Micropropagation medium composition on-line sensing system development

A plant micropropagation system has been developed with the capacity of allowing periodic, automatic, aseptic monitoring of culture medium composition. The system was able to sample automatically culture medium from a liquid/membrane bioreactor without compromising asepsis or disturbing plant tissue. Media samples were analyzed to determine sucrose and dextrose concentrations. Bioreactor media was automatically replenished to compensate for volumes lost due to sampling. For a 28-day culture period using Stage 1 Nicotiana tabacum (tobacco) explants as a model culture system, media sucrose concentration decreased with a corresponding increase in dextrose concentration and tissue fresh weight.This article is published as technical contribution No. 3484 from the South Carolina Agricultural Experiment Station, Clemson, USA. Mention of products and tradenames does not constitute endorsement by the authors, or the South Carolina Agricultural Experiment Station.     

Utilization of microbial community potential for removal of chlorpyrifos: a review

Chlorpyrifos (CP) is the most commonly used pesticide in agricultural fields worldwide. Exposure to CP and its metabolites creates severe neuron-disorders in human beings. Improper handling and uncontrolled application of CP by farmers have lead to the contamination of surface and ground water bodies. Biodegradation offers an efficient and cost effective method for the removal of CP and other toxic organophosphorus pesticides from the contaminated environment. The degradation of CP by various microorganisms has been investigated by several researchers over the past few years. This review presents a critical summary of the recent published results on the biodegradation of CP. A diverse range of bacterial species such as Agrobacterium sp., Alcaligenes faecalis, Enterobacter sp. Arthrobacter sp. Bacillus pumilus, Pseudomonas sp. etc., fungal species like Trichoderma viridae, Aspergillus niger, Verticillium sp., Acremonium sp. Cladosporium cladosporiodes, etc. and certain algal species viz. Chlorella vulgaris, Spirulina platensis, Synechocystis sp., etc., have been shown to degrade CP. The efficacy of these communities for CP degradation in batch and continuous modes has also been discussed but more studies are required on continuous reactors. Also, the available published information on kinetics of biodegradation of CP along with the available results on molecular biological approaches are discussed in this work.     

In-vitro liquid culture of the entomopathogenic nematode,Steinernema colombiense,in orbitally shaken flasks

Steinernema colombiense, an entomopathogenic nematode species (EPN) was grown in two types of orbitally shaken flasks at 130?rpm and 28°C, containing 10 or 20?mL, respectively of a complex culture medium with an initial EPN-concentration of 1,000 Infective Juveniles (IJ)/mL. At the 10th day, the EPN-concentration was 58,771 individuals/mL with 87% of them in the IJ stage. No significant differences were found between the EPN growth kinetics in both types of flasks. The nematode-population growth was modelled by a re-parameterized Gompertz equation of three-parameters with best-fit values of 3.8 days for the lag time, 33.8 day^-1 for the maximum growth rate, and 57.3 (dimensionless) for the maximum asymptotic growth.     

Development of a finite element/multi-body model of a newborn infant for restraint analysis and design

A finite element/multi-body model of a newborn infant has been developed by researchers at the University of Windsor. The geometry of this model is derived from a Nita newborn hospital training mannequin. It consists of 17 parts: eight upper and lower limb segments, the torso, head, and a seven-segment neck with seven translational and eight rotational joints. Anthropometry is consistent with hospital growth charts, measurements requested from health professionals and data from the open literature. The biomechanical properties of the model (i.e. joint stiffnesses) are implementations of data identified in the open literature. The model has been validated with respect to studies of the biomechanics of shaken baby syndrome, infant falls and the Q0 anthropomorphic testing device. A significant conclusion of this study is that the kinetics of the Q0 neck is not biofidelic. This model is currently used in an analysis of airway patency for infants in modern automotive child restraints.     

Specific oxygen uptake of the entomopathogenic nematode,Steinernema carpocapsae CABA01, in submerged culture

The specific oxygen uptake rate (qO₂) of stages of the entomopathogenic nematode (EPN) Steinernema carpocapsae CABA01 in liquid culture was measured. Nematodes were grown into previously pasteurised culture broths of their symbiotic bacterium, Xenorhabdus nematophila, in orbitally agitated flask cultures (V_L = 125 mL) at N = 150 rpm and T = 25°C. The basal medium contained 3% (w/v) soy trypticase broth and 0.5% (w/v) yeast extract. The EPNs developed from the egg stage to the adult stage exhibiting qO₂ values of 1.92, 5.48, 0.48, 0.28 and 0.0014 [10^?1 mmolO₂/(g_{nematode-wet base} h)] for the egg-Juvenile 1 (J1), J2, J3, J4 and the adult stages, respectively.     

