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Erschienen in: Malaria Journal 1/2007

Open Access 01.12.2007 | Research

Comparison of PCR-based detection of Plasmodium falciparum infections based on single and multicopy genes

verfasst von: Segun I Oyedeji, Henrietta O Awobode, Gamaliel C Monday, Eric Kendjo, Peter G Kremsner, Jürgen F Kun

Erschienen in: Malaria Journal | Ausgabe 1/2007

Abstract

PCR-based assays are the most sensitive and specific methods to detect malaria parasites.
This study compared the diagnostic accuracy of three PCR-based assays that do not only differ in their sequence target, but also in the number of copies of their target region, for the detection of Plasmodium falciparum in 401 individuals living in a malaria-endemic area in Nigeria. Compared to a composite reference generated from results of all the 3 PCR assays, the stevor gene amplification had a sensitivity of 100% (Kappa = 1; 95% CI = 1.000–1.000), 83% (Kappa = 0.718; 95% CI = 0.648–0.788) by SSUrRNA gene PCR and 71% (Kappa = 0.552; 95% CI = 0.478–0.627) by the msa-2 gene amplification.
Results from this study indicate that the stevor gene amplification is the most sensitive technique for the detection of P. falciparum. This assay may be an important reference standard, especially when a confirmatory technique with high sensitivity and specificity is needed for ruling out P. falciparum infection.
Hinweise

Authors' contributions

SIO contributed to the study design, data collection and analysis, interpretation of the results and preparation of the manuscript.
HOA contributed to the study design, data collection, interpretation of the results and preparation of the manuscript.
GCM contributed to the collection of data preparation of the draft manuscript.
EK contributed to the analysis of data and interpretation of the results.
PGK contributed to the study design, interpretation of the results and preparation of the manuscript.
JFK contributed to the study design, data analysis, interpretation of the results and preparation of the manuscript.
All authors read and approved the final manuscript.

Background

Prompt and accurate diagnosis is an important tool in the effective management and control of malaria, a disease which accounts for more than a million deaths annually [1, 2]. Microscopy remains the cheapest and most commonly used method for malaria diagnosis and relies on the microscopic examination of stained blood films. Microscopy, however, has its own limitations, even when performed by an expert. It is time-consuming and its sensitivity is limited, particularly when parasitaemia is low [3]. Sensitivity and specificity are of crucial importance in any identification method as a false negative result could result in the non-treatment of a potentially fatal disease and conversely a false positive result will expose individuals to unnecessary drug intake, with its associated side-effects and high cost, while leaving the true cause of the illness untreated. Misdiagnosis of malaria, however, could have fatal consequences [4]. It is, therefore, important that a reliable, rapid and efficient method for diagnosis, surveillance and epidemiological studies is identified.
Several new diagnostic techniques have been developed during the last decade, including antigen detection and fluorescence-based assays, which have the added advantage of moving the diagnosis of malaria nearer to the patient and even faster [5, 6]. These, also have drawbacks as their sensitivity decreases especially with low parasitaemia or they may have variability in performance: misdiagnosis with such methods have been reported [711].
PCR-based assays have been used in diagnosis and as reference or standard for the assessment of the sensitivity and specificity of microscopy and commonly deployed rapid diagnostic tests and may indeed, be of clinical value in some selected situations [1217]. Examples include: the PCR of stevor gene, a multiple copy sequence of the subtelomeric variable open reading frame of Plasmodium falciparum which were used in detection or measurement of very low parasitaemia [18, 19]; the genus and species-specific PCR, which was designed to amplify portions of the sequences coding for the small subunit ribosomal RNA (SSUrRNA) and has the added advantage of being able to detect all the four species of human Plasmodium, as well as the quantification of P. falciparum [20, 21]; and, the PCR typing of highly polymorphic merozoite surface antigens of P. falciparum, which has been particularly useful in determining genetic variability of parasite isolates [22] and in detection of microscopically undetected parasites in drug trials [23].
Despite the fact that PCR-based assays are meant to be highly sensitive, it is not known whether this sensitivity is equally good with all amplification techniques in use and high enough to justify their use as reference or standard in the diagnosis of P. falciparum infection. In this study, the sensitivity and specificity of three different PCR assays with target to mono- or multi-copy gene for the detection of P. falciparum were assessed.

Methods

Study site and sample collection

This study was conducted in Lafia, located within the Guinea savanna ecological zone in north-central Nigeria. Within this region, malaria transmission has formerly been described as stable and uniformly intense through most of the year [24, 25]. 401 children between 6 months and 8 years of age, from amongst those who reported for routine medical examination on complaint of fever, were enrolled into the study at the Dalhatu Araf Specialist Hospital (DASH), Lafia between November, 2005 and July, 2006. Informed consent was obtained from the parent or guardian of each child prior to being included in the study. Ethical approval for the study was granted by the Ethical Review Committees of the Nasarawa State Ministry of Health and the Dalhatu Araf Specialist Hospital, Lafia. Inclusion criteria are febrile conditions suggestive of malaria which includes chills, history of fever within the preceding 48 h or pyrexia at presentation (axillary temperature >37.5°C), or fever of unknown aetiology. Children with signs or symptoms of severe illness were excluded from the study.
Finger-prick blood samples were collected for thick and thin blood smear and on filter paper for PCR. Two drops of blood were spotted on each filter paper, air dried and individually sealed in plastic bags and stored at room temperature until DNA extraction. Giemsa-stained thick and thin blood films were examined for malaria parasites. Parasitaemia were quantified relative to 250 white blood cells (WBC) on thick films and estimated as parasites per μl assuming a mean WBC of 8,000 per μl of blood. Blood smears were labeled negative if no parasites were seen after examination of 200 oil immersion fields (×1,000) on a thick blood film.

