Study On Diversity Of Phage In Napahai

Napahai is a unique seasonal plateau wetland with low latitude and high altitude in China.It is an isolated and fragmented ecosystem that own distinct rainy and dry seasons,and showed characteristics of both land and water.Where there are a large number of phages were one of the most important factors to balance wetland ecosystem.Undoubtedly,it was very important to study the phage systematicly.In addition to the study on plants and animals,the research on phage was still rare in Napahai wetland.In this study,the diversity of phages was investigated by cultured and uncultured techniques in Napahai wetland.71 cultured bacteria and 23 phages were isolated from water samples sampled in Napahai wetland(E99�37'31",N27053'37")in 2011.The biological characteristics of physiology and biochemistry of Aeromonas sp.NPH-1 strain and its phage were studied.Phage abundance of water and soil samples sampled in 2013 was estimated by fluorescence microscopy and flow cytometry.It showed that mean abundance of phage in soil and water samples ranged from 1.32�106?3.69�106/ml and 2.05�106?4.97�107/ml in May,respectively,while 1.35�106-3.54�106/ml and 3.85�106?2.21�107/ml in December,respectively.Phage abundance of soil samples surface was the highest,and showed the most dramatic change,especially the wetland soil samples.There was no significant correlation of phage abundance to seasonal variations and other environmental factors,appeared to be positive correlated to chlorophyll a,total nitrogen and potassium,and negative correlated to pH by SPSS analysis.The diversity of uncultrued phage was investigated by PCR with degenerate primers of conservative g20,DNA polymerase and g23 genes from the water sample(E99037'22",N27�53'32")in 2013.The results showed that:the amplified products of g20 gene were similar with paddy cyanophage sequences came from various regions.It indicated that the cyanophage in Napahai wetland had a rich diversity.In the amplified products of DNA polymerase gene,only a phage could be similar with rare clones,and the rest clones clustered four groups were unknown.There may be some unique phages in Napahai wetland.Some amplified products of g23 gene were similar with T4-like phages,and the others clustered two groups were unknown sequences which had far evolutionary distances with the currently known phages sequences.So the unkown sequences may be the new paticular phages.In this study,many cold-active bacteria and phages were isolated in Napahai wetland.Phage abundance was estimated by fluorescence microscopy and flow cytometry,and phages diversity was investigated by molecular markers.The results showed the phages had high diversity and were different to known phages.The research will contribute to theoretical reference for the further study.