Activated packed bed bioreactor for rapid nitrification in brackish water hatchery systems

A packed bed bioreactor (PBBR) was developed for rapid establishment of nitrification in brackish water hatchery systems in the tropics. The reactors were activated by immobilizing ammonia-oxidizing (AMONPCU-1) and nitrite-oxidizing (NIONPCU-1) bacterial consortia on polystyrene and low-density polyethylene beads, respectively. Fluorescence in situ hybridization demonstrated the presence of autotrophic nitrifiers belong to Nitrosococcus mobilis, lineage of β ammonia oxidizers and nitrite oxidizer Nitrobacter sp. in the consortia. The activated reactors upon integration to the hatchery system resulted in significant ammonia removal (P < 0.01) culminating to its undetectable levels. Consequently, a significantly higher percent survival of larvae was observed in the larval production systems. With spent water the reactors could establish nitrification with high percentage removal of ammonia (78%), nitrite (79%) and BOD (56%) within 7 days of initiation of the process. PBBR is configured in such a way to minimize the energy requirements for continuous operation by limiting the energy inputs to a single stage pumping of water and aeration to the aeration cells. The PBBR shall enable hatchery systems to operate under closed recirculating mode and pave the way for better water management in the aquaculture industry.     

Microwell engineering characterization for mammalian cell culture process development

Experimentation in shaken microplate formats offers a potential platform technology for the rapid evaluation and optimization of cell culture conditions. Provided that cell growth and antibody production kinetics are comparable to those found in currently used shake flask systems then the microwell approach offers the possibility to obtain early process design data more cost effectively and with reduced material requirements. This work describes a detailed engineering characterization of liquid mixing and gas–liquid mass transfer in microwell systems and their impact on suspension cell cultures. For growth of murine hybridoma cells producing IgG1, 24‐well plates have been characterized in terms of energy dissipation (P/V) (via Computational Fluid Dynamics, CFD), fluid flow, mixing and oxygen transfer rate as a function of shaking frequency and liquid fill volume. Predicted k_La values varied between 1.3 and 29 h^?1; liquid‐phase mixing time, quantified using iodine decolorization experiments, varied from 1.7 s to 3.5 h; while the predicted P/V ranged from 5 to 35 W m^?3. CFD simulations of the shear rate predicted hydrodynamic forces will not be detrimental to cells. For hybridoma cultures however, high shaking speeds (>250 rpm) were shown to have a negative impact on cell growth, while a combination of low shaking speed and high well fill volume (120 rpm, 2,000 µL) resulted in oxygen limited conditions. Based on these findings a first engineering comparison of cell culture kinetics in microwell and shake flask formats was made at matched average energy dissipation rates. Cell growth kinetics and antibody titer were found to be similar in 24‐well microtiter plates and 250 mL shake flasks. Overall this work has demonstrated that cell culture performed in shaken microwell plates can provide data that is both reproducible and comparable to currently used shake flask systems while offering at least a 30‐fold decrease in scale of operation and material requirements. Linked with automation this provides a route towards the high throughput evaluation of robust cell lines under realistic suspension culture conditions. Biotechnol. Bioeng. 2010; 105: 260–275. © 2009 Wiley Periodicals, Inc.     

Microfluidic biolector—microfluidic bioprocess control in microtiter plates

In industrial‐scale biotechnological processes, the active control of the pH‐value combined with the controlled feeding of substrate solutions (fed‐batch) is the standard strategy to cultivate both prokaryotic and eukaryotic cells. On the contrary, for small‐scale cultivations, much simpler batch experiments with no process control are performed. This lack of process control often hinders researchers to scale‐up and scale‐down fermentation experiments, because the microbial metabolism and thereby the growth and production kinetics drastically changes depending on the cultivation strategy applied. While small‐scale batches are typically performed highly parallel and in high throughput, large‐scale cultivations demand sophisticated equipment for process control which is in most cases costly and difficult to handle. Currently, there is no technical system on the market that realizes simple process control in high throughput. The novel concept of a microfermentation system described in this work combines a fiber‐optic online‐monitoring device for microtiter plates (MTPs)—the BioLector technology—together with microfluidic control of cultivation processes in volumes below 1 mL. In the microfluidic chip, a micropump is integrated to realize distinct substrate flow rates during fed‐batch cultivation in microscale. Hence, a cultivation system with several distinct advantages could be established: (1) high information output on a microscale; (2) many experiments can be performed in parallel and be automated using MTPs; (3) this system is user‐friendly and can easily be transferred to a disposable single‐use system. This article elucidates this new concept and illustrates applications in fermentations of Escherichia coli under pH‐controlled and fed‐batch conditions in shaken MTPs. Biotechnol. Bioeng. 2010;107: 497–505. © 2010 Wiley Periodicals, Inc.