Greenhouses represent an important evolutionary niche for Alternaria alternata.

Alternaria alternata is a ubiquitous soil-borne fungus capable of causing diseases in a variety of plants and occasionally in humans. While populations of A. alternata from infected plants have received significant attention, relatively little is known about its soil populations, including its population genetic structure and antifungal susceptibilities. In addition, over the last two decades, greenhouses have become increasingly important for food and ornamental plant production throughout the world, but how greenhouses might impact microbial pathogens such as A. alternata populations remains largely unknown. Different from open crop fields, greenhouses are often more intensively cultivated, with each greenhouse being a relatively small and isolated space where temperature and humidity are higher than surrounding environments. Previous studies have shown that greenhouse populations of two common molds, Aspergillus fumigatus and A. alternata, within a small community in southwestern China were variably differentiated. However, the relative contribution of physical separation among local greenhouses to the large-scale population structure remains unknown. Here, we isolated strains of A. alternata from seven greenhouses in Shijiazhuang, northeast China. Their genetic diversity and triazole susceptibilities were analyzed and compared with each other and with 242 isolates from nine greenhouses in Kunming, southwest China. Results showed that the isolation of greenhouses located <1 km from each other locally contributed similarly to the overall genetic variation as that between the two distant geographic regions. In addition, our results indicate that greenhouses could be significant sources of triazole resistance, with greenhouses often differing in their frequencies of resistant strains to different triazoles. IMPORTANCE: Greenhouses have become increasingly important for food production and food security. However, our understanding of how greenhouses may contribute to genetic variations in soil microbial populations is very limited. In this study, we obtained and analyzed soil populations of the cosmopolitan fungal pathogen Alternaria alternata in seven greenhouses in Shijiazhuang, northeast China. Our analyses revealed high proportions of isolates being resistant to agricultural triazole fungicides and medical triazole drugs, including cross-resistance to both groups of triazoles. In addition, we found that greenhouse populations of A. alternata located within a few kilometers showed similar levels of genetic differentiation as those separated by over 2,000 km between northeast and southwest China. Our study suggests that greenhouse populations of this and potentially other fungal pathogens represent an important ecological niche and an emerging threat to food security and human health.

Characterization of Escherichia coli O157:non-H7 isolated from retail food in China and first report of mcr-1/IncI2-carrying colistin-resistant E. coli O157:H26 and E. coli O157:H4.

Escherichia coli O157 belongs to a diverse serogroup including different H serotypes. E. coli O157: H7 is the most common serotype that can cause acute gastroenteritis, hemorrhagic colitis (HC), and hemolytic-uremic syndrome (HUS) in humans. In recent years, some E. coli O157:non-H7 strains have been reported to cause sporadic cases and outbreaks of diarrheal diseases. However, the phenotypic and genotypic characteristics of E. coli O157:non-H7 are poorly understood. In this study, E. coli O157:non-H7 strains were isolated from retail food sold on markets in 13 cities in China during 2012-2016 and characterized systematically in terms of their H serotypes, virulence genes, antibiotic resistance, and genotypes. Six H serotypes (H26, H42, H11, H38, H4, and H5) were identified, of which, O157:H42 (31.4 %) and O157:H26 (28.6 %) were the most prevalent. Most of the isolates (82.9 %) carried virulence genes. Ten isolates (28.6 %) carried the eae gene and were identified as atypical enteropathogenic E. coli. Multilocus sequence typing showed that the E. coli O157:non-H7 strains demonstrated diverse sequence types with different evolutionary trends than E. coli O157:H7. All the isolates exhibited multidrug resistance. The isolates showed a high prevalence of resistance to AMC, AMP, CTX, CIP, T/S, TE, and FFC. The predominant antibiotic-resistance genes were TEM-1 (40.0 %), CTX-M-55 (34.3 %), aadA (74.3 %), sul2 (62.9 %), floR (91.4 %), tetA (85.7 %), qnrS (37.1 %), oqxA (62.9 %), and oqxB (62.9 %). For the first time, we identified IncI2 plasmid-mediated colistin-resistant strains (six O157:H26 and one O157:H4). These strains co-harbored plasmid-mediated mcr-1 gene, CTX-M-55, OXA-4, PMQR, and other resistant genes, which is of great concern. Colistin and cefotaxime are generally used as the last defense to treat complicated infections. The emergence of virulent and multidrug resistant strains in food poses a serious threat to human health. The strict monitoring and surveillance of multiple-drug resistant strains in food are needed to prevent their dissemination to humans.

