Toxin gene profiling of enterotoxic and emetic Bacillus cereus (original) (raw)
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Bacillus cereus is an opportunistic Food borne Pathogen that contaminates a wide variety of foods. There are mainly two types of illness caused by B. cereus in human beings namely diarrhoeal and emetic types. The emetic type is due to a single heat-stable toxin, and the diarrheal type to 3 or 4 heat-instable enterotoxins. In this research work, B. cereus spp were isolated from fermented rice noodle (Myanmar Traditional Food). For emetic toxin production, toxin of three isolated B. cereus were extracted by methanol extraction and subjected to LC-MS. Three isolated strains were genetically characterized by using gene specific primer for enterotoxin production. None of these strains showed emetic toxin production. But, all three strains tested had all the components of Haemolytic BL toxin (HBL), a haemolytic enterotoxin complex made of three proteins and Non Haemolytic Enterotoxin (NHE), a non-haemolytic enterotoxin complex made of three proteins and enterotoxin T genes. In addition, genes for the cytotoxin K were also found in two isolates, one isolate (E4) showed negative. From these results, the three isolates may be virulence because they have all HBL genes and all NHE genes. So, these strains seem to be enterotoxin producing strains.
Identification of emetic toxin producingBacillus cereusstrains by a novel molecular assay
Fems Microbiology Letters, 2004
Bacillus cereus causes two types of gastrointestinal diseases: emesis and diarrhea. The emetic type of the disease is attributed to the heat-stable depsipeptide cereulide and symptoms resemble Staphylococcus aureus intoxication, but there is no rapid method available to detect B. cereus strains causing this type of disease. In this study, a polymerase chain reaction (PCR) fragment of unknown function was identified, which was shown to be specific for emetic toxin producing strains of B. cereus. The sequence of this amplicon was determined and a PCR assay was developed on this basis. One hundred B. cereus isolates obtained from different food poisoning outbreaks and diverse food sources from various geographical locations and 29 strains from other species belonging to the B. cereus group were tested by this assay. In addition, 49 non-B. cereus group strains, with special emphasis on food pathogens, were used to show that the assay is specific for emetic toxin producing B. cereus strains. The presented PCR assay is the first molecular tool for the rapid detection of emetic toxin producing B. cereus strains.
The Food Poisoning Toxins of Bacillus cereus
Toxins
Bacillus cereus is a ubiquitous soil bacterium responsible for two types of food-associated gastrointestinal diseases. While the emetic type, a food intoxication, manifests in nausea and vomiting, food infections with enteropathogenic strains cause diarrhea and abdominal pain. Causative toxins are the cyclic dodecadepsipeptide cereulide, and the proteinaceous enterotoxins hemolysin BL (Hbl), nonhemolytic enterotoxin (Nhe) and cytotoxin K (CytK), respectively. This review covers the current knowledge on distribution and genetic organization of the toxin genes, as well as mechanisms of enterotoxin gene regulation and toxin secretion. In this context, the exceptionally high variability of toxin production between single strains is highlighted. In addition, the mode of action of the pore-forming enterotoxins and their effect on target cells is described in detail. The main focus of this review are the two tripartite enterotoxin complexes Hbl and Nhe, but the latest findings on cereulide...
Identification of emetic toxin producing Bacillus cereus strains by a novel molecular assay
FEMS microbiology …, 2004
Bacillus cereus causes two types of gastrointestinal diseases: emesis and diarrhea. The emetic type of the disease is attributed to the heat-stable depsipeptide cereulide and symptoms resemble Staphylococcus aureus intoxication, but there is no rapid method available to detect B. cereus strains causing this type of disease. In this study, a polymerase chain reaction (PCR) fragment of unknown function was identified, which was shown to be specific for emetic toxin producing strains of B. cereus. The sequence of this amplicon was determined and a PCR assay was developed on this basis. One hundred B. cereus isolates obtained from different food poisoning outbreaks and diverse food sources from various geographical locations and 29 strains from other species belonging to the B. cereus group were tested by this assay. In addition, 49 non-B. cereus group strains, with special emphasis on food pathogens, were used to show that the assay is specific for emetic toxin producing B. cereus strains. The presented PCR assay is the first molecular tool for the rapid detection of emetic toxin producing B. cereus strains.
Multiple-Locus Sequence Typing and Analysis of Toxin Genes in Bacillus cereus Food-Borne Isolates
Applied and Environmental Microbiology, 2008
In the present study we characterized 47 food-borne isolates of Bacillus cereus using multilocus sequence typing (MLST). Newly determined sequences were combined with sequences available in public data banks in order to produce the largest data set possible. Phylogenetic analysis was performed on a total of 296 strains for which MLST sequence information is available, and three main lineages-I, II, and III-within the B. cereus complex were identified. With few exceptions, all food-borne isolates were in group I. The occurrence of horizontal gene transfer (HGT) among various strains was analyzed by several statistical methods, providing evidence of widespread lateral gene transfer within B. cereus. We also investigated the occurrence of toxinencoding genes, focusing on their evolutionary history within B. cereus. Several patterns were identified, indicating a pivotal role of HGT in the evolution of toxin-encoding genes. Our results indicate that HGT is an important element in shaping the population structure of the B. cereus complex. The results presented here also provide strong evidence of reticulate evolution within the B. cereus complex.
