A new pentaplex-nested PCR to detect five pathogenic bacteria in free living amoebae

Calvo, Laura; Gregorio, I.; Garcı́a, Agustı́n Costa; Fernández, María Teresa; Goñi, Pilar; Clavel, Antonio; Peleato, M. Luisa; Fillat, Marı́a F.

doi:10.1016/j.watres.2012.09.039

Cited by 20 publications

(20 citation statements)

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“…The isolation of FLA naturally infected with Ps. aeruginosa [5,[26][27][28][29] demonstrated the role of amoebae and their cysts as vectors for these intracellular bacteria [30]. Our results showed the intracellular multiplication of Ps.…”

Section: Co-culture Of a Spelaea And Bacteriasupporting

confidence: 53%

Allovahlkampfia spelaea is a Potential Environmental Host for Pathogenic Bacteria

Mohamed¹,

Huseein²,

Farrag³

et al. 2016

J Bacteriol Parasitol

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Allovahlkampfia spelaea was identified for the first time in 2009. As a free living amoeba, it has been suggested to be a protective host for some bacterial pathogens against harsh environmental conditions and can transmit them to vulnerable hosts. We aimed in this study to test the interactions between Allovahlkampfia spelaea and some waterborne and foodborne bacteria and unravel if the tested bacteria can survive and multiply inside amoeba. We used a keratitis isolate of Allovahlkampfia spelaea grown in PYG medium containing proteose peptone, yeast extracts, and glucose. We examined amoeba interactions with Methicillin resistant Staphylococcus aureus, Escherichia coli 1, Klebsiella pneumoniae, Enterobacter aerogenes, Citrobacter cloaca, Proteus mirabilis, Raoultella terrigena, Raoultella ornitholytica, Aeromonas hydrophila and Pseudomonas aeruginosa using the co-culture assays. Amoebal survival rate with different bacterial strains were determined. With the exception of Proteus mirabilis that showed decreased survival rates inside amoebal cells, other bacterial isolates could survive and multiply inside Allovahlkampfia spelaea that was associated with decreased survival rates of the amoeba. Particularly, Pseudomonas aeruginosa, Aeromonas hydrophila and MRSA exhibited significantly increased multiplication rates inside amoeba. Our study demonstrated that Allovahlkampfia spelaea may act as a replicative host for pathogenic bacteria with environmental and clinical implications.

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Section: Co-culture Of a Spelaea And Bacteriasupporting

confidence: 53%

Allovahlkampfia spelaea is a Potential Environmental Host for Pathogenic Bacteria

Mohamed¹,

Huseein²,

Farrag³

et al. 2016

J Bacteriol Parasitol

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“…Many studies have evaluated the interaction between Acanthamoeba spp. and Pseudomonas spp., as well as investigated the presence of these bacterial genera as amoeba-resistant bacteria 3 , 8 , 11 .…”

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confidence: 99%

IDENTIFICATION OF Pseudomonas spp. AS AMOEBA-RESISTANT MICROORGANISMS IN ISOLATES OF Acanthamoeba

Maschio

Corção

Rott

2015

Rev. Inst. Med. trop. S. Paulo

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“…Asides from these PCR based, there is a nested PCR which has permitted the simultaneous detection of a group of potentially pathogenic bacteria, Legionella pneumophila , Mycobacterium spp and Pseudomonas spp, and adds the detection of toxic cyanobacteria of the genus Microcystis due to environmental and health implications of toxic blooms. Thus, the presence of these potentially pathogenic bacteria has been demonstrated in natural, recreational and drinking water supplies (Calvo et al ., ; Garcia et al ., ).…”

Section: Molecular Techniquesmentioning

confidence: 97%

“…The subsequent PCR to apply depend on the objective of the research, but the use of universal primers, 16sRNA type, can lead to mixed amplifications that cannot be resolved by sequencing. The multiplex‐nested PCR described by Calvo and colleagues () is simple to perform, requires only basic equipment and allows the detection of various genera of potentially pathogenic bacteria simultaneously with good sensitivity and specificity. The RT‐PCR techniques allow us to quantify amoebae and bacteria inside them, but they are complex and require expensive equipment and reagents.…”

Section: Molecular Techniquesmentioning

confidence: 99%

“…However, it is not always possible since the uncultivable bacteria or these in low number cannot be recovered, and in this case, molecular techniques may be used. The most common technique includes amplification of a fragment of the 16S rRNA gene by PCR or real-time PCR in which case quantification of the endosymbionts may be Calvo et al, 2013 Identification of amoeba endosymbionts 343 performed (Salah et al, 2009). If previous cell sorting had been performed to obtain individual cells, genetic studies to identify the bacteria included in amoebae could be made by amplification of 16S rRNA fragments, using bacterial universal primers (Lane, 1991;Horn and Wagner, 2004;Corsaro and Venditti, 2009;.…”

Section: Molecular Techniquesmentioning

confidence: 99%

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Identifying endosymbiont bacteria associated with free‐living amoebae

Goñi

Fernández

Rubio

2014

Environmental Microbiology

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SummaryThe association between free-living amoebae and pathogenic bacteria is an issue that has gained great importance due to the environmental and health consequences that it implies. In this paper, we analyse the techniques to follow an epidemiological study to identify associations between genera, species, genotypes and subgenotypes of amoebae with pathogenic bacteria, analysing their evolution and considering their usefulness. In this sense, we highlight the combination of microscopic and molecular techniques as the most appropriate way to obtain fully reliable results as well as the need to achieve the standardization of these techniques to allow the comparison of both environmental and clinical results.

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A new pentaplex-nested PCR to detect five pathogenic bacteria in free living amoebae

Cited by 20 publications

References 53 publications

Allovahlkampfia spelaea is a Potential Environmental Host for Pathogenic Bacteria

Allovahlkampfia spelaea is a Potential Environmental Host for Pathogenic Bacteria

IDENTIFICATION OF Pseudomonas spp. AS AMOEBA-RESISTANT MICROORGANISMS IN ISOLATES OF Acanthamoeba

Identifying endosymbiont bacteria associated with free‐living amoebae

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