Amplified Immunoassay ELISA-ELCA for Measuring Clostridium botulinum Type E Neurotoxin in Fish Fillets

1994 ◽  
Vol 57 (11) ◽  
pp. 985-990 ◽  
Author(s):  
MICHAEL G. ROMAN ◽  
JOHN Y. HUMBER ◽  
PAUL A. HALL ◽  
N. RUKMA REDDY ◽  
HAIM M. SOLOMON ◽  
...  
Keyword(s):  
Clostridium Botulinum ◽  
Toxin Production ◽  
Coagulation Cascade ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mouse Bioassay ◽  
Sampling Sequence ◽  
Botulinum Type ◽  
Catfish Fillets ◽  
Clostridium Botulinum Type E ◽  
Coagulation Assay

The measurement of Clostridium botulinum type E toxin in fish was accomplished using an amplified immunoassay (enzyme-linked immunosorbent assay-enzyme-linked coagulation assay [ELISA-ELCA]) based on the coagulation cascade. Fresh catfish fillets inoculated with a mixture of spores from five strains of C. botulinum type E were packaged in high barrier film with air, vacuum and modified atmosphere and stored at 4, 8 or 16°C for up to 75 days. Toxin production was monitored during storage by both mouse bioassay (trypsin and non-trypsin treated) and ELISA-ELCA on the non-trypsinized samples. All 26 inoculated products that were positive by the mouse bioassay were also positive by ELISA-ELCA. Of 35 uninoculated samples which were not toxic in mouse bioassay, none were positive by ELISA-ELCA; of 73 inoculated samples which were not toxic by mouse bioassay, 14 had toxin measurable by the ELISA-ELCA. The position of these immunoassay-positives in the sampling sequence indicated that the toxin was identified by the immunoassay before it was found in the mouse bioassay. These results suggest that the ELISA-ELCA technique is a usable alternative to the mouse bioassay for monitoring C. botulinum type E toxin production in fish challenge studies.

Toxin Production by Clostridium botulinum Type E in Packaged Channel Catfish

1997 ◽  
Vol 60 (11) ◽  
pp. 1358-1363 ◽  
Author(s):  
PING CAI ◽  
MARK A. HARRISON ◽  
YAO-WEN HUANG ◽  
JUAN L. SILVA
Keyword(s):  
Channel Catfish ◽  
Clostridium Botulinum ◽  
Modified Atmosphere Packaging ◽  
Toxin Production ◽  
Storage Conditions ◽  
Mouse Bioassay ◽  
Modified Atmosphere ◽  
Oxygen Barrier ◽  
Botulinum Type ◽  
Clostridium Botulinum Type E

Channel catfish were inoculated with 3 to 4 log spores/g of a mixed pool of four strains of C. botulinum type E (Beluga, Minnesota, G21-5, and 070) and were packaged with an oxygen-permeable overwrap, in an oxygen-barrier bag with a modified atmosphere of CO2-N2 (80:20) or in a master bag with the same modified atmosphere. Packaged fish were stored at either 4°C and sampled at intervals over 30 days or at 10°C and sampled at intervals over 12 days. An additional master bag treatment in which overwrap-packaged catfish was stored first at 4°C, then removed from the master bags and stored at 10°C, was sampled at intervals over 18 days. Toxin production was evaluated using the mouse bioassay. Aerobic psychrotrophic and anaerobic populations were enumerated, and product spoilage characteristics were noted. Under abusive storage conditions of 10°C, there was no difference among the potential for toxin production in the packaged fish, with botulinum toxin detected on fish from each package type by day 6. At 4°C, toxin production was detected on day 9 in the overwrapped packages, while it was on day 18 in the modified atmosphere packaging. No toxin was found in the master bags held continually at 4°C. Toxin was detected on day 18 from samples initially held at 4°C in the master bag and subsequently held at 10°C. Spoilage preceded toxin production for samples stored at 4°C for each type of packaging. At 10°C, spoilage and toxin detection times coincided.

