Impact of Acid on Survival of Vibrio vulnificus and Vibrio vulnificus Phage†

2000 ◽  
Vol 63 (8) ◽  
pp. 1049-1052 ◽  
Author(s):  
JAHEON KOO ◽  
ANGELO DePAOLA ◽  
DOUGLAS L. MARSHALL
Keyword(s):  
Vibrio Vulnificus ◽  
Limit Of Detection ◽  
Host Strain ◽  
Acid Sensitivity ◽  
Phage Strain ◽  
D Values

Three strains of Vibrio vulnificus and V. vulnificus phages were tested for acid sensitivity at 21°C. V. vulnificus strain 304 was more resistant to pH 4.0 than strains CVD-1 and A-9, whereas acid sensitivities of V. vulnificus strains at pH 3.0 and 2.0 were similar. V. vulnificus phage strain 110A-7 was more resistant to pH 4.0 than strain 153A-7, whereas acid sensitivities of phage strains at pH 3.5 and 3.0 were similar. Numbers of V. vulnificus and its phage were close to the limit of detection after 100 s at pH 2.0 and after 24 min at pH 3.0. Acid D-values at 21°C decreased as pH decreased for both V. vulnificus and phages. D-values of phage strains at pH 3.5 were 10-fold greater than those of host strain at pH 4.0. D-values of phage strains were slightly greater than those of host strain at pH 3.0. These results suggest that V. vulnificus and its phage were very sensitive to pH of less than 3.0, although V. vulnificus phages were more resistant to acid than their host.

Effect of Simulated Gastric Fluid and Bile on Survival of Vibrio vulnificus and Vibrio vulnificus Phage†

2000 ◽  
Vol 63 (12) ◽  
pp. 1665-1669 ◽  
Author(s):  
JAHEON KOO ◽  
ANGELO DePAOLA ◽  
DOUGLAS L. MARSHALL
Keyword(s):  
Small Intestine ◽  
Vibrio Vulnificus ◽  
Gastric Fluid ◽  
Simulated Gastric Fluid ◽  
D Values ◽  
Survival Differences ◽  
Phosphate Buffered Saline

Bacteria and phages may be exposed to acid conditions in the stomach and to bile in the intestine. Survival of three strains of Vibrio vulnificus and three strains of its phages was examined at 37°C after exposure to simulated gastric fluid at pH 3 to 4 or to 0, 1, and 2% bile in broth or buffer. Mean D-values (decimal reduction times) at pH 4 and 3 were 3.3 and 1.3 min for V. vulnificus and 97.8 and 0.7 min for its phages. No V. vulnificus survivors were found at pH 2.0. There were few survival differences among strains of V. vulnificus or its phages. Numbers of V. vulnificus increased 1 log in tryptic soy broth containing 1 or 2% bile after 3 h. Numbers of V. vulnificus and its phages remained constant in phosphate-buffered saline regardless of bile concentrations up to 3 h. Those V. vulnificus bacteria and phages that survive stomach acidity may proliferate in the small intestine, since they are resistant to bile.

scholarly journals Rapid and Sensitive Detection of Vibrio vulnificus Using CRISPR/Cas12a Combined With a Recombinase-Aided Amplification Assay

2021 ◽  
Vol 12 ◽  
Author(s):  
Xingxing Xiao ◽  
Ziqin Lin ◽  
Xianhui Huang ◽  
Jinfang Lu ◽  
Yan Zhou ◽  
...  
Keyword(s):  
Vibrio Vulnificus ◽  
Detection Method ◽  
Limit Of Detection ◽  
Sensitive Detection ◽  
Detection Methods ◽  
Farmed Fish ◽  
Safety Control ◽  
Amplification Assay ◽  
Sophisticated Instrument ◽  
Aquatic Pathogen

