Rapid and visual detection of Toxoplasma gondii oocyst in cat feces using loop-mediated isothermal amplification (LAMP) assay.

Toxoplasma gondii is an obligate parasitic protozoon that transmits to animals and humans via ingested food. Cats that act as T. gondii's final hosts play a critical role in T. gondii transmission by shedding millions of oocysts. Timely diagnosis of infected cats is essential for preventing toxoplasmosis because oocysts are a putative T. gondii source in epidemiology. We developed a new visual LAMP assay targeting the B1 gene to analyze single oocysts in cat feces in this study. The amplification result could be visually estimated based on the color change. LAMP assay analytical sensitivity was 101 copies/µL for the B1 gene plasmid, which was tenfold better than the PCR reaction. There were no cross-reactions with other parasites. The LAMP assay can detect a single T. gondii oocyst in 200 mg of cat feces. The LAMP assay detected a single oocyst in 200 mg cat feces at a higher rate than the PCR assay (83.3% vs. 50.0%).

Rapid visual detection of the allergen Dermatophagoides farinae in house dust by loop-mediated isothermal amplification.

Dermatophagoides farinae is considered to be an important factor causing some allergic diseases, such as urticaria, allergic rhinitis, asthma, and other interrelated diseases. Avoiding exposure to allergens is the most effective way to reduce allergic reactions. In this study, we successfully established a loop-mediated isothermal amplification (LAMP) method for the detection of D. farinae DNA target internal transcribed spacer (ITS) and D. farinae 1 allergen (Der f 1) genes. The turbidity-monitoring system and visual fluorescent reagents were used to verify the test results of LAMP assay. Following optimization of the primers and reaction temperatures, the amplification sensitivity, specificity, and efficiency of the method for detecting D. farinae were assessed. There was no cross-reaction with other arthropod species that are commonly found in indoor environmental dust, such as Dermatophagoides pteronyssinus, Alophagoides ovatus, Periplaneta americana, Anopheles sinensis, and Musca domestica. Furthermore, the sensitivity of LAMP assay for detecting D. farinae DNA was 10 times greater than that of conventional PCR. The positive detection rate by the LAMP method was greater than the conventional PCR for both single D. farinae mites and D. farinae mites in indoor dust. A new type of LAMP method for D. farinae based on the Der f 1 and ITS genes was, therefore, successfully established. This study is the first time to detect the D. farinae allergen using LAMP assay. This assay could be useful as a model for the rapid detection of allergens produced by other house dust mites in the future.

Detection and assessment of dust mite allergens in an indoor environment in Anhui, China.

Dust in the home environment is thought to be a potential trigger for increasing allergic diseases, such as allergic rash, rhinitis, asthma, and other conditions, associated with dust mites. To verify the status of dust mite prevalence in indoor surroundings, we collected 189 dust samples from the air conditioner filters (n = 75) and floors (n = 114) of households, schools, and hotels in the Anhui area, China. All samples were measured for dust mite breeding rate and breeding density under light microscopy and analyzed for dust mite species Dermatophagoides farinae 1 (Der f 1) and Dermatophagoides pteronyssinus 1 (Der p 1) allergen using enzyme-linked immunosorbent assay (ELISA). The dust mite breeding rates were 34.67% (26/75) and 20.18% (23/114), respectively, in the dust samples from the floor and air conditioning filters. The breeding density was the highest in households (10/g), followed by schools (9/g) and hotels (4/g). ELISA indicated that the allergen threshold (2.0 µg/g dust) of Der f 1 was exceeded in only two samples and Der p 1 in one sample. Additionally, a questionnaire was used to investigate the health knowledge on allergic diseases involved in indoor facilities, finding that most allergy sufferers were aware that indoor dust might be responsible for their conditions. The findings suggest that regular maintenance of indoor hygiene and cleaning of air-conditioning filters should reduce the risks of exposure to indoor allergens.