Growth and Production of Toxin of Clostridium botulinum Type E in Rainbow Trout under Various Storage Conditions

1995 ◽  
Vol 58 (8) ◽  
pp. 863-866 ◽  
Author(s):  
DONNA M. GARREN ◽  
MARK A. HARRISON ◽  
YAO-WEN HUANG
Keyword(s):  
Rainbow Trout ◽  
Botulinum Toxin ◽  
Clostridium Botulinum ◽  
Toxin Production ◽  
Most Probable Number ◽  
Mouse Bioassay ◽  
Oxygen Barrier ◽  
Probable Number ◽  
Botulinum Type ◽  
Clostridium Botulinum Type E

Rainbow trout (Oncorhynchus mykiss) were inoculated with 3 to 4 1og10 spores per g of fish of a mixed pool of four strains of Clostridium botulinum type E (Beluga, Minnesota, G21-5, and 070). The trout were vacuum-skin packaged with either oxygen-barrier or oxygen-permeable films. Trout packaged with oxygen-permeable film were stored at 4°C for 21 days, while trout packaged with oxygen-barrier film were stored either at 4°C for 21 days or at 10°C for 15 days. Storage at 10°C was used to simulate commercial temperature abuse. Clostridium botulinum outgrowth was determined by a most probable-number (MPN) method using (tryptone peptone yeast extract glucose trypsin) anaerobic broth. Toxin production was evaluated using a mouse bioassay. Psychrotrophic and anaerobic populations increased with time regardless of packaging type. After 6 days at l0°C, botulinum toxin was detected in the packaged trout; however, the fish was noticeably spoiled before that time. No botulinum toxin was detected in trout packaged with either barrier or permeable films and stored at 4°C for 21 days, although the product was considered spoiled by day 12.

Risk of Clostridium botulinum Type E Toxin Production in Blue Crab Meat Packaged in Four Commercial-Type Containers

1996 ◽  
Vol 59 (3) ◽  
pp. 257-260 ◽  
Author(s):  
MARK A. HARRISON ◽  
DONNA M. GARREN ◽  
YAO-WEN HUANG ◽  
KEITH W. GATES
Keyword(s):  
Botulinum Toxin ◽  
Blue Crab ◽  
Clostridium Botulinum ◽  
Storage Period ◽  
Toxin Production ◽  
Mouse Bioassay ◽  
Botulinum Type ◽  
Clostridium Botulinum Type E ◽  
Entire Storage Period ◽  
Crab Meat

The aim of this investigation was to determine if a risk of Clostridium botulinum growth and toxin production existed in four different packaged crabmeat products. Freshly picked blue crab meat was inoculated with 103 to 104 spores per g of a mixed pool of four strains of C. botulinum type E (Beluga, Minnesota, G21-5, and 070). The lump crabmeat was packaged in four different packaging containers: (i) 12-oz copolymer polyethylene cups currently used by most crab processors; (ii) 12-oz copolymer polyethylene cups with heat-shrink, tamper-evident low-density polypropylene seals; (iii) 8-oz copolymer polyethylene cups with easy-open aluminum ends; and (iv) 8-oz copolymer polypropylene cups with integral tamper-evident pull-tabs. The packages were stored at either 4°C for 21 days or 10°C for 15 days. Storage at 10°C was used to simulate temperature abuse. The mouse bioassay was used to detect the presence of C. botulinum toxin. Psychrotrophic and anaerobic populations were enumerated and were found to increase with time regardless of packaging type. No botulinum toxin was detected in any of the four packaging types stored at 4°C or 10°C throughout the entire storage period.