Vibrio vulnificus is an important zoonotic and aquatic pathogen and can cause vibriosis in humans and aquatic animals (especially farmed fish and shrimp species). Rapid and sensitive detection methods for V. vulnificus are still required to diagnose human vibriosis early and reduce aquaculture losses. Herein, we developed a rapid and sensitive diagnostic method comprising a recombinase-aided amplification (RAA) assay and the CRISPR/Cas12a system (named RAA-CRISPR/Cas12a) to detect V. vulnificus. The RAA-CRISPR/Cas12a method allows rapid and sensitive detection of V. vulnificus in 40 min without a sophisticated instrument, and the limit of detection is two copies of V. vulnificus genomic DNA per reaction. Meanwhile, the method shows satisfactory specificity toward non-target bacteria and high accuracy in the spiked blood, stool, and shrimp samples. Therefore, our proposed rapid and sensitive V. vulnificus detection method, RAA-CRISPR/Cas12a, has great potential for early diagnosis of human vibriosis and on-site V. vulnificus detection in aquaculture and food safety control.

Use of a Mutant Strain for Evaluating Processing Strategies to Inactivate Vibrio vulnificus in Oysters

1999 ◽  
Vol 62 (6) ◽  
pp. 592-600 ◽  
Author(s):  
CAROL S. DOMBROSKI ◽  
LEE-ANN JAYKUS ◽  
DAVID P. GREEN ◽  
BRIAN E. FARKAS
Keyword(s):  
Thermal Inactivation ◽  
Vibrio Vulnificus ◽  
Disease Risk ◽  
Capillary Tube ◽  
Control Strategies ◽  
Most Probable Number ◽  
Probable Number ◽  
Pure Cultures ◽  
Selective Plating ◽  
D Values

Vibrio vulnificus is a ubiquitous marine bacterium frequently isolated from shellfish and associated with severe and often fatal disease in humans. Various control strategies to reduce the disease risk associated with V. vulnificus contamination in shellfish have been proposed. However, evaluating the efficacy of these control strategies is complicated because of the difficulty in distinguishing V. vulnificus from the high levels of background environmental Vibrio spp. The purpose of this research was to develop a model indicator V. vulnificus strain that could be readily differentiated from background microflora and used to facilitate the evaluation of processing efficacy. A spontaneous nalidixic acid–resistant strain of V. vulnificus (Vv-NA) was prepared from a wild-type parent (Vv-WT) using selective plating techniques. Vv-NA was very similar to Vv-WT with respect to biochemical characteristics, appearance on selective plating media, detection limits using most probable number and polymerase chain reaction, and growth rate. In comparative freeze inactivation studies on pure cultures, Vv-WT and Vv-NA had similar freeze inactivation profiles at −20°C (conventional freezing), at −85°C (cold blast freezing), and in liquid nitrogen (cryogenic freezing). In oyster homogenates artificially inoculated with Vv-NA, the organism was inactivated 95 to 99% after freezing, irrespective of freezing temperature. Thermal inactivation comparisons of pure cultures of Vv-WT and Vv-NA using the capillary tube method revealed statistically significant differences in D values at 47°C (2.2 versus 3.0 min, respectively) and 50°C (0.83 versus 0.56 min, respectively), but nearly identical values at 52°C (0.21 versus 0.22 min, respectively). However, these D values were notably higher than those reported by other investigators and hence provided a conservative means by which to evaluate thermal inactivation. In oyster homogenates seeded with Vv-NA, D values of 1.3 ± 0.09 min and 0.41 ± 0.01 min were obtained at 46°C and 48°C, respectively. This study demonstrated that Vv-NA is readily enumerated and could be used as a surrogate for evaluating the degree of V. vulnificus inactivation provided by freezing and thermal treatments of oyster homogenates.

