In this study, it was aimed to observe the antiviral effects of two different concentrations (5 mg/kg and 10mg/kg) of six different microalgae and cyanobacteria species on IBV D274 strain. However, there are articles that review the antiviral and antimicrobial properties of secondary metabolites of microalgae (Chaisuwan et al., 2021), in this study, P. tricornutum, Scenedesmus sp. and Pseudanabaena sp., A. platensis, N. shiloi, Prasinococcus sp. strains were tried to determine the antiviral properties by using hemagglutation tests.
The antiviral activity of microalgae/cyanobacteria was determined by both evaluating embryo viability and HA activity. In the interpretation of the hemagglutination activity in the HA test, the observation of lace-like agglutination form as the HA positive well and the button form (the sinking of the erythrocytes) were taken into account as the HA negative well. The HA titers of the samples and control groups were calculated on log2-based, and the antiviral properties of the extracts were evaluated compared to the virus control group. (WHO, 2013; Grimmes, 2002).
The HA titer of P tricornutum for 5 mg/kg and 10 mg/kg concentrations was found 256. It is possible to say that the antiviral effect was observed for the extract, regardless of concentration. When compared with the control group, Phaeodactylum sp. extract reduced virus activity by 3-fold on log2-base.. Among the quite limited studies with P. tricornutum, the most important one was the experiment studied by Fabregas et al.. They investigated the in vitro antiviral effect of endocellular and exocellular extracts obtained from different marine microalgea on aemorrhagic septicaemia virus (VHSV) ve African swine fever virus (ASFV). For P. tricornutum, it did not observed any inhibitory effect on these viruses (Fabregas et al., 1999). The antiviral activity that was found in our in ovo study compared to control groups, can be attributed to extraction method or inhibition on target molecules of IBV by P. tricornutum.
Scenedesmus sp. is seen that the average HA titers were 256 in CAF samples taken from eggs inoculated with two concentrations of the extract. Considering the same value in all eggs, Scenedesmus sp. was observed that the antiviral effect of algal organism on IBV D274 strain was independent of concentration. Whether the antiviral effect is concentration dependent can be determined by lower and higher concentration trials. In vitro antiviral activity studies performed by S. obliquus belongs to Scenedesmus sp. showed a weak effect (Afify et al., 2018; Ibrahim et al., 2021). Some specific experiment were made on Huh 7.5, MA104, BGM (Afify et al., 2018), and Vero cell lines (Afify et al., 2018; Ibrahim et al., 2021). Afify investigated the mode of action of extract against Coxsackievirus B3 strain and attained that the inhibition were higher rate in viral attachment step than other viral cycle (Afify et al., 2018). It is thought in this study that observed weak antiviral activity of Scenedesmus sp result from rare inhibition of attachment protein of IBV.
A. Platensis has many therapeutic effects, including its antiviral properties due to the bioactive compounds it contains. In this study, A. Platensis was found to have antiviral properties as in the literature. Hayashi et al. reported that A. Platensis inhibited virus replication in their study (Hayashi et al., 1993). Based on this, it can be interpreted that A. platensis IBV D274 can inhibit the replication of the virus. The HA titer of A. platensis as a result of hemagglutination was determined as 128 for 5 mg/kg concentration. A titer of 256 HA was determined in embryonated eggs containing A. platensis extract at 10 mg/kg concentrations. The present values for this concentration can be interpreted as the presence of antiviral activity of A. platensis. In ovo antiviral activity results of P. tricornutum, Scenedesmus sp., A. platensis species were found similar. In a study conducted against HIV-1 in the literature, it was stated that both a polysaccharide-containing fraction and a tannin-free fraction were responsible for showing antiviral activity of A. Platensis. Antiviral properties for P. tricornutum and Scenedesmus sp. species can also be attributed to their polysaccharide content (Ayehunie et al., 1998).
HA titer of Prasinococcus sp. was determined as 128 for eggs with 5 mg/kg concentration obtained as a result of hemagglutination tests of embryonated eggs containing extract. A titer of 128 HA was also determined for eggs with a concentration of 10 mg/kg. The studies in the literature, it was mentioned about the therapeutic effects of polysaccharide contents of Prasinococcus sp. (Bavington and Moss, 2013). However, no study was found that investigated the antiviral properties of Prasinococcus sp. In this study, the effect of Prasinococcus sp. on IBV D274 was determined. It is thought that the reason for the effectiveness of Prasinococcus sp. on IBV D274 may be the important biochemical compounds it contains. It is thought that N. shiloi, which has a titer of 64 HA at both concentrations, may have an antiviral effect. N. shiloi, which was reported to have a high amount of lipid molecules in its content, was effective on the IBV virus. For N. shiloi is said to have an antiviral effect. Another unique value of this study is that it can provide information about the antiviral properties of N. shiloi.
Compared to other microalgae species, it was seen that the antiviral effect was found in Pseudanabaena sp. as a result of the HA test performed on the chorioallantoic fluid samples taken from the eggs in which the extract was inoculated.. Pseudanabaena sp. has exhibited more antiviral properties at the concentration of 10 mg/kg than at the concentration of 5 mg/kg. As a result of the HA test, it was found that the highest antiviral effect was in this sample. In the newly established study by Ceglowska et al., The various chromatographic extracts/proteins obtained from Pseudanabaena galeata were performed against West Nile virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and human coronavirus OC43 (HCoV-OC43) to evaluate antiviral activity. For some fractions from extracts, it was obtained the decreased cytopathic effect on the cell line which means the extracts prevent the propagation of the virus. Especially in the experiment performed on A549 cell lines which present ACE2 receptor that is important for attachment of Sars-CoV-2, it was observed significant activity (Cegłowska et al., 2022). IBV which was used in this experiment, is in the same family as Sars-CoV-2 and cause epidemic or pandemics, particularly on poultries. Similarly with Sars-CoV-2 and P. galeata study, extracts from Pseudanabaena sp. can be considered to target spike proteins. Other studies with extracts of this species may be good alternatives to develop antiviral drugs against respiratory tract viruses that cause possible epidemics.In vitro antioxidant activity studies have been conducted with Pseudanabaena sp. High levels of phycobiliproteins as well as carotenoids were determined in water and methanol extracts of Pseudanabaena sp. It can be thought that these metabolites, which are stated to be effective against antioxidant activity, are also the major components in terms of exhibiting antiviral activity (Paliwal et al., 2015).