DNA extraction and PCR

DNA was extracted from the dried blood spots on filter paper using the QIAamp® DNA Mini Kit (Qiagen, Hilden, Germany) following the manufacturer's protocol, and stored at -20°C until further analysis. 150 μl of distilled water was used to elute DNA. All PCR assays include a primary and nested reaction to enhance specificity. Amplification was performed using a BIOMETRA TB1 thermal cycler (Biotron, Göttingen Germany).

SSUrRNA gene PCR

In all reaction, 2.0 μl of DNA extract were amplified in a final volume of 25 μl containing 2.5 μl ×10 reaction buffer, 100 μM of each dNTPs (dATP, dGTP, dTTP, and dCTP), 0.5 pM of each primer (rPLU5/rPLU6 for the primary reaction and rFAL1/rFAL2 for the nested reaction) and 0.75 units of Taq DNA polymerase (Qiagen, Hilden, Germany). Primer sequences (Table 1) are based on the SSUrRNA sequences described elsewhere [26]. The PCR programme was: denaturation at 95°C for 5 min followed by 25 cycles (30 cycles in nested) of 1 min at 94°C, 2 min at 60°C and 2 min at 72°C and a final extension period of 5 min at 72°C.
Table 1
Nucleotide sequence of primers used in this study
Primer Name
Nucleotide sequence
rPLU5
5'-CCT GTT GTT GCC TTA AAC TTC-3'
rPLU6
5'-TTA AAA TTG TTG CAG TTA AAA CG-3'
rFAL 1
5'-TTA AAC TGG TTT GGG AAA ACC AAA TAT ATT-3'
rFAL 2
5'-ACA CAA TGA ACT CAA TCA TGA CTA CCC GTC-3'
P5
5'-GGG AAT TCT TTA TTT GAT GAA GAT G-3'
P18
5'-TTT CA(C/T) CAC CAA ACA TTT CTT-3'
P19
5'-AAT CCA CAT TAT CAC AAT GA-3'
P20
5'-CCG ATT TTA ACA TAA TAT GA-3'
P17
5'-ACA TTA TCA TAA TGA (C/T) CC AGA ACT-3'
P24
5'-GTT TGC AAT AAT TCT TTT TCT AGC-3'
MSA 2-1
5'-ATG AAG GTA ATT AAA ACA TTG TCT ATT ATA-3'
MSA 2-4
5'-TTA TAT GAA TAT GGC AAA AGA TAA AAC AAG-3'
MSA 2-2
5'-ACA TTC ATA AAC AAT GCT TAT AAT ATG AGT-3'
MSA 2-3
5'-GAT TAT TTC TAG AAC CAT GCA TAT GTC CAT-3'
FC 27-1
5'-GCA AAT GAA GGT TCT AAT ACT AAT AG-3'
FC 27-2
5'-GCT TTG GGT CCT TCT TCA GTT GAT TC-3'
3D7-1
5'-GCA GAA AGT AAG CCT TCT ACT GGT GCT-3'
3D7-2
5'-GAT TTG TTT CGG CAT TAT TAT GA-3'

stevor gene PCR

Primary amplification was performed with reaction mixture of 50 μl containing 5.0 μl ×10 reaction buffer, 200 μM of each dNTPs (dATP, dGTP, dTTP, and dCTP), 1.25 units of Taq DNA polymerase, 0.4 pM of each primer (P5, P18, P19 and P20) and 5.0 μl of DNA extract. The PCR programme was: denaturation at 93°C for 3 min followed by 25 cycles of 30 sec at 93°C, 50 sec at 50°C and 30 sec at 72°C and a final extension period of 3 min at 72°C. 2.0 μl of the first-round PCR product was used in the second round amplification which was performed with a reaction mixture of 50 μl containing 5.0 μl ×10 reaction buffer, 200 μM of each dNTPs, 1.25 units of Taq DNA polymerase and 0.4 pM of each primer (P17 and P24). The primer sequences for the stevor PCR are shown in Table 1. PCR conditions for the nested reaction was: denaturation at 93°C for 3 sec followed by 25 cycles of 30 sec at 93°C, 50 sec at 55°C and 30 sec at 72°C and a final extension period of 3 min at 72°C.

