# Technique Evolutions for Microorganism Detection in Complex Samples: A Review
## 3 Developments in Innovative Detection Methods
### Improvements and Developments in Analytical Methods Requiring Sampling
#### 3.2.1 Paper Sensors  
Paper sensors are strips of paper on which reagents, molecules, or ligands, can be grafted to make sensors. They can be used to detect the presence or absence of an analyte and can be useful in some diagnoses. The best-known example of the use of paper sensors is the pregnancy test. Paper sensors were initially intended for use in the field in developing countries where the installation of expensive and cumbersome equipment is impossible. However, their many advantages make them good laboratory tools too. With their low cost, ease of use, and portability (not very cumbersome and heavy) it becomes possible to multiply sampling for pathogen detection. During an industrial process using cell bioreactors or in the manufacturing of innovative therapy drugs, one can now easily imagine more controls at all critical steps of the process. Moreover, paper sensors can be integrated in a more complex device in order to automate the task by a robot and allow a quick reading of the result afterwards. Indeed, sampling takes time, it requires an operator and can lead to contamination. These constraints make it impossible to repeat the controls while they should be regular and representative of the whole sample. With the help of an automaton and paper sensors, these problems can be solved, in particular the problem of sterility breakage by the operator. Several techniques are possible, and several different ligands can be used with paper sensors. We review the main techniques. Paper sensors are affordable devices for food [71; 72; 73] or environmental quality control [74]. They can also be used for the detection of pathogens or the rapid and accurate detection of diseases [75; 76; 77]. Their versatility makes them useful for virus diagnosis [78] or detection of bacterial spores [79]. [PERSON] et al. reported an approach for the selective and sensitive multiplexed detection of _E. coli_ using a lab-on-paper test strip based on the activity of an intracellular enzyme. The objective is to trigger a rapid enzymatic hydrolysis of the substrate, causing a color change from colorless to blue or from yellow to red depending on the bacteria. With a step of preconcentration using immunomagnetic nanoparticles, the detection limit was about 10 CFU/mL for _E. coli_ in 30 min and without cell culture. Thus, these paper test strips may be suitable for the detection of viable pathogens in water, blood, or cell therapy products. In addition, if a culture step is added, it allows the detection of less than 1 CFU in 100 mL in 8 h, making the paper test strips useful for the detection of multiple pathogens in complex samples. However, a filtration step is required to remove enzymes from the medium before lysis of the microorganism. This allows selective detection of viable bacteria. The test strips can be adapted to different bacteria, or different intracellular enzyme markers by varying the nature of the reagents printed on the paper test strip. [PERSON] et al. presented a simple all-in-one paper-based sensor for the detection of _E. coli_ using a composite ink [77]. The paper consists of a fluorogenic DNAzyme probe for bacterial recognition and signal generation. It uses lysoszymes that lyse the bacteria and sugars that stabilize the printed bioactive molecules. A fluorescent signal can be detected within 5 min of contact between the bacteria and the paper probe. The detection limit is \(10^{2}\) cells per mL, in different sample matrices, without sample enrichment (Figure 3). In recent years, many efforts have been made to develop easy-to-use and inexpensive detection methods using paper sensors. Paper is readily available and can be modified by printing several types of reagents. Biorecognition items can be antibodies, nucleic acids, or proteins to detect a variety of analytes. The availability of high-throughput printers combined with microfabrication techniques allows the precise design of small diagnostic devices.