Microbiota of sliced cooked ham packaged in modified atmosphere throughout the shelf life: Microbiota of sliced cooked ham in MAP

https://doi.org/10.1016/j.ijfoodmicro.2018.09.017Get rights and content

Highlights

  • The microbiota of 19 MAP sliced cooked ham lots was monitored during the shelf life.

  • Plate counts, RAPD-PCR fingerprinting, and 16S rRNA gene profiling were applied.

  • Lactobacillus sakei and Leuconostoc carnosum were dominant cultivable LAB species.

  • Presence of recurrent strains suggests contamination by production plant bacteria.

Abstract

Fourteen lots of cooked ham in modified atmosphere packaging (CH) were analyzed within a few days from packaging (S) and at the end of the shelf-life (E), after storage at 7 °C to simulate thermal abuse. Five more lots, rejected from the market because spoiled (R), were included in the study. Quality of the products was generally compromised during the shelf life, with only 4 lots remaining unaltered. Analysis of 16S rRNA gene amplicons resulted in 801 OTUs. S samples presented a higher diversity than E and R ones. At the beginning of the shelf life, Proteobacteria and Firmicutes dominated the microbiota, with Acinetobacter, Brochothrix, Carnobacterium, Lactobacillus, Prevotella, Pseudomonas, Psychrobacter, Weissella, Vibrio rumoiensis occurring frequently and/or abundantly.

E and R samples were dominated by Firmicutes mostly ascribed to Lactobacillales. It is noteworthy the appearance of abundant Leuconostoc, negligible in S samples, in some E and R samples, while in other LAB were outnumbered by V. rumoiensis or Brochothrix thermosphacta. The microbiota of spoiled and R samples could not be clustered on the basis of specific defects (discoloration, presence of slime, sourness, and swollen packages) or supplemented additives. LAB population of S samples, averaging 2.9 log10(cfu/g), increased to 7.7 log10(cfu/g) in the E and R samples. Dominant cultivable LAB belonged to the species Lactobacillus sakei and Leuconostoc carnosum. The same biotypes ascribed to different species where often found in the corresponding S and R samples, and sometime in different batches provided from the same producer, suggesting a recurrent contamination from the plant of production. Consistently with growth of LAB, initial pH (6.26) dropped to 5.74 in E samples. Volatiles organic compound (VOCs) analysis revealed that ethanol was the major metabolite produced during the shelf life. The profile of volatile compounds got enriched with other molecules (e.g. 2-butanone, ethyl acetate, acetic acid, acetoin, butanoic acid, ethyl ester, butanoic acid, and 2,3-butanediol) mainly ascribed to microbial metabolism.

Introduction

Modified Atmosphere Packaging (MAP) is widely utilized to reduce the addition of exogenous preservatives and to extend the shelf life of food without altering the physical and chemical properties. However, sliced cooked ham in MAP (hereinafter referred to as CH) still represents a perishable product, very sensitive to bacterial spoilage (Borch et al., 1996; Samelis et al., 1998; Vasilopoulos et al., 2015). The main undesired defects caused by microbial growth include pH decrease, gas and slime production, blowing, discoloration, purge, and off-flavors formation. In some cases, the defects can result in the premature spoilage and in the reduction of shelf-life (Korkeala and Björkroth, 1997).

Manufacturing of cooked ham ends with cooking at a core temperature up to 70 °C, thus killing most of vegetative microbes. However, CH undergoes a massive bacterial proliferation toward the end of the shelf life despite the combination of hygienic precautions, preservative procedures such as chilling, micro-aerophilic conditions, and presence of NaCl and nitrites. The bacterial community in CH can reach 107–109 cfu/ g in few weeks after packaging, with a composition depending on many factors, including packaging, gas atmosphere, product composition, hygienic conditions throughout the processing line, and the temperature during both the distribution and the storage in consumer fridge (Audenaert et al., 2010; Kreyenschmidt et al., 2010; Samelis et al., 1998; Vasilopoulos et al., 2008). Previous studies reported a preferential growth of psychrotrophic lactic acid bacteria (LAB), with Leuconostoc carnosum, Leuconostoc gelidum, Lactobacillus sakei, Lactobacillus curvatus, Carnobacterium divergens, Carnobacterium maltaromaticum as the most recurrent species, which dominated the bacterial community at the end of the shelf life (Audenaert et al., 2010; Geeraerts et al., 2017; Vasilopoulos et al., 2008). Nonetheless, information on the microbiota composition in CH is still scarce with respect to the bacterial groups other than LAB.

In order to obtain a wide description of the microbiota composition and diversity, CH samples coming from 10 producers in 6 European countries were analyzed. They presented different characteristics in terms of MAP composition, time of storage, and presence of additives. Microbiota was studied throughout the shelf life of CH by 16S rRNA gene profiling and by a culture-dependent method to specifically isolate and trace the LAB biotypes, with the perspective to select candidates for CH biopreservation. Dominant LAB were isolated, genotyped by RAPD-PCR fingerprinting, and were given a taxonomic designation by partial sequencing of 16S rRNA gene. CH samples were analyzed at the packaging time and at the end of shelf life, during which they were stored at 7 °C. The set of samples included also a group of CH that were rejected as spoiled. For all the samples, the occurrence of defects was evaluated and the prevalent volatile compounds (VOCs) were determined by solid phase micro extraction (SPME) coupled with GC–MS.

Section snippets

Sample collection and experimental design

Eleven commercial CH were received from 10 producers spread over 6 European countries. The additives and the gas mixture, composed by N2 and CO2, are reported in Table S1. As a whole, 19 batches of production were studied. On 14 batches, microbiological and chemical analyses were performed at the start of the shelf life (S) and at the expiration date (E). The initial measurements were generally performed 2–4 days after packaging but, because of logistic management, in some case the analyses

Spoilage assessment

Fourteen CH lots were evaluated both within a few days (mean 5 days) from the date of packaging (hereinafter referred to as S samples) and at the end of a shelf-life of 23–45 days (mean 30 days), during which they were stored at 7 °C (E samples). The remaining 5 lots, rejected from market because spoiled (R samples), were analyzed on average after 30 days from the date of packaging.

All the S samples were tasty and did not present any alteration. Only 4 CH lots reached the use-by date in very

Conclusions

The market of ready-to-eat food products is growing, with consumption of CH that has been increasing rapidly and has further growth potentials. Microbiology of CH is a relevant issue, since it has a bearing on both safety and quality of the product. Whereas the outbreaks have been rare, premature spoilage of CH can occur, and at least some sensorial defects generally affect the product, approaching to the end of the shelf life. A comprehensive overview of CH microbial communities was required.

Funding

This study was partially supported by Christian Hansen, Denmark, by grants to AM from the Spanish Ministry of Economy and Competitiveness (SAF2015-65878-R) and Generalitat Valenciana (PrometeoII/2014/065), and by the European Regional Development Fund (ERDF).

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