Evaluation of a modified semi-automated immunomagnetic separation technique for the detection of Cryptosporidium oocysts in human faeces

2008 ◽  
Vol 75 (1) ◽  
pp. 139-141 ◽  
Author(s):  
G. Robinson ◽  
J. Watkins ◽  
R.M. Chalmers
Keyword(s):  
Immunomagnetic Separation ◽  
Separation Technique ◽  
Human Faeces ◽  
Cryptosporidium Oocysts

Enumeration of Cryptosporidium oocysts from surface water concentrates by laser-scanning cytometry

2000 ◽  
Vol 41 (7) ◽  
pp. 197-202 ◽  
Author(s):  
F. Zanelli ◽  
B. Compagnon ◽  
J. C. Joret ◽  
M. R. de Roubin
Keyword(s):  
Surface Water ◽  
Water Samples ◽  
Laser Scanning ◽  
Immunomagnetic Separation ◽  
Entire Surface ◽  
Cryptosporidium Oocysts ◽  
Different Types ◽  
Laser Scanning Cytometry ◽  
Direct Microscopic Observation

The utilization of the ChemScan® RDI was tested for different types of water concentrates. Concentrates were prepared by cartridge filtration or flocculation, and analysed either without purification, or after Immunomagnetic separation (IMS) or flotation on percoll-sucrose gradients. Theenumeration of the oocysts was subsequently performed using the ChemScan® RDI Cryptosporidium application. Enumeration by direct microscopic observation of the entire surface of the membrane was carried out as a control, and recoveries were calculated as a ratio between the ChemScan® RDI result and the result obtained with direct microscopic enumeration. The Chemscan enumeration technique proved reliable, with recoveries yielding close to 100% in most cases (average 125%, range from 86 to 467%) for all the concentration/purification techniques tested. The quality of the antibodies was shown to be critical, with antibodies from some suppliers yielding recoveries a low as 10% in some cases. This difficulty could, however, be overcome by the utilization of the antibody provided by Chemunex. These data conclusively prove that laser scanning cytometry, which greatly facilitates the microscopic enumeration of Cryptosporidium oocysts from water samples and decreases the time of observation by four to six times, can be successfully applied to water concentrates prepared from a variety of concentration/purification techniques.

Determination of the Recovery Efficiency of Cryptosporidium Oocysts and Giardia Cysts from Seeded Bivalve Mollusks

2013 ◽  
Vol 76 (1) ◽  
pp. 93-98 ◽  
Author(s):  
FRANCISKA M. SCHETS ◽  
HAROLD H. J. L. van den BERG ◽  
ANA MARIA de RODA HUSMAN
Keyword(s):  
Quantitative Assessment ◽  
Intestinal Parasites ◽  
Cryptosporidium Oocyst ◽  
Immunomagnetic Separation ◽  
Giardia Cyst ◽  
Bivalve Mollusks ◽  
Recovery Efficiency ◽  
Risk Of Infection ◽  
Cryptosporidium Oocysts

The intestinal parasites Cryptosporidium and Giardia are transmitted by water and food and cause human gastroenteritis. Filter-feeding bivalve mollusks, such as oysters and mussels, filter large volumes of water and thus concentrate such pathogens, which makes these bivalves potential vectors of disease. To assess the risk of infection from consumption of contaminated bivalves, parasite numbers and parasite recovery data are required. A modified immunomagnetic separation (IMS) procedure was used to determine Cryptosporidium oocyst and Giardia cyst numbers in individually homogenized oysters (Crassostrea gigas) and mussels (Mytilus edulis). About 12% of the commercial bivalves were positive, with low (oo)cyst numbers per specimen. The recovery efficiency of the IMS procedure was systematically evaluated. Experiments included seeding of homogenized bivalves and whole animals with 100 to 1,000 (oo)cysts. Both seeding procedures yielded highly variable recovery rates. Median Cryptosporidium recoveries were 7.9 to 21% in oysters and 62% in mussels. Median Giardia recoveries were 10 to 25% in oysters and 110% in mussels. Giardia recovery was significantly higher than Cryptosporidium recovery. (Oo)cysts were less efficiently recovered from seeded whole animals than from seeded homogenates, with median Cryptosporidium recoveries of 5.3% in oysters and 45% in mussels and median Giardia recoveries of 4.0% in oysters and 82% in mussels. Both bivalve homogenate seeding and whole animal seeding yielded higher (oo)cyst recovery in mussels than in oysters, likely because of the presence of less shellfish tissue in IMS when analyzing the smaller mussels compared with the larger oysters, resulting in more efficient (oo)cyst extraction. The data generated in this study may be used in the quantitative assessment of the risk of infection with Cryptosporidium or Giardia associated with the consumption of raw bivalve mollusks. This information may be used for making risk management decisions.

scholarly journals Development of a Method for Detection of Giardia duodenalis Cysts on Lettuce and for Simultaneous Analysis of Salad Products for the Presence of Giardia Cysts and Cryptosporidium Oocysts

2007 ◽  
Vol 73 (22) ◽  
pp. 7388-7391 ◽  
Author(s):  
N. Cook ◽  
R. A. B. Nichols ◽  
N. Wilkinson ◽  
C. A. Paton ◽  
K. Barker ◽  
...  
Keyword(s):  
Internal Control ◽  
Giardia Duodenalis ◽  
Immunomagnetic Separation ◽  
Internal Controls ◽  
Food Sample ◽  
Simultaneous Analysis ◽  
Natural Food ◽  
Cryptosporidium Oocysts ◽  
Natural Foods ◽  
Texas Red