Effects of Desiccation Practices of Cultured Atlantic Oysters (Crassostrea virginica) on Vibrio spp. in Portersville Bay, Alabama, USA

2017 ◽  
Vol 80 (8) ◽  
pp. 1280-1287 ◽  
Author(s):  
Stephanie M. Grodeska ◽  
Jessica L. Jones ◽  
Covadonga R. Arias ◽  
William C. Walton
Keyword(s):  
Gulf Of Mexico ◽  
Vibrio Vulnificus ◽  
Limit Of Detection ◽  
Ambient Air ◽  
Most Probable Number ◽  
Air Exposure ◽  
Probable Number ◽  
Background Levels ◽  
And Control ◽  
Vibrio Spp

ABSTRACT The expansion of off-bottom aquaculture to the Gulf of Mexico has raised public health concerns for human health officials. High temperatures in the Gulf of Mexico are associated with high levels of Vibrio parahaemolyticus and Vibrio vulnificus. Routine desiccation practices associated with off-bottom aquaculture expose oysters to ambient air, allowing Vibrio spp. to proliferate in the closed oyster. Currently, there is limited research on the length of time needed for Vibrio spp. levels in desiccated oysters to return to background levels, defined as the levels found in oysters that remain continually submersed and not exposed to ambient air. This study determined the time needed to return V. parahaemolyticus, V. vulnificus, and Vibrio cholerae levels to background levels in oysters exposed to the following desiccation practices: 3-h freshwater dip followed by 24-h ambient air exposure, 27-h ambient air exposure, and control. All oysters were submerged at least 2 weeks prior to the beginning of each trial, with the control samples remaining submerged for the duration of each trial. Vibrio spp. levels were enumerated from samples collected on days 0, 1, 2, 3, 7, 10, and 14 after resubmersion using a three-tube most-probable-number enrichment followed by BAX PCR. V. cholerae levels were frequently (92%) below the limit of detection at all times, so they were not statistically analyzed. V. parahaemolyticus and V. vulnificus levels in the 27-h ambient air exposure and the 3-h freshwater dip followed by 24-h ambient air exposure samples were significantly elevated compared with background samples. In most cases, the Vibrio spp. levels in oysters in both desiccation treatments remained elevated compared with background levels until 2 or 3 days post-resubmersion. However, there was one trial in which the Vibrio spp. levels did not return to background levels until day 7. The results of this study provide scientific support that oyster farmers should be required to implement a minimum 7-day resubmersion regimen. This length of time allowed the Vibrio spp. levels to become not significantly different across all treatments.

scholarly journals Validation of a Green Fluorescent Protein-Labeled Strain of Vibrio vulnificus for Use in the Evaluation of Postharvest Strategies for Handling of Raw Oysters

2006 ◽  
Vol 72 (11) ◽  
pp. 7205-7211 ◽  
Author(s):  
S. L. Drake ◽  
D. Elhanafi ◽  
W. Bang ◽  
M. A. Drake ◽  
D. P. Green ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Vibrio Vulnificus ◽  
Limit Of Detection ◽  
Acid Tolerance ◽  
Refrigerated Storage ◽  
Population Declines ◽  
Wild Type ◽  
Sodium Pyruvate ◽  
Green Fluorescent

