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BACKGROUND: Parasite clearance time after artemisinin-based combination therapy (ACT) may be increasing in Asian and African settings. The association between parasite clearance following ACT and transmissibility is currently unknown. METHODS: We determined parasite clearance dynamics by duplex quantitative polymerase chain reaction (qPCR) in samples collected in the first 3 days after treatment of uncomplicated malaria with ACT. Gametocyte carriage was determined by Pfs25 quantitative nucleic acid sequence-based amplification assays; infectiousness to mosquitoes by membrane-feeding assays on day 7 after treatment. RESULTS: Residual parasitemia was detected by qPCR in 31.8% (95% confidence interval [CI], 24.6-39.8) of the children on day 3 after initiation of treatment. Residual parasitemia was associated with a 2-fold longer duration of gametocyte carriage (P = .0007), a higher likelihood of infecting mosquitoes (relative risk, 1.95; 95% CI, 1.17-3.24; P = .015), and a higher parasite burden in mosquitoes (incidence rate ratio, 2.92; 95% CI, 1.61-5.31; P < .001). Children with residual parasitemia were also significantly more likely to experience microscopically detectable parasitemia during follow-up (relative risk, 11.25; 95% CI, 4.08-31.01; P < .001). CONCLUSIONS: Residual submicroscopic parasitemia is common after ACT and is associated with a higher transmission potential. Residual parasitemia may also have consequences for individual patients because of its higher risk of recurrent parasitemia.

Item does not contain fulltext Mycobacterium genavense is an opportunistic non-tuberculous mycobacterium previously mostly associated with HIV-infected patients with CD4 counts below 100/muL. In this retrospective observational study of medical charts we studied all Dutch patients in whom M. genavense was detected between January 2002 and January 2010. Of the 14 patients identified, 13 (93%) showed clinically relevant M. genavense disease. All patients with M. genavense disease were severely immunocompromised, including HIV-infected patients, solid organ transplant recipients, those with chronic steroid use in combination with other immune modulating drugs, recipients of chemotherapy for non-Hodgkin lymphoma, and those with immunodeficiency syndromes. Two patients had non-disseminated pulmonary M. genavense disease. Of the 12 patients treated, eight (75%) showed a favourable outcome. Four patients died in this study, three despite treatment for M. genavense disease. We conclude that M. genavense is a clinically relevant pathogen in severely immunocompromised patients that causes predominantly disseminated disease with serious morbidity and mortality. M. genavense is increasingly seen among non-HIV immunocompromised patients.

In order to switch from IS 6110 and polymorphic GC-rich repetitive sequence (PGRS) restriction fragment length polymorphism (RFLP) to 24-locus variable-number tandem-repeat (VNTR) typing of Mycobacterium tuberculosis complex isolates in the national tuberculosis control program in The Netherlands, a detailed evaluation on discriminatory power and agreement with findings in a cluster investigation was performed on 3,975 tuberculosis cases during the period of 2004 to 2008. The level of discrimination of the two typing methods did not differ substantially: RFLP typing yielded 2,733 distinct patterns compared to 2,607 in VNTR typing. The global concordance, defined as isolates labeled unique or identically distributed in clusters by both methods, amounted to 78.5% ( n = 3,123). Of the remaining 855 cases, 12% ( n = 479) of the cases were clustered only by VNTR, 7.7% ( n = 305) only by RFLP typing, and 1.8% ( n = 71) revealed different cluster compositions in the two approaches. A cluster investigation was performed for 87% ( n = 1,462) of the cases clustered by RFLP. For the 740 cases with confirmed or presumed epidemiological links, 92% were concordant with VNTR typing. In contrast, only 64% of the 722 cases without an epidemiological link but clustered by RFLP typing were also clustered by VNTR typing. We conclude that VNTR typing has a discriminatory power equal to IS 6110 RFLP typing but is in better agreement with findings in a cluster investigation performed on an RFLP-clustering-based cluster investigation. Both aspects make VNTR typing a suitable method for tuberculosis surveillance systems.

Controlled human malaria infection with sporozoites is a standardized and powerful tool for evaluation of malaria vaccine and drug efficacy but so far only applied by exposure to bites of Plasmodium falciparum (Pf)-infected mosquitoes. We assessed in an open label Phase 1 trial, infection after intradermal injection of respectively 2,500, 10,000, or 25,000 aseptic, purified, vialed, cryopreserved Pf sporozoites (PfSPZ) in three groups (N = 6/group) of healthy Dutch volunteers. Infection was safe and parasitemia developed in 15 of 18 volunteers (84%), 5 of 6 volunteers in each group. There were no differences between groups in time until parasitemia by microscopy or quantitative polymerase chain reaction, parasite kinetics, clinical symptoms, or laboratory values. This is the first successful infection by needle and syringe with PfSPZ manufactured in compliance with regulatory standards. After further optimization, the use of such PfSPZ may facilitate and accelerate clinical development of novel malaria drugs and vaccines.

