Evaluation of quantitative polymerase chain reaction assays targeting Mycobacterium avium, M. intracellulare, and M. avium subspecies paratuberculosis in drinking water biofilms

Mycobacterium avium (MA), Mycobacterium intracellulare (MI), and Mycobacterium avium subsp. paratuberculosis (MAP) are difficult to culture due to their slow growing nature. A quantitative polymerase chain reaction (qPCR) method for the rapid detection of MA, MI, and MAP can be used to provide data supporting drinking water biofilms as potential sources of human exposure. The aim of this study was to characterize two qPCR assays targeting partial 16S rRNA gene sequences of MA and MI and use these assays, along with two previously reported MAP qPCR assays (IS900 and Target 251), to investigate Mycobacterium occurrence in kitchen faucet biofilms. MA and MI qPCR assays demonstrated 100% specificity and sensitivity when evaluated against 18 non-MA complex, 76 MA, and 17 MI isolates. Both assays detected approximately 1,000 cells from a diluted cell stock inoculated on a sampling swab 100% of the time. DNA analysis by qPCR indicated that 35.3, 56.9 and 11.8% of the 51 kitchen faucet biofilm samples collected contained MA, MI, and MAP, respectively. This study introduces novel qPCR assays designed to specifically detect MA and MI in biofilm. Results support the use of qPCR as an alternative to culture for detection and enumeration of MA, MI, and MAP in microbiologically complex samples.

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Polymerase chain reaction–restriction fragment length polymorphism of the rpoB gene for identification of Mycobacterium avium subsp. paratuberculosis and differentiation of Mycobacterium avium subspecies

Diagnostic Microbiology and Infectious Disease ◽

10.1016/j.diagmicrobio.2011.01.014 ◽

2011 ◽

Vol 70 (1) ◽

pp. 65-71 ◽

Cited By ~ 5

Author(s):

Jake Whang ◽

Byung Soo Lee ◽

Go-Eun Choi ◽

Sang-Nae Cho ◽

Park Young Kil ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Restriction Fragment Length Polymorphism ◽

Mycobacterium Avium ◽

Length Polymorphism ◽

Fragment Length Polymorphism ◽

Rpob Gene ◽

Chain Reaction ◽

Polymerase Chain ◽

Mycobacterium Avium Subsp Paratuberculosis ◽

Restriction Fragment Length

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Comparison of polymerase chain reaction and microscopy for the detection of Fasciola spp. in the fecal matter of domestic bovines in Kalasin Province, Thailand

Veterinary World ◽

10.14202/vetworld.2021.2878-2882 ◽

2021 ◽

pp. 2878-2882

Author(s):

Sirikanda Thanasuwan ◽

Anupong Tankrathok

Keyword(s):

Polymerase Chain Reaction ◽

False Negative ◽

Economic Losses ◽

Rrna Gene ◽

28S Rrna ◽

Chain Reaction ◽

Fecal Matter ◽

Specificity And Sensitivity ◽

Polymerase Chain ◽

Fasciola Spp

Background and Aim: Fasciola spp. are important foodborne trematodes and waterborne zoonotic parasites that cause health problems and economic losses worldwide, including in Thailand. Fasciola spp. are usually detected by sedimentation or the formalin-ethyl acetate concentration technique (FECT) under microscopy, which is less specific and sensitive. Accurate detection is important to detect real incidence for protection against and elimination of fasciolosis in the area. This study aimed to determine the distribution of Fasciola spp. and compare the specificity and sensitivity of FECT under microscopy to that of polymerase chain reaction (PCR) in cattle feces. Materials and Methods: The study was conducted in Kalasin Province, Thailand. Feces of 46 cattle were investigated for infection with Fasciola spp. To detect infection, FECT under microscopy and PCR amplification of the 28S rRNA gene of Fasciola spp. were used to identify egg parasites. Results: Feces of 16 of 46 (34.78%) cattle were positive for Fasciola spp. using FECT under microscopy, whereas PCR showed that 67.39% (31 of 46) were positive for Fasciola spp. False-negative results were as high as 32.61% when diagnosed under microscopy. Conclusion: This study confirmed the infection of cattle with Fasciola spp. in Kalasin Province, indicating that PCR demonstrated higher sensitivity and specificity when diagnosing infection. FECT under microscopy can still be used as a primary and traditional method for diagnosis. However, relapse cases of Fasciola spp. and Paramphistomum spp. should be diagnosed by microscopy combined with PCR. This is the first report on the molecular distribution of fecal samples in cattle in Kalasin Province.

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