Direct detection of shigella in stool specimens by use of a metagenomic approach

The underestimation of Shigella species as a cause of childhood diarrhea disease has become increasingly apparent with quantitative PCR (qPCR)-based diagnostic methods versus culture. We sought to confirm qPCR-based detection of Shigella via a metagenomics approach. Three groups of samples were selected from diarrheal cases from the Global Enteric Multicenter Study: nine Shigella culture-positive and qPCR-positive (culture+ qPCR+) samples, nine culture-negative but qPCR-positive (culture- qPCR+) samples, and nine culture-negative and qPCR-negative (culture- qPCR-) samples. Fecal DNA was sequenced using paired-end Illumina HiSeq, whereby 3.26 × 108 ± 5.6 × 107 high-quality reads were generated for each sample. We used Kraken software to compare the read counts specific to Shigella among the three groups. The proportions of Shigella-specific nonhuman sequence reads between culture+ qPCR+ (0.65 ± 0.42%) and culture- qPCR+ (0.55 ± 0.31%) samples were similar (Mann-Whitney U test, P = 0.627) and distinct from the culture- qPCR- group (0.17 ± 0.15%, P \u3c 0.05). The read counts of sequences previously targeted by Shigella/enteroinvasive Escherichia coli (EIEC) qPCR assays, namely, ipaH, virA, virG, ial, ShET2, and ipaH3, were also similar between the culture+ qPCR+ and culture- qPCR+ groups and distinct from the culture- qPCR- groups (P \u3c 0.001). Kraken performed well versus other methods: its precision and recall of Shigella were excellent at the genus level but variable at the species level. In summary, metagenomic sequencing indicates that Shigella/EIEC qPCR-positive samples are similar to those of Shigella culture-positive samples in Shigella sequence composition, thus supporting qPCR as an accurate method for detecting Shigella

Intermittent applied mechanical loading induces subchondral bone thickening that may be intensified locally by contiguous articular cartilage lesions

Objectives: Changes in subchondral bone (SCB) and cross-talk with articular cartilage (AC) have been linked to osteoarthritis (OA). Using micro-computed tomography (micro-CT) this study: (1) examines changes in SCB architecture in a non-invasive loading mouse model in which focal AC lesions are induced selectively in the lateral femur, and (2) determines any modifications in the contralateral knee, linked to changes in gait, which might complicate use of this limb as an internal control. Methods: Right knee joints of CBA mice were loaded: once with 2weeks of habitual use (n=7), for 2weeks (n=8) or for 5weeks (n=5). Both left (contralateral) and right (loaded) knees were micro-CT scanned and the SCB and trabecular bone analysed. Gait analysis was also performed. Results: These analyses showed a significant increase in SCB thickness in the lateral compartments in joints loaded for 5weeks, which was most marked in the lateral femur; the contralateral non-loaded knee also showed transient SCB thickening (loaded once and repetitively). Epiphyseal trabecular bone BV/TV and trabecular thickness were also increased in the lateral compartments after 5 weeks of loading, and in all joint compartments in the contralateral knee. Gait analysis showed that applied loading only affected gait in the contralateral himd-limb in all groups of mice from the second week after the first loading episode. Conclusions: These data indicate a spatial link between SCB thickening and AC lesions following mechanical trauma, and the clear limitations associated with the use of contralateral joints as controls in such OA models, and perhaps in OA diagnosis

Quality of anti-malarial drugs provided by public and private healthcare providers in south-east Nigeria

BACKGROUND: There is little existing knowledge about actual quality of drugs provided by different providers in Nigeria and in many sub-Saharan African countries. Such information is important for improving malaria treatment that will help in the development and implementation of actions designed to improve the quality of treatment. The objective of the study was to determine the quality of drugs used for the treatment of malaria in a broad spectrum of public and private healthcare providers. METHODS: The study was undertaken in six towns (three urban and three rural) in Anambra state, south-east Nigeria. Anti-malarials (225 samples), which included artesunate, dihydroartemisinin, sulphadoxine-pyrimethamine (SP), quinine, and chloroquine, were either purchased or collected from randomly selected providers. The quality of these drugs was assessed by laboratory analysis of the dissolution profile using published pharmacopoeial monograms and measuring the amount of active ingredient using high performance liquid chromatography (HPLC). FINDINGS: It was found that 60 (37%) of the anti-malarials tested did not meet the United States Pharmacopoeia (USP) specifications for the amount of active ingredients, with the suspect drugs either lacking the active ingredients or containing suboptimal quantities of the active ingredients. Quinine (46%) and SP formulations (39%) were among drugs that did not satisfy the tolerance limits published in USP monograms. A total of 78% of the suspect drugs were from private facilities, mostly low-level providers, such as patent medicine dealers (vendors). CONCLUSION: This study found that there was a high prevalence of poor quality drugs. The findings provide areas for public intervention to improve the quality of malaria treatment services. There should be enforced checks and regulation of drug supply management as well as stiffer penalties for people stocking substandard and counterfeit drugs

Comparison of Bioavailability Between the Most Available Generic Tablet Formulation Containing Artemether and Lumefantrine on the Tanzanian Market and the Innovator's Product.