ABSTRACT In this paper we describe a biological indicator which can be used to study the behavior of Vibrio vulnificus, an important molluscan shellfish-associated human pathogen. A V. vulnificus ATCC 27562 derivative that expresses green fluorescent protein (GFP) and kanamycin resistance was constructed using conjugation. Strain validation was performed by comparing the GFP-expressing strain (Vv-GFP) and the wild-type strain (Vv-WT) with respect to growth characteristics, heat tolerance (45°C), freeze-thaw tolerance (−20o and −80°C), acid tolerance (pH 5.0, 4.0, and 3.5), cold storage tolerance (5°C), cold adaptation (15°C), and response to starvation. Levels of recovery were evaluated using nonselective medium (tryptic soy agar containing 2% NaCl) with and without sodium pyruvate. The indicator strain was subsequently used to evaluate the survival of V. vulnificus in oysters exposed to organic acids (citric and acetic acids) and various cooling regimens. In most cases, Vv-GFP was comparable to Vv-WT with respect to growth and survival upon exposure to various biological stressors; when differences between the GFP-expressing and parent strains occurred, they usually disappeared when sodium pyruvate was added to media. When V. vulnificus was inoculated into shellstock oysters, the counts dropped 2 log10 after 11 to 12 days of refrigerated storage, regardless of the way in which the oysters were initially cooled. Steeper population declines after 12 days of refrigerated storage were observed for both iced and refrigerated products than for slowly cooled product and product held under conservative harvest conditions. By the end of the refrigeration storage study (22 days), the counts of Vv-GFP in iced and refrigerated oysters had reached the limit of detection (102 CFU/oyster), but slowly cooled oysters and oysters stored under conservative harvest conditions still contained approximately 103 and >104 CFU V. vulnificus/oyster by day 22, respectively. The Vv-GFP levels in the oyster meat remained stable for up to 24 h when the meat was exposed to acidic conditions at various pH values. Ease of detection and comparability to the wild-type parent make Vv-GFP a good candidate for use in studying the behavior of V. vulnificus upon exposure to sublethal stressors that might be encountered during postharvest handling of molluscan shellfish.

Machinery Mold (Galactomyces geotrichum) Survival following Thermal and Hydrostatic Pressure Processing

2019 ◽  
Vol 82 (6) ◽  
pp. 1034-1038 ◽  
Author(s):  
SHIYU CAI ◽  
ABIGAIL B. SNYDER
Keyword(s):  
Limit Of Detection ◽  
Statistical Significance ◽  
Bacterial Pathogen ◽  
High Pressure Processing ◽  
Treatment Level ◽  
Production Environment ◽  
Log Reduction ◽  
Vegetable Products ◽  
D Values ◽  
D Value

ABSTRACT Machinery mold is a foodborne fungus associated with spoilage of fruit and vegetable products, as well as colonization of the food production environment. Galactomyces spp. are not considered heat-resistant molds, although their sensitivity to mild heat treatments, like blanching and pasteurization, as well as to nonthermal treatments, like high pressure processing (HPP), remains unknown. Cultures of two Galactomyces spp. isolates, Galactomyces geotrichum and Galactomyces candidum, were heat treated at 50, 53, and 55°C and HPP treated at 4°C for 90 s at 300, 400, and 600 MPa. The 2-day-old G. geotrichum had a D-value of 0.99 ± 0.04 min and 28-day-old G. geotrichum had a D-value of 1.28 ± 0.07 min at 55°C. Meanwhile, 2-day-old G. candidum had a D-value of 3.13 ± 0.20 min and 28-day-old G. candidum had a D-value of 1.60 ± 0.01 min at 55°C. Overall, the significant differences in D-values between the two cultivation times were modest, and statistical significance was not consistent across all three temperatures. However, the differences were more pronounced at higher processing temperatures. G. geotrichum was only reduced by 0.32 ± 0.07 log CFU/mL after 90 s at 300 MPa. However, increasing the treatment level to 400 MPa for 90 s decreased the counts for G. geotrichum to below the limit of detection (<1 log CFU/mL) following HPP treatment. Meanwhile, the counts for G. candidum survivors at 400 MPa were 1.36 ± 0.08 log CFU/mL and were under the limit of detection at 600 MPa for 90 s. Based on the findings in this study, machinery mold contaminants would be readily inactivated by the processing conditions used to target a 5-log reduction in the bacterial pathogen of concern. HIGHLIGHTS

scholarly journals Detection of Vibrio vulnificus in Seafood With a DNAzyme-Based Biosensor

2021 ◽  
Vol 12 ◽  
Author(s):  
Shihui Fan ◽  
Chao Ma ◽  
Xiaopeng Tian ◽  
Xiaoyi Ma ◽  
Mingcan Qin ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Vibrio Vulnificus ◽  
Detection Method ◽  
Limit Of Detection ◽  
Fluorescent Sensor ◽  
Specific Detection ◽  
Aquatic Products ◽  
Specificity And Sensitivity ◽  
Marine Products

