scholarly journals Highly Sensitive and Specific Detection of Staphylococcal Enterotoxins SEA, SEG, SEH, and SEI by Immunoassay

Toxins ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 130
Author(s):  
Cécile Féraudet Tarisse ◽  
Céline Goulard-Huet ◽  
Yacine Nia ◽  
Karine Devilliers ◽  
Dominique Marcé ◽  
...  
Keyword(s):  
Food Poisoning ◽  
Cross Reactivity ◽  
Threat Detection ◽  
Enzyme Immunoassays ◽  
Staphylococcal Enterotoxins ◽  
Strain Characterization ◽  
Highly Sensitive ◽  
Genes Encoding ◽  
Contaminated Food ◽  
First Time

Staphylococcal food poisoning (SFP) is one of the most common foodborne diseases worldwide, resulting from the ingestion of staphylococcal enterotoxins (SEs), primarily SE type A (SEA), which is produced in food by enterotoxigenic strains of staphylococci, mainly S. aureus. Since newly identified SEs have been shown to have emetic properties and the genes encoding them have been found in food involved in poisoning outbreaks, it is necessary to have reliable tools to prove the presence of the toxins themselves, to clarify the role played by these non-classical SEs, and to precisely document SFP outbreaks. We have produced and characterized monoclonal antibodies directed specifically against SE type G, H or I (SEG, SEH or SEI respectively) or SEA. With these antibodies, we have developed, for each of these four targets, highly sensitive, specific, and reliable 3-h sandwich enzyme immunoassays that we evaluated for their suitability for SE detection in different matrices (bacterial cultures of S. aureus, contaminated food, human samples) for different purposes (strain characterization, food safety, biological threat detection, diagnosis). We also initiated and described for the first time the development of monoplex and quintuplex (SEA, SE type B (SEB), SEG, SEH, and SEI) lateral flow immunoassays for these new staphylococcal enterotoxins. The detection limits in buffer were under 10 pg/mL (0.4 pM) by enzyme immunoassays and at least 300 pg/mL (11 pM) by immunochromatography for all target toxins with no cross-reactivity observed. Spiking studies and/or bacterial supernatant analysis demonstrated the applicability of the developed methods, which could become reliable detection tools for the routine investigation of SEG, SEH, and SEI.

scholarly journals Prevalence and Molecular Characterization of Enterotoxin-Producing Strains of Staphylococcus Aureus Isolated from Serbian Dairy Cows

2016 ◽  
Vol 66 (4) ◽  
pp. 466-477 ◽  
Author(s):  
Marija Pajić ◽  
Stanko Boboš ◽  
Branko Velebit ◽  
Zoran Rašić ◽  
Vera Katić ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Protein A ◽  
Food Poisoning ◽  
Clinical Mastitis ◽  
Staphylococcal Protein A ◽  
Phylogenetic Groups ◽  
Gene Encoding ◽  
Staphylococcal Enterotoxins ◽  
Genes Encoding ◽  
The Republic

AbstractStaphylococcus aureus is known worldwide as a frequent cause of mastitis in dairy cattle. Due to the production of heath resistant enterotoxins, this pathogen is also a major cause of food poisoning among humans, with symptoms of often severe vomiting and diarrhea. The aim of our study was to determine the prevalence of enterotoxinproducing strains of S. aureus originating from samples of cows with subclinical and clinical mastitis in the Republic of Serbia. Furthermore, we analyzed the type of staphylococcal enterotoxin they produce and phylogenetic relatedness among the S. aureus isolates recovered from milk in this study. Production of staphylococcal enterotoxins A, B, C, D and E was determined by commercial immunoenzyme assay VIDAS® SET2, and presence of corresponding genes encoding enterotoxin synthesis in positive isolates confi rmed by Polymerase Chain Reaction. Enterotoxin production was determined in 5 out of 75 (6.67%) isolates of S. aureus and all of them produced staphylococcal enterotoxins C. After analyzing the nucleotide sequence of the gene encoding the synthesis of staphylococcal protein A, S. aureus isolates were assigned into 2 phylogenetic groups, including 7 clusters. All S. aureus isolates with the presence of sec gene formed one cluster even dough they originated from milk samples from different farms.