Genetic Structure and Triazole Antifungal Susceptibilities of Alternaria alternata from Greenhouses in Kunming, China.

Alternaria alternata is an opportunistic human fungal pathogen and a ubiquitous phytopathogen capable of causing diseases to >100 agricultural crops and ornamental plants. To control plant diseases caused by A. alternata, triazole fungicides have been widely used both in open crop and vegetable fields and in indoor growth facilities such as greenhouses. At present, the effect of fungicide use on triazole resistance development in A. alternata populations is not known. Here, we isolated 237 A. alternata strains from nine greenhouses around metropolitan Kunming in Yunnan, southwest China, determined their genotypes using 10 short tandem repeat markers, and quantified their susceptibility to four triazoles (difenoconazole, tebuconazole, itraconazole, and voriconazole). Abundant allelic and genotypic diversities were detected among these A. alternata strains. Significantly, over 17% of the strains were resistant to difenoconazole, and both known and new drug-resistance mutations were found in the triazole target gene cyp51. Our findings of high-level genetic variation of A. alternata in greenhouses coupled with high-frequency fungicide resistance call for greater attention to continued monitoring and to developing alternative plant fungal disease management strategies in greenhouses. IMPORTANCE Alternaria alternata is among the most common fungi in our environments, such as indoor facilities, the soil, and outdoor air. It can cause diseases in >100 crop and ornamental plants. Furthermore, it can cause human infections. However, our understanding of its genetic diversity and antifungal susceptibility is very limited. Indeed, the critical threshold values for resistance have not been defined for most antifungal drugs in this species. Greenhouses are known to have heavy applications of agricultural fungicides. In this study, we analyzed strains of A. alternata from nine greenhouses near metropolitan Kunming in southwestern China. Our study revealed very high genetic diversity and identified strains with high MIC values against two agricultural and two medical triazole antifungals within each of the nine greenhouses. Our study calls for greater attention to this emerging threat to food security and human health.

Genetic Diversity and Azole Resistance Among Natural Aspergillus fumigatus Populations in Yunnan, China.

The emergence and spread of azole resistance alleles in clinical and environmental isolates of Aspergillus fumigatus is a global human health concern and endangers the "One Health" approach in our fight against antifungal resistance (AFR) in this pathogen. A major challenge to combat AFR in A. fumigatus is the massive aerial dispersal ability of its asexual spores. Our recent fine-scale survey of greenhouse populations of A. fumigatus near Kunming, Yunnan, China, suggested that the use of azole fungicides for plant protection was likely a major driver of the high-frequency azole-resistant A. fumigatus (ARAF) in greenhouses. Here, we investigated the potential spread of those ARAF and the structure of geographic populations of A. fumigatus by analyzing 452 isolates from 19 geographic locations across Yunnan. We found lower frequencies of ARAF in these outdoor populations than those in greenhouses near Kunming, but there were abundant new alleles and new genotypes, including those associated with azole resistance, consistent with multiple independent origins of ARAF across Yunnan. Interestingly, among the four ecological niches, the sediments of a large lake near Kunming were found to have the highest frequency of ARAF (~ 43%). While most genetic variations were observed within the 19 local populations, statistically significant genetic differentiations were found between many subpopulations within Yunnan. Furthermore, similar to greenhouse populations, these outdoor populations of A. fumigatus in Yunnan were significantly different from those in other parts of the world. Our results call for increased attention to local and regional studies of this fungal pathogen to help develop targeted control strategies against ARAF.

High prevalence of multidrug-resistant Escherichia coli and first detection of IncHI2/IncX4-plasmid carrying mcr-1 E. coli in retail ready-to-eat foods in China.