Vibrio vulnificus is an important pathogenic bacterium that is often associated with seafood-borne illnesses. Therefore, to detect this pathogen in aquatic products, a DNAzyme-based fluorescent sensor was developed for the in vitro detection of V. vulnificus. After screening and mutation, a DNAzyme that we denominated “RFD-VV-M2” exhibited the highest activity, specificity, and sensitivity. The limit of detection was 2.2 × 103 CFU/ml, and results could be obtained within 5–10 min. Our findings suggested that the target of DNAzyme RFD-VV-M2 was a protein with a molecular weight between 50 and 100 kDa. The proposed biosensor exhibited an excellent capacity to detect marine products contaminated with V. vulnificus. Therefore, our study established a rapid, simple, sensitive, and highly specific detection method for V. vulnificus in aquatic products.

Influence of corneal biomechanical properties on IOP indices in patients with keratoconus

The Eye ◽  
2019 ◽  
Vol 21 (128) ◽  
pp. 15-19
Author(s):  
Irina Bubnova ◽  
Veronica Averich ◽  
Elena Belousova
Keyword(s):  
Disease Progression ◽  
Corneal Thickness ◽  
Central Zone ◽  
Biomechanical Properties ◽  
Resistance Factor ◽  
Corneal Resistance Factor ◽  
Mean Values ◽  
Coefficient Of Elasticity ◽  
D Values ◽  
Corneal Biomechanical Properties

Purpose: Evaluation of corneal biomechanical prop¬erties and their influence on IOP indices in patients with keratoconus. Material and methods. The study included 194 eyes with keratoconus (113 patients aged from 23 to 36 years old). Corneal refraction in central zone varied from 48.25 to 56.75 D, values of corneal thickness ranged from 279 to 558 μm. Patients were divided into 4 groups according to Amsler classification: I stage – 40 eyes; II stage – 78 eyes; III stage – 54 eyes and IV stage – 22 eyes. Standard ophthal¬mological examination was carried out including pneumo¬tonometry. IOP indices and values of biomechanical prop¬erties were evaluated by dynamic bidirectional pneumatic applanation and pneumatic impression. Results. Study of corneal biomechanical properties in patients with keratoconus showed a decrease of such biomechanical indices as corneal hysteresis (CH) on aver¬age to 8.42±1.12 mm Hg, corneal resistance factor (CRF) – to 7.45±0.96 mm Hg, coefficient of elasticity (CE) – 5.35± 0.87 mm Hg. Values of these indices strongly depended on the stage of keratoconus. In the whole sample, the aver¬age corneal compensated IOP (IOPcc) amounted to 15.08± 2.43 mm Hg, Goldman IOP (IOPg) was 11.61±2.37 mm Hg and pneumatic tonometry IOP (IOPp) was 10.13±2.94 mm Hg. IOPcc indices didn’t have any statistically significant differ¬ence in dependence on the stage of keratoconus (р>0.473), while in process of disease progression IOPg and IOPp indi¬ces showed statistically significant decrease of mean values. Conclusion. Progression of keratoconus led to a de¬crease in corneal biomechanical properties which deter¬mine reduction of such indices as IOPg and IOPp in contrast to IOPcc.

SERUM TRIIODOTHYRONINE DETERMINATION BY SATURATION ANALYSIS

1973 ◽  
Vol 72 (4) ◽  
pp. 714-726 ◽  
Author(s):  
A. Burger ◽  
B. Miller ◽  
C. Sakoloff ◽  
M. B. Vallotton
Keyword(s):  
Ionic Strength ◽  
Limit Of Detection ◽  
Improved Method ◽  
Accuracy Of Measurement ◽  
Tracer Amount ◽  
The Mean ◽  
Physiological Values ◽  
Hyperthyroid Patients ◽  
Solvent Blank

