Detection of free-living amoebae by using multiplex quantitative PCR

Calvez, Thomas Le; Trouilhé, Marie-Cécile; Humeau, Philippe; Moletta-Denat, Marina; Frère, Jacques; Héchard, Yann

doi:10.1016/j.mcp.2012.03.003

Cited by 41 publications

(27 citation statements)

References 25 publications

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“…During the last few years, molecular techniques (PCR) are increasingly being used in the identification of the genotype (Booton et al 2005, Caumo et al 2009, Edagawa et al 2009, Goldschmidt et al 2012, Howe et al 1997, Itahashi et al 2011, Jain and Tilak 2011, Laummaunwai et al 2012, Le Calvez et al 2012. Our results confirmed the usefulness of PCR for the early, rapid and sensitive diagnosis of pathogenic Acanthamoeba spp.…”

Section: Discussionsupporting

confidence: 79%

Comparative analyses of different genetic markers for the detection of Acanthamoeba spp. isolates

Derda

Wojtkowiak-Giera

Hadaś

2014

Acta Parasitologica

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Acanthamoeba are widespread free-living amoebae which may cause granulomatous amoebic encephalitis (GAE), keratitis, skin ulcerations and disseminated tissue infection. An important diagnostic and prognostic factor for the treatment of infection is a quick and correct diagnosis of amoebae strains. The aim of our study was to develop a rapid method for detection and identification of pathogenic Acanthamoeba spp. strains from diagnostic material collected from water. In this study we analysed five amplification-based genetic markers (Aca 16S, Ac6/210, GP, JDP, Nelson) used for identification of pathogenic Acanthamoeba spp. strains isolated in water sources in Poland, Iceland and Sweden. Our results demonstrated the presence of pathogenic Acanthamoeba strains in tap water. PCR assay appeared to be a more rapid and sensitive method to detect the presence of amoebae than the limited conventional techniques. Based on our observations, we can confirm that the use of four out of five genetic markers (Aca 16S, Ac 6/210, JDP, GP, Nelson) may be helpful in identification of Acanthamoeba spp. strains, but only one Aca 16S primer pair is a highly specific marker that distinguishes between pathogenic strains of Acanthamoeba and other free-living amoeba families.

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Section: Discussionsupporting

confidence: 79%

Comparative analyses of different genetic markers for the detection of Acanthamoeba spp. isolates

Derda

Wojtkowiak-Giera

Hadaś

2014

Acta Parasitologica

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“…All DNA samples from water and soil were also screened for the presence of free-living amoebae. PCR analyses were performed by amplification of the 18S rRNA gene with primers specific for Amoebozoan (Amo_1400_F/Amo-1540-R) and Vahlkampfiidae (Vahl_ 560_F/Vahl_730_R), as described previously (25).…”

Section: Methodsmentioning

confidence: 99%

Assessment of Chlamydia psittaci Shedding and Environmental Contamination as Potential Sources of Worker Exposure throughout the Mule Duck Breeding Process

Hulin

Bernard

Vorimore

et al. 2016

Appl Environ Microbiol

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g Chlamydia psittaci is an obligate intracellular bacterium responsible for avian chlamydiosis, otherwise known as psittacosis, a zoonotic disease that may lead to severe atypical pneumonia. This study was conducted on seven mule duck flocks harboring asymptomatic birds to explore the circulation and persistence of C. psittaci during the entire breeding process and assess the potential sources of worker exposure. Cloacal swabs and air samples were taken on each occasion requiring humans to handle the birds. In parallel, environmental samples, including dust, water, and soil, were collected. Specific real-time PCR analyses revealed the presence of C. psittaci in all flocks but with three different shedding patterns involving ducks about the age of 4, 8, and 12 weeks with heavy, moderate, and low excretion levels, respectively. Air samples were only positive in flocks harboring heavy shedders. Dust in flocks with heavy or moderate shedders carried chlamydial loads strongly associated with the loads detected in avian and soil samples. Environmental contamination, significantly correlated with shedding dynamics, was considered to be the most probable source of exposure. The high prevalence of bacteriophage Chp1 in all flocks, mostly jointly present with chlamydia, suggests an important factor in C. psittaci persistence, thus creating a greater risk for humans. A survey conducted in these flocks regarding farming practices and activities showed that disinfection seems to be the most promising practice for reducing C. psittaci prevalence in ducks and that the place and the duration of action during operations seem to be potential risk factors. Strict adherence to good practices is strongly recommended.

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“…Further refinements in the PCR technique have also been developed to yield higher sensitivities. Le Calvez et al [27] recently described a multiplexed quantitative PCR method which is able to distinguish individual Acanthamoeba species in free-living mixed flora samples. Laummaunwai et al [28] developed an algorithm for DNA extraction which is able to produce viable DNA from a single Acanthamoeba cyst; previously cystic DNA extraction has been the Achilles heel of the technique.…”

Section: Diagnosismentioning

confidence: 99%