msa-2 gene PCR

The primary reaction was designed to amplify the entire coding region of the msa-2 gene using the msa-2-1 and msa-2-4 primers pairs (Table 1). The reaction mixture was performed in a final volume of 50 μl containing 5.0 μl of DNA extract, 5.0 μl ×10 reaction buffer, 200 μM of each dNTPs (dATP, dGTP, dTTP, and dCTP), 1.25 units of Taq DNA polymerase and 0.5 pM of each primer. The PCR programme was: denaturation at 94°C for 5 min followed by 35 cycles of 10 sec at 94°C, 30 sec at 57°C and 40 sec at 72°C and a final extension period of 3 min at 72°C. This was followed by two sets of nested reactions using allelic family-specific primers (FC27 and 3D7). A third reaction was performed to amplify the entire central variable region with the primer pairs msa-2-2 and msa-2-3 (Table 1), in order to detect sequences that may not be allelic family-specific. All nested reactions were performed in a final volume of 50 μl containing 2.0 μl of PCR product from primary reaction, 5.0 μl ×10 reaction buffer, 200 μM of each dNTPs, 0.5 pM of each primer and 1.25 units of Taq DNA polymerase. The PCR programme was: denaturation at 94°C for 5 sec followed by 30 cycles of 10 sec at 94°C, 30 sec at 57°C and 40 sec at 72°C and a final extension period of 3 min at 72°C.
PCR products were subjected to electrophoresis on 1.5–2% agarose gels and visualized by transillumination with ultraviolet light after staining with SYBR® Green. Fragment sizes were calculated relative to the standard size marker (100 bp DNA ladder) using the BioDocAnalyze (Biometra, Göttingen, Germany) computer software package.
All reactions were run in parallel with DNA from negative controls (non-exposed European samples) confirmed negative by the three PCR assays, and DNA from FCR-3 and 3D7 strains as positive controls. This strategy was used to enhance specificity and to ensure that the reaction does not spuriously amplify homologous sequences. For a reaction to remain valid, all negative controls in a run must remain negative while the positive controls must remain positive.

Statistical analysis

Data were entered into SPSS for Windows software package, version 11.0.0 (SPSS Inc., 2001). Sensitivity, specificity and Cohen's kappa coefficient (κ) with their 95% confidence intervals (CI) were determined using Stata version 9.2 (StataCorp, College Station, Texas). Cohen's kappa coefficient is a measure of the agreement between two tests beyond that expected by chance, where 0 is chance agreement and 1 is perfect agreement [27].

Results

Among the 401 children enrolled in this study, 169 presented with microscopically confirmed P. falciparum. The mean (± SD) age and weight of the children were 40.4 (± 19.2) months and 13.6 (± 3.1) Kg respectively. The baseline characteristics of the study children are shown in Table 2. A composite reference was generated and used as the gold standard to assess the sensitivity of each PCR assay. This was defined as a positive test by at least one of the three PCR assays relative to the positive and negative controls. This method was adopted in order to avoid errors in measuring the sensitivity of the techniques if a particular assay was chosen as the "gold standard" when it does not have 100% sensitivity. Sensitivity was defined as the probability that a truly infected individual will test positive and was assessed relative to the FCR-3 and 3D7 strains as positive controls. Specificity was defined as the probability that a truly uninfected individual will test negative and was assessed in 86 unexposed European samples.
Table 2
Baseline characteristics of children enrolled into the study
Number of subjects
401
Mean age (months)
40.4 (± 19.2)*
Sex (male/female)
203/198
Mean temperature (°C)
38.4 (± 1.2)*
Mean haematocrit (%)
28 (± 3.1)*
Mean weight (Kg)
13.1 (± 3.2)*
% positive by microscopy
42.1% (169/401)
% positive by stevor PCR
71.1% (285/401)
% positive by SSUrRNA PCR
59.1% (237/401)
% positive by msa-2 PCR
50.1% (201/401)
* ± Standard deviation in parentheses
A total of 285 patients were detected to be infected with P. falciparum by the composite reference. Cohen's kappa coefficient showed that the stevor PCR has a complete agreement with the composite reference having a Kappa (95%CI) of 1.00 (1.000–1.000), followed by the SSUrRNA PCR κ = 0.718 (0.648–0.788) and the msa-2 PCR κ = 0.552 (0.478–0.627). The results of the detection of P. falciparum by the three PCR assays are shown in Table 3. All the 86 unexposed European samples tested negative with the three PCR assays thus, implying 100% specificity (Table 3). The specificity of microscopy could not be determined from the unexposed European samples because their slides were not available for microscopy.
Table 3
Sensitivity of the PCR-based assays for the detection of P. falciparum
 
stevor PCR
SSUrRNA PCR
msa-2 PCR
Sensitivity
100%
83%
71%
Specificity
100%
100%
100%
Correctly classified
100%
87.2%
77.6%
Agreement (Kappa)a, b
1.00 (1.000–1.000)
0.718 (0.648–0.788)
0.552 (0.478–0.627)
aComposite reference compared with study assay; Kappa = 1 would indicate complete agreement.
b95% confidence intervals in parentheses.
To determine reproducibility, all samples that did not show identical result with the three PCR techniques were classified as discordant and were re-amplified. In all, 84 samples were re-examined and the PCR results obtained were 100% reproducible.
The diagnostic accuracy of the stevor PCR was significantly different to the other two PCR assays (P < 0.001) and, likewise, that of the SSUrRNA PCR was significantly different from the msa-2 PCR (P < 0.001).