ABSTRACT An improved method for the determination of serum triiodothyronine (T3) has been developed. After addition of a tracer amount of the hormone, T3 was extracted from 1 ml serum under conditions of pH and ionic strength which favoured T3 extraction (89%) over thyroxine (T4) extraction (58%). Chromatography of the extracted material on Sephadex LH-20 separated T3 completely from residual T4. The T3 eluate was dried, then re-dissolved in 0.5 ml NaOH 0.04 n. To 0.2 ml duplicate aliquots, a standard amount of TBG was added for the competitive protein analysis. After one hour incubation at 4°C, separation of bound from free T3 was achieved on small Sephadex G-25 columns. Overall recovery was 67 ± 10.8% and correction for the loss was made. The solvent blank was 37 ± 27 (sd) ng/100 ml. Accuracy of measurement of known quantities of T3 added to serum was 98.4%. The coefficient of variation within the assay was 6.2% and between the assays it was 11.4%. The limit of detection (0.1 ng) corresponded to a concentration of 25 ng/100 ml. T4 added to serum did not interfere with T3 determination until high non-physiological values were reached. The mean ± sd serum T3 in 54 euthyroid subjects was 153 ± 58 ng/100 ml and in 24 hyperthyroid patients it was 428 ±186 ng/100 ml; 4 out of the 24 hyperthyroid values were within 2 sd of the mean euthyroid group. All the values found in the euthyroid group were well above the limit of detection of the method.

Position Paper on Treatment of Hepatitis C in Romania, 2017. Part One

2017 ◽  
Vol 26 (2) ◽  
pp. 171-181 ◽  
Author(s):  
Liana Gheorghe ◽  
Ioan Sporea ◽  
Speranţa Iacob ◽  
Roxana Şirli ◽  
Anca Trifan ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Liver Disease ◽  
Hepatitis C ◽  
Limit Of Detection ◽  
Position Paper ◽  
Hcv Infection ◽  
Fixed Dose ◽  
Diagnostic Tools ◽  
End Stage Liver Disease ◽  
End Stage

Background & Aims: Hepatitis C Virus (HCV) infection is a common condition with endemic prevalence in some areas of the world. In Romania, the mean prevalence is about 3%. New treatments became available on the market in recent years and new drugs are in the pipeline. A re-evaluation of HCV therapy was considered mandatory. The Romanian Society of Gastroenterology and Hepatology undertook this task for the practitioners of this country.Methodology: A group of recognized experts was created who screened the available literature and the major available guidelines. A list of items requiring attention has been created. These items were discussed and rated. Decisions were taken by consensus.Recommendations: We present here the first of the two parts of our Society’s recommendations for chronic HCV infection treatment. An agreement was reached regarding the diagnostic tools, the assessment of severity and the up-dated therapy schedules.Conclusions: This Position Paper represents a guide for the assessment and the therapy of HCV infection. The recommendations are in concordance with other guidelines but are applied to the real-life conditions in this country.Abbreviations: DAAs: Direct-acting antivirals; DDIs: Drug-drug interactions; ESLD: End-stage liver disease; ESRD: End-stage renal disease; eGFR: Estimated glomerular filtration rate; EASL: European Association for the Study of the Liver; EMA: European Medicines Agency; FDA: US Food and Drug Administration; FDC: Fixed-dose combination; GT: Genotype; GRADE: Grading of Recommendations Assessment, Development and Evaluation; HCV: Hepatitis C virus; HCC: Hepatocellular carcinoma; LT: Liver transplantation; LLD: Lower limit of detection; MELD score: Mayo-Clinic End-Stage Liver Disease score; ANMDM: National Agency of Medicines and Medical Devices; PPIs: Proton pump inhibitors; PWID: People who inject drugs; RCT: Randomized controlled trial; RDT: Rapid diagnostic test; RAS: Resistance-associated substitution; SRGH: Romanian Society of Gastroenterology and Hepatology; SAE: serious adverse events; SPC: Summary of Product Characteristics; SVR: Sustained virologic response.

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