Antibiotic-resistant bacteria in food pose an important threat to public health. Multidrug-resistant strains in ready-to-eat (RTE) foods can be transferred to humans through diet, which increases their health risk. This study systematically investigated antibiotic resistance and antibiotic resistance genes in E. coli isolated from retail RTE foods and characterized plasmid-mediated colistin-resistant E. coli strains. A total of 1118 RTE food samples were collected from markets in 39 cities in China, and 126 E. coli strains, >95% of which were multidrug-resistant, were isolated. The isolates showed a high prevalence of resistance to tetracycline (95.24%), ampicillin (82.54%), trimethoprim-sulfamethoxazole (77.78%), nalidixic acid (74.60%), cephalothin (72.22%), chloramphenicol (66.67%), and streptomycin (53.97%). Twenty-two extended-spectrum ß-lactamase (ESBL)-producing E. coli and four colistin-resistant E. coli were identified. The resistance genes TEM, CTX-M, tetA, sul2, strA/strB, aadA, and qnrS were the most frequently detected. CTX-M-55 and CTX-M-14 were the predominant CTX-M types. All the four colistin-resistant E. coli isolates were positive for mcr-1. The mcr-1 gene can be transferred to E. coli C600 through conjugation and transformation. Whole-genome sequencing revealed that the mcr-1 genes were found in IncX4 and IncHI2 plasmids. To the best of our knowledge, this is the first report of IncHI2/IncX4 plasmid-bearing mcr-1-positive E. coli strains in RTE foods sold in markets, and the first report of the isolation of the international epidemic E. coli clone ST101 and mcr-1-carrying ESBL-producing E. coli from RTE foods. These results provide valuable information for assessing antibiotic-resistant E. coli infections and controlling antibiotic-resistant E. coli.

Local anesthetic levobupivacaine induces ferroptosis and inhibits progression by up-regulating p53 in non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is a common malignancy with high mortality and poor prognosis. Levobupivacaine is a widely used local anesthetic and presents potential anti-tumor activity. Nevertheless, the function of levobupivacaine in the NSCLC development remains elusive. Here, we tried to investigate the impact of levobupivacaine on the NSCLC progression and the underlying mechanism. Significantly, we revealed that levobupivacaine could inhibit the proliferation and induce the apoptosis of NSCLC cells. Levobupivacaine was able to attenuate the invasion and migration in the cells. Meanwhile, the treatment of levobupivacaine enhanced the erastin-induced inhibition of cell growth of NSCLC cells. The treatment of levobupivacaine remarkably increased the levels of ROS, iron, and Fe2+ in NSCLC cells. Mechanically, levobupivacaine up-regulated the expression of p53 and induced ferroptosis by regulating p53 in NSCLC cells. Moreover, tumorigenicity analysis in nude mice showed that the treatment of levobupivacaine significantly repressed the tumor growth of NSCLC cells in vivo. In summary, we concluded that the local anesthetic levobupivacaine inhibits the progression and induces ferroptosis of NSCLC by up-regulating p53. Our finding provides new insights into the mechanism by which levobupivacaine attenuates the development of NSCLC. Levobupivacaine may serve as a potential anti-tumor candidate for the therapeutic strategy of NSCLC.

The Local Anesthetic Bupivacaine Inhibits the Progression of Non-Small Cell Lung Cancer by Inducing Autophagy Through Akt/mTOR Signaling.

Non-small cell lung cancer (NSCLC) is a prevalent malignancy with high mortality and poor prognosis. Bupivacaine serves as a widely used local anesthetic and presents potential anti-tumor activity. Nevertheless, the function of bupivacaine in the NSCLC development remains elusive. Here, we tried to investigate the impact of bupivacaine on the NSCLC progression. Significantly, we revealed that bupivacaine was able to reduce the proliferation and induce the apoptosis of NSCLC cells. Bupivacaine could attenuate the invasion and migration in the cells. Mechanically, the treatment of bupivacaine increased the expression ratio of light chain 3B-II (LC3B-II)/LC3B-I and the expression of Beclin-1 in the NSCLC cells. Meanwhile, the expression of the autophagic adaptor protein p62 was decreased by bupivacaine treatment in the cells. The treatment of bupivacaine attenuated the phosphorylation of AKT and mTOR in the NSCLC cells. The AKT activator SC79 and autophagy inhibitor 3-methyladenine (3-MA) reversed the bupivacaine-inhibited phosphorylation of AKT and mTOR and bupivacaine-induced autophagy, as well as the bupivacaine-attenuated NSCLC progression in the cells. Bupivacaine could inhibit the tumor growth in vivo. In conclusion, we discovered that the local anesthetic bupivacaine inhibited the progression of NSCLC by inducing autophagy through Akt/mTOR signaling. Our finding provides new insights into the mechanism by which bupivacaine attenuates the development of NSCLC. Bupivacaine may serve as a potential anti-tumor candidate for the therapeutic strategy of NSCLC.

Isolation and characterization of a novel Escherichia coli Kayfunavirus phage DY1.