ABSTRACT We report a method for detecting Giardia duodenalis cysts on lettuce, which we subsequently use to examine salad products for the presence of Giardia cysts and Cryptosporidium oocysts. The method is based on four basic steps: extraction of cysts from the foodstuffs, concentration of the extract and separation of the cysts from food materials, staining of the cysts to allow their visualization, and identification of cysts by microscopy. The concentration and separation steps are performed by centrifugation, followed by immunomagnetic separation using proprietary kits. Cyst staining is also performed using proprietary reagents. The method recovered 46.0% ± 19.0% (n = 30) of artificially contaminating cysts in 30 g of lettuce. We tested the method on a variety of commercially available natural foods, which we also seeded with a commercially available internal control, immediately prior to concentration of the extract. Recoveries of the Texas Red-stained Giardia cyst and Cryptosporidium oocyst internal controls were 36.5% ± 14.3% and 36.2% ± 19.7% (n = 20), respectively. One natural food sample of organic watercress, spinach, and rocket salad contained one Giardia cyst 50 g−1 of sample as an indigenous surface contaminant.

scholarly journals Effects of pH and Magnetic Material on Immunomagnetic Separation of Cryptosporidium Oocysts from Concentrated Water Samples

2002 ◽  
Vol 68 (4) ◽  
pp. 2066-2070 ◽  
Author(s):  
Ryan C. Kuhn ◽  
Channah M. Rock ◽  
Kevin H. Oshima
Keyword(s):  
Magnetic Material ◽  
Water Samples ◽  
Environmental Water ◽  
Immunomagnetic Separation ◽  
Deionized Water ◽  
Ph Values ◽  
Cryptosporidium Oocysts ◽  
Optimum Ph ◽  
Effects Of Ph ◽  
Optimal Ph

ABSTRACT In this study, we examined the effect that magnetic materials and pH have on the recoveries of Cryptosporidium oocysts by immunomagnetic separation (IMS). We determined that particles that were concentrated on a magnet during bead separation have no influence on oocyst recovery; however, removal of these particles did influence pH values. The optimal pH of the IMS was determined to be 7.0. The numbers of oocysts recovered from deionized water at pH 7.0 were 26.3% higher than those recovered from samples that were not at optimal pH. The results indicate that the buffers in the IMS kit did not adequately maintain an optimum pH in some water samples. By adjusting the pH of concentrated environmental water samples to 7.0, recoveries of oocysts increased by 26.4% compared to recoveries from samples where the pH was not adjusted.

Optimization of the Envirochek capsule method and immunomagnetic separation procedure for the detection of low levels of Cryptosporidium in large drinking water samples

2000 ◽  
Vol 41 (7) ◽  
pp. 111-117 ◽  
Author(s):  
A. Pezzana ◽  
Ph. Vilaginès ◽  
F. Bordet ◽  
D. Coquard ◽  
B. Sarrette ◽  
...  
Keyword(s):  
Drinking Water ◽  
Water Samples ◽  
Magnetic Beads ◽  
Tap Water ◽  
Immunomagnetic Separation ◽  
Sample Volume ◽  
Separation Procedure ◽  
Cryptosporidium Oocysts ◽  
Low Levels ◽  
Drinking Water Samples

The method for concentration of Cryptosporidium oocysts in large drinking water samples using the Envirocheck capsule has been optimized for the detection of low levels of oocysts. Elution from the filter by contact time and vortex agitation gave 68% oocyst recovery. Centrifugation (1,250 g; 30 min; 4°C) improved recovery to 94% without morphological damage of the oocysts. Increasing the ratio of magnetic beads to sample volume in the IMS procedure led to 69% efficiency. In these conditions, the overall recovery of the procedure was 49% as assessed with low oocysts spike doses in 100 litres tap water samples. The methodology described allows the detection of 0.1 oocyst per litre when 100 litres samples are processed.

scholarly journals Cryptosporidium in small water systems in Puerto Rico: a pilot study

2015 ◽  
Vol 13 (3) ◽  
pp. 853-858 ◽  
Author(s):  
Guy Robinson ◽  
Harvey A. Minnigh ◽  
Paul R. Hunter ◽  
Rachel M. Chalmers ◽  
Graciela I. Ramírez Toro
Keyword(s):  
Drinking Water ◽  
Puerto Rico ◽  
Pilot Study ◽  
Immunomagnetic Separation ◽  
Water Systems ◽  
Small Water ◽  
Cryptosporidium Oocysts ◽  
Small Water Systems ◽  
Potential Public Health ◽  
Drinking Water Systems

A pilot study was undertaken to investigate the occurrence of Cryptosporidium in four very small drinking water systems supplying communities in rural Puerto Rico. Water samples (40 L) were collected and oocysts were concentrated by calcium carbonate flocculation, recovered by immunomagnetic separation and detected by immunofluorescence microscopy. Cryptosporidium oocysts were identified in all four systems. This is the first report of evidence of the potential public health risk from this chlorine-resistant pathogen in Puerto Rican small water systems. Further work is warranted to fully assess the health risks that Cryptosporidium and other protozoa pose to populations served by community-managed small drinking water systems.

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