| Microbenthos are important components in benthic microbial food web. However, there is little attention to the research of microbenthic ecology because of their complexity in biodiversity and ecological functions and the limitation of research methods. In the present study, we investigated the community structure, distribution, and the qualitative and quantitative importance of microbenthos in the Yellow Sea sediment based on samples collected from 82 stations during the cruises in 2007 and 2008. Benthic environmental variables including water temperature, salinity, water depth and sediment organic matter, chlorophyll a, phaeophytin a, and sediment median grain size were analyzed and discussed their possible relationships with the community structure and distribution of microbenthos.Firstly, we tested the effects of different storage manners and preservation duration on enumeration of microbenthos. The method of epifluorescence microscopy with DAPI has been widely used in the quantitative research on planktonic bacteria and protists, but scarcely applied in benthic study. We tested the effects of cold and freezing storage and preservation duration on the enumeration of marine benthic bacteria, cyanobacteria, and protists including diatoms, phototrophic nanoflagellates (PNF) and heterotrophic nanoflagellates (HNF) by DAPI epifluorescence microscopy. The test samples were collected from 0?2 cm and 2?5 cm sediment layers in the Yellow Sea, preserved with 2% glutaraldehyde, and stored at 4°C and ?20°C in the dark, respectively. The results suggested no significant differences in abundance between the 0?2 cm surface samples stored at 4°C and ?20°C in two stations, while in one station the abundances of PNF (2?5μm, 5?10μm) stored at 4°C were higher than that at ?20°C. The preservation duration effect was further tested using sediment samples collected from 0?2 cm and 2?5 cm sediment layers and stored at 4°C in the dark. The abundances of bacteria, cyanobacteria, diatoms, PNF and HNF were counted after one and four months. Statistical analyses show that storage time had no significant effect on the enumeration of benthic bacteria, cyanobacteria, PNF (5?10μm), PNF (>10μm), HNF (>10μm) and diatom. However, the abundances of PNF (2?5μm), HNF (2?5μm) and HNF (5?10 μm) were distinctly decreased after four months. Our experiments suggested that preservation types and duration as well as sediment types could influence the enumeration efficiency of marine benthic microorganisms. Thus, it is recommended that marine benthic samples can be temporarily stored at 4°C in the dark and should be enumerated as soon as possible.Using epifluorescence microscopy and the Ludox density centrifugation combining the quantitative protargol staining techniques, we investigated the spatial distribution and composition of microbenthos in the seafloor sediments from 48 stations of the Yellow Sea during the cruise on June 15-28, 2007. The microbenthic abundance was on average (1.15 ? 0.38) ? 109 cells/cm3 and biomass was 120 ? 54μg C/cm3 (? 6009 ? 2673μg C/10 cm2) in the upper 5 cm of the sediments. The abundance and biomass of microbenthos in the northern Yellow Sea were separately 12% and 34% higher than those in the southern Yellow Sea. Similarly, the abundance and biomass of microbenthos in the Yellow Sea Cold Water Mass (YSCWM) were separately 12% and 17% higher than those in the area outside the YSCWM. The bacterial abundance ranged from 2.36 ? 108 to 1.85 ? 109 cells/cm3 wet sediment, about three orders of magnitude higher than that of phototrophic (PNF, from 6.43 ? 105 to 8.80 ? 106 cells/cm3 ) and heterotrophic nanoflagellates (HNF, from 5.78 ? 104 to 5.91 ? 106 cells/cm3), and about four orders of magnitude higher than that of cyanobacteria (from 2.31 ? 104 to 2.31 ? 106 cells/cm3) in the upper 5 cm of the sediments. The diatom abundance highly varied from 3 to 1.06 ? 105 cells/cm3 in the upper 8 cm of the sediments, whereas that of heterotrophic microflagellates (HMF, 1?182 cells/cm3) and ciliates (1?221 cells/cm3) was less variable and lower. The biomass partitioning indicated the primary importance of benthic bacteria (on average 50.26 ?g C/cm3), followed by PNF (40.73 ?g C/cm3), HNF (19.34 ?g C/cm3), and cyanobacteria (8.84 ?g C/cm3). Benthic diatoms (0.84 ?g C/cm3), ciliates (0.15 ?g C/cm3) and HMF (0.03 ?g C/cm3) contributed relatively small proportions to the total biomass of microbenthos. About 95% of diatoms, 77% of ciliates and 56% of HMF were distributed in the upper 2 cm of the sediments, while no distinct vertical distributions were observed for bacteria, cyanobacteria, PNF and HNF. The statistical analyses show that the abundance of ciliates and diatoms was significantly positively correlated with sediment Chl-a concentrations while the diatom abundance was also negatively correlated with water depth. The HMF abundance was significantly negatively correlated with bottom water salinity, organic matter contents and sediment median grain size. The microbenthos were quantitatively important in the shallow sea floor, where their main components were about one to four, on average three orders of magnitude higher in abundance than corresponding planktonic ones in the same sea area. The comparison of the direct count of microphytobenthos and the estimation based on the ratio of biomass to Chl a indicated that pico-sized phytobenthos might contribute a large proportion to the primary production. Benthic ciliates and heterotrophic flagellates contributed about 90? to the estimated combined metabolic rate of micro- and meiobenthic consumers in the whole sea area, with nanoheterotrophs accounting for the majority. Benthic bacteria were 3.5 times higher in biomass than that of meiobenthos while it was even 17.5 times higher in production than that of meiobenthos. The data suggest the potential for rapid primary and secondary