Item does not contain fulltext Rationale: The dosage of 10 mg/kg/d rifampin, as currently used in the treatment of tuberculosis (TB), is not an optimal dose. Shortening of treatment duration might be achievable using an increased rifampin dose. Objectives: Determination of optimal rifampin dosage in mice, resulting in maximum therapeutic effect and without adverse effects. Assessment of associated pharmacokinetic parameters and pharmacokinetic/pharmacodynamic indices. Methods: A murine TB infection using a Beijing genotype Mycobacterium tuberculosis strain was established by intratracheal bacterial instillation followed by proper inhalation, while keeping mice in a vertical position. We assessed dose-dependent activity of rifampin in single-drug treatment during 3 weeks. The maximum tolerated dosage, pharmacokinetic parameters, and pharmacokinetic/pharmacodynamic index were determined. Therapeutic efficacy of a range of rifampin (R) dosages added to a regimen of isoniazid (H) and pyrazinamide (Z) was assessed. Measurements and Main Results: Maximum tolerated dosage of rifampin in the murine TB was 160 mg/kg/d. Pharmacokinetic measurement in HR(10)Z and HR(160)Z therapy regimens showed for rifampin a Cmax of 16.2 and 157.3 mg/L, an AUC0-24h of 132 and 1,782 h.mg/L, and AUC0-24h/minimum inhibitory concentration ratios of 528 and 7129, respectively. A clear dose-effect correlation was observed for rifampin after 3-week single-drug treatment. Administration of HR(80)Z allowed 9-week treatment duration to be effective without relapse of infection. Conclusions: Our findings indicate that the currently used rifampin dosage in the therapy of TB is too low. In our murine TB model a rifampin dosage of 80 mg/kg/d enabled a significant reduction in therapy duration without adverse effects.

Item does not contain fulltext BACKGROUND: 'Covering your cough' reduces droplet number, but its effect on airborne pathogen transmission is less clear. The World Health Organization specifically recommends cough etiquette to prevent the spread of Mycobacterium tuberculosis, but implementation is generally poor and evidence supporting its value is lacking. METHODS: We constructed a model to assess 'real life' transmission risk by counting viable pathogens from aerosols produced by coughing patients, thus allowing the assessment of outward protection measures in a standardised fashion. During the validation process, we focused on rod-shaped bacteria as surrogates for M. tuberculosis. RESULTS: The Cough Cylinder enabled us to sample Pseudomonas aeruginosa, Escherichia coli and mycobacteria from aerosols produced by patients with cystic fibrosis, primary ciliary dyskinesia and tuberculosis. Pathogens in droplets and in airborne particles could be sampled. Delayed air sampling allowed specific measurement of persistent airborne particles. CONCLUSION: This novel experimental system allows measurement of aerosol pathogen spread in a highly standardised fashion. It also offers the possibility to assess the impact of different interventions to limit aerosol transmission. 01 januari 2013 8 p.

Item does not contain fulltext Co-trimoxazole (SXT), a combination of sulfamethoxazole and trimethoprim, has shown in vitro activity against Mycobacterium tuberculosis. However, the pharmacokinetic and pharmacodynamic parameters of SXT in multidrug-resistant (MDR) tuberculosis (TB) are, thus far, lacking. Therefore, we evaluated its pharmacokinetics and drug susceptibility, along with its tolerability during treatment. Based on drug susceptibility testing, MDR-TB patients received SXT as a part of their MDR treatment. The pharmacokinetic parameters of sulfamethoxazole, the effective component of SXT against M. tuberculosis, were evaluated. The ratio of the area under the curve from 0 to 24 h (AUC0-24) to minimum inhibitory concentration (MIC) was used as the best pharmacokinetic/pharmacodynamic parameter to predict the efficacy of sulfamethoxazole. Adverse effects of SXT were also evaluated. 10 patients with MDR-TB (one of whom had extensively drug-resistant TB) received 480 mg of SXT with a median dosage of 6.5 mg.kg(-1) of SXT (range 6.1-6.8 mg.kg(-1)) once daily for a median treatment period of 381 days (range 129-465 days). In two patients, the dose was escalated to 960 mg. The free AUC0-24/MIC of sulfamethoxazole exceeded 25 in only one patient. SXT was safe and well-tolerated, except for one patient who had gastrointestinal side-effects after receiving 960 mg of SXT. Additional studies are needed to find the pharmacokinetic and pharmacodynamic targets, and consequently to set the optimal dose, of SXT for MDR-TB treatment. 01 augustus 2013