Discussion

Routine microscopic examination of Giemsa-stained blood smears is usually considered as the gold standard for malaria diagnosis. However, this technique requires highly skilled personnel and the information obtained by microscopy is limited when parasite levels are very low or when parasite morphology is altered. This has prompted the development of rapid diagnostic assays based on the detection of parasite antigens in whole blood and DNA based methods for the detection of parasites, mainly the clinically important P. falciparum infection. Rapid diagnostic tests, however, have low sensitivity at parasitaemia below 100 parasites/μl and have insufficient accuracy (Rubio, 2001).
PCR-based methods have been consistently shown to be powerful tool for malaria diagnosis [2831]. Despite the fact that PCR-based assays have better sensitivity than conventional microscopy and antigen-based diagnostic tests [32], observations from this study show that their level of sensitivity could vary depending on the approach employed and the characteristic of the target sequence of the chosen assay. In this study, the stevor PCR was found to have the highest sensitivity for detecting P. falciparum, with 100% sensitivity and could correctly classify 100% of the population. This was followed by the SSUrRNA PCR with a sensitivity of 83% and could correctly classify 87% of the population, and then the msa-2 gene PCR with 71% sensitivity and could correctly classify 78% of the study population. Differences in sensitivity could thus, be correlated with the copy number of the amplified region although, sequence variations in target region of primers used might be a contributing factor. The stevor-based PCR is therefore, at a vantage position since it was designed to amplify fragments of a gene that has between 30 to 40 copies per haploid genome [33, 34] compared to the SSUrRNA gene which has about four copies per haploid genome [35, 36] or the msa-2 gene which is a single copy gene [37, 38].
One limitation of this study though, is that it focuses on P. falciparum infections and does not address other species of human Plasmodium. However, P. falciparum demands particular attention because, it is the species of human malaria parasite that causes the most severe form of the disease and can kill with stunning speed.

Conclusion

Despite having higher sensitivity and specificity in detecting Plasmodium infections, the use of PCR-based techniques in routine diagnosis is limited because of their logistical and technical difficulties [15]. They are labour-intensive and costly to maintain. However, PCR assays are invaluable tool in reference laboratories where quality assurance is required and, for public health surveillance in detecting submicroscopic asymptomatic infections. They could be particularly important in critical situations where a confirmatory assay with high sensitivity and specificity is needed for ruling out P. falciparum infection especially in non-immune travellers. The level of sensitivity of an assay though, could vary depending on the characteristics of the target sequence and the approach employed. This study shows that PCR assays with target to multiple copy sequences are more sensitive, the larger the copy number. As the scale of field studies grow, PCR diagnosis of malaria will play an increasing role in epidemiology with the development of high-throughput techniques that could facilitate large-scale analysis of samples within a short period.

Acknowledgements

We sincerely appreciate Andrea Weierich and Mr. Kyari (DASH) for their technical support in carrying out this study. We are grateful to the children in our study area and their parents for participating in this study. SIO was supported by a Deutscher Akademischer Austausch Dienst (DAAD) Research Fellowship.
Open Access This article is published under license to BioMed Central Ltd. This is an Open Access article is distributed under the terms of the Creative Commons Attribution License ( https://​creativecommons.​org/​licenses/​by/​2.​0 ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Authors' contributions

