## 3 Discussion
flavus_ was cultured on a medium with a high amount of NaCl. These results taken together indicate that _AfsaK_ might play an essential role in asexual development more especially in the survival of conidia and conidiophores under harsh conditions. This corroborates the findings in the earlier study by [PERSON] et al. [19] which reported that _A. nidulans sakA_ plays a role in the asexual development where it regulates the genes that are involve in sustention of conidia and their persistence. Slerotia are the sexual reproductive and survival structures which are produced by different fungi which helps fungi to adapt to unsuitable environmental conditions [33]. Our results indicated that _A. flavus_ produced sclerotia in the absence of stress and the deletion of _AfsaK_ increased sclerotia production in contrast to other strains. This might be due to inactivation of _AfsaK_ affected other signal pathway that are involved in normalization of sexual development in _A. flavus_. Further, the qRT-PCR results of the expression level of _nsdC_ and _nsdD_ genes related to sclerotial development indicated that the expression levels of both genes were significantly up-regulated in \(\Delta\)_AfsaK_ than that in \(\Delta\)_AfsaK_::_AfsaK_ and WT. Putting all these results together, we can confirm that _AfsaK_ regulates the sexual development of _A. flavus_ and the hyperosmotic stress may have been preventing, delaying or inhibiting sclerotia formation. Our results showed that under osmotic stress, sclerotia could not be produced in all strains after 7 days of incubation at 37 \({}^{\circ}\)C. Other studies also confirmed that salt stress have inhibitory effect on sclerotia production in various fungal species viz. _Sclerotitium rolfsii, Sclerotitium_ and _Rhizcotonia solani_[15; 34]. Moreover, stressed fungi with different types of salts produced fewer sclerotia in contrast to the unaltered control [34] after 15 d of incubation. In 1999, [PERSON] et al. [37] also reported that _A. ochraceus_ treated with different levels of water activity exhibits different levels of sclerotia formation. They went on and indicated that sclerotia were freely produced in _A. ochraceus_ treated with more than 0.99a\({}_{\mathrm{w}}\), while only initials were formed for _A. ochraceus_ treated with 0.975a\({}_{\mathrm{w}}\) and no sclerotia at all were produced for _A. ochraceus_ treated less than 0.975a\({}_{\mathrm{w}}\). The characterization of the function of _AfsaK_A in aflatoxin production was also performed. Obtained data proved that \(\Delta\)_AfsaK_ considerably increased the production of AFB\({}_{1}\) compared to WT and \(\Delta\)_AfsaK_::_AfsaK_ when the agar medium was induced by hyperosmotic stress agents. This finding is consistent with the observed increase of the transcription levels of _afR_, _afS_, _afQ_ and _afQ_ genes which are related to aflatoxin biosynthesis. Therefore, we can conclude that _sakA_ gene acts by negatively regulating the aflatoxin formation while the osmotic stress affects the biosynthesis of aflatoxin in _A. flavus_. However, the details on the function of _sakA_ gene in AFB\({}_{1}\) biosynthesis under hyperosmotic are still unknown in other world of fungi. In a recent study, [PERSON] et al. confirmed that _sakA_ and _mpkC_ collaborate during _A. fumigatus_ virulence in neutropemic mice [13]. An earlier study by [PERSON] et al. [38] indicated that not only _sakA_ involved in the cellular homeostasis regulation but also participates in a coordinated response to H\({}_{2}\)O\({}_{2}\). Moreover, it has been demonstrated that _sakA_ homologue, that is, _hogT_ gene, has a major function in virulence and stress response in _Cryptococcus neoformans_[39]. In the present study, we examined the role of _AfsaK_ in pathogenicity by infecting both the maize kernels and peanuts seeds. Results revealed that \(\Delta\)_AfsaK_ strain on the infected crop seeds appeared to produce a higher number of dark-green conidia than the WT and complemented strains in _A.