Evaluation of three PCR schemes for detection of Bartonella bacilliformis in blood samples: sensitivity, specificity and applicability

2.50
Hdl Handle:
http://hdl.handle.net/10757/347086
Title:
Evaluation of three PCR schemes for detection of Bartonella bacilliformis in blood samples: sensitivity, specificity and applicability
Authors:
Gomes, C.S.P.; Silva, W.; Tinco, C.; Martinez Puchol, S.; Pons, Maria J ( 0000-0001-8384-2315 ) ; Bazan, Jorge; Del Valle Mendoza, Juana; Ruiz, J.
Publisher:
Elsevier B.V.
Issue Date:
24-Mar-2015
URI:
http://hdl.handle.net/10757/347086
Abstract:
Background: Bartonella bacilliformis is the etiological agent of Carrion’s disease, a neglected illness with a febrile lethal stage and a warty benign phase, being the human the only known reservoir. The diagnostic by microscopy in endemic areas is several times erroneous. Furthermore, the culture of this bacterium is time-consuming, being the diagnostic by PCR the easiest way to perform a correct diagnostic. The objective of this study was to evaluate the detection limit of three PCR schemes, designed to detect B.bacilliformis, both in blood and filter papers to test their potential use for transferring samples from endemic areas to reference centers. Moreover, the specificity was also observed as well as the applicability of the technique with clinical samples from different stages of the disease. Methods & Materials: Fragments of 16SrRNA and fla genes were amplified as well as the variable-intergenic region (its). The detection limit was determined by bacterial quantification with flow cytometry and performing dilutions (106cfu/ml-10cfu/ml) both in blood and filter papers. DNA was extracted and PCRs were performed. Specificity was tested by processing other bacteraemia microorganisms. Clinical samples, 12 from febrile patients, 13 from warty and 71 from healthy asymptomatic individuals living in endemic area(Mandinga-Cajamarca) were also processed. Results: The 16SrRNA PCR scheme showed the lower detection limit (5 cfu from blood and filter paper) being the PCR scheme chosen to be tested in clinical samples. All febrile patients’ samples were positive, whereas in warty individuals only 3(23%) faint bands were obtained. No amplification was obtained in samples from healthy people. Fainter bands were always obtained when PCRs were made of filter papers. All PCRs were specific for B.bacilliformis. Conclusion: The 16SrRNA PCR seems to be the best technique to detect feverish patients. However, the applicability to identify asymptomatic carriers was undetermined. Filter paper may be an alternative for easy transportation of samples but is need to consider the decreasing sensitivity of the results. It is critical to develop rapid, sensitive and specific technique capable of being applied in endemic rural areas, to avoid misdiagnosis and facilitate the detection of asymptomatic carriers that will allow progress towards the eradication of this disease.
Type:
info:eu-repo/semantics/conferenceObject
Rights:
info:eu-repo/semantics/openAccess
Language:
eng
Description:
Revisión por pares; 16th International Congress on Infectious Diseases (ICID), 2014. 2-5 de Abril 2014. Cape Town, South Africa
ISSN:
1201-9712

Full metadata record

DC FieldValue Language
dc.contributor.authorGomes, C.S.P.es_PE
dc.contributor.authorSilva, W.es_PE
dc.contributor.authorTinco, C.es_PE
dc.contributor.authorMartinez Puchol, S.es_PE
dc.contributor.authorPons, Maria Jes_PE
dc.contributor.authorBazan, Jorgees_PE
dc.contributor.authorDel Valle Mendoza, Juanaes_PE
dc.contributor.authorRuiz, J.es_PE
dc.creatorUniversidad Peruana de Ciencias Aplicadas (UPC)es_PE
dc.date.accessioned2015-03-24T19:41:23Zes_PE
dc.date.available2015-03-24T19:41:23Zes_PE
dc.date.issued2015-03-24es_PE
dc.identifier.issn1201-9712es_PE
dc.identifier.urihttp://hdl.handle.net/10757/347086es_PE
dc.descriptionRevisión por pareses_PE
dc.description16th International Congress on Infectious Diseases (ICID), 2014. 2-5 de Abril 2014. Cape Town, South Africaes_PE
dc.description.abstractBackground: Bartonella bacilliformis is the etiological agent of Carrion’s disease, a neglected illness with a febrile lethal stage and a warty benign phase, being the human the only known reservoir. The diagnostic by microscopy in endemic areas is several times erroneous. Furthermore, the culture of this bacterium is time-consuming, being the diagnostic by PCR the easiest way to perform a correct diagnostic. The objective of this study was to evaluate the detection limit of three PCR schemes, designed to detect B.bacilliformis, both in blood and filter papers to test their potential use for transferring samples from endemic areas to reference centers. Moreover, the specificity was also observed as well as the applicability of the technique with clinical samples from different stages of the disease. Methods & Materials: Fragments of 16SrRNA and fla genes were amplified as well as the variable-intergenic region (its). The detection limit was determined by bacterial quantification with flow cytometry and performing dilutions (106cfu/ml-10cfu/ml) both in blood and filter papers. DNA was extracted and PCRs were performed. Specificity was tested by processing other bacteraemia microorganisms. Clinical samples, 12 from febrile patients, 13 from warty and 71 from healthy asymptomatic individuals living in endemic area(Mandinga-Cajamarca) were also processed. Results: The 16SrRNA PCR scheme showed the lower detection limit (5 cfu from blood and filter paper) being the PCR scheme chosen to be tested in clinical samples. All febrile patients’ samples were positive, whereas in warty individuals only 3(23%) faint bands were obtained. No amplification was obtained in samples from healthy people. Fainter bands were always obtained when PCRs were made of filter papers. All PCRs were specific for B.bacilliformis. Conclusion: The 16SrRNA PCR seems to be the best technique to detect feverish patients. However, the applicability to identify asymptomatic carriers was undetermined. Filter paper may be an alternative for easy transportation of samples but is need to consider the decreasing sensitivity of the results. It is critical to develop rapid, sensitive and specific technique capable of being applied in endemic rural areas, to avoid misdiagnosis and facilitate the detection of asymptomatic carriers that will allow progress towards the eradication of this disease.eng
dc.formatapplication/pdfes_PE
dc.language.isoenges_PE
dc.publisherElsevier B.V.es_PE
dc.rightsinfo:eu-repo/semantics/openAccesses_PE
dc.sourceUniversidad Peruana de Ciencias Aplicadas (UPC)es_PE
dc.sourceRepositorio Académico - UPCes_PE
dc.titleEvaluation of three PCR schemes for detection of Bartonella bacilliformis in blood samples: sensitivity, specificity and applicabilityes_PE
dc.typeinfo:eu-repo/semantics/conferenceObjectes_PE
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