SIO contributed to the study design, data collection and analysis, interpretation of the results and preparation of the manuscript.
HOA contributed to the study design, data collection, interpretation of the results and preparation of the manuscript.
GCM contributed to the collection of data preparation of the draft manuscript.
EK contributed to the analysis of data and interpretation of the results.
PGK contributed to the study design, interpretation of the results and preparation of the manuscript.
JFK contributed to the study design, data analysis, interpretation of the results and preparation of the manuscript.
All authors read and approved the final manuscript.
Literatur
1.
Zurück zum Zitat WHO: Strategic Orientation Paper on Prevention and Control of Malaria. WHO, Geneva. 2005 WHO: Strategic Orientation Paper on Prevention and Control of Malaria. WHO, Geneva. 2005
2.
Zurück zum Zitat WHO: World Malaria Report 2005. WHO and UNICEF, Geneva. 2005 WHO: World Malaria Report 2005. WHO and UNICEF, Geneva. 2005
3.
Zurück zum Zitat Coleman RE, Maneechai N, Rachaphaew N, Kumpitak C, Miller RS, Soyseng V, Thimasarn K, Sattabongkot J: Comparison of field and expert laboratory microscopy for active surveillance for asymptomatic Plasmodium falciparum and Plasmodium vivax in western Thailand. Am J Trop Med Hyg. 2002, 67: 141-144.PubMed Coleman RE, Maneechai N, Rachaphaew N, Kumpitak C, Miller RS, Soyseng V, Thimasarn K, Sattabongkot J: Comparison of field and expert laboratory microscopy for active surveillance for asymptomatic Plasmodium falciparum and Plasmodium vivax in western Thailand. Am J Trop Med Hyg. 2002, 67: 141-144.PubMed
4.
Zurück zum Zitat Amexo M, Tolhurst R, Barnish G, Bates I: Malaria misdiagnosis: effects on the poor and vulnerable. Lancet. 2004, 364: 1896-1898. 10.1016/S0140-6736(04)17446-1.CrossRefPubMed Amexo M, Tolhurst R, Barnish G, Bates I: Malaria misdiagnosis: effects on the poor and vulnerable. Lancet. 2004, 364: 1896-1898. 10.1016/S0140-6736(04)17446-1.CrossRefPubMed
5.
Zurück zum Zitat Makler MT, Palmer CJ, Ager AL: A review of practical techniques for the diagnosis of malaria. Ann Trop Med Parasitol. 1998, 92: 419-433. 10.1080/00034989859401.CrossRefPubMed Makler MT, Palmer CJ, Ager AL: A review of practical techniques for the diagnosis of malaria. Ann Trop Med Parasitol. 1998, 92: 419-433. 10.1080/00034989859401.CrossRefPubMed
7.
Zurück zum Zitat Grobusch MP, Alpermann U, Schwenke S, Jelinek T, Warhurst DC: False-positive rapid tests for malaria in patients with rheumatoid factor. Lancet. 1999, 353: 297-10.1016/S0140-6736(05)74930-8.CrossRefPubMed Grobusch MP, Alpermann U, Schwenke S, Jelinek T, Warhurst DC: False-positive rapid tests for malaria in patients with rheumatoid factor. Lancet. 1999, 353: 297-10.1016/S0140-6736(05)74930-8.CrossRefPubMed
8.
Zurück zum Zitat Iqbal J, Khalid N, Hira PR: Comparison of two commercial assays with expert microscopy for confirmation of symptomatically diagnosed malaria. J Clin Microbiol. 2002, 40: 4675-4678. 10.1128/JCM.40.12.4675-4678.2002.PubMedCentralCrossRefPubMed Iqbal J, Khalid N, Hira PR: Comparison of two commercial assays with expert microscopy for confirmation of symptomatically diagnosed malaria. J Clin Microbiol. 2002, 40: 4675-4678. 10.1128/JCM.40.12.4675-4678.2002.PubMedCentralCrossRefPubMed
9.
Zurück zum Zitat Iqbal J, Siddique A, Jameel M, Hira PR: Persistent histidine-rich protein 2, parasite lactate dehydrogenase, and panmalarial antigen reactivity after clearance of Plasmodium falciparum monoinfection. J Clin Microbiol. 2004, 42: 4237-4241. 10.1128/JCM.42.9.4237-4241.2004.PubMedCentralCrossRefPubMed Iqbal J, Siddique A, Jameel M, Hira PR: Persistent histidine-rich protein 2, parasite lactate dehydrogenase, and panmalarial antigen reactivity after clearance of Plasmodium falciparum monoinfection. J Clin Microbiol. 2004, 42: 4237-4241. 10.1128/JCM.42.9.4237-4241.2004.PubMedCentralCrossRefPubMed
10.
Zurück zum Zitat Happi CT, Gbotosho GO, Sowunmi A, Falade CO, Akinboye DO, Oladepo O, Oduola AM: Malaria diagnosis: false negative parasight-F tests in falciparum malaria patients in Nigeria. Afr J Med Med Sci. 2004, 33: 15-18.PubMed Happi CT, Gbotosho GO, Sowunmi A, Falade CO, Akinboye DO, Oladepo O, Oduola AM: Malaria diagnosis: false negative parasight-F tests in falciparum malaria patients in Nigeria. Afr J Med Med Sci. 2004, 33: 15-18.PubMed
11.
