Caries clusters at lesion-severity thresholds: A Sierra Leone case study.

OBJECTIVES: Despite being almost entirely preventable, globally, dental caries is extremely prevalent. Moreover, dental caries will continue to present an even larger challenge for lower income countries, particularly those in the African context, as they transition to a more Western diet. Hence, epidemiological data providing insight into disease patterns and trends is critical to inform public health action. The purpose of this study was to examine dental caries clusters by caries detection threshold among 15-year-old adolescents in Sierra Leone, using data from the latest national survey, and to explore associated sociodemographic factors. METHODS: This paper presents a secondary analysis of oral health data on 490 15-year-olds from the Sierra Leone national oral health survey of schoolchildren. Hierarchical cluster analysis of dental caries experience was conducted across all surfaces at four decay detection thresholds using the International Caries Detection and Assessment System (ICDAS) (clinical: ICDAS 2-6, cavitated: ICDAS 3-6, obvious: ICDAS 4-6 and extensive obvious: ICDAS 5-6 decay) across the four regions of Sierra Leone. Ordered logistic regression was used to estimate the association of sociodemographic factors with generated clusters relating to clinical and obvious decay experience. These are of both clinical and epidemiological relevance. RESULTS: A 3-cluster decay pattern representing a 'low' to 'high' decay experience distribution was observed under each decay detection threshold across surfaces. For clinical decay (including visual enamel caries), 28.8% had low, 55.1% medium and 15.9% high caries status. In the adjusted model, the only significant risk factor across obvious and clinical decay thresholds was region, with adolescents outside the Western region more likely to experience decay. CONCLUSION: This study suggests that adolescents in Sierra Leone fall into three distinct caries clusters: low, medium to high decay experience distribution, regardless of decay threshold. It reinforces the importance of recognizing dental caries detection thresholds and the use of contemporary epidemiological methodology. This suggests that adolescents outside the Western region are likely to have higher caries experience. The data also provides insight to the nature of adolescents in each cluster and should help to inform policy and planning of the integration of oral health into primary care and school systems.

Novel developments in rapid mycotoxin detection.

Rapid antibody-based mycotoxin screening techniques are designed to be used outside a laboratory environment, at the place of sampling. Results are expected immediately, so that commodities can be further processed without delay. Because they are used for mycotoxin analysis, very low levels (ppb and ppt range) should be detected. A further requirement is that the obtained results are accurate with a false negative rate of <5% at the level of interest.At first, plastic microtiter plates were used as solid phase materials for immobilizing antibodies (enzyme-linked immunosorbent assays). However, to increase speed and user-friendliness, plastics were replaced by microporous membranes. As an example a flow-through enzyme immunoassay for the detection of fumonisins in cornflakes with a cut-off value of 275 µg/kg is described. No false negative results were observed and the false positive rate was 18%. However, enzyme labels, used to enable visual evaluation of results, did not seem to be completely satisfactory in terms of stability and repeatability of the generated signal. Therefore microparticle labels such as colloidal gold particles are used more and more,e.g. in a lateral flow dipstick immunoassay. When applied to the detection of aflatoxin B1 in pig feed a cut-off value of 5 µg/kg could be reached with no false negative results and a false positive rate of only 10%. Sample pretreatment for screening techniques should be rapid and simple. Preferably a simple solvent extraction is used, followed by a filtration and dilution step. However, for strongly coloured or complex food matrices, this did not seem to work. The combination of clean-up and detection in one single test device is a new approach. When using this clean-up tandem assay column for the detection of ochratoxin A in roasted coffee, a cut-off value of 6 µg/kg was reached. No false positive results were obtained, however, the false negative rate was 8%.

Development of a flow-through enzyme immunoassay and application in screening green coffee samples for ochratoxin A with confirmation by high-performance liquid chromatography.

A flow-through enzyme immunoassay has been developed for the screening of green coffee bean samples for ochratoxin A (OA) and was later used in a survey on OA in green coffee from different countries. The test has a sensitivity of 8 ng/g, and calculated recoveries ranged from 70 to 89% and from 86 to 95% for spiked and naturally contaminated samples, respectively. There were no significant differences in within-day and between-day assay performance (P > 0.05). Green coffee samples (15 Arabica and 7 Robusta) received from an international coffee trader were analyzed for intrinsic fungal contamination, screened for OA, and subsequently confirmed by high-performance liquid chromatography (HPLC). All 22 samples were contaminated by fungal species of the genus Aspergillus, while Penicillium species were isolated from a mere 13.6% of the total number of samples. Isolates were tested for their ability to produce OA, and only 3.9% were positive. There was no correlation between occurrence of OA-producing isolates and levels of OA in contaminated samples. Results of the screening procedure showed that 4 of the 22 samples were contaminated with 8 ng/g or higher. The HPLC method confirmed that the OA levels ranged from 27 to 168 ng/g. A fifth sample, which was shown to be negative during screening, had an OA concentration of 4 ng/g. There were no false negatives or positives recorded, and the flow-through enzyme immunoassay results correlated with those obtained by HPLC.

