Loop Mediated Isothermal Amplification for Detection of<i>Trypanosoma brucei gambiense</i>in Urine and Saliva Samples in Nonhuman Primate Model

Ngotho, Maina; Kagira, John; Gachie, Beatrice; Karanja, Simon; Waema, Maxwell W.; Maranga, Dawn Nyawira; Maina, Naomi

doi:10.1155/2015/867846

Cited by 17 publications

(11 citation statements)

References 28 publications

(32 reference statements)

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“…Due to the specific nature of the action of LAMP primers, the amount of DNA produced is considerably higher than PCR based amplification. The corresponding release of pyrophosphate results in visible turbidity which allows easy visualization of amplification products [39]. LAMP has been observed to be less sensitive than PCR to inhibitors in complex samples such as blood, possibly due to the use of a different DNA polymerase (Bst DNA polymerase rather than Taq polymerase as in PCR).…”

Section: Resultsmentioning

confidence: 99%

Detecting Mycobacterium ulcerans in Clinical Specimens from Ghana: Comparative Evaluation of BU-LAMP and IS2404 PCR

Aninagyei¹,

Rufai²,

Dede³

et al. 2020

Int J Trop Dis

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Background: Conventional and nested polymerase chain reaction (PCR), and loop-mediated isothermal amplification (LAMP) have been used to identify Mycobacterium ulcerans in separate studies and different specimens. However, the sensitivities of these three techniques have not been compared in a single study. Objective: This study compared the performance of two variant PCR techniques and LAMP assays to detect M. Ulcerans in same clinical specimens. Methods: Samples were collected from patients suspected of Buruli ulcer disease (BUD) in Southern Ghana. Ulcerative and non-ulcerative forms of the disease were swabbed and aspirated respectively. Insertion sequence 2404 (IS2404) M. ulcerans targets were detected in each sample using conventional polymerase chain reaction (PCR), nested PCR and loop-mediated isothermal amplification (LAMP) assay. Results: In all, 141 suspected BUD patients were sampled (Amasaman, n = 52; Obom, n = 17; Paakro, n = 31; Nkawie, n = 21 and Tepa, n = 20). The reference technique, nested PCR, detected M. ulcerans in 122 (86.5%) whereas conventional PCR and BU-LAMP detected M. ulcerans in 104 (73.7%) and 119 (84.4%) samples respectively. Compared to nested PCR, conventional PCR performed poorly (x 2 = 19.7, p < 0.01; κ = 0.58; % agreement = 86.62) while BU-LAMP was as good as nested PCR (x 2 = 0.457, p = 0.459; κ = 0.88; % agreement = 97.16). Sensitivity of conventional PCR and BU-LAMP were 84.6% (95% CI: 76.9-90.4) and 97.5% (95% CI: 92.9-99.5) respectively. BU-LAMP and nested PCR detected M. ulcerans in ulcerative forms of the disease and category II lesions better while the three techniques did not differ in sensitivities in other clinical forms and lesion categories. Conclusion: BU-LAMP assay is very comparable to nested PCR in detecting M. ulcerans from clinical specimens. More importantly, LAMP assay is user-friendly, fast, requires less instrumentation and easy to use in resource-limited laboratory with minimal user training.

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Section: Resultsmentioning

confidence: 99%

Detecting Mycobacterium ulcerans in Clinical Specimens from Ghana: Comparative Evaluation of BU-LAMP and IS2404 PCR

Aninagyei¹,

Rufai²,

Dede³

et al. 2020

Int J Trop Dis

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“…b . gambiense DNA in several biofluids including serum and saliva [46]. Up to 77 days post infection (dpi) TgsGP LAMP recorded 100% detection in the saliva samples [46].…”

Section: Discussionmentioning

confidence: 99%

“…gambiense DNA in several biofluids including serum and saliva [46]. Up to 77 days post infection (dpi) TgsGP LAMP recorded 100% detection in the saliva samples [46]. While LAMP recorded 33% detection in the serum 180 dpi, TgsGP LAMP failed to detect trypanosomes in saliva after 140 days.…”

Section: Discussionmentioning

confidence: 99%

Using detergent-enhanced LAMP for African trypanosome detection in human cerebrospinal fluid and implications for disease staging