scholarly journals Production and Characterization of Antibodies against Each of the Three Subunits of the Bacillus cereus Nonhemolytic Enterotoxin Complex

2005 ◽  
Vol 71 (12) ◽  
pp. 8214-8220 ◽  
Author(s):  
Richard Dietrich ◽  
Maximilian Moravek ◽  
Christine Bürk ◽  
Per Einar Granum ◽  
Erwin Märtlbauer
Keyword(s):  
Bacillus Cereus ◽  
Rabbit Antiserum ◽  
Food Poisoning ◽  
Cross Reactivity ◽  
Terminal Part ◽  
Complete Set ◽  
Different Strains ◽  
First Time ◽  
Three Components

ABSTRACT The nonhemolytic enterotoxin (Nhe) is one of the two three-component enterotoxins which are responsible for diarrheal food poisoning syndrome caused by Bacillus cereus. To facilitate the detection of this toxin, consisting of the subunits NheA, NheB, and NheC, a complete set of high-affinity antibodies against each of the three components was established and characterized. A rabbit antiserum specific for the C-terminal part (15 amino acids) of NheC was produced using a respective synthetic peptide coupled to a protein carrier for immunization. Using purified B. cereus exoprotein preparations as immunogens, one monoclonal antibody against NheA and several antibodies against NheB were obtained. No cross-reactivity with other proteins produced by different strains of B. cereus was observed. Antibodies against the NheB component were able to neutralize the cytotoxic activity (up to 98%) of Nhe. Based on indirect enzyme immunoassays, the antibodies developed in this study were successfully used in the characterization of the enterotoxic activity of several B. cereus strains. For the first time, it could be shown that strains carrying the nhe genes usually express the complete set of the three components, including NheC. However, the amount of toxin produced varies considerably between the different strains.

Reproducibility Study for the Detection of Staphylococcal Enterotoxins in Dairy Products between Official Italian National Laboratories

2014 ◽  
Vol 77 (6) ◽  
pp. 999-1004 ◽  
Author(s):  
D. M. BIANCHI ◽  
F. INGRAVALLE ◽  
D. ADRIANO ◽  
S. GALLINA ◽  
M. GRAMAGLIA ◽  
...  
Keyword(s):  
Food Poisoning ◽  
Reference Laboratory ◽  
Potential Hazard ◽  
Consumer Health ◽  
National Network ◽  
Staphylococcal Enterotoxins ◽  
Reproducibility Study ◽  
Ring Trial ◽  
Contaminated Food ◽  
European Union Legislation

Staphylococcal food poisoning is a common foodborne disease caused by the ingestion of staphylococcal enterotoxins (SEs) produced mainly by enterotoxigenic strains of Staphylococcus aureus. To date, 21 SEs and/or enterotoxin-like types have been identified, several of which represent a potential hazard for consumers. To protect consumer health and to reduce the amount of SE-contaminated food entering the market, European Union legislation regulating food safety requires testing for SEs. The Italian National Reference Laboratory organized a ring trial to test technical and analytical proficiency in the national network of official food laboratories. Twenty-four laboratories took part, and each received and analyzed 24 blind dairy samples. Reproducibility of the results from the laboratories was assessed by the Cohen κ index, and accuracy (sensitivity and specificity) was evaluated according to the International Organization for Standardization definition (ISO 16140:2003). Trial results revealed partially satisfactory agreement: 254 of 276 possible paired participants (92%) reached a κ value >0.60, which is conventionally recognized as satisfactory. Accuracy was deemed satisfactory; 100% sensitivity and 100% specificity were achieved by 22 and 18 of the 24 laboratories, respectively.