Zurück zum Zitat Belizario VY, Pasay CJ, Bersabe MJ, de Leon WU, Guerrero DM, Bugaoisan VM: Field evaluation of malaria rapid diagnostic tests for the diagnosis of P. falciparum and non-P. falciparum infections. Southeast Asian J Trop Med Public Health. 2005, 36: 552-561.PubMed Belizario VY, Pasay CJ, Bersabe MJ, de Leon WU, Guerrero DM, Bugaoisan VM: Field evaluation of malaria rapid diagnostic tests for the diagnosis of P. falciparum and non-P. falciparum infections. Southeast Asian J Trop Med Public Health. 2005, 36: 552-561.PubMed
12.
Zurück zum Zitat Zhong KJ, Kain KC: Evaluation of a colorimetric PCR-based assay to diagnose Plasmodium falciparum malaria in travelers. J Clin Microbiol. 1999, 37: 339-341.PubMedCentralPubMed Zhong KJ, Kain KC: Evaluation of a colorimetric PCR-based assay to diagnose Plasmodium falciparum malaria in travelers. J Clin Microbiol. 1999, 37: 339-341.PubMedCentralPubMed
13.
Zurück zum Zitat Laoboonchai A, Kawamoto F, Thanoosingha N, Kojima S, Scott Miller RR, Kain KC, Wongsrichanalai C: PCR-based ELISA technique for malaria diagnosis of specimens from Thailand. Trop Med Int Health. 2001, 6: 458-462. 10.1046/j.1365-3156.2001.00736.x.CrossRefPubMed Laoboonchai A, Kawamoto F, Thanoosingha N, Kojima S, Scott Miller RR, Kain KC, Wongsrichanalai C: PCR-based ELISA technique for malaria diagnosis of specimens from Thailand. Trop Med Int Health. 2001, 6: 458-462. 10.1046/j.1365-3156.2001.00736.x.CrossRefPubMed
14.
Zurück zum Zitat Rubio JM, Buhigas I, Subirats M, Baquero M, Puente S, Benito A: Limited level of accuracy provided by available rapid diagnosis tests for malaria enhances the need for PCR-based reference laboratories. J Clin Microbiol. 2001, 39: 2736-2737. 10.1128/JCM.39.7.2736-2737.2001.PubMedCentralCrossRefPubMed Rubio JM, Buhigas I, Subirats M, Baquero M, Puente S, Benito A: Limited level of accuracy provided by available rapid diagnosis tests for malaria enhances the need for PCR-based reference laboratories. J Clin Microbiol. 2001, 39: 2736-2737. 10.1128/JCM.39.7.2736-2737.2001.PubMedCentralCrossRefPubMed
15.
Zurück zum Zitat Hanscheid T, Grobusch MP: How useful is PCR in the diagnosis of malaria?. Trends Parasitol. 2002, 18: 395-398. 10.1016/S1471-4922(02)02348-6.CrossRefPubMed Hanscheid T, Grobusch MP: How useful is PCR in the diagnosis of malaria?. Trends Parasitol. 2002, 18: 395-398. 10.1016/S1471-4922(02)02348-6.CrossRefPubMed
16.
Zurück zum Zitat Singer LM, Newman RD, Diarra A, Moran AC, Huber CS, Stennies G, Sirima SB, Konate A, Yameogo M, Sawadogo R, Barnwell JW, Parise ME: Evaluation of a malaria rapid diagnostic test for assessing the burden of malaria during pregnancy. Am J Trop Med Hyg. 2004, 70: 481-485.PubMed Singer LM, Newman RD, Diarra A, Moran AC, Huber CS, Stennies G, Sirima SB, Konate A, Yameogo M, Sawadogo R, Barnwell JW, Parise ME: Evaluation of a malaria rapid diagnostic test for assessing the burden of malaria during pregnancy. Am J Trop Med Hyg. 2004, 70: 481-485.PubMed
17.
Zurück zum Zitat Bell D, Peeling RW: Evaluation of rapid diagnostic tests: malaria. Nat Rev Microbiol. 2006, 4: S34-8. 10.1038/nrmicro1524.CrossRefPubMed Bell D, Peeling RW: Evaluation of rapid diagnostic tests: malaria. Nat Rev Microbiol. 2006, 4: S34-8. 10.1038/nrmicro1524.CrossRefPubMed
18.
Zurück zum Zitat Cheng Q, Lawrence G, Reed C, Stowers A, Ranford-Cartwright L, Creasey A, Carter R, Saul A: Measurement of Plasmodium falciparum growth rates in vivo: a test of malaria vaccines. Am J Trop Med Hyg. 1997, 57: 495-500.PubMed Cheng Q, Lawrence G, Reed C, Stowers A, Ranford-Cartwright L, Creasey A, Carter R, Saul A: Measurement of Plasmodium falciparum growth rates in vivo: a test of malaria vaccines. Am J Trop Med Hyg. 1997, 57: 495-500.PubMed
19.
Zurück zum Zitat Sylla EH, Kun JF, Kremsner PG: Mosquito distribution and entomological inoculation rates in three malaria-endemic areas in Gabon. Trans R Soc Trop Med Hyg. 2000, 94: 652-656. 10.1016/S0035-9203(00)90219-0.CrossRefPubMed Sylla EH, Kun JF, Kremsner PG: Mosquito distribution and entomological inoculation rates in three malaria-endemic areas in Gabon. Trans R Soc Trop Med Hyg. 2000, 94: 652-656. 10.1016/S0035-9203(00)90219-0.CrossRefPubMed
20.
Zurück zum Zitat Snounou G, Viriyakosol S, Jarra W, Thaithong S, Brown KN: Identification of the four human malaria parasite species in field samples by the polymerase chain reaction and detection of a high prevalence of mixed infections. Mol Biochem Parasitol. 1993, 58: 283-292. 10.1016/0166-6851(93)90050-8.CrossRefPubMed Snounou G, Viriyakosol S, Jarra W, Thaithong S, Brown KN: Identification of the four human malaria parasite species in field samples by the polymerase chain reaction and detection of a high prevalence of mixed infections. Mol Biochem Parasitol. 1993, 58: 283-292. 10.1016/0166-6851(93)90050-8.CrossRefPubMed