productions of microbenthos and detrital utilization in the shallow sea floor sediments of the Yellow Sea.A total of 198 morphospecies, representing 16 classes/subclasses, 27 orders, and 89 genera, were identified from the samples collected in 2007. Prostomatea was the most predominant group accounting for 45% of total abundance and 58% of total biomass. The carnivores constituted the primary feeding type occupying about 64% of total biomass, followed by the bacterivores (21%), algivores (12%), and omnivores (3%). Statistical analyses show that the species number was significantly positively correlated with bottom water temperature and negatively with bottom water salinity, water depth, and median grain size. Besides, both Margalef and Shannon-Wiener diversity and Pielou evenness indices showed significantly negative correlation with bottom water salinity, while Shannon-Wiener diversity and Pielou evenness indices also had negative correlation with water depth and organic matter and positive with median grain size. The BIOENV analysis show that bottom water temperature affecting the ciliate community structure. Based on the estimated ciliate bacterivory and herbivory, we indicate that ciliate ingestion had only a marginal influence on bacterial abundance but possibly had an important impact on diatoms in the sediments of the Yellow Sea.The abundance of microbenthos was on average (6.43 ? 2.54) ? 108 cells/cm3 and biomass was 64.14 ? 49.78 ?g C/cm3 in the upper 5 cm of the sediments from 33 stations in the Yellow Sea at the late stage of E. prolifera bloom in summer 2008, which was about 45% lower in standing crops than that in 2007. The microbenthos standing crops decreased more obviously in the stations heavily affected by the E. prolifera bloom. The abundance and biomass of microbenthos in the northern Yellow Sea were separately 15% and 54% higher than those in the southern Yellow Sea. While the abundance and biomass were separately 19% and 35% higher inside than outside the YSCWM. The abundance of bacteria was on average 6.40 ? 108 cells/cm3, about two orders of magnitude higher than that of phototrophic (PNF, on average 1.75 ? 106 cells/cm3) and heterotrophic nanoflagellates (HNF, 0.66 ? 106 cells/cm3) and three orders of magnitude higher than that of cyanobacteria (2.72 ? 105 cells/cm3) in the upper 5 cm of the sediments. The abundance of diatoms was about 2.12 ? 103 cells/cm3 in the upper 8 cm of the sediments, but distinctly higher than that of heterotrophic microflagellates (HMF, 6 cells/cm3) and ciliates (25 cells/cm3). The biomass partitioning indicate the primary importance of benthic bacteria (on average 28.17 ?g C/cm3), followed by PNF (21.27 ?g C/cm3), HNF (11.18 ?g C/cm3), and cyanobacteria (2.83 ?g C/cm3). Benthic diatoms (0.52 ?g C/cm3), ciliates (0.11 ?g C/cm3) and HMF (0.07 ?g C/cm3) contributed relatively small fractions to the total biomass of microbial communities. About 87% of HMF, 55% of diatoms and 65% of ciliates were distributed in the surface 0?2 cm sediment layer, while there were no distinct vertical distributions for bacteria, cyanobacteria, PNF and HNF. The statistical analyses show that the correlations between microbenthos abundance and environmental factors in the northern Yellow Sea were different from those in the southern Yellow Sea. The microbenthic abundance in the northern Yellow Sea had no correlation with environmental factors except that the negative correlations between HMF abundance and sediment Chl-a concentrations and betweet PNF abundance and bottom water Chl concentrations, while in the southern Yellow Sea, the correlations between ciliate abundance and environmental factors were distinct from those between diatom abundance and environmental factors. The abundance and biomass of microbenthos assemblages were lower than those in 2007 except of HMF, indicating inhibition of microbenthos growth caused by the green algal bloom of E. prolifera.A total of 132 morphospecies, representing 13 classes/subclasses, 24 orders, and 55 genus were identified from the samples collected in summer 2008. Prostomatea was also the most predominant group accounting for 71% of total abundance and 64% of total biomass. The carnivores constituted about 53% of total biomass, followed by the bacterivores (33%), algivores (11%), and omnivores (1%). Statistical analyses show that ciliate abundance had positively correlated with bottom water salinity, water depth, and sediment organic matter content. Margalef diversity showed only positively correlation with bottom water temperature. Both Shannon-Wiener diversity and Pielou evenness indices showed negative correlations with water depth and organic matter content and positive with sediment median grain size, while the latter also showed significantly negative correlation with bottom water salinity. The BIOENV analysis show that bottom water temperature and salinity and sediment median grain size affecting the ciliate community structure. Except of Margalef diversity index, the standing crops, species number, Shannon diversity, and Pielou evenness indices of ciliates during the cruise in 2008 were lower than those in 2007, indicating inhibition of ciliate community structure and diversity caused by the green algal bloom of E. prolifera.Overall, the standing crops of microbenthos in the upper 5 cm of the sediments of the Yellow Sea were about two to four orders of magnitude higher than those of corresponding planktonic ones in the same sea area. The quantitively importance of microbenthos might relate with their functional importance, which indicate their important role in matter cycle and the transfer to higher trophic levels. An in-depth study of the standing crops and diversity dynamics of microbenthos will contribute to the understanding of the structure and function of benthic ecosystem.
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