Analysis of the VIDAS® Staph Enterotoxin III (SET3) for Detection of Staphylococcal Enterotoxins G, H, and I in Foods

2018 ◽  
Vol 101 (5) ◽  
pp. 1482-1489 ◽  
Author(s):  
Jennifer M Hait ◽  
Angela T Nguyen ◽  
Sandra M Tallent
Keyword(s):  
Detection Limit ◽  
Sensitivity And Specificity ◽  
Food Poisoning ◽  
Cross Reactivity ◽  
Ease Of Use ◽  
Aureus Strain ◽  
The Novel ◽  
Staphylococcal Enterotoxins ◽  
Concurrent Testing ◽  
Sensitivity Specificity

Abstract Background: Staphylococcal food poisoning (SFP) frequently causes illnesses worldwide. SFP occurs from the ingestion of staphylococcal enterotoxins (SEs) preformed in foods by enterotoxigenic strains of Staphylococcus species, primarily S. aureus. SEG, SEH, and SEI induce emesis and have been implicated in outbreaks. Immunological-based methods are deemed the most practical methods for the routine analysis of SEs in foods given their ease of use, sensitivity, specificity, and commercial availability. These kits are routinely used to test for SEA-SEE. However, only recently has a kit been developed to detect SEG, SEH, and SEI. Objective: Our research examined the performance of the novel VIDAS® Staph Enterotoxin III (SET3) for the detection of staphylococcal enterotoxins SEG, SEH, and SEI in foods. Methods: Here we assess the sensitivity and specificity of SET3 using duplicate test portions of six foods at varying concentrations of inclusivity and exclusivity inocula: pure SEG, SEH, SEI, S. aureus strain extracts positive for seg, seh, and sei, as well as SEA, SEB, SEC, SED, and SEE. Results: The overall detection limit was less than 2.09 ng/mL for foods inoculated with SEG, SEH, and SEI, with no cross reactivity observed. Highlights: Integrating concurrent testing to detect the presence of SEA–SEE and SEG–SEI utilizing the SET3 along with the VIDAS SET2, Ridascreen® SET total, or other comparable kits will be instrumental for the future food assessments in our laboratory and may become the new standard for SE analysis of foods.

scholarly journals Human Kallikrein 8: Immunoassay Development and Identification in Tissue Extracts and Biological Fluids

2003 ◽  
Vol 49 (1) ◽  
pp. 87-96 ◽  
Author(s):  
Tadaaki Kishi ◽  
Linda Grass ◽  
Antoninus Soosaipillai ◽  
Chigusa Shimizu-Okabe ◽  
Eleftherios P Diamandis
Keyword(s):  
Dynamic Range ◽  
Biological Fluids ◽  
Expression System ◽  
Baculovirus Expression System ◽  
Cross Reactivity ◽  
Tissue Extracts ◽  
Highly Sensitive ◽  
Sandwich Type ◽  
Chromatographic Procedure ◽  
First Time

Abstract Background: The serine protease human kallikrein 8 (hK8; neuropsin), a new member of the human kallikrein family, was predicted to be secreted; thus, it is expected to be present in biological fluids. The aim of this study was to develop a sensitive and specific immunoassay for hK8 (hK8-ELISA) and establish the distribution of hK8 in tissue extracts and biological fluids. Methods: Recombinant hK8 was produced in a baculovirus expression system and purified with a three-step chromatographic procedure. Purified hK8 was injected into mice and rabbits for antibody generation. A highly specific and sensitive sandwich-type immunoassay (ELISA) was developed using the rabbit and mouse antisera to hK8. The hK8-ELISA was then used to study the distribution of hK8 in various biological fluids and tissue extracts. Results: The dynamic range of the hK8-ELISA was 0.2 (detection limit) to 20 μg/L, and imprecision (CV) was <10% within this range. This hK8-ELISA was specific for hK8 and had no detectable cross-reactivity with other members of the human kallikrein family. With this assay, hK8 was detected in tissue extracts of esophagus (highest concentrations), skin, testis, tonsil, kidney, breast, and salivary gland and in the biological fluids breast milk (highest concentrations), amniotic fluid, seminal plasma, and serum. Furthermore, in some cancer cell lines, the concentration of hK8 was regulated by steroid hormones. Conclusions: We report for the first time production of recombinant hK8 protein, generation of antibodies, and development of a highly sensitive and specific immunoassay for quantification of hK8 in tissue extracts and biological fluids. This assay can be used to explore the potential of hK8 as a marker of cancer or other conditions.

scholarly journals Structure Revision of Isocereulide A, an Isoform of the Food Poisoning Emetic Bacillus cereus Toxin Cereulide

