University of Khartoum

Morphological, Cytogenetic and Molecular Characterization of Simulium Damnosum (Diptera: Simliidae) In Abu Hamed Locality, Sudan

Morphological, Cytogenetic and Molecular Characterization of Simulium Damnosum (Diptera: Simliidae) In Abu Hamed Locality, Sudan

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Title: Morphological, Cytogenetic and Molecular Characterization of Simulium Damnosum (Diptera: Simliidae) In Abu Hamed Locality, Sudan
Author: Ahmed, Abdelrahim Mubarak Ali
Abstract: A morphological, cytogenetic and molecular approach was taken to study black flies of the Simulium species complex. Around 600 adult and 200 larvae of black flies were collected from Abu Hamed (River Nile State, Sudan) during February and March of each of the years 2011, 2012 and 2014. Sixty (60) black flies were subjected to morphometric examinations. The thorax length (TL), antenna length (AL), wing length (WL), wing width (WW) and femur length (FL) were measured and the data obtained was analysed. Somatic chromosomes were prepared from adult ganglion cells while polytene chromosomes were prepared from female ovarian nurse cells. The cells were squashed, stained with geimsa or aceto-orcein and the resulting karyogrames were photographed. Karyotype was constructed from somatic cells while cytological analysis of the banding patterns was performed using polytene chromosomes. Genomic DNA was extracted and 16S ribosomal RNA primers were selected. Molecular characterization was done using Polymerase Chain Reaction Amplification (PCR) of fly genomic DNA. Ten of the resulting PCR isolates were sequenced and analyzed using BioEdit software. Sequences were aligned with relevant sequences from the GenBank using Blastn tool. Phylogenetic trees were constructed using Neighborhood bootstrap. Results of morphometric studies showed significant differences (P<0.01) between flies collected on the three seasons regarding thorax length, antenna length, wing width and femur length with wing length during both 2012 and 2014 years than that reported during 2011. Flies collected in both 2012 and 2014 years had higher means of the thorax/antenna mean ratio than during 2011; thorax length/femur length (T/F) was significantly higher (P<0.01) in 2012 and 2014 than in 2011. There was strong (r between.0.6-0.9) and significant (P<0.01) correlation between TL and both T/W and T/WW. There was significant (P<0.01) relationship between WL and both W/A, and W/F. Results of cytogenetic study showed a mitotic chromosome number of 2n=6 in all specimens, with somatic pairing in metaphase chromosomes. Band and inter-band regions, puffs and inversions were detected. DNA amplified from all specimens resulted in a characteristic 556 bp band or fragment indicating the amplification of Simulium complex member species. The phylogenetic tree separated the S. damnosum samples into three clades with polymorphism.The morphological, cytogenetic and molecular analysis supports the presence of a diverse species structure of black flies at Abu Hamad. It is recommended that further studies on morphometric characters of black flies and larvae as well as studing the internal transcribed spacers (ITS) and the polymorphic microsatalites. The use of polymorphic microsatellites is also recommended for differentiating populations of Simulium spp. Complex. Further studies of molecular phylogeny using the sequences of the second internal transcribed spacer (ITS2) of ribosomal DNAx (rDNA) is also recommended.
URI: http://khartoumspace.uofk.edu/123456789/25537


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