Journal of Applied and Natural Science, 2022
Bacillus cereus produces pore-forming toxins responsible for diarrhoea; therefore, rapidly detecting these toxins in food retailed for consumption is needed. The genomic DNA of 100 B. cereus isolates recovered from some retailed foods was extracted and used as a template for enterotoxin detection. The detection of genes of non-haemolyticnonhemolytic enterotoxin (nheA, nheB, nheC), hemolysin BL (hblA, hblC, hblD), entFM, cytK and bceT by the isolates was carried out with PCR using primers specific for the targeted genes, while the production of Nhe and Hbl enterotoxins in fifty of the randomly chosen isolates was detected with a Duopath Cereus Enterotoxin kit. Ninety-five percent of the isolates carried one or more components of the NHE complex, while 56% had one or more components of HBL. Sixteen out of the 100 isolates carried all the genes for NHE and HBL complex genes. The entFM, cytK and bceT genes were detected in 85%, 74% and 60% of B. cereus isolates, respectively. Starchy f...
Molecular Characterization and Enterotoxin Genes Typing of Local Strains of Bacillus Cereus
2009
Abstract: Two virulent strains of Bacillus cereus coded as BC13 and BC37 isolated from corn snacks collected from Egyptian market were identified using biochemical and staining methods. Further confirmation was done by determining their cellular protein pattern compared to a standard culture of B. cereus NRRL 569. The cellular protein profile revealed 97.55 to 99% similarity between NRRL 569 reference strain and local strains (BC13, BC37). Random amplification polymerase DNA analysis of the two tested isolates compared to standard culture using eleven arbitrary primers showed similarities ranging from 75.5% to 77.09% between the tested isolates. Separation of extracellular proteins of both tested isolates using SDS-PAGE revealed the presence of protein bands with molecular weights between 34 and 54 kDa in both tested isolates, suspected as enterotoxins. To ensure the presence of suspected enterotoxins, two pairs of primers newly designed and reported during year 2008 (FHblC and RHbl...
Determination of the toxic potential ofBacillus cereusisolates by quantitative enterotoxin analyses
Fems Microbiology Letters, 2006
Haemolysin BL (HBL) and nonhaemolytic enterotoxin (Nhe), each consisting of three components, represent the major enterotoxins produced by Bacillus cereus. To evaluate the expression of these toxins, a set of 100 B. cereus strains was examined. Molecular biological characterization showed that 42% of the strains harboured the genes for HBL and 99% for Nhe. The production of all Nhe and HBL components were analyzed using specific antibodies and, in culture supernatants, detectable levels of HBL and Nhe were found for 100% of hbl-positive and 96% of nhe-positive strains. The concentrations of the HBL-L 2 and NheB component ranged from 0.02 to 5.6 mg mL À1 and from 0.03 to 14.2 mg mL À1 , respectively. Comparison of the amount of NheB produced by food poisoning and food/environmental strains revealed that the median value for all food poisoning strains was significantly higher than for the food/environmental isolates. The data presented in this study provide evidence that specific and quantitative determination of the enterotoxins is necessary to evaluate the toxic potential of B. cereus. In particular, the level of Nhe seems to explain most of the cytotoxic activity of B. cereus isolates and may indicate a highly diarrheic potential.
International Journal of Food Microbiology, 2011
The major objectives of this study were to determine the incidence, diversity and characteristics of Bacillus cereus group spp. isolated from food products marketed in Belgium. The food products investigated in this study included cooked pasta, lasagna, béchamel sauce, bolognaise sauce, fresh minced beef, fresh-cut vegetables and raw basmati rice. B. cereus group spp. were detected in 56.3% (324 of 575) of the samples giving rise to 380 strains. The highest incidence (100%) occurred in the raw basmati rice. Although only 10 (2.6%) of the 380 isolates were determined to be psychrotolerant (able to grow at ≤ 7°C), 25 (6.2%), 189 (49.7%) and 334 (87.9%) isolates were able to grow at mild temperature abuse conditions of 8°C, 9°C and 10°C, respectively. The large diversity of the isolates obtained (overall and between isolates obtained from the same product type) was highlighted by the results of the (GTG) 5 PCR fingerprinting of 80 selected isolates. Sixty-one of these 80 isolates belonged to 15 distinct clusters (≥ 85% Pearson correlation) whereas the remaining 19 were each clustered separately. Further diversity was also found in the distribution of toxin genes as 16 different profiles were observed in the 80 selected isolates. Whilst none of 80 selected strains harboured the ces gene required for the production of the emetic toxin cereulide, 42 strains (52.5%) carried all seven genes required for the production of the diarrhoeal enterotoxins: haemolytic BL, non-haemolytic enterotoxin and cytotoxin K. The results of this study highlight not only the omnipresence but also the highly diverse ecology of B. cereus spp. within and across several food product types available on the retail market in Belgium. They should also provide the impetus for more studies to enable detailed risk assessment studies to be performed.