Identification of a Yersinia pestis-specific DNA probe with potential for use in plague surveillance

McDonough, Kathleen A.; Schwan, Tom G.; Thomas, Rex E.; Falkow, Stanley

doi:10.1128/jcm.26.12.2515-2519.1988

Cited by 26 publications

(13 citation statements)

References 33 publications

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“…EcoH\ and Pst\ under the manufacturers' suggested conditions. Desired fragments were isolated from low-melting-point agarose as previously described (McDonough et al. 1988).…”

Section: Bacterial Strains and Growth Conditionsmentioning

confidence: 99%

A Yersinia pestis‐specific DNA fragment encodes temperature‐dependent coagulase and fibrinolysin associated phenotypes

McDonough

Falkow

1989

Molecular Microbiology

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The effect of temperature on coagulase and fibrinolysin expression (Pla) by Yersinia pestis has been implicated in the transmission of plague by fleas. In an attempt to improve our understanding of this process, we have cloned, sequenced and characterized the gene encoding the Pla phenotypes in Y. pestis, and examined its temperature-dependent regulation. The coding region for this gene overlaps a 900bp Y. pestis-specific DNA fragment that we have previously shown to be capable of detecting plague bacilli in fleas. The pla gene contains a single open reading frame encoding 312 amino acids with a predicted molecular weight of 34.7 kD and a putative signal sequence of 20 amino acids. This coding region appears to be sufficient for both coagulase and fibrinolytic activities. In Y. pestis, modulation between coagulase and fibrinolytic activities is temperature-dependent: coagulase activity is most evident at temperatures below 30 degrees C but fibrinolytic activity increases with higher temperatures (greater than 30 degrees C), regardless of the temperature at which the bacteria are grown. Our results lead us to believe that this regulation occurs post-translationally. It is possible that the alternative forms of the Pla protein are essential to 'flea blockage' and subsequent transmission of the plague bacillus to animals.

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“…EcoH\ and Pst\ under the manufacturers' suggested conditions. Desired fragments were isolated from low-melting-point agarose as previously described (McDonough et al. 1988).…”

Section: Bacterial Strains and Growth Conditionsmentioning

confidence: 99%

A Yersinia pestis‐specific DNA fragment encodes temperature‐dependent coagulase and fibrinolysin associated phenotypes

McDonough

Falkow

1989

Molecular Microbiology

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“…Recently, molecular techniques have been proposed as a means of more rapidly identifying plague bacteria in fleas. DNA hybridization probes were developed but were unable to reliably detect fewer than 10 5 plague bacteria (13,17). More recently, several PCR assays have been developed for plague diagnosis (1,7,11).…”

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confidence: 99%

PCR Detection of Yersinia pestis in Fleas: Comparison with Mouse Inoculation

et al. 1999

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The “gold standard” for identifying Yersinia pestis-infected fleas has been inoculation of mice with pooled flea material. Inoculated mice are monitored for 21 days, and those that die are further analyzed for Y. pestis infection by fluorescent-antibody assay and/or culture. PCR may provide a more rapid and sensitive alternative for identifying Y. pestis in fleas. To compare these assays, samples were prepared from 381 field-collected fleas. Each flea was analyzed individually by both PCR and mouse inoculation. Sixty of the 381 flea samples were positive forY. pestis by PCR; 48 of these PCR-positive samples caused death in mice (80.0% agreement). None of the 321 PCR-negative samples caused death. Among the 12 mice that survived inoculation with PCR-positive samples, 10 were later demonstrated by serology or culture to have been infected with Y. pestis. This suggests that death of inoculated mice is less reliable than PCR as an indicator of the presence of Y. pestis in flea samples. Mouse inoculation assays produce results that are comparable to PCR only when surviving as well as dead mice are analyzed for infection. The rapidity and sensitivity (10 to 100 CFU of Y. pestis) of PCR suggest that it could serve as a useful alternative to mouse inoculation for routine plague surveillance and outbreak investigations.

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“…These methods are reliable but require weeks for their completion and the use of laboratory animals. Recently, a DNA probe hybridization method was developed to detect Y pestis in fleas (11,20). However, the DNA probe method only detects fleas infected with 105 or more bacteria (11,20).…”

mentioning

confidence: 99%

“…Recently, a DNA probe hybridization method was developed to detect Y pestis in fleas (11,20). However, the DNA probe method only detects fleas infected with 105 or more bacteria (11,20). We describe here a rapid and sensitive alternative method for the direct detection of as few as 10 Y pestis cells in infected fleas using the polymerase chain reaction (PCR).…”

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confidence: 99%

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New method for plague surveillance using polymerase chain reaction to detect Yersinia pestis in fleas

1993

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Yersinia pestis, the plague bacillus, infects a variety of mammals throughout the world and is transmitted by fleas. We developed a polymerase chain reaction (PCR) test using primers designed from the Y. pestis plasminogen activator gene to directly detect plague-infected fleas. As few as 10 Y. pestis cells were detected, even in the presence of flea tissue, by PCR and then agarose gel electrophoresis and ethidium bromide staining.

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Identification of a Yersinia pestis-specific DNA probe with potential for use in plague surveillance

Cited by 26 publications

References 33 publications

A Yersinia pestis‐specific DNA fragment encodes temperature‐dependent coagulase and fibrinolysin associated phenotypes

A Yersinia pestis‐specific DNA fragment encodes temperature‐dependent coagulase and fibrinolysin associated phenotypes

PCR Detection of Yersinia pestis in Fleas: Comparison with Mouse Inoculation

New method for plague surveillance using polymerase chain reaction to detect Yersinia pestis in fleas

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