Survey of Fusarium moniliforme (F. verticillioides) and production of fumonisin B1 in cereal grains and oilseeds in Zimbabwe.

In a national survey carried out in 1995 and 1996, the distribution of Fusarium moniliforme (= F. verticillioides) and fumonisin B1 (FB1) levels in cereals and oilseeds from Zimbabwe were analyzed. The results of this study showed that the incidence of F. moniliforme and other Fusarium species and levels of FB1 generally decreased from regions with high rainfall and annual moderate temperatures to low rainfall regions. There was no Fusarium contamination and FB1 was not detected in sunflowers and soybeans. The incidence of F. moniliforme and its metabolite exhibited a substrate preference with high incidences of Fusarium species and high FB1 levels being recorded for maize followed by wheat, rapoko and sorghum.

Detection of type III secretion system genes in animal isolates of Bordetella bronchiseptica.

A cosmid clone bank of Bordetella bronchiseptica genomic DNA was screened for the presence of type III secretion (TTS) genes using a probe derived from the TTS system genes of Ralstonia solanacearum. A 3.35kb PstI fragment, sub-cloned from a hybridising cosmid clone, was sequenced and found to contain a 97bp overlap with the previously reported B. bronchiseptica bscIJKLNO TTS gene cluster. DNA and predicted protein homology analysis suggests that a bscPQRST cluster lies immediately downstream of bscIJKLNO. A PCR amplification assay indicated that the bscT locus was present in 27 B. bronchiseptica animal isolates tested (100%). Dot-blot DNA hybridisation using probes for bscT and bscP confirmed the presence of these loci in six canine isolates associated with a variety of clinical signs. Although TTS has been implicated in the pathogenicity of B. bronchiseptica, it is likely that different clinical manifestations may be due to variations in gene expression or host factors, rather than the absence or presence of TTS genes.

Detection of T-2 toxin in different cereals by flow-through enzyme immunoassay with a simultaneous internal reference.

In most previously described membrane-based immunoassays a separate negative control assay is always carried out to evaluate the performance of the assay. To overcome this problem, a membrane-based flow-through enzyme immunoassay with an internal control has been developed for the detection of T-2 toxin in cereals (patent pending). An Immunodyne ABC membrane was coated with 2 microL of goat anti-horseradish peroxidase (HRP) (internal control spot) (1:1000) and 2 microL of rabbit anti-mouse (test spot) (undiluted) immunoglobulins, and the free binding sites were blocked. In addition to the antibody-coated Immunodyne ABC membrane, the assay also comprises a plastic snap-fit device, absorbent cotton wool, mouse anti-T-2 monoclonal antibodies (Mab), and T-2-HRP conjugate. The color intensity (Delta) of the internal control and that of the negative sample showed no difference (P > 0.05), whereas there was a significant difference between the internal control and positive samples (P < 0.05). The minimum detectable limit that could be visually detected with confidence was 50 ng of T-2 per gram of cereal sample.

Development of a portable field immunoassay for the detection of aflatoxin M1 in milk.

Many methods for AFM1 detection exist, but most are time consuming, employ expensive equipment and require experienced personnel. To overcome these problems a membrane-based flow-through enzyme immunoassay has been developed (patent pending). The assay comprised a nylon Immunodyne ABC membrane spotted with anti-mouse antibodies, a plastic snap-fit device, absorbent cotton wool, mouse anti-AFM1 monoclonal antibodies (Mab), and AFB1-horseradish peroxidase (HRP) conjugate. This assay was coupled to an immunoaffinity column (IAC). The visual detection limit was 0.05 ng/g AFM1 in milk. Assay time for IAC clean-up was 12 min, and that for the flow-through assay was 18 min, hence the total assay time was 30 min. This method allows for a rapid screening of milk consignments which do not conform to the maximum permissible limits of 0.05 ng/g AFM1, hence enabling the rejection of such at the farm level. Laboratory validation was done using certified reference materials (CRM) with AFM1 concentrations of < 0.05, 0.09 and 0.76 ng/g. Precision of the assay was high as shown by the high repeatability of the assay results. There were no significant differences in recoveries between standard in buffer and CRM (P > 0.05), and assay responses for these two were highly correlated (99.63%).