Despite their high clinical and socioeconomic impacts, there is currently no approved antiviral therapy for the prophylaxis or treatment of enterovirus infections. Here we report on a novel inhibitor of enterovirus replication, compound 1, 2-fluoro-4-(2-methyl-8-(3-(methylsulfonyl)benzylamino)imidazo[1,2- a ]pyrazin-3-yl)phenol. This compound exhibited a broad spectrum of antiviral activity, as it inhibited all tested species of enteroviruses and rhinoviruses, with 50% effective concentrations ranging between 4 and 71 nM. After a lengthy resistance selection process, coxsackievirus mutants resistant to compound 1 were isolated that carried substitutions in their 3A protein. Remarkably, the same substitutions were recently shown to provide resistance to inhibitors of phosphatidylinositol 4-kinase IIIβ (PI4KIIIβ), a lipid kinase that is essential for enterovirus replication, suggesting that compound 1 may also target this host factor. Accordingly, compound 1 directly inhibited PI4KIIIβ in an in vitro kinase activity assay. Furthermore, the compound strongly reduced the PI 4-phosphate levels of the Golgi complex in cells. Rescue of coxsackievirus replication in the presence of compound 1 by a mutant PI4KIIIβ carrying a substitution in its ATP-binding pocket revealed that the compound directly binds the kinase at this site. Finally, we determined that an analogue of compound 1, 3-(3-fluoro-4-methoxyphenyl)-2-methyl- N -(pyridin-4-ylmethyl)imidazo[1,2- a ]pyrazin-8-amine, is well tolerated in mice and has a dose-dependent protective activity in a coxsackievirus serotype B4-induced pancreatitis model.

Item does not contain fulltext Tuberculosis (TB) has re-emerged over the past two decades: in industrialized countries in association with immigration, and in Africa owing to the human immunodeficiency virus epidemic. Drug-resistant TB is a major threat worldwide. The variable and uncertain impact of TB control necessitates not only better tools (diagnostics, drugs, and vaccines), but also better insights into the natural history and epidemiology of TB. Molecular epidemiological studies over the last two decades have contributed to such insights by answering long-standing questions, such as the proportion of cases attributable to recent transmission, risk factors for recent transmission, the occurrence of multiple Mycobacterium tuberculosis infection, and the proportion of recurrent TB cases attributable to re-infection. M. tuberculosis lineages have been identified and shown to be associated with geographical origin. The Beijing genotype is strongly associated with multidrug resistance, and may have escaped from bacille Calmette-Guerin-induced immunity. DNA fingerprinting has quantified the importance of institutional transmission and laboratory cross-contamination, and has helped to focus contact investigations. Questions to be answered in the near future with whole genome sequencing include identification of chains of transmission within clusters of patients, more precise quantification of mixed infection, and transmission probabilities and rates of progression from infection to disease of various M. tuberculosis lineages, as well as possible variations in vaccine efficacy by lineage. Perhaps most importantly, dynamics in the population structure of M. tuberculosis in response to control measures in high-prevalence areas should be better understood. 01 oktober 2013

Despite great effort by health organizations worldwide in fighting tuberculosis (TB), morbidity and mortality are not declining as expected. One of the reasons is related to the evolutionary development of Mycobacterium tuberculosis , in particular the Beijing genotype strains. In a previous study, we showed the association between the Beijing genotype and an increased mutation frequency for rifampin resistance. In this study, we use a Beijing genotype strain and an East-African/Indian genotype strain to investigate with our mouse TB model whether the higher mutation frequency observed in a Beijing genotype strain is associated with treatment failure particularly during noncompliance therapy. Both genotype strains showed high virulence in comparison to that of M. tuberculosis strain H37Rv, resulting in a highly progressive infection with a rapid lethal outcome in untreated mice. Compliance treatment was effective without relapse of TB irrespective of the infecting strain, showing similar decreases in the mycobacterial load in infected organs and similar histopathological changes. Noncompliance treatment, simulated by a reduced duration and dosing frequency, resulted in a relapse of infection. Relapse rates were correlated with the level of noncompliance and were identical for Beijing infection and East African/Indian infection. However, only in Beijing-infected mice, isoniazid-resistant mutants were selected at the highest level of noncompliance. This is in line with the substantial selection of isoniazid-resistant mutants in vitro in a wide isoniazid concentration window observed for the Beijing strain and not for the EAI strain. These results suggest that genotype diversity of M. tuberculosis may be involved in emergence of resistance and indicates that genotype-tailor-made treatment should be investigated.

Item does not contain fulltext OBJECTIVE: The introduction of molecular typing methods in the 1990s to study the epidemiology of tuberculosis (TB) has significantly improved the possibilities of quantifying transmission of Mycobacterium tuberculosis in different human settings. The purpose of this study was to investigate transmission of TB in 35 family-households in Poland. METHODS: Two PCR-based genotyping methods: spoligotyping and mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) typing were used. RESULTS: Of 78 patients, 49 (63%), could be assigned to intra-household transmission on the basis of identical DNA fingerprints upon a combined typing approach. However, if a single spoligotype spacer or a single MIRU-VNTR locus variation was tolerated in the cluster definition, the intra-household transmission raised to 85% of all patients. For 12 patients in 6 households, the M. tuberculosis isolates were clearly distinct in either spoligotyping or VNTR typing or in both genotyping methods, suggesting that these patients were infected by the sources in the community. CONCLUSIONS: This study is the first to provide the results of a molecular epidemiological investigation performed within family-households in Poland. It shows the household setting as an important reservoir of M. tuberculosis transmission, and thus argues in favor of routine and extensive screening of the family contacts of TB patients. 01 juni 2012

