<p>Loop-mediated isothermal amplification for detection of <em>Legionella pneumophila</em> in respiratory specimens of hospitalized patients in Ahvaz, southwest Iran</p>

Abstract

Background Legionnaires’ disease is an important public health problem that can cause substantial mortality and morbidity. Legionnaires’ disease-risk estimation may be compromised by uncertainties in Legionella -detection methods. The aim of this study was the detection of L. pneumophila in respiratory specimens of hospitalized patients with respiratory symptoms by culture, PCR, and loop-mediated isothermal amplification (LAMP) methods. Methods Sputum and bronchoalveolar lavage samples were obtained from patients with pneumonia admitted to teaching hospitals in Ahvaz, Iran from June 2016 to March 2017. Isolation of Legionella spp. was done by culturing the samples directly onto buffered charcoal–yeast extract and modified Wadowsky–Yee agar medium. Then, PCR and LAMP assays were performed for detection of L. pneumophila via its mip gene in respiratory specimens. Results A total of 100 respiratory specimens were collected. Our results showed that 1% of the samples were culture positive for Legionella spp., and 3% and 7% of samples were positive for L. pneumophila using the mip gene on PCR and LAMP assays, respectively. Conclusion Legionnaires’ disease should be considered in the diagnosis of pulmonary infectious diseases. Also, the LAMP assay is a faster method with higher sensitivity and specificity than conventional methods, such as PCR and culture, for laboratory diagnosis of Legionnaires’ disease.

<p>Lateral flow biosensor combined with loop-mediated isothermal amplification for simple, rapid, sensitive, and reliable detection of <em>Brucella</em> spp</p>

“…To overcome the drawbacks posed by PCR-based assays, various other techniques, on the basis of isothermal amplification of nucleic acid, have been designed, which are able to rapidly synthesize DNA in large amounts without the use of any sophisticated apparatus 11. Among these isothermal amplification methods, loop-mediated isothermal amplification (LAMP) is one of the most widely used techniques, which has been employed for rapid, sensitive, and specific detection of many bacterial pathogens including Brucella spp 1,12–16.…”

Occurrence of the Legionella species in the respiratory samples of patients with pneumonia symptoms from Ahvaz, Iran; first detection of Legionella cherrii

“…To date, several studies have investigated the prevalence of different species of Legionella in clinical and environmental samples in various regions of Iran through conventional and modern methods including culture, urinary antigen test, PCR, and LAMP [13,14,17]. However, according to the available information, no study has been conducted to investigate the presence of different species of Legionella by the real-time PCR gene method targeting the 23S-5S rRNA gene spacer region and sequencing of this gene in clinical samples in Iran.…”

“…The colonies that resembled the Legionella spp were selected for further evaluation by standard bacteriological tests including Gram-staining, catalase, oxidase, and other biochemical tests. Besides, a number of colonies were cultured on a conventional unselective culture media such as sheep blood agar and MacConkey agar that lacked L-cysteine and incubated under previous conditions [13,14]. The isolates that were found to be Gram-negative rods under the microscope and were not able to grow on cysteine-free media were considered as Legionella species.…”

“…One portion was used for culture method and the other for real-time PCR assay. The specimens of molecular methods were kept at −70 ºC until analysis [14].…”

“…Over 90% of legionellosis infection being caused by L. pneumophila belonging to 16 serological groups, of which Lp 1 contributed to 80% of cases [9][10][11]. So far, various methods including enzyme-linked immunosorbent assays (ELISA), polymerase chain reaction (PCR), loop-mediated isothermal amplification (LAMP), and real-time PCR have been introduced to detect the Legionella species in clinical and environmental samples, however, the culture method is still considered as the gold standard diagnostic method [9][10][11][12][13][14]. In recent decades, due to the disadvantages of the culture method, including time-consuming and need for specific media, reliance on more modern methods such as nucleic acid amplification tests (NAATs) and whole genome sequencing has increased [5,6,9].…”