21.
Zurück zum Zitat Schoone GJ, Oskam L, Kroon NC, Schallig HD, Omar SA: Detection and quantification of Plasmodium falciparum in blood samples using quantitative nucleic acid sequence-based amplification. J Clin Microbiol. 2000, 38: 4072-4075.PubMedCentralPubMed Schoone GJ, Oskam L, Kroon NC, Schallig HD, Omar SA: Detection and quantification of Plasmodium falciparum in blood samples using quantitative nucleic acid sequence-based amplification. J Clin Microbiol. 2000, 38: 4072-4075.PubMedCentralPubMed
22.
Zurück zum Zitat Contamin H, Fandeur T, Bonnefoy S, Skouri F, Ntoumi F, Mercereau-Puijalon O: PCR typing of field isolates of Plasmodium falciparum. J Clin Microbiol. 1995, 33: 944-951.PubMedCentralPubMed Contamin H, Fandeur T, Bonnefoy S, Skouri F, Ntoumi F, Mercereau-Puijalon O: PCR typing of field isolates of Plasmodium falciparum. J Clin Microbiol. 1995, 33: 944-951.PubMedCentralPubMed
23.
Zurück zum Zitat Ayala E, Lescano AG, Gilman RH, Calderon M, Pinedo VV, Terry H, Cabrera L, Vinetz JM: Polymerase chain reaction and molecular genotyping to monitor parasitological response to anti-malarial chemotherapy in the Peruvian Amazon. Am J Trop Med Hyg. 2006, 74: 546-553.PubMed Ayala E, Lescano AG, Gilman RH, Calderon M, Pinedo VV, Terry H, Cabrera L, Vinetz JM: Polymerase chain reaction and molecular genotyping to monitor parasitological response to anti-malarial chemotherapy in the Peruvian Amazon. Am J Trop Med Hyg. 2006, 74: 546-553.PubMed
25.
Zurück zum Zitat Molineaux L, Gramiccia G: The Garki project: Research on the epidemiology and control of malaria in the Sudan savanna of West Africa. 1980, , World Health Organization, Geneva Molineaux L, Gramiccia G: The Garki project: Research on the epidemiology and control of malaria in the Sudan savanna of West Africa. 1980, , World Health Organization, Geneva
26.
Zurück zum Zitat Snounou G, Viriyakosol S, Zhu XP, Jarra W, Pinheiro L, do Rosario VE, Thaithong S, Brown KN: High sensitivity of detection of human malaria parasites by the use of nested polymerase chain reaction. Mol Biochem Parasitol. 1993, 61: 315-320. 10.1016/0166-6851(93)90077-B.CrossRefPubMed Snounou G, Viriyakosol S, Zhu XP, Jarra W, Pinheiro L, do Rosario VE, Thaithong S, Brown KN: High sensitivity of detection of human malaria parasites by the use of nested polymerase chain reaction. Mol Biochem Parasitol. 1993, 61: 315-320. 10.1016/0166-6851(93)90077-B.CrossRefPubMed
27.
Zurück zum Zitat Landis JR, Koch GG: The measurement of observer agreement for categorical data. Biometrics. 1977, 33: 159-174. 10.2307/2529310.CrossRefPubMed Landis JR, Koch GG: The measurement of observer agreement for categorical data. Biometrics. 1977, 33: 159-174. 10.2307/2529310.CrossRefPubMed
28.
Zurück zum Zitat Coleman RE, Sattabongkot J, Promstaporm S, Maneechai N, Tippayachai B, Kengluecha A, Rachapaew N, Zollner G, Miller RS, Vaughan JA, Thimasarn K, Khuntirat B: Comparison of PCR and microscopy for the detection of asymptomatic malaria in a Plasmodium falciparum/vivax endemic area in Thailand. Malar J. 2006, 5: 121-10.1186/1475-2875-5-121.PubMedCentralCrossRefPubMed Coleman RE, Sattabongkot J, Promstaporm S, Maneechai N, Tippayachai B, Kengluecha A, Rachapaew N, Zollner G, Miller RS, Vaughan JA, Thimasarn K, Khuntirat B: Comparison of PCR and microscopy for the detection of asymptomatic malaria in a Plasmodium falciparum/vivax endemic area in Thailand. Malar J. 2006, 5: 121-10.1186/1475-2875-5-121.PubMedCentralCrossRefPubMed
29.
Zurück zum Zitat Berry A, Fabre R, Benoit-Vical F, Cassaing S, Magnaval JF: Contribution of PCR-based methods to diagnosis and management of imported malaria. Med Trop (Mars). 2005, 65: 176-183. Berry A, Fabre R, Benoit-Vical F, Cassaing S, Magnaval JF: Contribution of PCR-based methods to diagnosis and management of imported malaria. Med Trop (Mars). 2005, 65: 176-183.
30.
Zurück zum Zitat Cox-Singh J, Mahayet S, Abdullah MS, Singh B: Increased sensitivity of malaria detection by nested polymerase chain reaction using simple sampling and DNA extraction. Int J Parasitol. 1997, 27: 1575-1577. 10.1016/S0020-7519(97)00147-1.CrossRefPubMed Cox-Singh J, Mahayet S, Abdullah MS, Singh B: Increased sensitivity of malaria detection by nested polymerase chain reaction using simple sampling and DNA extraction. Int J Parasitol. 1997, 27: 1575-1577. 10.1016/S0020-7519(97)00147-1.CrossRefPubMed