Phage therapy has been revitalized since antibiotic resistance in bacteria is increasing. Compared with antibiotics, phages can target specific bacteria precisely, which requires more understanding of phage-host interactions by investigating different phages. Escherichia coli is a common pathogen with very high diversity. Based on the O antigens, E. coli can be classified into at least 183 serotypes and existing phages are far from being able to lyse all E. coli. Therefore, a novel phage specific to E. coli, named DY1, was identified and characterized to enhance our understanding of the phages of E. coli and expand the phage library. Phage DY1 belongs to the family Autographiviridae which is derived from Podoviridae. The genome of DY1 was determined to be 39,817 bp and comprises 54 putative open reading frames. Comparative genome and phylogenetic analysis demonstrated that DY1 was highly similar to phages belonging to the genus Kayfunavirus; however, the highest average nucleotide identity (ANI) values of DY1 with known phages was 0.82 suggesting that DY1 was a novel phage. Through stability tests, DY1 was very stable at temperatures ranging from 20 to 50 °C and pH levels from 5 to 11. Taken together, we report that phage DY1 is a novel Kayfunavirus phage with the potential for phage therapy.

Comparative Analyses of Mitochondrial Genomes Provide Evolutionary Insights Into Nematode-Trapping Fungi.

Predatory fungi in Orbiliaceae (Ascomycota) have evolved a diversity of trapping devices that enable them to trap and kill nematodes, other small animals, and protozoans. These trapping devices include adhesive hyphae, adhesive knobs, adhesive networks, constricting rings, and non-constricting rings. Their diversity and practical importance have attracted significant attention from biologists, making them excellent model organisms for studying adaptative evolution and as biological control agents against parasitic nematodes. The putative origins and evolutionary relationships among these carnivorous fungi have been investigated using nuclear protein-encoding genes, but their patterns of mitogenome relationships and divergences remain unknown. Here we analyze and compare the mitogenomes of 12 fungal strains belonging to eight species, including six species representing all four types of nematode trapping devices and two from related but non-predatory fungi. All 12 analyzed mitogenomes were of circular DNA molecules, with lengths ranging from 146,101 bp to 280,699 bp. Gene synteny analysis revealed several gene rearrangements and intron transfers among the mitogenomes. In addition, the number of protein coding genes (PCGs), GC content, AT skew, and GC skew varied among these mitogenomes. The increased number and total size of introns were the main contributors to the length differences among the mitogenomes. Phylogenetic analyses of the protein-coding genes indicated that mitochondrial and nuclear genomes evolved at different rates, and signals of positive selection were found in several genes involved in energy metabolism. Our study provides novel insights into the evolution of nematode-trapping fungi and shall facilitate further investigations of this ecologically and agriculturally important group of fungi.

Isolation and Characterization of Non-O157 Shiga Toxin-Producing Escherichia coli in Foods Sold at Retail Markets in China.

ABSTRACT: Non-O157 Shiga toxin-producing Escherichia coli (STEC) strains are significant foodborne pathogens that can cause acute diarrhea in humans. This study was conducted to investigate the contamination by non-O157 STEC in different types of food sold at retail markets in the People's Republic of China and to characterize non-O157 STEC strains. From May 2012 to April 2014, 1,200 retail food samples were collected from markets in 24 cities in China. Forty-four non-O157 isolates were recovered from 43 STEC-positive samples. Of the isolates, 22 and 19 carried the stx1 and stx2 genes, respectively, and 3 harbored both stx1 and stx2. stx1a and stx2a were the most prevalent stx subtypes. Other virulence genes, ent, hlyA, astA, eae, espB, iha, subAB, and tia, were commonly detected. Diverse O serogroups were identified among these isolates. Multilocus sequence typing indicated the high genetic diversity of the isolates. Thirty-two sequence types (STs) were identified among the 44 isolates, with ST383 (9.09%), ST134 (6.82%), and ST91 (6.82%) the most prevalent. Nine new STs were found. The isolates had a high prevalence of resistance to cephalothin, ampicillin, tetracycline, trimethoprim-sulfamethoxazole, nalidixic acid, streptomycin, and chloramphenicol. Twenty isolates (45.45%) were resistant to at least three antibiotics. This study provides updated surveillance data for non-O157 STEC isolates from foods sold at retail markets. Virulent and multidrug-resistant non-O57 STEC strains were isolated from all types of food. Our findings highlight the need for increased monitoring of non-O157 STEC in retail foods.

Prevalence and Characterization of Atypical Enteropathogenic Escherichia coli Isolated from Retail Foods in China.

