Papers by Keyword: Polymerase Chain Reaction (PCR)

Abstract: nfectious microbial pathogens constitute the largest cause of morbidity and mortality worldwide. Early diagnosis and rapid infection control measures can lead to improved outcomes, earlier discharges and reduced nosocomial infections. Conventional diagnostic methods for infectious diseases such as microscopy, culture, and immunological methods, in most cases, are not universally applicable, less sensitive and could take from days to months to complete depending on the pathogen. Molecular assays based on nucleic acids such as polymerase chain reaction (PCR) have improved the sensitivity, specificity and turn-around time in diagnostic microbiology laboratories. These tests are particularly important to detect very low levels of pathogens in clinical samples, and for organisms that have long half-lives or are non-culturable. However, individual molecular tests are available for only a limited number of the more common infectious agents. Moreover, infectious disease events arising from novel pathogens or genetic variants have significantly increased, recently, for which, routine diagnostic methods are not yet available. Therefore, molecular methods and technologies capable of detecting multiple pathogens in a single test have become available over the last few years. Although, these methods are based on the conventional nucleic acid amplification and hybridization chemistry, enhanced multiplexing capability has been achieved through innovations in nucleic acid labeling techniques, and post-amplification analytic methods and instrumentation. The availability of these test kits brought a new level of convenience to the physicians ordering practices, and to the laboratory personnel, as they require very little hands on time. However, these tests are yet unaffordable to many laboratories, and in many cases, the sensitivity is poor compared to that of single-target, real-time PCR assays. Looking into the future, the revolutionary, next generation sequencing (NGS) technology is now being considered as a potential method for rapid identification of hundreds of pathogens, in an unbiased manner, with a single test that could significantly benefit patients who are critically ill with undiagnosed disease.

Abstract: This paper reviews several suitable microvalves for polymerase chain reaction (PCR). First, three type micro-valves in PCR chips are discussed, including pneumatic, servomotor-controlled and passive plug microvalves. Then we present our servomotor-controlled microvalves, with the structure of long passive plug. This valve had many obvious advantages such as simple fabrication and operation, perfect sealing, the ability to withstand relatively high pressure. Furthermore, the microvalve can be operated in a self-actuated mode.

Abstract: The on-chip platinum micro-heater prototypes for thermal cycling equipped with platinum temperature sensor are fabricated. The device has been designed, fabricated and characterized to explore the feasibility of the micro-heater for a fully integrated disposable lab-on-a-chip with the PCR module. The on-chip micro-heater demonstrates that the temperature transitions are shorter by comparison with the conventional PCR temperature routines.

Abstract: Genetic determinants for heavy metal resistance could be exploited in the design of bioprocesses for environmental cleanup. The removal of Ni(II), Co(II), Cr(VI) and Mn(II) was investigated using Serratia marcescens strains C-1, C4, 16 and Kluyvera sp. Nic3 isolated from nickel deposits in Moa, Holguín (Cuba). The high nickel and cobalt resistance of S. marcescens C-1 is based on the NreB- genetic determinant ncrABC, which encodes histidine-rich proteins, NcrA, NcrB and NcrC. NcrC is a Ni(II)/Co(II) uptake protein. In this work the presence of ncrAB fragment in S. marcescens C4, 16 and Kluyvera sp. Nic3 was determined by PCR. ncrAB fragment was efficiently amplified from genomic DNA of the strains C4 and 16 but not from strain Nic3. Strains C4, 16 and C-1 showed the highest resistance to Ni(II) and Co(II). The Ni(II)/Co(II) removal capacities of C4, 16 and C-1 strains were two times higher than that by Nic3 (12.88 mg of Ni(II)/g of biomass and 9.77 mg of Co(II)/g of biomass). Uptake of Cr(VI) and Mn(II) was not observed for any of these strains. pH value has an important influence on the Ni(II) and Co(II) removal capacity of S. marcescens C-1. Additional studies are currently in progress aimed to check the metal removal efficiency of strain C1 in batch reactors.

Abstract: The largest lignite mining area in Europe is located 150 km southeast of Berlin. Acidic lakes exist in this area, known to be caused by marcasite oxidation. Thirty-two samples from the open-pit brown coal-mine Jaenschwalde were analyzed for microorganisms. Cell numbers determined after separation from sand particles revealed concentrations of 102 to 107 microorganisms per g sample. In samples exposed to the air within an hour, up to 4x107 cells were counted. Measurement of metabolic activity by microcalorimetry showed for such samples up to 50 µW per g sand, whereas in heap samples (with low moisture) low or even no activity was measurable. DNA extraction was successful for 28 samples. In 26 samples microbial 16S rRNA genes were amplified by PCR. Acidithiobacillus ferrooxidans and At. thiooxidans specific amplificates were detected by nested PCR in 23 and 10 cases, respectively. A specific signal indicating Leptospirillum ferrooxidans was obtained with nine samples. Random samples were sequenced and showed 96 to 99 % identity with published data of all three species. Surprisingly, in four samples archaeal 16S rRNA genes were amplified by PCR. Sequencing of two samples showed 99 % identity with unidentified or uncultured archaea found in NCBI-databases. Molecular biology results for At. ferrooxidans as well as for At. thiooxidans were supported by successful isolations of pure cultures in 23 cases. Cultivation of the archaea failed so far. These data indicate that iron- and sulfur-oxidizing microorganisms occur at these sites in large numbers. If in addition the evidence for archaea can become verified, a screening for hot spots as the sites of their occurrence would become interesting.

Abstract: Biochip is an emerging technology and has evoked great research interests in recent years. In this paper, a novel air-driven loop-type microfluidic biochip was investigated. Differing from conventional micro channels, this chip has a micro loop-channel and 3 sets of driving conduits with valveless design in their intersections so that the microfluid can be driven smoothly in unidirectional circular movements. The driving efficiency reaches the highest if the entry angle of driving conduits is in the tangent direction of the loop-channel. However, the smaller the included angle, the impact area the larger, leading to comparatively serious reflow phenomenon. Furthermore, the microfluid can be controlled to stop almost instantaneously in the loop segment. Therefore, this loop-type biochip is suitable for biochemical reactions under repeated multiple temperature operations such as polymerase chain reaction. A full circular movement completes a cycle of PCR amplification. Besides, this biochip has its merits including simpler chip design, shorter channel length, and flexible controllability for biochemical reactions.