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Using Rhizopus sp. against the growth of Aspergillus carbonarius as a potential biological control method of ochratoxin A contamination in raw cocoa beans
a Department of Food Science and Technology, University of Nangui Abrogoua, 02 Bp 801 Abidjan 02, Côte d'Ivoire b UMR Qualisud, CIRAD - TA-B 95/16, 73 rue JF Breton, 34398 Montpellier Cedex 5, France
Didier Kra Brou KEDJEBOa, Tagro Simplice GUEHIa, Maï Koumba KONEa, Pascaline ALTERb, Noël DURANDb, Renaud BOULANGERb, Angélique FONTANAb, Didier MONTETb
Ochratoxin A (OTA) is a carcinogenic and nephrotoxic mycotoxin produced by fungal species belonging to
Aspergillus and Penicillium genus. OTA occurrence in foodstuffs depends on the climate during harvest
and on the post-harvest storage conditions. Among known ochratoxins, OTA currently contaminates
crops. Due to its prevalence and toxicity, OTA occurrence development conditions in raw cocoa beans
were previously studied during post-harvest treatments. This research aimed to investigate on the
biological control of OTA excretion in cocoa.
Material and MethodsCocoa CocoapodsusedwerethevarietyofcloneCLM99harvestedatTrinidadTobagoin2013.
Analyzes •Cocoapodinocula-onbyRhizopusRhizopussp.isolatedfromroAencocoapodsopenedaBer7daysdelayandA.carbonariusisolatedfromcoffeecherrieswereseparatelygrewonPDAagarfor5daysat25°C(Figure1).Differentconidiainoculawerepreparedfromcalibratedsuspensionsat102,104and106conidiapermillileter.Then1mLofeachpreparedinoculumwasasepPcallyinjectedusingasyringeinsidehealthcocoapods.InoculatedcocoapodswereincubatedinaclimaPcchamberat30°C,60%relaPvehumidityfor4or8days.•OchratoxinAcontentmeasurementOTAwasextractedfromcocoabeanssourcedfrominoculatedpodsandthenseparatedbyimmuno-affinity,idenPfiedbyHPLCandquanPfiedbyspectro-fluorescencedetector.
ResultsTableIpresentsthephotographsofbeansinsidecocoapodsaBerinoculaPonwithvariousconcentraPonsofA.carbonariusandRhizopussp.conidiasuspensions.Figure2indicatesOTAcontentforcocoabeanssourcedfrom:
•Podsinoculatedwith104conidiapermLofA.carbonariusand104conidiapermLofRhizopussp.-incubatedfor4days:noOTAproducPonwasobserved- incubatedfor8days:OTAproducPonof37µg.kg-1washighlightedwhencontrolpresentednoOTAproducPon.
•Podsinoculatedwith104conidiapermLofA.carbonariusand106conidiapermLofRhizopussp.-incubatedfor4days:noOTAproducPonwasnoPced-incubatedfor8days:OTAproducPonof5.6µg.kg-1wasobtainedagainst0µgperkgforthecontrol.
•Podsinoculatedwith104conidiapermLofRhizopussp.and106conidiapermLofA.carbonarius-incubatedfor4days:36.3µg.kg-1ofOTAwasrecorded-incubatedfor8days:6.2µg.kg-1ofOTAwasobserved
Conclusion
Durand,N.,2012.
Dynamiquedesp
opulationsmicrobi
ennes
au cours du trai
tementpost-réco
lte du café et r
elations
interspécifiques e
ntre souches och
ratoxinogènes. Th
èse de
Doctorat, 243 P.
Université de Mo
ntpellierII, Scienc
es et
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Ponsone,M.L.,Ch
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Rhizopus sp appeared as a promising biological control fungal agent for OTA
production in raw cocoa beans. As the fungal growth in raw cocoa beans constituted
a serious qualitative default, further investigation must be done on the identification
of the metabolites leading to the control of OTA excretion in cocoa.
Figure 1. Sampling during post-harvest processing of cocoa
Figure 1. Cultures of Aspergillus carbonarius (A) and Rhizopus sp. (B) on PDA agar used for cocoa beans inoculation inside pods
Table I. Photographs of beans inside cocoa pods inoculated with suspensions of various concentrations in A. carbonarius and Rhizopus sp. conidia
Corresponding author: [email protected]
Concentration of fungal conidia (conidia.mL-1)
Incubation
time (days)
A.carbonarius Rhizopus sp A.carbonarius Rhizopus sp A.carbonarius Rhizopus sp A.carbonarius Rhizopus sp
0 104 106 104 104 106
4
8
Figure 2. OTA content of cocoa beans from pods inoculated with various
concentrations of A. carbonarius and Rhizopus
sp conidia suspensions of