Atypical enteropathogenic Escherichia coli (aEPEC) is an emerging pathogen that has been implicated in outbreaks of diarrhea worldwide. The objective of this study was to investigate the occurrence of aEPEC in retail foods at markets in the People's Republic of China and to characterize the isolates for virulence genes, intimin gene ( eae) subtypes, multilocus sequence types (STs), and antimicrobial susceptibility. From May 2014 to April 2015, 1,200 food samples were collected from retail markets in China, and 41 aEPEC isolates were detected in 2.75% (33 of 1,200) of the food samples. The virulence genes tir, katP, etpD, efa/lifA, ent, nleB, and nleE were commonly detected in these isolates. Nine eae subtypes were detected in the isolates, among which θ (23 isolates) and ß1 (6 isolates) were the most prevalent. The 41 isolates were divided into 27 STs by multilocus sequence typing. ST752 and ST10 were the most prevalent. Antibiotic susceptibility testing revealed high resistance among isolates to streptomycin (87.80%), cephalothin (73.16%), ampicillin (51.22%), tetracycline (63.42%), trimethoprim-sulfamethoxazole (43.90%), and kanamycin (43.90%). Thirty isolates (73.17%) were resistant to at least three antibiotics, and 20 (53.66 %) were resistant to five or more antibiotics. Our results suggest that retail foods in markets are important sources of aEPEC. The presence of virulent and multidrug-resistant aEPEC in retail foods poses a potential threat to consumers. Surveillance of aEPEC contamination and prudent use of antibiotics is strongly recommended in China.

Characterization of Extended-Spectrum ß-Lactamase-Producing Enterobacteriaceae From Retail Food in China.

In this study, we characterized the ß-lactamase genes and phenotypic resistance of cephalosporin-resistant Enterobacteriaceae isolated from retail foods in China. Of 1,024 Enterobacteriaceae isolates recovered from raw meat products, aquatic products, raw vegetables, retail-level ready-to-eat (RTE) foods, frozen foods, and mushrooms from 2011 to 2014, 164 (16.0%) showed cefotaxime (CTX) and/or ceftazidime (CAZ) cephalosporin resistance, and 96 (9.4%) showed the extended-spectrum ß-lactamase (ESBL) phenotype. More than 30% isolates were resistant to all antimicrobial agents except carbapenems (MEM 3.1% and IPM 5.2%), cefoxitin (FOX 6.3%), and amoxicillin/clavulanic acid (AMC 26%), and 94.8% of the strains were resistant to up to seven antibiotics. Polymerase chain reaction analysis showed that blaTEM (81.9%) was the most common gene, followed by blaCTX-M (68.1%) and blaSHV (38.9%). Moreover, 16.8% (72/429) of food samples contained ESBL-positive Enterobacteriaceae, with the following patterns: 32.9% (23/70) in frozen foods, 27.2% (5/29) in mushrooms, 17.6% (24/131) in raw meats, 13.3% (4/30) in fresh vegetables, 11.1% (8/72) in RTE foods, and 9.3% (9/97) in aquatic products. In addition, 24 of 217 foods collected in South China (11.1%), 25 of 131 foods collected in North of the Yangtze River region (19.1%), and 23 of 81 foods collected in South of the Yangtze River region (28.4%) were positive for ESBL- Enterobacteriaceae. Conjugation experiments demonstrated that the 22 of 72 isolates were transconjugants that had received the ß-lactamase gene and were resistant to ß-lactam antibiotics as well as some non-ß-lactam antibiotics. These findings demonstrated that retail foods may be reservoirs for the dissemination of ß-lactam antibiotics and that resistance genes could be transmitted to humans through the food chain; and the predominant ESBL-producing Enterobacteriaceae in China was isolated from in frozen chicken-meat, followed by frozen pork, cold noodles in sauce, cucumber, raw chicken meat, frozen pasta, brine-soaked chicken and tomato.

Phenotypic and Genotypic Characterization of Klebsiella pneumoniae Isolated From Retail Foods in China.

