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Transcriptomic analysis of C. jejuni grown in anaerobic jar with air mix and in Modular Atmospheric Controlled System

ABSTRACT: Transcriptomic analysis of C. jejuni HPC5 grown in anaerobic jar supplemented with air mix and in Modular Atmospheric Controlled System (MAC). C. jejuni HPC5 is a Campylobacter strain isolated from broiler chickens. Majority of the Campylobacter strains are cultured in an atmosphere containing 5-10% (v/v) oxygen or 5% hydrogen and 5-10% (v/v) carbon dioxide and the rest with nitrogen. To set up a jar (3.5L, Oxoid) to the required microaerophilic conditions, a vacuum is created to -22 psi and then air mixture of 85% v/v nitrogen, 10% v/v carbon dioxide and 5% v/v hydrogen (Air Products, Crewe, UK) is introduced into the jar. This resulted in a microaerobic atmosphere containing approximately 5.6% oxygen, 3.6% hydrogen, 7.3% carbon dioxide and 83% nitrogen. The Modular Atmospheric Controlled System (MAC; Don Whitely Scientific) is an anaerobic chamber where air mixture of 85% nitrogen, 10% carbon dioxide and 5% oxygen is used for growing Campylobacter. Differences in growth pattern were observed when Campylobacters were grown in it and transcriptomic analysis was done to prove the difference in the regulation of genes in the two conditions.

ORGANISM(S): Campylobacter jejuni subsp. jejuni NCTC 11168 = ATCC 700819 Campylobacter jejuni RM1221 Campylobacter coli RM2228 Campylobacter jejuni

PROVIDER: GSE17801 | GEO | 2011/04/01

SECONDARY ACCESSION(S): PRJNA118335

REPOSITORIES: GEO

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Project description:Modified atmosphere packaging (MAP) is a common strategy to selectively prevent the growth of certain species of meat spoiling bacteria. While studies on the effectiveness of MAP are still scarce on a putative control over the population of photobacteria detected as meat spoilers, they could develop means to enhance safety and quality of raw meat. This study aims to determine the impact on photobacteria of two modified atmospheres: with high oxygen concentration (red and white meats), and free oxygen MAP (white meats and seafood). We have conducted growth experiments of the two main species found on meat, Photobacterium carnosum (P.) and P. phosphoreum, on a meat simulation media under different gas mixtures of nitrogen, oxygen and carbon dioxide representing air-, high oxygen- and vacuum-like conditions with and without carbon dioxide present. Growth was monitored based on optical density, and samples were taken during exponential growth for a comparative proteomic analysis that allowed the determination of the effects of the different gases and their synergy. Growth under air atmosphere appears optimal particularly for P. carnosum, with enhancement of energy metabolism, respiration, oxygen consuming reactions, and a predicted preference for lipids as carbon source. However, all the other atmospheres show some degree of growth reduction. An increase in oxygen concentration leads to an increase in enzymes counteracting oxidative stress for both species, and enhancement of heme utilization and iron-sulfur cluster assembly proteins for P. phosphoreum. Absence of oxygen appears to switch the metabolism towards fermentative pathways, where either ribose (P. phosphoreum), or glycogen (P. carnosum) appear to be the preferred substrates. Additionally, it promotes the use of alternative electron donors/acceptors, mainly formate and nitrate/nitrite. Stress response is manifested as enhanced expression of enzymes able to produce ammonia (e.g. carbonic anhydrase, hydroxylamine reductase) and regulate osmotic stress. Our results suggest that photobacteria do not sense the environmental levels of carbon dioxide but rather adapt to their own anaerobic metabolism. The regulation in presence of carbon dioxide is limited and strain-specific under anaerobic conditions. However, when oxygen at air-like concentration is present together with carbon dioxide the oxidative stress appears enhanced compared to air conditions (very low carbon dioxide), explained if both gases have a synergistic effect. This is further supported by the increase in oxygen concentration in presence of carbon dioxide. The atmosphere is able to fully inhibit P. carnosum, heavily reduce P. phosphoreum growth in vitro and trigger diversification of energy production with higher energetic cost, highlighting the importance of concomitant bacteria for their growth on raw meat under said atmosphere.

Project description:Oxygen and carbon dioxide are common protective gases used in modified atmosphere packaging (MAP) of meat. Within the package, they selectively suppress members of the spoilage microbiome, reshaping it to adapted species concomitantly growing upon MAP. Thus, this species must exhibit adaptation mechanisms to withstand the inhibitory effect of carbon dioxide and oxygen, and cope with selective nutrition on MAP meat. In order to uncover these mechanisms, the typical representative meat-spoiling bacteria Brochothrix (B.) thermosphacta TMW2.2101 and four lactic acid bacteria (LAB) Carnobacterium (C.) divergens TMW2.1577, C. maltaromaticum TMW2.1581, Leuconostoc (L.) gelidum subsp. gelidum TMW2.1618 and L. gelidum subsp. gasicomitatum TMW2.1619 were grown in a meat simulation medium under a controlled, sterile environment, aerated constantly with either air, 100%_N2, 30%_CO2/70%_O2 or 30%_CO2/70%_N2. Growth dynamics were monitored and a label-free quantitative mass spectrometric approach was employed to determine changes within the bacterial proteomes in response to the different gas atmospheres. Revealed bacterial tolerance to modified atmospheres (MA) comprise two possible scenarios: Either bacteria were intrinsically adapted to MA, exhibiting no proteomic regulation of enzymes (L. gelidum subsp. gelidum and gasicomitatum) or, tolerance was provided by varying specific metabolic adaptation (B. thermosphacta, C. divergens, C. maltaromaticum). In detail, metabolic adaptation mechanisms to oxygen comprised an enhanced oxidative stress reduction response, adjustment of the pyruvate metabolism and catabolic oxygen consuming reactions. Adaptation to carbon dioxide was characterized by an upregulation of proteins involved in intracellular pH homeostasis, maintenance of osmotic balance and alteration of the fatty acid composition of the cell membrane. We furthermore predict species-specific strategies for different and preferential carbon source utilization enabling a non-competitive coexistence on meat and resulting in a synergistic spoilage. We conclude that a gas atmosphere containing 30%_CO2/70%_O2 has no inhibitory effect on the analyzed prominent meat-spoiling bacteria whereas 30%_CO2/70%_N2 predictively inhibits C. divergens TMW21577 and B. thermosphacta TMW2.2101 but not the other three species. This gives a mechanistically explanation of their acknowledged status as typical spoilage organisms on packaged meats.

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Transcriptomic analysis of C. jejuni grown in anaerobic jar with air mix and in Modular Atmospheric Controlled System

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