Item does not contain fulltext Phylogenetic analyses on the basis of multiple house-keeping genes and whole genome sequences have offered new insights in the phylogeny of the genus Mycobacterium. This genus yields obligate pathogens, the M. tuberculosis complex and M. leprae, as well as opportunistic pathogens (e.g. M. avium, M. intracellulare, M. kansasii, M. marinum, M. malmoense) and saprophytes (e.g. M. phlei, M. sphagni, M. gordonae). The most virulent mycobacteria, the M. tuberculosis complex, M. leprae and the M. kansasii-M. szulgai-M. marinum-M. ulcerans group are phylogenetically related and infections by these organisms are better treatable than those caused by less virulent and phylogenetically more distantly related Mycobacterium species. The most virulent Mycobacterium species are also characterized by high levels of natural drug susceptibility. In this paper, we review studies of phylogeny, drug susceptibility, and clinical significance to support our hypothesis that drug susceptibility in mycobacteria is acquired and reflects the low level of competition in -and adaptation to- a closer-to-human (environmental) niche. In turn, mycobacteria that inhabit the most competitive environmental niches are the least adapted to humans, thus of low clinical significance, but most tolerant to antibiotics derived from microbes with which they share their habitat, lowering the chances of cure in case of infection. 01 juni 2012

Upon the invasion of the host by microorganisms, innate immunity is triggered through pathogen recognition by pattern recognition receptors (PRRs). Toll-like receptors (TLRs) are the best-studied class of PRRs, and they recognize specific pathogen-associated molecular patterns (PAMPs) from various microorganisms. A large number of studies have shown that genetic variation in TLRs may influence susceptibility to infections. We assessed the genetic variation of TLR2 , which encodes one of the most important TLRs, in various populations around the globe and correlated it with changes in the function of the molecule. The three best-known nonsynonymous TLR2 polymorphisms (1892C>A, 2029C>T, and 2258G>A) were assessed in different populations from the main continental masses: Romanians, Vlax-Roma, Dutch (European populations), Han Chinese (East Asia), Dogon, Fulani (Africa), and Trio Indians (America). The 2029C>T polymorphism was absent in both European and non-European populations, with the exception of the Vlax-Roma, suggesting that this polymorphism most likely arose in Indo-Aryan people after migration into South Asia. The 1892C>A polymorphism that was found exclusively in European populations, but not in Asian, African, or American volunteers, probably occurred in proto-Indo-Europeans. Interestingly, 2258G>A was present only in Europeans, including Vlax-Roma, but at a very low frequency. The differential pattern of the TLR2 polymorphisms in various populations may explain some of the differences in susceptibility to infections between these populations.

Contains fulltext : 110971.pdf (Publisher’s version ) (Open Access) To determine differences in the ability of Mycobacterium tuberculosis strains to withstand antituberculosis drug treatment, we compared the activity of antituberculosis drugs against susceptible Beijing and East-African/Indian genotype M. tuberculosis strains. Beijing genotype strains showed high rates of mutation within a wide range of drug concentrations, possibly explaining this genotype's association with multidrug-resistant tuberculosis. 01 april 2012

Item does not contain fulltext Mycobacterium species, including Mycobacterium tuberculosis and Mycobacterium leprae, are among the most potent human bacterial pathogens. The discovery of cytosolic mycobacteria challenged the paradigm that these pathogens exclusively localize within the phagosome of host cells. As yet the biological relevance of mycobacterial translocation to the cytosol remained unclear. In this current study we used electron microscopy techniques to establish a clear link between translocation and mycobacterial virulence. Pathogenic, patient-derived mycobacteria species were found to translocate to the cytosol, while non-pathogenic species did not. We were further able to link cytosolic translocation with pathogenicity by introducing the ESX-1 (type VII) secretion system into the non-virulent, exclusively phagolysosomal Mycobacterium bovis BCG. Furthermore, we show that translocation is dependent on the C-terminus of the early-secreted antigen ESAT-6. The C-terminal truncation of ESAT-6 was shown to result in attenuation in mice, again linking translocation to virulence. Together, these data demonstrate the molecular mechanism facilitating translocation of mycobacteria. The ability to translocate from the phagolysosome to the cytosol is with this study proven to be biologically significant as it determines mycobacterial virulence. 01 augustus 2012

Contains fulltext : 107786.pdf (Publisher’s version ) (Closed access) Mycobacterium bovis isolates from the Iberian Peninsula are dominated by strains with spoligotype patterns deleted for spacer 21. Whole-genome sequencing of three Spanish strains with spacer 21 missing in their spoligotype pattern revealed a series of SNPs and subsequent screening of a selection of these SNPs identified one in gene guaA that is specific to these strains. This group of strains from the Iberian Peninsula missing spoligotype spacer 21 represents a new clonal complex of M. bovis, defined by the SNP profile with a distinct spoligotype signature. We have named this clonal complex European 2 (Eu2) and found that it was present at low frequency in both France and Italy and absent from the British Isles. 01 juni 2012

