The Investigation Of Survival Rate And Antibiotic Resistance Of Listeria Spp.in Response To The Various Environment Stress During The Food Processing

Listeria monocytogenes is an important foodborne pathogen,and is widely recognized as a significant zoonotic agent among Listeria Spp..Listeria monocytogenes can cause serious diseases such as central nervous system diseases with a high mortality rate,which has gradually attracted the attention by the Chinese media.Recently,Chinese foodborne pathogen monitoring system is slowly developed.More and more in vitro diagnostic kits for Listeria monocytogenes have been reported in China.However,due to the lack of national reference materials,a comprehensive,objective,and scientific evaluation standard for pathogen detection remains lacking,which is a bottleneck for healthy development of the market.In addition,Listeria monocytogenes distributed widely in the environment,but few knowledge is available for Listeria species other than Listeria monocytogenes.The objectives of this study are to(1)Collect Listeria monocytogenes with clear background through systematic characterization and keep these records as the national reference;(2)Comparative analysis of the survival rate of six different Listeria species with the treatment under different environmental stresses;and(3)Investigate of the antimicrobial resistance changes of Listeria monocytogenes before and after the treatment under different stress conditions.1.Preparation of Chinese National Reference strains of Listeria monocytogenesA collection of clinical Listeria monocytogenes(n=13),reference strains(n=25),negative strains(other 5 Listeria species)and some foodborne pathogens(Escherichia coli,Streptococcus)were retrieved from a total of 458 Listeria isolates.We prepared the Chinese National Reference strains of Listeria monocytogenes through a variety of characterization and identification methods,including conventional biochemical identification,molecular identification,mass spectrometry identification,and antibiotic resistance analysis by MIC assay,whole genome sequencing and bioinformatics analysis by predicting this pllenotype and genotype association.2.The survival rate of Listeria Spp.under different environmental stressesVarious stress conditions exist in the food processing environment,including heat,acid-base,salt and ultraviolet stresses.We selected some Listeria monocytogenes(n=11,including 6 different serotypes)as a group,and non-Listeria monocytogenes(n=10,including 5 other common species)as a group,and investigated the comparative survival rate of bacteria after a particular stress to evaluate their stress adaptability.The results showed that Listeria spp.were more sensitive to heat stress.21 strains of Listeria began to die after being treated at 65? for 4 minutes.The pH range of 4-10 had no effect on the survival rate of Listeria between two groups or among strains.When the environment reaches extremely alkaline(pH=12)and extremely acidic(pH=2),the survival rate of bacteria was highly dependent on the treatment time.At pH=12,the treatment time was up to 30 minutes,and none of bacteria isolates can be detected by plate culture method.However,when treated with pH=2 for 10 minutes,8 strains could not be detected.Furthermore,the ability of the Listeria monocytogenes group to resist strong acidic stress was significantly different from that of non-Listeria monocytogenes group.10%salt stress can effectively inhibit the growth of Listeria strains.However,with the extension of the treatment time,the downward trend of bacterial survival rate began to slow down.Moreover,the ability of the Listeria monocytogenes group to resist salt stress showed a significant difference compared to the non-Lis ter ia monocytogenes.Listeria strains were sensitive to ultraviolet rays,and none of them can be detected after being irradiated for 100 seconds.3.Antibiotic resistance of Listeria spp.after different stress treatmentThe antibiotic resistance of Listeria Spp.can be changed when exposed to the aforementioned stress conditions for a certain period of time.The most obvious observation is that under all examined stress conditions,the resistance to the sulfonamide compound trimethoprim showed an increased in antibiotic resistance MIC values.Under 10%salt stress,bacteria that can survive for a long time,become to show increased MIC values to multiple antibiotics.Regardless of whether it is in the acid or alkali stress environment,the resistance to the above antibiotics were identical.Under the salt stress condition,it was observed that among these kinds of changed antibiotic resistance,the change of MIC value of the Listeria monocytogenes group was significantly larger than that of the non-Listeria monocytogenes group.By collaboration with the Chinese Food and Drug Administration,we have established a reliable panel of Listeria monocytogenes collection as the national reference,which can provide essential guideline for in vitro diagnostic methods.We examined the survival rate and drug resistance of Listeria Spp.under different conditions of sterilization and bacteriostatics,which is aim to provide reference for the food processing industry.