Isolation,Identification,Antialgal Characteristics And Mechanism Of One Broad-spectrum Antialgal Bacterium

The eutrophication of water bodies and the outbreak of cyanobacteria l blooms is one of the global environmental problems.Many large and medium-sized shallow lakes in China,such as Taihu Lake,Chaohu Lake,Dianchi Lake,etc.,have been encounting the occurrence of cyanobacterial blooms with large scales and high intensities,causing disruption of the structure and function of aquatic ecosystem and affecting aquaculture,lands tourism,and other activities.Microcystis dominated blooms in China are characterized with the highest frequency of occurrence,the widest range of d istribution,longest duration,and the greatest harm.Blooms dominated by Anabaena,a filamentous cyanobacterial group,are the next to Microcystis blooms in frequency and scale,while Anabaena species have much higher diversity in the morphology,physiolo gical ecology and harmful products?such as toxins and odor substances?.Cylindrospermopsis,another filamentous cyanobacterial genus,has been widely concerned due to its bloom formation and globally wide invasion.The technology by using microbes to control algal blooms has various advantages such as low cost,environmental friendliness and no secondary pollution,which becomes a research hotspot in related fields.In this study,an algae-lysing bacterium isolated from soil in Xinzhou District,Wuhan City,was identified through analyzing the morphological,phylogenetic,physiological and biochemical characteristics.Microcystis aeruginosa PCC 7806,Anabaena eucompacta CHAB 1799 and Cylindrospermopsis raciborskii CHAB 151 were used to test its algicidal abilities.Upon the correct identification of the algae-lysing bacterial strain,the algicidal characteristics,the algae-lysing way,the algae-lysing active substances and the influence of environmental factors on the algae-lysing effect were further examined in the three cyanobacterial species.The strain was specifically applied to treat the cylindrospermopsin?CYN?producing C.raciborskii strain CHAB 3438,to study the algae-lysing mechanism and the effect on the production and distribution of CYN.The main findings in this study are described as follows:1.A strain with strong algae-lysing capacity on PCC 7806,CHAB 1799,and CHAB 151 was isolated from soil in Xinzhou District,Wuhan City and named as WP.The strain was rod-shaped and was about 1.3¹.7?m,with a width of 0.45⁰.55?m,spore-forming,spherical,swollen,and Gram-positive,mostly appearing as single cell.The 16S rRNA gene sequence homology of the strain WP with that of Lysinibacillus fusiformis NRS-350 reached 99.6%,so it was identified as belonging to the genus Lysinibacillus.2.The cell density thresholds of the strain WP inhibiting the growth of PCC 7806,CHAB 1799 and CHAB 151 were shown as less than or equal to 6×10² cfu/mL,6×10³ and 6×10⁴ cfu/mL,respectively.The strongest inhibitory effect on PCC 7806 by the strain WP compared with the reported algicidal bacteria,implied that strain WP is an algicidal strain with a great potential for control Microcystis.The strain WP has a strong algae-lysing effect on many kinds of bloom-forming cyanobacteria,with different the algae-lysing algae efficiency among different species and strains,but no obvious algicidal effect on the green algae.The physiological state of cyanobacteria and the density of cyanobacterial cells are important factors affecting the algae-lysing effect,and the physiological state of the strain WP hardly affects its algae-lysing ability.When the temperature ranged from 10 to 35?,the strain WP showed strongest algicidal activity against PCC 7806 and CHAB 151.While the light intensity ranged from 10 to60?mol·m^-2·s^-1,Lysinibacillus sp.WP was shown to have all higher algae-lysing efficiency to the three types of strains PCC 7806,CHAB 1799 and CHAB 151.The algae-lysing modes of Lysinibacillus sp.WP towards PCC 7806,CHAB 1799,and CHAB 151 were all indirectly algae-lysing.When co-cultivating strain WP with cyanobacterial strains,cyanobacterial medium or metabolites could be used as the carbon and nitrogen sources to the strain WP.Metabolism,growth,reproduction,and production of algae-lysing substances have the effect of inhibiting the growth of algae.The algicidal active substance secreted by the strain has high thermal stability and acid-base stability.It is speculated that the substance may be a non-protein material with a relative molecular mass of less than 3 KDa.3.The optimum growth temperature of PCC 7806 was 35?,and 25³5? for CHAB 151.PCC 7806 and CHAB 1799 grow best when the light intensity is 30?mol·m^-2·s^-1,and the optimum light intensity of CHAB 151 ranges from 30 to 60?mol·m^-2·s^-1.Compared to CHAB 1799 and CHAB 151,PCC 7806 can adapt to a wider range of light intensity and temperature.4.Strain WP may secrete extracellular algae-lysing active substance to dissolve C.raciborskii CHAB 3438 through initailly destroying the gas vesicles causing its cells to sink,and then affect photosynthetic pigments from allophycocyanin?APC?to Chlorophyll a?Chl-a?and photosynthetic efficiency.Eventually C.raciborskii CHAB3438 could not normally perform photosynthesis,and its growth was inhibited and gradually died.CYN is distributed both intracellularly and extracellularly.After cell-free filtrate of the strain WP treated CHAB 3438,the total amount of CYN in the treatment did not show significant change,but lower than the control.On the other hand,the extracellular CYN was much higher than intracellular CYN,indicating the cell-free filtrate of the strain WP caused CHAB 3438 to released intracellular CYN into culture solution along the lysis process.The WP strain did not show the ability to degrade CYN.The present study also focused new types of water bloom forming species A.eucompacta and the CYN-producing C.raciborskii,and a broad-spectrum algae-lysing bacterium was successfully obtained,filling the gap in algae-lysing bacteria.The strain WP may secrete extracellular algae-lysing active substances to dissolve C.raciborskii CHAB 3438 through destroying the gas vesicles firstly and controlling the total amount of CYN with increase the proportion of extracellular CYN.Such a result provided a new way to algae-lysing method.The toxin release in algae-lysing process should be concerned,and combined application of such algae-lysing bacteria with CYN or other cyanotoxin-degrading bacteria,may be proposed in practical measures for both removals of cyanobacterial blooms and cyanotoxins.