Geographical Distribution and Molecular Characterization of aToxigenic and a toxigenic Isolates of Aspergillus Flavus and Aspergillus Parasiticus in kenyax
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Date
2013-03-21
Authors
Okun, Daniel
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Abstract
Grainsof important food and export crops in Africa, such as maize, cassava and peanuts,
are susceptible and prone to contamination with different toxin producing moulds.
Aflatoxinsare mycotoxins associated with liver damage and cancer in humans and animals.
Thesetoxic substances are produced by fungi (such as Aspergillus flavus and Aspergillus
parasiticus)in food and feed commodities that are exposed to poor conditions during crop
cultivationor during storage and processing of the harvest. The presence of aflatoxins
especiallyin maize and peanuts in Kenya is of great concern. Reports of aflatoxin poisoning
inmaizeand peanuts in Kenya are an indication of the high level of exposure of people
whoselives depend on these crops as their staple foods. Recent developments in the
applicationofatoxigenic strains of these fungi as biological control agents against toxigenic
strainscould provide a solution to the problem. In Kenya the distribution and
characterizationof toxigenic' and atoxigenic Aspergillus strains is not known. The main
objectiveof this project 'was to isolate, identify and characterize atoxigenic and toxigenic
strainsof A. jlavus and A. i/arasiticus in Kenya. Fungal isolates from soils under maize and
peanutcultivation from various locations in Kenya were examined to determine the
distributionsof toxigenic Aspegillus species and to identify endemic atoxigenic strains. In
thisstudy,soils samples were collected randomly from Kenyan fields located in seven agroecologicalzones
(AEZ) (CL3, CL4, L3, L4, LM3, LM4 and LM5) and characterized by use
ofmorphological,physiological and molecular techniques. In this study, Aspergillus section
Flaviwasdetected in all the 57 soil samples. In total, 220 isolates belonging to A. flavus and
A.parasiticus were obtained. Aspergillus section Flavi L-strain was the most commonly
isolatedmember (54%), followed by Aspergillus section Flavi S-starins (35%). Aspergillus
jlavus was the most predominant (63.2%), followed by A. parasitic us (27.7%), A. tamari
(5.5%)and A. nomius (2.7%). The mean Colony Forming Units (CFU) of the Aspergillus
coloniesper gram of soil was extremely variable among the districts, ranging from 3.0xl03
to1.72x106(p<0.05). The mean temperatures across the collection sites also varied (from 24
toWC) according to the respectiveAgroecological zones AEZ. The pH varied from 5.5 -
6.8, whichis within the optimal pH requirement for the members of section Flavi. Each of
theregionshad atoxigenic strains of potential value which can be employed as biological
controlagents in the management of afl~toxicoses. For molecular characterization in the
presentstudy, internal transcribed spacer (ITS 4 and ITS 5 primers) targeting the 5.8S
regionof the rDNA and AB28rrW81 primers targeting 18S ribosomal DNA region of
Aspergillusspp were used. The two primers generated a specific target band ::::600bp that
wassubsequently sequenced. The sequences obtained were edited by Bioedit and other
analysisperformed using Mega 5. The ITS-PCR and AB 28ffW 81-PCR products were
digestedusing HinjI endonuclease and generated polymorphic bands that ranged from= 100
to350bp.Aspergillus section Flavi sequences for the ITSl- 5.8S-ITS2 rDNA genes were
comparedagainst all other sequences available online with the basic local alignment search
tool algoritlim (BLAST, National Center for Biotechnology Information). Sequence
homologysearches within the Aspergillus section Flavi revealed that the majority of the
isolates(36.8%) were Aspergillus flavus , followed by A. terreus ( 15.8%), A. insuetus
(15.8%), A. parasiticus (10.5%), A. oryzae (5.2%), A. ochraceus (5.2%), A. variecolor
(5.2%)and A. calidoustus (5.2%). These results strongly support the need for using
molecularmarkers as an auxiliary tool in differentiating fungal species.
Description
QK 625.M7O38