Klebsiella pneumoniae is not only a major hospital-acquired pathogen but also an important food-borne pathogen that can cause septicaemia, liver abscesses, and diarrhea in humans. The phenotypic and genotypic characteristics of K. pneumoniae in retail foods have not been thoroughly investigated in China. The objective of this study was to characterize K. pneumoniae isolates through biotyping, serotyping, determination of virulence factors, antibiotic resistance testing, enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR), and (GTG)5-PCR molecular typing. From May 2013 to April 2014, a total of 61 K. pneumoniae isolates were collected from retail foods in China. Using API 20E test strips, five different biotype profiles were identified among these isolates. The majority of isolates belonged to biochemical profile "5215773" (50 isolates, 80.6%). The capsular serotypes of the 61 K. pneumoniae isolates and one reference strain were determined by PCR. Of the seven capsular serotypes tested, four different capsular serotypes were identified. Serotypes K1, K20, K57, and K2 were detected in two, three, two, and one isolates, respectively. Serotypes K3, K5, and K54 were not detected. The presence of 11 virulence genes was assessed by PCR. The most common virulence genes were fimH (85.5%), ureA (79.0%), wabG (77.4%), uge (56.5%), and kfuBC (29.0%). ERIC-PCR and (GTG)5-PCR molecular typing indicated high genetic diversity among K. pneumoniae isolates. We identified 60 different ERIC patterns and 56 distinct (GTG)5 patterns. Genotypic results indicated that isolates carrying similar virulence factors were generally genetically related. Some isolates from the same geographic area have a closer relationship. The isolates showed high levels of resistance to ampicillin (51/62, 82.2%). Resistance to streptomycin (11/62, 17.7%) and piperacillin (10/62, 16.1%) was also common. The presence of virulent and antibiotic-resistant K. pneumoniae in foods poses a potential health hazard for consumers. Our findings highlight the importance of surveillance of K. pneumoniae in foods.

Antibiotic-Resistant Extended Spectrum ß-Lactamase- and Plasmid-Mediated AmpC-Producing Enterobacteriaceae Isolated from Retail Food Products and the Pearl River in Guangzhou, China.

We conducted a survey in 2015 to evaluate the presence of extended spectrum ß-lactamase (ESBL)- and plasmid-mediated AmpC-producing Enterobacteriaceae in retail food and water of the Pearl River in Guangzhou, China, as well as their antibiotic resistance profiles. Samples (88 fresh food samples and 43 water samples) from eight different districts were analyzed by direct plating and after enrichment. Multidrug-resistant strains were found in 41.7 and 43.4% of food and water samples, respectively. ESBLs were found in 3.4 and 11.6% of food and water samples, respectively, and AmpC producers were found in 13.6 and 16.3% of food and water samples, respectively. Molecular characterization revealed the domination of blaCTX-M genes; plasmidic AmpC was of the type DHA-1 both in food and water samples. Thirteen of Fifty one ß-lactamase-producing positive isolates were detected to be transconjugants, which readily received the ß-lactamase genes conferring resistance to ß-lactam antibiotics as well as some non-ß-lactam antibiotics. These findings provide evidence that retail food and the river water may be considered as reservoirs for the dissemination of ß-lactam antibiotics, and these resistance genes could readily be transmitted to humans through the food chain and water.

Prevalence, molecular characterization, and antibiotic susceptibility of Cronobacter spp. in Chinese ready-to-eat foods.

Cronobacter spp. are foodborne pathogens that cause rare but life-threatening diseases in neonates and infants; they can also cause disease in adults. Cronobacter spp. contamination of ready-to-eat (RTE) foods has been reported previously. However, to date, the prevalence and contamination levels of these bacteria in RTE foods in China have not yet been determined. Therefore, the aim of this study was to investigate the prevalence of Cronobacter spp. in RTE foods marketed in China. Two-hundred and eighty RTE food samples were collected from different producers and retailers and analyzed using quantitative methods. The isolates obtained were identified to the species level based on fusA sequences, and were subtyped using a PCR-based serotyping technique. Selected isolates were further characterized by multilocus sequence typing (MLST) and antimicrobial sensitivity determination. Of 280 samples tested, 52 (18.6%) were positive for Cronobacter spp. The contamination levels were less than 110 MPN/g for 78.8% (41/52) of the samples. The results of the O-antigen serotyping for 111 isolates showed that Cronobacter sakazakii serogroup O2 (28 isolates) was the most prevalent serotype. MLST analyses produced 41 sequence types (STs), including 20 novel STs. ST8 was the most prevalent ST (9 isolates) followed by ST4 (5 isolates). Antimicrobial sensitivity testing showed that 84.5% and 46.5% of the isolates were resistant to penicillin G and cephalothin, respectively; in contrast, all of the tested isolates were susceptible to cefotaxime, ciprofloxacin, tetracycline, and nalidixic acid. To the best of our knowledge, this is the first report on Cronobacter spp. prevalence in RTE foods in China, and the findings of our study nonetheless suggested that Cronobacter spp. contamination of Chinese RTE foods poses a potential risk for the consumer. Thus, the study highlights the significance of developing more effective control strategies during the manufacturing process.