The Rapid Detection Of Microbes Based On Piezoelectric Sensor

Infectious diseases caused by microorganism have been a serious threat to human health,especially for bloodstream infection which has a high mortality.Timely and accurate detection of pathogens is very important for reducing the mortality of infectious diseases.At present,the automated microbial detection systems such as BACTEC products and Bac T/Alert products have been used for fast and exact detection of clinical microbes.However,the increase of CO2 in the process of detection caused by nonbacterial factors may lead to false positive results.Moreover,the radioactive contamination exists in the BACTEC 460.Besides,the expensive cost of these imported instructions increases the burden of patients.It is difficult to promote the widespread use of these instruments in poor or developing countries and can not meet the more clinical needs.Therefore,it is necessary to develop a novel automatic blood culture system for widespread application.Series piezoelectric sensor can sensitively detect the change of electrical parameters of conductive electrode and convert it into frequency change.By virtue of the advantages of simple operation,low cost,and real-time online monitoring,the series piezoelectric sensor has been widely used in analytical test.But it has not achieve selective and multiple samples detection.In this paper,several new sensors were constructed by introducing new materials and technologies on the basis of series piezoelectric quartz crystal to solve the current problems in microbial detection.The main works are shown as follows:1.A new aptamer-single walled carbon nanotubes-interdigital gold electrode-series piezoelectric quartz crystal(Apt-SWCNT-IDE-SPQC)sensor was first developed for rapid,selective detection of Group A Streptococcus(GAS).The selected aptamer was acted as recognition probe,the sensitive IDE-SPQC was used as detection platform,SWCNT with high specific surface area and good electrical conductivity were used as linker for signal amplification.The Apt modified on SWCNT-IDE-SPQC could specifically bind to GAS,this process leaded to the change of electric parameters of IDE and caused sensitive shift in frequency.The developed sensor could quantitative detection of GAS and the detection limit was 12 cfu/m L.The test could complete within 40 min.The new method was simple,fast,sensitive,high selectivity and low cost.It would be applied in the areas of environment,food and clinic for rapid detection and identification of GAS.2.Blood culture is the "gold standard" of clinical diagnosis of bloodstream infection.The traditional manual culture method is laborious and easy to be contaminated.The automated blood culture systems are convenient,rapid and accurate compared with the manual culture method.But the cost of these equipments and consumables is expensive that is unable to achieve the wide application.To meet clinical need,a new automated multi-channel piezoelectric blood culture system was developed based on the series piezoelectric sensor combing with the controller local area network(CAN)technology.The new system was simple and stable,and could detect multiple samples.In addition,the detection software was also improved to make the system be easily operated.For the microbial detection,the detection results were consistent with the Bac T/ Alert 3D,and the sensitivity was as low as 10cfu/m L.The new developed piezoelectric blood culture instrument is expected to reduce the medical cost and promote the development of the domestic blood culture system.3.Antibiotic has inhibitory effect on the growth of microorganism.The presence of antibiotics in detected samples is an important reason why low positive rate and prolonged time of blood culture.Therefore,the detection of clinical blood samples should consider the possible existence of antibiotics.To improve the positive detection rate and shorten the detection time,mixed resins were used to culture medium for adsorption of antibiotics.The ability of CAD-40,HPD-950,X-5,AB-8,HZ-90 resins and their combination to adsorb commonly used antibiotics was studied and the optimized mixed resins(0.20 g CAD-40,0.25 g HPD-950 and 0.35 g HZ-90 resins)were chosen by orthogonal test.The mixed resins(MR)were used to detect the simulated blood samples containing bacteria and antibiotics.The results proved that the medium containing MR could effectively enhance the accuracy rate when detection of bacteria in samples containing antibiotics.4.A novel method was developed for rapid and universal detection of microbes by pleurocidin-SWCNT-IDE-MSPQC sensor.The response mechanism of the sensor was based on the specific interaction between pleurocidin and microbial cell which could trigger the pleurocidin to detach from the SWCNT-IDE and cause the sensitive frequency shift response.Proposed method could successfully detect the common microbes in blood infections within 15 minutes.It would be applied for rapid microbial detection.5.In chapter 6,a free modified IDE-MSPQC combining with magnetic bead-aptamer-Poly(A)DNA was designed for selective detection of Pseudomonas aeruginosa.The complex of magnetic bead-aptamerPoly(A)DNA had a similar sandwich structure,magnetic bead was used as carrier for immobilization of the aptamer,Poly(A)DNA acted as signal probe which was modified onto magnetic beads by the complementary base pairing with aptamer.The response mechanism of the developed strategy was that the Pseudomonas aeruginosa specifically reacted with the aptamer immobilized on magnetic bead which made the Poly(A)DNA drop off,the Poly(A)DNA could bind to Au-IDE and trigger the sensitively frequency shift response of the MSPQC.The detection could finish within 10 min and the detection limit was 9 cfu/m L.The recovery rates of Pseudomonas aeruginosa in simulatd blood samples were 88 ~ 104%.It is expected to apply the sensor for rapid identification of Pseudomonas aeruginosa in the fields of clinical diagnosis,food and environment monitoring.