Molecules ◽  
2021 ◽  
Vol 26 (5) ◽  
pp. 1360
Author(s):  
Veronika Walser ◽  
Markus Kranzler ◽  
Monika Ehling-Schulz ◽  
Timo D. Stark ◽  
Thomas F. Hofmann
Keyword(s):  
Bacillus Cereus ◽  
Food Industry ◽  
Chemical Structure ◽  
Food Poisoning ◽  
2D Nmr ◽  
Food Samples ◽  
2D Nmr Spectroscopy ◽  
Safety Hazard ◽  
Contaminated Food ◽  
First Time

The emetic Bacillus cereus toxin cereulide presents an enormous safety hazard in the food industry, inducing emesis and nausea after the consumption of contaminated foods. Additional to cereulide itself, seven structurally related isoforms, namely the isocereulides A–G, have already been elucidated in their chemical structure and could further be identified in B. cereus contaminated food samples. The newly performed isolation of isocereulide A allowed, for the first time, 1D- and 2D-NMR spectroscopy of a biosynthetically produced isocereulide, revealing results that contradict previous assumptions of an l-O-Leu moiety within its chemical structure. By furthermore applying posthydrolytical dipeptide analysis, amino acid and α-hydroxy acid analysis by means of UPLC-ESI-TOF-MS, as well as MSn sequencing, the structure of previously reported isocereulide A could be corrected. Instead of the l-O-Leu as assumed to date, one l-O-Ile unit could be verified in the cyclic dodecadepsipeptide, revising the structure of isocereulide A to [(d-O-Leu-d-Ala-l-O-Val-l-Val)2(d-O-Leu-d-Ala-l-O-Ile-l-Val)].

scholarly journals Innovative Application of Mass Spectrometry for the Characterization of Staphylococcal Enterotoxins Involved in Food Poisoning Outbreaks

2008 ◽  
Vol 75 (3) ◽  
pp. 882-884 ◽  
Author(s):  
Jacques-Antoine Hennekinne ◽  
Virginie Brun ◽  
Marie-Laure De Buyser ◽  
Alain Dupuis ◽  
Annick Ostyn ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Food Poisoning ◽  
Quantitative Mass Spectrometry ◽  
Staphylococcal Enterotoxins ◽  
Common Food ◽  
Food Borne ◽  
First Time

ABSTRACT Staphylococcal poisoning is a common food-borne disease for which immunoassays to detect enterotoxins were developed, but these assays often lead to false diagnoses due to interferences or lack of specificity. Absolute quantitative mass spectrometry was for the first time successfully applied to an investigation of a staphylococcal outbreak due to coconut pearls.

scholarly journals The Assessment of IL-21 and IL-22 at the mRNA Level in Tumor Tissue and Protein Concentration in Serum and Peritoneal Fluid in Patients with Ovarian Cancer

2021 ◽  
Vol 10 (14) ◽  
pp. 3058
Author(s):  
Aleksandra Mielczarek-Palacz ◽  
Celina Kruszniewska-Rajs ◽  
Marta Smycz-Kubańska ◽  
Jarosław Strzelczyk ◽  
Wojciech Szanecki ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Peritoneal Fluid ◽  
Tumor Tissue ◽  
Mrna Level ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
First Time ◽  
Ovarian Serous Cancer

The aim of the analysis was for the first time to assess the expression of genes encoding IL-21 and IL-22 at the mRNA level in ovarian tumor specimens and the concentration of these parameters in serum and peritoneal fluid in patients with ovarian serous cancer. The levels of IL-21 and IL-22 transcripts were evaluated with the use of the real-time RT-qPCR. Enzyme-linked immunosorbent assay (ELISA) was used to determine the concentration of proteins. Quantitative analysis of IL-21 gene mRNA in the tumor tissue showed the highest activity in the G1 degree of histopathological differentiation and was higher in G1 compared to the control group. The concentration of IL-21 and IL-22 in the serum and in the peritoneal fluid of women with ovarian cancer varied depending on the degree of histopathological differentiation of the cancer and showed statistical variability compared to controls. The conducted studies have shown that the local and systemic changes in the immune system involving IL-21 and IL-22 indicate the participation of these parameters in the pathogenesis of ovarian cancer, and modulation in the IL-21/IL-22 system may prove useful in the development of new diagnostic and therapeutic strategies used in patients, which require further research.

