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Updated: Nov 29, 2025
High-throughput Detection of Respiratory Pathogens in Animal Specimens by Nanoscale PCR
Published on: November 28, 2016
Triplex Hybridization-Based Nanosystem for the Rapid Screening of Pneumocystis Pneumonia in Clinical Samples
Luis Pla1,2,3, Anna Aviñó3,4, Ramón Eritja3,4
1Instituto Interuniversitario de Investigación de Reconocimiento Molecular y Desarrollo Tecnológico, Universitat Politècnica de València, Universitat de València, Camino de Vera s/n, 46022 Valencia, Spain.
Abstract:
Pneumocystis pneumonia (PcP) is a disease produced by the opportunistic infection of the fungus Pneumocystis jirovecii. As delayed or unsuitable treatments increase the risk of mortality, the development of rapid and accurate diagnostic tools for PcP are of great importance. Unfortunately, current standard methods present severe limitations and are far from adequate. In this work, a time-competitive, sensitive and selective biosensor based on DNA-gated nanomaterials for the identification of P. jirovecii is presented. The biosensor consists of a nanoporous anodic alumina (NAA) scaffold which pores are filled with a dye reporter and capped with specific DNA oligonucleotides. In the presence of P. jirovecii genomic DNA, the gated biosensor is open, and the cargo is delivered to the solution where it is monitored through fluorescence spectroscopy. The use of capping oligonucleotides able to form duplex or triplex with P. jirovecii DNA is studied. The final diagnostic tool shows a limit of detection (LOD) of 1 nM of target complementary DNA and does not require previous amplification steps. The method was applied to identify DNA from P. jirovecii in unmodified bronchoalveolar lavage, nasopharyngeal aspirates, and sputum samples in 60 min. This is a promising alternative method for the routinely diagnosis of Pneumocystis pneumonia.
Insights
A new DNA-gated nanomaterial biosensor rapidly detects *Pneumocystis jirovecii*, the fungus causing Pneumocystis pneumonia (PcP). This sensitive tool offers a promising alternative for timely PcP diagnosis and treatment.
Area of Science:
- Biotechnology
- Nanomaterials Science
- Molecular Diagnostics
Background:
- Pneumocystis pneumonia (PcP) is a serious opportunistic infection caused by *Pneumocystis jirovecii*.
- Delayed or inadequate treatment of PcP increases mortality risk.
- Current diagnostic methods for PcP are limited and often inadequate, necessitating improved tools.
Purpose of the Study:
- To develop a rapid, sensitive, and selective biosensor for identifying *P. jirovecii*.
- To utilize DNA-gated nanomaterials for efficient detection of the pathogen.
- To provide a viable alternative for routine PcP diagnosis.
Main Methods:
- Fabrication of a biosensor using a nanoporous anodic alumina (NAA) scaffold.
- Capping NAA pores with DNA oligonucleotides specific to *P. jirovecii* DNA.
- Monitoring fluorescence spectroscopy to detect reporter dye release upon target DNA binding.
Main Results:
- The biosensor demonstrated high sensitivity with a limit of detection (LOD) of 1 nM for target DNA.
- No prior amplification steps were required for DNA detection.
- The method successfully identified *P. jirovecii* DNA in clinical samples (bronchoalveolar lavage, nasopharyngeal aspirates, sputum) within 60 minutes.
Conclusions:
- A novel DNA-gated nanomaterial biosensor offers a rapid and sensitive method for *P. jirovecii* detection.
- This biosensor presents a promising alternative to current diagnostic techniques for Pneumocystis pneumonia.
- The developed tool can aid in timely diagnosis and treatment of PcP.