Effect of Salt Level and Nitrite on Toxin Production by Clostridium botulinum Type E Spores in Smoked Great Lakes Whitefish1

1987 ◽  
Vol 50 (3) ◽  
pp. 212-217 ◽  
Author(s):  
S. L. CUPPETT ◽  
J. I. GRAY ◽  
J. J. PESTKA ◽  
A. M. BOOREN ◽  
J. F. PRICE ◽  
...  
Keyword(s):  
Great Lakes ◽  
Water Activity ◽  
Salt Concentration ◽  
Clostridium Botulinum ◽  
Toxin Production ◽  
Water Phase ◽  
Salt Level ◽  
Botulinum Type ◽  
Clostridium Botulinum Type E ◽  
Average Water

The effect of salt level and nitrite on botulinal safety of smoked whitefish was investigated. An average water-phase (wp) salt concentration of 4.4% inhibited outgrowth of Clostridium botulinum type E spores (103 spores/g) for over 35 d in temperature-abused (27°C) smoked whitefish. Incorporation of nitrite (220 mg/kg) during brining to the smoked salted (4.4%, wp) whitefish inhibited toxin production for 56 d at 27°C. An average salt concentration of 6.2% (wp), with or without nitrite, totally inhibited toxin production for the duration of the study (83 d). The effect of pH and water activity in temperature-abused smoked whitefish as a means of controlling toxin production by C. botulinum type E spores was evaluated.

TOXIN PRODUCTION BY CLOSTRIDIUM BOTULINUM, TYPE E, IN VACUUM-PACKED, IRRADIATED FRESH FISH IN RELATION TO CHANGES OF THE ASSOCIATED MICROFLORA

1965 ◽  
Vol 11 (3) ◽  
pp. 523-529 ◽  
Author(s):  
Kerstin Abrahamsson ◽  
N. N. De Silva ◽  
N. Molin
Keyword(s):  
Clostridium Botulinum ◽  
Toxin Production ◽  
Aerobic Bacteria ◽  
Prolonged Storage ◽  
Fresh Fish ◽  
Oxygen Scavenging ◽  
Botulinum Type ◽  
Clostridium Botulinum Type E

Thawed herring fillets were inoculated with spores of C. botulinum, type E, and then packed in laminated polythene bags. Some samples were vacuum-packed, others not, but the bags were sealed, while a third group was packed in open bags. The samples were stored at 20 °C and examined for the presence of toxin.The vacuum-packing considerably increased the strength of toxin present after 48 hours. On further prolonged storage at 20 °, however, the concentration of the toxin became roughly equal in all three cases.Changes in the associated spoilage flora lent further support to the view that oxygen scavenging by aerobic bacteria favors the growth of Clostridia. The flora surviving after irradiation with 0.6 Mrad consisted solely of micrococci, which were, however, sensitive to antibiotics such as penicillin and oxytetracycline.

Inhibition of Clostridium botulinum Type E Toxin Formation by Potassium Chloride and Sodium Chloride in Hot-Process (Smoked) Whitefish (Coregonus clupeaformis)

1985 ◽  
Vol 48 (11) ◽  
pp. 971-975 ◽  
Author(s):  
G. A. PELROY ◽  
A. SCHERER ◽  
M. E. PETERSON ◽  
R. PARANJPYE ◽  
M. W. EKLUND
Keyword(s):  
Ionic Strength ◽  
Clostridium Botulinum ◽  
Chloride Ion ◽  
Toxin Production ◽  
Ion Concentration ◽  
Coregonus Clupeaformis ◽  
Smoked Fish ◽  
Botulinum Type ◽  
Clostridium Botulinum Type E ◽  
Toxin Formation

Whitefish steaks were brined in NaCl, KCl, or equimolar NaCl:KCl to contain similar chloride ion concentration and inoculated intramuscularly with 10 or 100 spores of Clostridium botulinum type E per gram of fish. Steaks were then heated in a simulated (i.e., without smoke) hot-smoke process to internal temperatures of 62.8° to 76.7°C (145°–170°F) for the final 30 min of a 2- to 3-h process, packaged under vacuum in oxygen-impermeable film, and stored at abuse temperature of 25°C. During 7 d of storage, toxin production was inhibited in steaks containing more than 0.66 ionic strength NaCl, 0.64 KCl, or 0.71 equimolar NaCl:KCl. The results indicate that it is feasible to substitute KCl for NaCl in hot-process smoked fish for inhibition of outgrowth and toxin production by Clostridium botulinum type E.

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