scholarly journals Designing P. aeruginosa synthetic phages with reduced genomes

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Diana P. Pires ◽  
Rodrigo Monteiro ◽  
Dalila Mil-Homens ◽  
Arsénio Fialho ◽  
Timothy K. Lu ◽  
...  
Keyword(s):  
Galleria Mellonella ◽  
Antibacterial Properties ◽  
Genetically Engineered ◽  
In Vivo Studies ◽  
Infection Model ◽  
Promising Therapeutic Approach ◽  
Genes Encoding ◽  
First Time

AbstractIn the era where antibiotic resistance is considered one of the major worldwide concerns, bacteriophages have emerged as a promising therapeutic approach to deal with this problem. Genetically engineered bacteriophages can enable enhanced anti-bacterial functionalities, but require cloning additional genes into the phage genomes, which might be challenging due to the DNA encapsulation capacity of a phage. To tackle this issue, we designed and assembled for the first time synthetic phages with smaller genomes by knocking out up to 48% of the genes encoding hypothetical proteins from the genome of the newly isolated Pseudomonas aeruginosa phage vB_PaeP_PE3. The antibacterial efficacy of the wild-type and the synthetic phages was assessed in vitro as well as in vivo using a Galleria mellonella infection model. Overall, both in vitro and in vivo studies revealed that the knock-outs made in phage genome do not impair the antibacterial properties of the synthetic phages, indicating that this could be a good strategy to clear space from phage genomes in order to enable the introduction of other genes of interest that can potentiate the future treatment of P. aeruginosa infections.

scholarly journals Evaluation of mobile real-time polymerase chain reaction tests for the detection of severe acute respiratory syndrome coronavirus 2

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Chukwunonso Onyilagha ◽  
Henna Mistry ◽  
Peter Marszal ◽  
Mathieu Pinette ◽  
Darwyn Kobasa ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Real Time ◽  
Point Of Care ◽  
Middle Eastern ◽  
Turnaround Time ◽  
Cross Reactivity ◽  
Clinical Samples ◽  
Diarrhea Virus ◽  
Highly Sensitive ◽  
Transmissible Gastroenteritis

AbstractThe coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), calls for prompt and accurate diagnosis and rapid turnaround time for test results to limit transmission. Here, we evaluated two independent molecular assays, the Biomeme SARS-CoV-2 test, and the Precision Biomonitoring TripleLock SARS-CoV-2 test on a field-deployable point-of-care real-time PCR instrument, Franklin three9, in combination with Biomeme M1 Sample Prep Cartridge Kit for RNA 2.0 (M1) manual extraction system for rapid, specific, and sensitive detection of SARS-COV-2 in cell culture, human, and animal clinical samples. The Biomeme SARS-CoV-2 assay, which simultaneously detects two viral targets, the orf1ab and S genes, and the Precision Biomonitoring TripleLock SARS-CoV-2 assay that targets the 5′ untranslated region (5′ UTR) and the envelope (E) gene of SARS-CoV-2 were highly sensitive and detected as low as 15 SARS-CoV-2 genome copies per reaction. In addition, the two assays were specific and showed no cross-reactivity with Middle Eastern respiratory syndrome coronavirus (MERS-CoV), infectious bronchitis virus (IBV), porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis (TGE) virus, and other common human respiratory viruses and bacterial pathogens. Also, both assays were highly reproducible across different operators and instruments. When used to test animal samples, both assays equally detected SARS-CoV-2 genetic materials in the swabs from SARS-CoV-2-infected hamsters. The M1 lysis buffer completely inactivated SARS-CoV-2 within 10 min at room temperature enabling safe handling of clinical samples. Collectively, these results show that the Biomeme and Precision Biomonitoring TripleLock SARS-CoV-2 mobile testing platforms could reliably and promptly detect SARS-CoV-2 in both human and animal clinical samples in approximately an hour and can be used in remote areas or health care settings not traditionally serviced by a microbiology laboratory.

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