# In Vitro Antibacterialential Activity of Marine Microalgae Extract against _Vibrio harveyi_
## 4 Discussion
Methanolic extract in the current study inhibited _V. harveyi_ moderately, whereas the antibiotic synergism with the methanolic extract of _C. vulgaris_ exerted enhanced anti-bacterial activity in _E. coli_. [41]. Therefore, the selection of a solvent is also important for the further utilisation of post-extracted microalgal biomass. According to [PERSON] et al., 2017 [26], it was suggested that fatty acids could be involved in the antibacterial activity due to increased activity from green microalga extracts obtained with non-polar solvents compared to polar solvents. In the present study, microalgae extraction by saline water and DMSO demonstrated a comparable less effective antibacterial activity against _V. harveyi_ based on the concentration of _V. harveyi_. Nevertheless, it was reported that the extraction of antimicrobial compounds from _Scenedesmus subspicatus_ using DMSO inhibited the growth of _Klebsiella pneumoniae_ and _Escherichia coli_[42]. As observed in previous studies, water extracts of _Dunaliella_ sp. had lower bioactive properties than the ones obtained from organic solvents [43]. Hence, the selection of a solvent is important for optimisation of the extraction process and generation of its bioactive compounds, which have diverse applications not only in aquaculture but also against human pathogens. According to [PERSON] et al., 2012 [36] no antimicrobial activity was detected in the aqueous extracts, which was probably because of the low polar nature of the active components. The number of _V. harveyi_ increased exponentially, especially in the first 24 h during the experimental period, without the presence of microalgae cells (control) demonstrating that the bacteria cells were able to utilise the growth medium of the microalgae cultures. The inhibition of _V. harveyi_ in co-culture assay with _N. oceanica_, _Isochryssi_ sp. and _T. weissflogii_ indicated the production of antibacterial compounds by these microalgae cells. Previous research has shown that _Isochryssi_ sp. and _Nannochloropsis_ exhibit antibacterial action against the majority of vibriosis pathogens, including _V. alginolyticus_, _V. lentus_, _V. splendidus_, _V. scophthalmi_, _V. parhaemolyticus_, and _V. anguillarum_[44]. The microalgae species _Nannochloropsis_ sp. and _Isochryssi_ sp. have been demonstrated to synthesise short-chain fatty acids, which are implicated in antibacterial activity [45; 46]. Thus, microalgae biomass is regarded as a source of valuable chemical elements that can be used in animals, particularly farmed fish, as pharmaceuticals in aquaculture industries. Another element that could explain these discrepancies is that antibacterial activity is linked to the culture conditions and growth phase of the microalgae cultures. Although all microalgae species were at the same concentration in all trials in this study, the growth phase for each microalgae culture could have been different and may have influenced the results. Previous studies suggested that modifying the culture conditions of green microalgae exhibited differences in antibacterial activity [47; 48; 49], whereas modifications on microalgae culture conditions could be stimulated to produce secondary metabolites with antibacterial activity, as well as potentially larger quantities of these secondary metabolites [50]. This may include modifications on media composition, pH, light, and temperature [51; 52]. As an example, the highest light intensity (4800 lux) exposure effectively induced the production of antibacterial compounds by _Dunaliella_ sp. and suggested that the higher antimicrobial effect was related to the content of bioactive compound produced in stress conditions [39]. Other than the culture environment, the period of microalgae cultivation also plays an important parameter in producing bioactive compounds [39; 53]. Therefore, the culture environment could be artificially manipulated in order to allow for increased production of antibacterial compounds in microalgae.