Design and Research on Artificial Rumen Continuous Culture System

[Objective] The paper was to study the dynamic changes of forage nutrient substance fermentation in rumen, and a set of continuous culture system of artificial rumen was designed. [Method] With in vivo as control, the simulating rumen fer- mentation effect in vitro culture system was evaluated. [Result] The simulation rumen fermentation test needed adaptive phase of 2-3 d, and the fermentation state was relatively stable within 3-9 d, with good effects. The test showed certain regularity variation with index value of rumen in vivo. [Conclusion] The continuous culture sys- tem of artificial rumen could be used as the ideal model to study the rumen fermen- tation in vivo.

STABILITY ANALYSIS OF EQUILIBRIUM FOR MICROORGANISMS IN CONTINUOUS CULTURE

The problem of equilibrium of non-linear dynamic system for microorganism in continuous culture is considered in this paper. The existence of equilibrium point is proved. The conclusion that the equilibrium is the continuous function of the dilution rate and substrate concentration in medium in a certain range is concluded. And the stability criterion of equilibrium is obtained. In theory, it explains the multiplicity shown in laboratory.

Evaluation of ruminal methane and ammonia formation and microbiota composition as affected by supplements based on mixtures of tannins and essential oils using Rusitec

Background Dietary supplements based on tannin extracts or essential oil compounds(EOC)have been repeatedly reported as a promising feeding strategy to reduce the environmental impact of ruminant husbandry.A previous batch culture screening of various supplements identified selected mixtures with an enhanced potential to mitigate ruminal methane and ammonia formation.Among these,Q-2(named after quebracho extract and EOC blend 2,composed of carvacrol,thymol,and eugenol)and C-10(chestnut extract and EOC blend 10,consisting of oregano and thyme essential oils and limonene)have been investigated in detail in the present study with the semi-continuous rumen simulation technique(Rusitec)in three independent runs.For this purpose,Q-2 and C-10,dosed according to the previous study,were compared with a non-supplemented diet(negative control,NC)and with one supplemented with the commercial EOC-based Agolin^(R) Ruminant(positive control,PC).Results From d 5 to 10 of fermentation incubation liquid was collected and analysed for pH,ammonia,protozoa count,and gas composition.Feed residues were collected for the determination of ruminal degradability.On d 10,samples of incubation liquid were also characterised for bacterial,archaeal and fungal communities by high-throughput sequencing of 16S rRNA and 26S ribosomal large subunit gene amplicons.Regardless of the duration of the fermentation period,Q-2 and C-10 were similarly efficient as PC in mitigating either ammonia(-37%by Q-2,-34%by PC)or methane formation(-12%by C-10,-12%by PC).The PC was also responsible for lower feed degradability and bacterial and fungal richness,whereas Q-2 and C-10 effects,particularly on microbiome diversities,were limited compared to NC.Conclusions All additives showed the potential to mitigate methane or ammonia formation,or both,in vitro over a period of 10 d.However,several differences occurred between PC and Q-2/C-10,indicating different mechanisms of action.The pronounced defaunation caused by PC and its suggested consequences apparently determined at least part of the mitigant effects.Although the depressive effect on NDF degradability caused by Q-2 and C-10 might partially explain their mitigation properties,their mechanisms of action remain mostly to be elucidated.

Effects of ambient DIN:DIP ratio on the nitrogen uptake of harmful dinoflagellate Prorocentrum minimum and Prorocentrum donghaiense in turbidistat