Item does not contain fulltext The development of a vaccine against malaria has public health priority. In a controlled setting, preliminary data on the efficacy of Plasmodium falciparum vaccine candidates can be obtained by exposing immunized human volunteers to the bites of laboratory-reared P. falciparum-infected mosquitoes. Using empirical data, we show that these trials, with small numbers of volunteers, are sufficiently powered to detect protective biological effects induced by preerythrocytic and/or blood-stage candidate vaccines if parasitemia is measured daily by quantitative polymerase chain reaction. Sporozoite challenge trials are thus a powerful tool for early selection of candidates that warrant efficacy of trials in the field. 01 augustus 2012

The immune regulatory mechanisms involved in the acquisition of Mycobacterium tuberculosis infection in children are largely unknown. We investigated the influence of parasitic infections, malnutrition and plasma cytokine profiles on tuberculin skin test (TST) positivity in Warao Amerindians in Venezuela. Pediatric household contacts of sputum smear-positive tuberculosis (TB) cases were enrolled for TST, chest radiograph, plasma cytokine analyses, QuantiFERON-TB Gold In-Tube (QFT-GIT) testing and stool examinations. Factors associated with TST positivity were studied using generalized estimation equations logistic regression models. Of the 141 asymptomatic contacts, 39% was TST-positive. After adjusting for age, gender and nutritional status, TST positivity was associated with Trichuris trichiura infections (OR 3.5, 95% CI 1.1-11.6) and low circulating levels of T helper 1 (Th1) cytokines (OR 0.51, 95% CI 0.33-0.79). Ascaris lumbricoides infections in interaction with Th2- and interleukin (IL)-10-dominated cytokine profiles were positively associated with TST positivity (OR 3.1, 95% CI 1.1-8.9 and OR 2.4,95% CI 1.04-5.7, respectively). A negative correlation of QFT-GIT mitogen responses with Th1 and Th2 levels and a positive correlation with age were observed (all p

Contains fulltext : 110533.pdf (Publisher’s version ) (Closed access) In this study, nonchromogenic mycobacteria were isolated from pulmonary samples of three patients in the Netherlands. All isolates had identical, unique 16S rRNA gene and 16S-23S ITS sequences, which were closely related to those of Mycobacterium chimaera and Mycobacterium marseillense. The biochemical features of the isolates differed slightly from those of M. chimaera, suggesting that the isolates may represent a possible separate species within the Mycobacterium avium complex (MAC). However, the cell-wall mycolic acid pattern, analysed by HPLC, and the partial sequences of the hsp65 and rpoB genes were identical to those of M. chimaera. We concluded that the isolates represent a novel variant of M. chimaera. The results of this analysis have led us to question the currently used methods of species definition for members of the genus Mycobacterium, which are based largely on 16S rRNA or rpoB gene sequencing. Definitions based on a single genetic target are likely to be insufficient. Genetic divergence, especially in the MAC, yields strains that cannot be confidently assigned to a specific species based on the analysis of a single genetic target. 01 september 2012

F ive Mycobacterium tuberculosis isolates were obtained from three body sites from a Dutch patient. The isolates displayed a single genotype by 24-locus MIRU-VNTR typing (except for a single locus not amplified from one isolate) but were differentiated by small variations in IS 6110 fingerprints, spoligotypes, 6 hypervariable MIRU-VNTR loci, and/or DiversiLab profiles, revealing patterns of microevolution in a clonal infection.

Item does not contain fulltext 01 december 2012

To the Editors: Nontuberculous mycobacteria (NTM) are opportunistic pathogens that are widely present in our environment, i.e. in the soil and in natural and processed water. This group of organisms, with varying biological properties and clinical relevance, has gained notoriety over the past two decades due to their capability to cause severe disease in patients with immunodeficiency and/or chronic lung disease [1]. Despite their rising isolation frequency and growing clinical importance, NTM infections receive little scientific or public health attention. This partly results from the dogma that these NTM infections are not transmitted from male-to-male [1]. During our previous research of Mycobacterium malmoense infection in the Netherlands [2], we studied two patients with M. malmoense pulmonary infection with a strong epidemiological link. Human transmission was suspected because an acid-fast bacilli (AFB) smear-positive patient was in close contact with an immunocompromised patient. Herein, we present details of the cases and the results of the molecular diagnostics that were performed. The patients gave informed consent; ethical approval was not required for this retrospective study. A 38-yr-old male with a medical history of chronic obstructive pulmonary disease and spontaneous pneumothorax reported to the respiratory physician with progressive dyspnoea, productive cough, fever, night sweats, malaise and weight loss. A chest radiograph revealed infiltrates with cavities in the upper lobes. Laboratory diagnostics showed a raised erythrocyte sedimentation rate (90 mm·h−1) and leukocytosis (white blood cell count …

