Search

Abstract Background Leprosy continues to be a health problem in endemic areas. More than 200,000 new cases of leprosy per year suggest that transmission of the disease is still ongoing, presumably as airborne infection through nasal droplets. Late diagnosis supports continued transmission and increases the individual risk for functional disabilities. Laboratory tools are considered beneficial to facilitate early detection and clinical assessment of cases. The aim of this study was to validate molecular tools allowing detection, quantification and assessment of viability of M. leprae from nasal swab samples which are easy to obtain without the need of any invasive procedures. Methods Validation of two real-time PCRs detecting M. leprae DNA (RLEP qPCR) and RNA (16S rRNA RT qPCR) was conducted on “must not detect”/“must detect” samples and 160 pre-treatment nasal swab samples from 20 clinically diagnosed multibacillary (MB) leprosy patients from Togo. Results Both assays were 100% M. leprae specific and showed analytical sensitivities of three templates each. Out of 20 clinically diagnosed MB leprosy patients, 15 (75.0%) had a positive RLEP qPCR result from nasal swab samples. The 16S rRNA RT qPCR detected viable bacilli in nasal swab samples of ten out of these 15 RLEP positive patients (66.7%). Conclusion The combined RLEP/16S rRNA (RT) qPCR assay provides a sensitive and specific tool to determine the bacterial load and viability of M. leprae from nasal swab samples and is applicable for early diagnosis, monitoring treatment response and investigating the role of nasal carriage of M. leprae in human-to-human transmission through aerosol infection.

Abstract Background Onchocerciasis is a parasitic disease caused by the filarial nematode Onchocerca volvulus. In endemic areas, the diagnosis is commonly confirmed by microscopic examination of skin snip samples, though this technique is considered to have low sensitivity. The available melting-curve based quantitative real-time PCR (qPCR) using degenerated primers targeting the O-150 repeat of O. volvulus was considered insufficient for confirming the individual diagnosis, especially in elimination studies. This study aimed to improve detection of O. volvulus DNA in clinical samples through the development of a highly sensitive qPCR assay. Methods A novel hydrolysis probe based qPCR assay was designed targeting the specific sequence of the O. volvulus O-5S rRNA gene. A total of 200 clinically suspected onchocerciasis cases were included from Goma district in South-west Ethiopia, from October 2012 through May 2013. Skin snip samples were collected and subjected to microscopy, O-150 qPCR, and the novel O-5S qPCR. Results Among the 200 individuals, 133 patients tested positive (positivity rate of 66.5%) and 67 negative by O-5S qPCR, 74 tested positive by microscopy (37.0%) and 78 tested positive by O-150 qPCR (39.0%). Among the 133 O-5S qPCR positive individuals, microscopy and O-150 qPCR detected 55.6 and 59.4% patients, respectively, implying a higher sensitivity of O-5S qPCR than microscopy and O-150 qPCR. None of the 67 individuals who tested negative by O-5S qPCR tested positive by microscopy or O-150 qPCR, implying 100% specificity of the newly designed O-5S qPCR assay. Conclusions The novel O-5S qPCR assay is more sensitive than both microscopic examination and the existing O-150 qPCR for the detection of O. volvulus from skin snip samples. The newly designed assay is an important step towards appropriate individual diagnosis and control of onchocerciasis.

Small discs punched out from leaves of tobacco (Nicotiana tabacum L.) were exposed for 3 rain in the dark to a high temperature of around 48°C. This caused a progressive inhibition of the gas-exchange (oxygen evolution) type photoacoustic signal, resulting' finally in phase inversion (i.e. leading to negative values), which indicates that oxygen uptake replaces the normal oxygen evolution. This effect was also observed in various other plant species. Oxygen uptake was rapidly reversed (within ca. 4-5 min) to a certain low value of oxygen evolution (about 20% of the control) by continuous illumination with relatively strong white light (minimum 55 W m-2). However, a few minutes in darkness following this heat treatment induced reappearence of the uptake signal. This photoacoustically detected oxygen uptake after heat shock may be interpreted as reflecting stimulated oxygen photoreduction (Mehler reaction) caused by (light dependent) inactivation of the Calvin cycle by heat, suggesting that oxygen may act as a major photosynthetic electron acceptor under stress conditions. Leaves suffering from such heat shock effects were completely restored to normal behaviour after a 24-h incubation at room temperature (25°C). Analysis of the modulation frequency and wavelength dependence of the photoacoustic signals showed that heat shock-induced oxygen uptake is a very complex phenomenon, composed of at least two components differing in kinetics and sensitivity to DCMU. [ABSTRACT FROM AUTHOR]

Exposure of tomato plants to a mild chilling temperature and relatively low ambient photon flux density for an extended period (10°C and 400 μmol photons m-2s-1 d and 5°C night for 6 d) resulted in a significant decrease in the variable chlorophyll fluorescence, the quantum yield of oxygen evolution and the amount of total absorbed energy stored in photochemical intermediates, but not in the chlorophyll concentration or in the activity of ribulose biphosphate carboxylase. These results indicate that photochemical processes involving PSII were affected, and might reflect photoinhibitory effects on the photosynthetic apparatus. Chilling treatment had relatively small influence on the maximal extent of the Emerson effect. This observation, together with the sharp decrease found in the quantum yield of oxygen evolution, could be reconciled with the above results only if some dependency between the two photosystems was assumed. On the basis of this interpretation, it was concluded that the strong Emerson effect after chilling still reflects the typical imbalance between PSI and PSII centers, even though populations of such unaffected pairs are smaller than in the untreated plants. The relatively new photoacoustic technique employed in this study is shown to be useful both as a diagnostic tool and as a means of investigating changes in photochemical activity in the study of environmental stress effects on photosynthesis. The results support the view that photoinhibition can play an important role in limiting photosynthetic activity, and therefore productivity, in chilling-sensitive plants such as the tomato under the natural conditions that prevail during the winter in Mediterranean climates. [ABSTRACT FROM AUTHOR]