31.
Zurück zum Zitat Di Santi SM, Kirchgatter K, Brunialti KC, Oliveira AM, Ferreira SR, Boulos M: PCR -- based diagnosis to evaluate the performance of malaria reference centers. Rev Inst Med Trop Sao Paulo. 2004, 46: 183-187.CrossRefPubMed Di Santi SM, Kirchgatter K, Brunialti KC, Oliveira AM, Ferreira SR, Boulos M: PCR -- based diagnosis to evaluate the performance of malaria reference centers. Rev Inst Med Trop Sao Paulo. 2004, 46: 183-187.CrossRefPubMed
32.
Zurück zum Zitat Tham JM, Lee SH, Tan TM, Ting RC, Kara UA: Detection and species determination of malaria parasites by PCR: comparison with microscopy and with ParaSight-F and ICT malaria Pf tests in a clinical environment. J Clin Microbiol. 1999, 37: 1269-1273.PubMedCentralPubMed Tham JM, Lee SH, Tan TM, Ting RC, Kara UA: Detection and species determination of malaria parasites by PCR: comparison with microscopy and with ParaSight-F and ICT malaria Pf tests in a clinical environment. J Clin Microbiol. 1999, 37: 1269-1273.PubMedCentralPubMed
33.
Zurück zum Zitat Blythe JE, Surentheran T, Preiser PR: STEVOR--a multifunctional protein?. Mol Biochem Parasitol. 2004, 134: 11-15. 10.1016/j.molbiopara.2003.09.011.CrossRefPubMed Blythe JE, Surentheran T, Preiser PR: STEVOR--a multifunctional protein?. Mol Biochem Parasitol. 2004, 134: 11-15. 10.1016/j.molbiopara.2003.09.011.CrossRefPubMed
34.
Zurück zum Zitat Cheng Q, Cloonan N, Fischer K, Thompson J, Waine G, Lanzer M, Saul A: stevor and rif are Plasmodium falciparum multicopy gene families which potentially encode variant antigens. Mol Biochem Parasitol. 1998, 97: 161-176. 10.1016/S0166-6851(98)00144-3.CrossRefPubMed Cheng Q, Cloonan N, Fischer K, Thompson J, Waine G, Lanzer M, Saul A: stevor and rif are Plasmodium falciparum multicopy gene families which potentially encode variant antigens. Mol Biochem Parasitol. 1998, 97: 161-176. 10.1016/S0166-6851(98)00144-3.CrossRefPubMed
35.
Zurück zum Zitat Dame JB, McCutchan TF: The four ribosomal DNA units of the malaria parasite Plasmodium berghei. Identification, restriction map, and copy number analysis. J Biol Chem. 1983, 258: 6984-6990.PubMed Dame JB, McCutchan TF: The four ribosomal DNA units of the malaria parasite Plasmodium berghei. Identification, restriction map, and copy number analysis. J Biol Chem. 1983, 258: 6984-6990.PubMed
36.
Zurück zum Zitat Langsley G, Hyde JE, Goman M, Scaife JG: Cloning and characterisation of the rRNA genes from the human malaria parasite Plasmodium falciparum. Nucleic Acids Res. 1983, 11: 8703-8717. 10.1093/nar/11.24.8703.PubMedCentralCrossRefPubMed Langsley G, Hyde JE, Goman M, Scaife JG: Cloning and characterisation of the rRNA genes from the human malaria parasite Plasmodium falciparum. Nucleic Acids Res. 1983, 11: 8703-8717. 10.1093/nar/11.24.8703.PubMedCentralCrossRefPubMed
37.
Zurück zum Zitat Fenton B, Clark JT, Khan CM, Robinson JV, Walliker D, Ridley R, Scaife JG, McBride JS: Structural and antigenic polymorphism of the 35- to 48-kilodalton merozoite surface antigen (MSA-2) of the malaria parasite Plasmodium falciparum. Mol Cell Biol. 1991, 11: 963-971.PubMedCentralCrossRefPubMed Fenton B, Clark JT, Khan CM, Robinson JV, Walliker D, Ridley R, Scaife JG, McBride JS: Structural and antigenic polymorphism of the 35- to 48-kilodalton merozoite surface antigen (MSA-2) of the malaria parasite Plasmodium falciparum. Mol Cell Biol. 1991, 11: 963-971.PubMedCentralCrossRefPubMed
38.
Zurück zum Zitat Weisman S, Wang L, Billman-Jacobe H, Nhan DH, Richie TL, Coppel RL: Antibody responses to infections with strains of Plasmodium falciparum expressing diverse forms of merozoite surface protein 2. Infect Immun. 2001, 69: 959-967. 10.1128/IAI.69.2.959-967.2001.PubMedCentralCrossRefPubMed Weisman S, Wang L, Billman-Jacobe H, Nhan DH, Richie TL, Coppel RL: Antibody responses to infections with strains of Plasmodium falciparum expressing diverse forms of merozoite surface protein 2. Infect Immun. 2001, 69: 959-967. 10.1128/IAI.69.2.959-967.2001.PubMedCentralCrossRefPubMed
Metadaten
Titel
Comparison of PCR-based detection of Plasmodium falciparum infections based on single and multicopy genes
verfasst von
Segun I Oyedeji
Henrietta O Awobode
Gamaliel C Monday
Eric Kendjo
Peter G Kremsner
Jürgen F Kun
Publikationsdatum
01.12.2007
Verlag
BioMed Central
Erschienen in
Malaria Journal / Ausgabe 1/2007
Elektronische ISSN: 1475-2875
DOI
https://doi.org/10.1186/1475-2875-6-112

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