Contains fulltext : 124358.pdf (Publisher’s version ) (Open Access) Using population-based data from rural Vietnam, we assessed tuberculosis (TB) transmission within and outside of households. Eighty-three percent of persons with recent household TB were infected by different strains of Mycobacterium tuberculosis than were their household members. This result argues against the effectiveness of active TB case finding among household members. 01 september 2010

We compared 14 molecular assays for their ability to detect the Mycobacterium tuberculosis complex in bronchoalveolar lavage fluid samples. Three approaches were followed. First, by using DNA from Mycobacterium bovis BCG, we determined the detection limits of the assays using routine molecular methods. Second, in order to determine the analytical sensitivities of the assays, we added one of four M. tuberculosis isolates with various numbers of the insertion sequence IS 6110 to N -acetyl- l -cysteine (NALC)-NaOH-treated bronchoalveolar lavage fluid samples in dilutions of 1:10 to 1:10,000,000. Third, intertest variabilities were measured and defined by the standard deviations for the quantitation cycle (Cq) values of three positive test results per dilution per assay. The 14 assays tested had similar analytical sensitivities, except for GeneXpert, which had an analytical sensitivity that was 10- to 100-fold lower than that of the other assays. The MP MTB/NTM test and the in-house TaqMan-10 revealed the best performances for the detection limit and had the highest analytical sensitivities. Most of the tests performed well regarding detection limit and analytical sensitivity for the detection of the M. tuberculosis complex in serial dilutions, and the differences were small. The MP MTB/NTM and the in-house TaqMan-10 assays revealed the best, and GeneXpert the worst, overall performances.

One of the most widespread clades of Mycobacterium tuberculosis worldwide, the Beijing genotype family, consists of ancient (atypical) and modern (typical) strains. Modern Beijing strains outcompete ancient strains in terms of prevalence, while reserving a higher degree of genetic conservation. We hypothesize that their selective advantage lies in eliciting a different host immune response. Bead-disrupted lysates of a collection of different M. tuberculosis strains of the modern ( n = 7) or ancient ( n = 7) Beijing genotype, as well as the Euro-American lineage ( n = 6), were used for induction of ex vivo cytokine production in peripheral blood mononuclear cells (PBMCs) from 10 healthy individuals. Hierarchical clustering and multivariate regression analyses were used to study possible differences in production of nine cytokines. Modern and ancient M. tuberculosis Beijing genotypes induced different cytokine signatures. Overall induction of interleukin-1β (IL-1β), gamma interferon (IFN-γ), and IL-22 was 38 to 40% lower after stimulation with modern Beijing strains (corrected P values of

Item does not contain fulltext One of the bottlenecks in the development of a whole sporozoite malaria vaccine is the route and method of sporozoite administration. Immunization and challenge of human volunteers by mosquito bites is effective, but cannot be used as a vaccine. Intravenous immunization with sporozoites is effective in rodents and non-human primates, and being studied in humans, but is not yet used for licensed vaccines for infectious diseases. Intradermal and subcutaneous immunization regimens show a strong decrease in protective efficacy, which in rodents, is associated with a decreased degree of parasite liver infection during immunization. The objective of this study was to explore alternative routes of sporozoite administration to increase efficiency of liver infection. Using in vivo imaging, we found that IM injection of sporozoites resulted in a greater parasite liver load compared to ID and SC injection. The use of small inoculation volumes and multiple injections further increased the subsequent liver load. These observations were corroborated in a Plasmodium yoelii model using cryopreserved sporozoites administered ID. Our findings provide a rationale for the design of clinical trials to optimize needle and syringe administration of Plasmodium falciparum sporozoites.

Sequence variation in the asparagine/aspartate-rich domain of pfmdr1 in 215 isolates of Plasmodium falciparum from three African countries was compared with published data. The role of this domain in modulating antimalarial sensitivity has not been established. The pfmdr1 86Y allele was significantly associated with different configurations of the Asn/Asp-rich domain in West and East Africa. In Kenya, a specific form of the Asn/Asp-rich domain was significantly linked to the 86Y, 184Y, and 1246Y haplotype of pfmdr1 .

BACKGROUND: School surveys provide an operational approach to assess malaria transmission through parasite prevalence. There is limited evidence on the comparability of prevalence estimates obtained from school and community surveys carried out at the same locality. METHODS: Concurrent school and community cross-sectional surveys were conducted in 46 school/community clusters in the western Kenyan highlands and households of school children were geolocated. Malaria was assessed by rapid diagnostic test (RDT) and combined seroprevalence of antibodies to bloodstage Plasmodium falciparum antigens. RESULTS: RDT prevalence in school and community populations was 25.7% (95% CI: 24.4-26.8) and 15.5% (95% CI: 14.4-16.7), respectively. Seroprevalence in the school and community populations was 51.9% (95% CI: 50.5-53.3) and 51.5% (95% CI: 49.5-52.9), respectively. RDT prevalence in schools could differentiate between low (39%, 95% CI: 25-49%) transmission areas in the community and, after a simple adjustment, were concordant with the community estimates. CONCLUSIONS: Estimates of malaria prevalence from school surveys were consistently higher than those from community surveys and were strongly correlated. School-based estimates can be used as a reliable indicator of malaria transmission intensity in the wider community and may provide a basis for identifying priority areas for malaria control.