Existence of anti-malarial generic drugs with low bioavailability marketed on sub-Saharan Africa has raised a concern on patients achieving therapeutic concentrations after intake of these products. This work compared bioavailability of one generic tablet formulation with innovator's product. Both were fixed dose combination tablet formulations containing artemether and lumefantrine.MethodologyThe study was conducted in Dar Es Salaam, Tanzania, in which a survey of the most abundant generic containing artemether-lumefantrine tablet formulation was carried out in retail pharmacies. The most widely available generic (Artefan(R), Ajanta Pharma Ltd, Maharashtra, India) was sampled for bioavailability comparison with Coartem(R) (Novartis Pharma, Basel, Switzerland) - the innovator's product. A randomized, two-treatment cross-over study was conducted in 18 healthy Tanzanian black male volunteers. Each volunteer received Artefan(R) (test) and Coartem(R) (as reference) formulation separated by 42 days of drug-free washout period. Serial blood samples were collected up to 168 hours after oral administration of a single dose of each treatment. Quantitation of lumefantrine plasma levels was done using HPLC with UV detection. Bioequivalence of the two products was assessed in accordance with the US Food and Drug Authority (FDA) guidelines. The most widely available generic in pharmacies was Artefan(R) from India. All eighteen enrolled volunteers completed the study and both test and reference tablet formulations were well tolerated. It was possible to quantify lumefantrine alone, therefore, the pharmacokinetic parameters reported herein are for lumefantrine. The geometric mean ratios for Cmax, AUC0-t and AUC0-[infinity] were 84% in all cases and within FDA recommended bioequivalence limits of 80% -- 125%, but the 90% confidence intervals were outside FDA recommended limits (CI 49--143%, 53 - 137%, 52 - 135% respectively). There were no statistical significant differences between the two formulations with regard to PK parameters (P > 0.05). Although the ratios of AUCs and Cmax were within the acceptable FDA range, bioequivalence between Artefan(R) and Coartem(R) tablet formulations was not demonstrated due to failure to comply with the FDA 90 % confidence interval criteria. Based on the observed total drug exposure (AUCs), Artefan(R) is likely to produce a similar therapeutic response as Coartem(R)

Determinants and Impact of Giardia Infection in the First 2 Years of Life in the MAL-ED Birth Cohort

Background. Giardia are among the most common enteropathogens detected in children in low-resource settings. We describe here the epidemiology of infection with Giardia in the first 2 years of life in the Etiology, Risk Factors, and Interactions of Enteric Infections and Malnutrition and the Consequences for Child Health and Development Project (MAL-ED), a multisite birth-cohort stu dy. Methods. From 2089 children, 34 916 stool samples collected during monthly surveillance and episodes of diarrhea were tested for Giardia using an enzyme immunoassay. We quantified the risk of Giardia detection, identified risk factors, and assessed the associations with micronutrients, markers of gut inflammation and permeability, diarrhea, and growth using multivariable linear regression. Results. The incidence of at least 1 Giardia detection varied according to site (range, 37.7%–96.4%) and was higher in the sec - ond year of life. Exclusive breastfeeding (HR for first Giardia detection in a monthly surveillance stool sample, 0.46 [95% confidence interval (CI), 0.28–0.75]), higher socioeconomic status (HR, 0.74 [95% CI, 0.56–0.97]), and recent metronidazole treatment (risk ratio for any surveillance stool detection, 0.69 [95% CI, 0.56–0.84]) were protective. Persistence of Giardia (consecutive detections) in the first 6 months of life was associated with reduced subsequent diarrheal rates in Naushahro Feroze, Pakistan but not at any other site. Giardia detection was also associated with an increased lactulose/mannitol ratio. Persistence of Giardia before 6 months of age was associated with a −0.29 (95% CI, −0.53 to −0.05) deficit in weight-for-age z score and −0.29 (95% CI, −0.64 to 0.07) deficit in length-for-age z score at 2 years. Conclusions. Infection with Giardia occurred across epidemiological contexts, and repeated detections in 40% of the children suggest that persistent infections were common. Early persistent infection with Giardia , independent of diarrhea, might contribute to intestinal permeability and stunted growth

Guidelines for Field Surveys of the Quality of Medicines: A Proposal

Paul Newton and colleagues propose guidelines for conducting and reporting field surveys of the quality of medicines

Cross-sectional study of availability and pharmaceutical quality of antibiotics requested with or without prescription (Over The Counter) in Surabaya, Indonesia

Contains fulltext : 87881.pdf (publisher's version ) (Open Access

Medicines informal market in Congo, Burundi and Angola: counterfeit and sub-standard antimalarials

BACKGROUND: The presence of counterfeits and sub-standards in African medicines market is a dramatic problem that causes many deaths each year. The increase of the phenomenon of pharmaceutical counterfeiting is due to the rise of the illegal market and to the impossibility to purchase branded high cost medicines. METHODS: In this paper the results of a quality control on antimalarial tablet samples purchased in the informal market in Congo, Burundi and Angola are reported. The quality control consisted in the assay of active substance by means of validated liquid chromatographic methods, uniformity of mass determination, disintegration and dissolution tests. Moreover, a general evaluation on label and packaging characteristics was performed. RESULTS: The results obtained on thirty antimalarial tablet samples containing chloroquine, quinine, mefloquine, sulphadoxine and pyrimethamine showed the presence of different kinds of problems: a general problem concerning the packaging (loose tablets, packaging without Producer name, Producer Country and sometimes without expiry date); low content of active substance (in one sample); different, non-declared, active substance (in one sample); sub-standard technological properties and very low dissolution profiles (in about 50% of samples). This last property could affect the bioavailability and bioequivalence in comparison with branded products and could be related to the use of different excipients in formulation or bad storage conditions. CONCLUSION: This paper evidences that the most common quality problem in the analysed samples appears to be the low dissolution profile. Here it is remarked that the presence of the right active substance in the right quantity is not a sufficient condition for a good quality drug. Dissolution test is not less important in a quality control and often evidences in vitro possible differences in therapeutic efficacy among drugs with the same active content. Dissolution profile can be dramatically affected by the choice of excipients in the oral solid formulation and, in many cases, is out of specifications due to the absence of formulation studies by producers of developing countries