogenic agars were evaluated for enumerating Listeria monocytogenes.

Abstract

In the summer of 2014, a multistate outbreak of listeriosis associated with contaminated stone fruit (peach and nectarine) was reported. A serotype 4b variant Listeria monocytogenes (Lm) strain of singleton Sequence Type (ST) 382 was isolated from clinical samples and stone fruit associated with the outbreak. A serotype 1/2b Lm strain of ST5, Clonal Complex 5 was isolated only from outbreak-associated stone fruit, not from clinical samples. Here we investigated the fate of the serotype 4b and 1/2b strains, at two inoculation levels (high level at 3.7 logCFU/fruit and low level at 2.7 logCFU/fruit), on the surfaces of white peach, yellow peach and yellow nectarine stored at 4 °C for 26 days. After rinsing the fruits, we determined the Lm levels in the rinsates and on the peels. We enumerated Lm using a direct plating method and compared two chromogenic agars. The Lm populations rapidly declined in the first 3 days and then declined more slowly until Day 19/21. The maximum decline was 1.6 logCFU/fruit on yellow peach inoculated with serotype 4b at high level. For fruits inoculated with high-level Lm, the lowest level of Lm (1.7 logCFU/fruit) was observed on for white peach inoculated with serotype 1/2b, and the highest level of Lm (2.6 logCFU/fruit) on Day 19/21 was observed on yellow peach inoculated with the serotype 1/2b strain. For fruits inoculated with low-level Lm, the lowest level of Lm (1.3 logCFU/fruit) was observed on yellow nectarine inoculated with either the serotype 4b or 1/2b strain, and the highest level of Lm (1.7 logCFU/fruit) on Day 19/21 was observed on yellow peach inoculated with ST382. The D-values ranged from 15 days to 28 days. Lm remained viable until the end of storage (Day 26), but the levels were not significantly different from those on Day 19/21. The types of stone fruit and Lm strain did not significantly affect the survival of Lm. These results demonstrate that contaminated stone fruit can carry a potential risk for causing listeriosis in susceptible populations. Comparison of direct plating results using two chromogenic agars showed that RAPID' L. mono and Agar Listeria Ottavani & Agosti performed equivalently for enumerating Lm on stone fruit. The fruit rinsing recovered 80% to 84% of Lm from fruit surfaces.

Introduction

Listeria monocytogenes (Lm) is recognized as one of the most dangerous foodborne pathogens, especially when it is present in ready-to-eat (RTE) foods that support their growth, as they have a case fatality rate of up to 20% (Kathariou, 2002). In recent years, fresh fruits contaminated with Lm have been linked to outbreaks, sporadic cases, and recalls (Garner and Kathariou, 2016; Kase et al., 2017). It had long been assumed that any contamination of intact fruit by microorganisms was limited to the fruit's external surface. Moreover, it had generally been assumed that if pathogenic bacteria entered the interior of some fruits, the inherent acidity of the pulp may prevent bacterial growth; however, the contamination, internalization, survival, and growth of Lm within intact fruits have been recently documented (Chen et al., 2016a, Chen et al., 2016b; Macarisin et al., 2019; Macarisin et al., 2017).

During the summer of 2014, detection of Lm contamination prompted a recall of stone fruit (peach, plum, and nectarine) produced in the U.S., and a related multistate outbreak of listeriosis occurred. The outbreak strain was sequence type (ST) 382, which belongs to the clone of singleton ST382 and serotype 4b variant (i.e. serotype 4b by traditional serotyping but atypical by PCR serotyping). In addition, a second strain of ST5, which belongs to the clone of Clonal Complex (CC) 5 and serotype 1/2b, was also recovered from those fruits. The pathogen enumeration on 7 lots of incriminated fruits revealed that the levels of Lm ranged from 0.7 to 3.5 log CFU/fruit for peach (i.e., yellow peach and white peach) and from 0.7 to 1.9 log CFU/fruit for nectarine (yellow nectarine and while nectarine). In 2019, another recall due to Lm contamination in stone fruit produced in Chile occurred (Food Safety News, 2019). Additionally, in a survey in South Africa, Listeria spp., indicator organism for Lm, was found on peach and in a peach processing environment (Duvenage and Korsten, 2017). These incidences highlight the need to study the fate of Lm on stone fruit. Collignon and Korsten demonstrated that Lm ATCC 19115 (serotype 4b) could attach and colonize surfaces of freshly-harvested peach, grow at 21 °C and survive through simulated export chain where peach was stored mostly at cold temperatures (Collignon and Korsten, 2010). The authors recommended future studies focusing on the risk of fruit contamination at the end of supply chain.

Given that whole stone fruit can serve as a vehicle of listeriosis, we evaluated the fate of Lm strains, isolated from naturally contaminated stone fruit, on several types of stone fruit under conditions simulating postharvest supply chain. After fruit leaves the packing houses, the temperature during transportation, distribution, retail settings and household refrigeration can be ≥5 °C (Cantwell and Reid, 2002; Kou et al., 2015). Thus, the objectives of the present study were to 1) investigate the fate of two Lm strains, isolated from stone fruit implicated in the 2014 investigation, on the surface of peach (white and yellow) and yellow nectarine stored at 4 °C; 2) evaluate the efficacy of a previously-employed rinsing method in recovering Lm from stone fruit surface; and 3) compare two chromogenic agars for enumerating Lm recovered from stone fruit. Results of the study can contribute to further assessments of the risk associated with Lm contamination of low acidity fruit.