Volunteers immunized under chloroquine chemoprophylaxis with Plasmodium falciparum sporozoites (CPS) develop complete, long-lasting protection against homologous sporozoite challenge. Chloroquine affects neither sporozoites nor liver-stages, but kills only asexual forms in erythrocytes once released from the liver into the circulation. Consequently, CPS immunization exposes the host to antigens from both preerythrocytic and blood stages, and induced immunity might target either of these stages. We therefore explored the life cycle stage specificity of CPS-induced protection. Twenty-five malaria-naïve volunteers were enrolled in a clinical trial, 15 of whom received CPS immunization. Five immunized subjects and five controls received a sporozoite challenge by mosquito bites, whereas nine immunized and five control subjects received an i.v. challenge with P. falciparum- infected erythrocytes. The latter approach completely bypasses preerythrocytic stages, enabling a direct comparison of protection against either life cycle stage. CPS-immunized subjects (13 of 14) developed anticircumsporozoite antibodies, whereas only one volunteer generated minimal titers against typical blood-stage antigens. IgG from CPS-immunized volunteers did not inhibit asexual blood-stage growth in vitro. All CPS-immunized subjects (5 of 5) were protected against sporozoite challenge. In contrast, nine of nine CPS-immunized subjects developed parasitemia after blood-stage challenge, with identical prepatent periods and blood-stage multiplication rates compared with controls. Intravenously challenged CPS-immunized subjects showed earlier fever and increased plasma concentrations of inflammatory markers D-dimer, IFN-γ, and monokine induced by IFN-γ than i.v. challenged controls. The complete lack of protection against blood-stage challenge indicates that CPS-induced protection is mediated by immunity against preerythrocytic stages. However, evidence is presented for immune recognition of P. falciparum -infected erythrocytes, suggesting memory responses unable to generate functional immunity.