et al. 2019

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Objective Where human African trypanosomiasis (HAT) patients are seen, failure to microscopically diagnose infections by Trypanosoma brucei gambiense in blood smears and/or cerebrospinal fluid (CSF) in the critical early stages of the disease is the single most important factor in treatment failure, a result of delayed treatment onset or its absence. We hypothesized that the enhanced sensitivity of detergent-enhanced loop-mediated isothermal amplification (LAMP) will allow for point of care (POC) detection of African trypanosomes in the CSF of HAT patients where the probability for detecting a single parasite or parasite DNA molecule in 1 μL of CSF sample is negligible by current methods. Methodology We used LAMP targeting the multicopy pan- T . brucei repetitive insertion mobile element (RIME LAMP) and the Trypanosoma brucei gambiense 5.8S rRNA-internal transcribed spacer 2 gene (TBG1 LAMP). We tested 1 μL out of 20 μL sham or Triton X-100 treated CSFs from 73 stage-1 and 77 stage-2 HAT patients from the Central African Republic and 100 CSF negative controls. Results Under sham conditions, parasite DNA was detected by RIME and TBG1 LAMP in 1.4% of the stage-1 and stage-2 gambiense HAT CSF samples tested. After sample incubation with detergent, the number of LAMP parasite positive stage-2 CSF’s increased to 26%, a value which included the 2 of the 4 CSF samples where trypanosomes were identified microscopically. Unexpected was the 41% increase in parasite positive stage-1 CSF’s detected by LAMP. Cohen’s kappa coefficients for RIME versus TBG1 LAMP of 0.92 (95%CI: 0.82–1.00) for stage-1 and 0.90 (95%CI: 0.80–1.00) for stage-2 reflected a high level of agreement between the data sets indicating that the results were not due to amplicon contamination, data confirmed in χ 2 tests (p<0.001) and Fisher’s exact probability test (p = 4.7e -13 ). Conclusion This study detected genomic trypanosome DNA in the CSF independent of the HAT stage and may be consistent with early CNS entry and other scenarios that identify critical knowledge gaps for future studies. Detergent-enhanced LAMP could be applicable for non-invasive African trypanosome detection in human skin and saliva or as an epidemiologic tool for the determination of human (or animal) African trypanosome prevalence in areas where chronically low parasitemias are present.

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“…The sole evidence on the presence of genetic material in urine comes from an experimental infection of vervet monkeys by T. brucei. In this model, Trypanosoma DNA could be amplified from urine, with LAMP as early as 17 days postinfection [173]. Biochemical changes associated with trypanosome infection have been published on animal models infected by various Trypanosoma species.…”

Section: Urinementioning

confidence: 99%

Noninvasive Biological Samples to Detect and Diagnose Infections due to Trypanosomatidae Parasites: A Systematic Review and Meta-Analysis

Sereno

Akhoundi

Sayehmri

et al. 2020

IJMS

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Unicellular eukaryotes of the Trypanosomatidae family include human and animal pathogens that belong to the Trypanosoma and Leishmania genera. Diagnosis of the diseases they cause requires the sampling of body fluids (e.g., blood, lymph, peritoneal fluid, cerebrospinal fluid) or organ biopsies (e.g., bone marrow, spleen), which are mostly obtained through invasive methods. Body fluids or appendages can be alternatives to these invasive biopsies but appropriateness remains poorly studied. To further address this question, we perform a systematic review on clues evidencing the presence of parasites, genetic material, antibodies, and antigens in body secretions, appendages, or the organs or proximal tissues that produce these materials. Paper selection was based on searches in PubMed, Web of Science, WorldWideScience, SciELO, Embase, and Google. The information of each selected article (n = 333) was classified into different sections and data were extracted from 77 papers. The presence of Trypanosomatidae parasites has been tracked in most of organs or proximal tissues that produce body secretions or appendages, in naturally or experimentally infected hosts. The meta-analysis highlights the paucity of studies on human African trypanosomiasis and an absence on animal trypanosomiasis. Among the collected data high heterogeneity in terms of the I 2 statistic (100%) is recorded. A high positivity is recorded for antibody and genetic material detection in urine of patients and dogs suffering leishmaniasis, and of antigens for leishmaniasis and Chagas disease. Data on conjunctival swabs can be analyzed with molecular methods solely for dogs suffering canine visceral leishmaniasis. Saliva and hair/bristles showed a pretty good positivity that support their potential to be used for leishmaniasis diagnosis. In conclusion, our study pinpoints significant gaps that need to be filled in order to properly address the interest of body secretion and hair or bristles for the diagnosis of infections caused by Leishmania and by other Trypanosomatidae parasites.

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Loop Mediated Isothermal Amplification for Detection ofTrypanosoma brucei gambiensein Urine and Saliva Samples in Nonhuman Primate Model

Cited by 17 publications

References 28 publications

Detecting Mycobacterium ulcerans in Clinical Specimens from Ghana: Comparative Evaluation of BU-LAMP and IS2404 PCR

Detecting Mycobacterium ulcerans in Clinical Specimens from Ghana: Comparative Evaluation of BU-LAMP and IS2404 PCR

Using detergent-enhanced LAMP for African trypanosome detection in human cerebrospinal fluid and implications for disease staging

Noninvasive Biological Samples to Detect and Diagnose Infections due to Trypanosomatidae Parasites: A Systematic Review and Meta-Analysis

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