The effects of varying nitrogen （N）： phosphorus （P） ratios on the growth and N-uptake and assimilation of the harmful dinoflagellates Prorocentrum minimum and Prorocentrum donghaiense were examined in turbidistat culture experiments. Algal cultures were supplied with media containing PO4^3- in various concentrations to obtain a wide range of N：P ratios. Experiments to determine rates of N uptake and assimilation of different N sources （NO^3-, NH4^＋, urea and glycine by P. minimum and NO3^-, NH4^＋ by P. donghaiense） were conducted using ^15-N tracer techniques at each N：P ratio. The growth rates suggested nutrient limitation at both high and low N：P ratios relative to the Redfield ratio. On a diel basis, the growth of both species was regulated by the light-dark cycle, which may be a result of regulation of both lightdependent growth and light-independent nutrient uptake. Maximum growth rates of both species always occurred at the beginning of light phase. In P-rich medium （low N：P ratio）, both species had higher N assimilation rates, suggesting N limitation. Low assimilation coefficients at high N：P ratios suggested P limitation of N uptake and assimilation. NO3 ^-and NH4^＋ contributed more than 90% of the total N uptake of P. minimum. Reduced N sources were more quickly assimilated than NO3^-. Highest average daily growth rates were recorded near an N：P ratio of 12 for both species. The N uptake rates of cultures at N：P ratios near Redfield ratio were more balanced with growth rates. The linkage between growth rates and N uptake/assimilation rates were conceptually described by the variation of cell N quota. The N：P ratios affect the N uptake and growth of Prorocentrum spp., and may regulate their bloom progression in eutrophic ecosystems.

Environmental control of growth and BChl a expression in an aerobic anoxygenic phototrophic bacterium, Erythrobacter longus (DSMZ6997)

anoxygenic phototrophic bacteria （AAPB）, which form a unique functional group of heterotrophic bacteria, have the abilit,to utilize light energy. The impact of carbon source and light intensity on the growth and bacteriochlorophyn a （ BChl a） expression of a typical strain of AAPB, Erythrobacter longus strain DSMZ6997 was examined during batch culture and continuous culture. The results showed that the expression of BChl a in DSMZ6997 was regulated by both carbon-source and light conditions, and was stimulated by low availability of carbon but inhibited by light to a certain extent at 300 lx and completely at 1 500 lx. In contrast, cell abundance, and even cell size of this strain, was substantially enhanced under light/dark cycle cultivation conditions over dark conditions, indicating the promotion of growth by light. These results led to the conclusion that utilization of light through BChl a helps AAPB to survive under carbon stress, while light at high intensity is harmful to the synthesis of BChl a in AAPB.

PARAMETER IDENTIFICATION AND TERMINAL STEADY-STATE OPTIMIZATION FOR S SYSTEM IN MICROBIAL CONTINUOUS FERMENTATION

In this paper, we propose a special S system to describe the process of continuous glycerol fermentation to 1,3-propanediol by Klebsiella pneurnoniae. Some properties of solution for S system are discussed. Moreover, in order to identify the values of parameters such that S system can simulate the fermentation as exactly as possible, we develop a param- eter identification model, and prove the identifiability of parameters. Numerical results show that the average relative error between computational value and experimental data is 20.0875% lower than 24.4225% in previous work, which demonstrates that S system is better in describing continuous fermentation. Finally, we present a terminal steady- state optimization model with state constraints. Optimization results show that the maximum volume yields of 1,3-propanediol at terminal moment increased from 114.3 to 124.911 mmol L-1 h-1. The results provide reference for the commercial production of 1,3-propanediol.

Ultra-thin temperature controllable microwell array chip for continuous real-time high-resolution imaging of living single cells

Single-cell imaging,a powerful analytical method to study single-cell behavior,such as gene expression and protein profiling,provides an essential basis for modern medical diagnosis.The coding and localization function of microfluidic chips has been developed and applied in living single-cell imaging in recent years.Simultaneously,chip-based living single-cell imaging is also limited by complicated trapping steps,low cell utilization,and difficult high-resolution imaging.To solve these problems,an ultra-thin temperature-controllable microwell array chip(UTCMA chip)was designed to develop a living single-cell workstation in this study for continuous on-chip culture and real-time high-resolution imaging of living single cells.The chip-based on ultra-thin ITO glass is highly matched with an inverted microscope(or confocal microscope)with a high magnification objective(100×oil lens),and the temperature of the chip can be controlled by combining it with a home-made temperature control device.High-throughput single-cell patterning is realized in one step when the microwell array on the chip uses hydrophilic glass as the substrate and hydrophobic SU-8 photoresist as the wall.The cell utilization rate,single-cell capture rate,and microwell occupancy rate are all close to 100%in the microwell array.This method will be useful in rare single-cell research,extending its application in the biological and medical-related fields,such as early diagnosis of disease,personalized therapy,and research-based on single-cell analysis.