German Federal Ministry of Education and Research A significant knowledge gap exists concerning the geographical distribution of nontuberculous mycobacteria (NTM) isolation worldwide.To provide a snapshot of NTM species distribution, global partners in the NTM-Network European Trials Group (NET) framework (www.ntm-net.org), a branch of the Tuberculosis Network European Trials Group (TB-NET), provided identification results of the total number of patients in 2008 in whom NTM were isolated from pulmonary samples. From these data, we visualised the relative distribution of the different NTM found per continent and per country.We received species identification data for 20182 patients, from 62 laboratories in 30 countries across six continents. 91 different NTM species were isolated. Mycobacterium avium complex (MAC) bacteria predominated in most countries, followed by M. gordonae and M. xenopi. Important differences in geographical distribution of MAC species as well as M. xenopi, M. kansasii and rapid-growing mycobacteria were observed.This snapshot demonstrates that the species distribution among NTM isolates from pulmonary specimens in the year 2008 differed by continent and differed by country within these continents. These differences in species distribution may partly determine the frequency and manifestations of pulmonary NTM disease in each geographical location. Radboud Univ Nijmegen, Med Ctr, Dept Pulm Dis, NL-6500 HB Nijmegen, Netherlands Radboud Univ Nijmegen, Med Ctr, Dept Med Microbiol, NL-6500 HB Nijmegen, Netherlands Amiens Teaching Hosp, Dept Resp Dis, Amiens, France Karolinska Inst, Dept Clin Microbiol, Stockholm, Sweden Hop Purpan, Lab Cent Microbiol, Toulouse, France CHU Nantes, Inst Biol, Serv Bacteriol Hyg, F-44035 Nantes 01, France Natl Hlth Lab Serv, Braamfontein, South Africa Univ Hosp Getafe, Dept Microbiol, Getafe, Spain Aurum Inst Hlth Res, Johannesburg, South Africa Adolfo Lutz Inst, Secao Bacteriol, Setor Micobacterias, São Paulo, Brazil Hosp Clin San Carlos, Dept Clin Microbiol, Madrid, Spain Hosp Univ Puerta de Hierro, Dept Microbiol, Madrid, Spain Natl Jewish Hlth, Div Mycobacterial & Resp Infect, Denver, CO USA Hosp Univ Princesa, Dept Microbiol, Madrid, Spain UK Hlth Protect Agcy, Natl Mycobacterium Reference Lab, London, England IIS Fdn Jimenez Diaz, Dept Clin Microbiology, Madrid, Spain Sci Inst Publ Hlth, Natl Reference Ctr TB & Mycobacteria, Brussels, Belgium Statens Serum Inst, Int Reference Lab Mycobacteriol, DK-2300 Copenhagen, Denmark St James Hosp, Dublin 8, Ireland Univ Dublin Trinity Coll, Dublin 2, Ireland Hosp Ramon & Cajal, E-28034 Madrid, Spain Hosp Univ Principe Asturias, Madrid, Spain Asklepios Facbkliniken, Supranat Reference Lab, Inst Microbiol & Lab Med, Gauting, Germany Natl Taiwan Univ Hosp, Dept Lab Med, Taipei, Taiwan Natl Taiwan Univ Hosp, Dept Internal Med, Taipei 100, Taiwan Austrian Agcy Hlth & Food Safety, Vienna, Austria Univ Warsaw, Fac Biol, Inst Microbiol, Dept Appl Microbiol, Warsaw, Poland Ontario Minist Hlth & Long Term Care, Publ Hlth Labs Branch, Toronto, ON, Canada Univ Zagreb Med Sch, Univ Hosp Ctr, Dept Resp Dis, Zagreb, Croatia Univ Witwatersrand, Clin HIV Res Unit, Johannesburg, South Africa St Jamess Hosp Dublin, Dublin, Ireland Sungkyunkwan Univ Sch Med, Samsung Med Ctr, Dept Med, Div Pulm & Crit Care Med, Seoul, South Korea Univ Hosp Freiburg, Ctr Infect Dis & Travel Med, Freiburg, Germany Universidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Imunol & Parasitol, Disciplina Microbiol, São Paulo, Brazil Adm Reg Saude Lisboa & Vale Tejo, Dept Publ Hlth, Publ Hlth Lab Mycobacteriol TB, Lisbon, Portugal Norwegian Inst Publ Hlth, Div Infect Dis Control, Oslo, Norway Univ Toronto, Univ Hlth Network, Joint Div Respirol, Toronto, ON, Canada Univ Toronto, Mt Sinai Hosp, Dept Med, Toronto, ON M5G 1X5, Canada Univ Victor Segalen Bordeaux, Lab Bacteriol, Bordeaux, France Guys & St Thomas NHS Fdn Trust, Dept Resp Med, London, England Koranyi Natl Inst TB & Pulmonol, Budapest, Hungary Hosp Dr Antonio Cetrangolo, Programa Control TB Prov Buenos Aires, Lab Referencia, Buenos Aires, Brazil Fukujuji Hosp, Japan AntiTB Assoc, Tokyo, Japan Sotiria Chest Dis Hosp, Natl Mycobacteria Reference Lab, Athens, Greece Hosp 12 Octubre, Dept Microbiol, E-28041 Madrid, Spain Hosp Severo Ochoa, Madrid, Spain United Hosp, Reg Reference Mycobacteria Lab, Ancona, Italy Natl Inst TB Resp Dis & Chest Surg, Vysne Hagy, Slovakia Inst Pasteur Guadeloupe, Unite TB & Mycobacteries, WHO Supranat TB Reference Lab, Abymes, Guadeloupe Natl Reference Ctr Mycobacteria, Borstel, Germany Hosp Gregorio Maranon, Serv Microbiol Clin & Enfermedades Infecciosas, Madrid, Spain Gen Directorate Hlth Lisbon, Amadora, Portugal Lung Dis Ctr Venda Nova, Amadora, Portugal Univ Witwatersrand, Johannesburg, South Africa Refik Saydam Natl Publ Hlth Agcy, Natl TB Reference Lab, Ankara, Turkey Natl Inst Publ Hlth & Environm, NL-3720 BA Bilthoven, Netherlands Koranyi Natl Inst TB & Resp Med, Budapest, Hungary Greenslopes Private Hosp, Gallipoli Med Res Ctr, Brisbane, Qld, Australia Hosp Val dHebron, Barcelona, Spain Ist Sci San Raffaele, Emerging Bacterial Pathogens Unit, I-20132 Milan, Italy Natl Jewish Hlth, Adv Diagnost Labs, Dept Mycobacteriol, Denver, CO USA Univ Calgary, Prov Lab Publ Hlth, Calgary, AB, Canada Natl Inst Hlth & Welf, Epidemiol Surveillance & Response Unit, Dept Infect Dis Surveillance & Control, Helsinki, Finland Med Univ Warsaw, Dept Internal Med Pneumol & Allergol, Warsaw, Poland Oregon Hlth & Sci Univ, Div Infect Dis Publ Hlth & Prevent Med, Portland, OR 97201 USA Univ Freiburg, Dept Internal Med Infect Dis 2, D-79106 Freiburg, Germany Universidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Imunol & Parasitol, Disciplina Microbiol, São Paulo, Brazil German Federal Ministry of Education and Research: BMBF 01 EO 0803 Web of Science

Contains fulltext : 117695.pdf (Publisher’s version ) (Open Access) OBJECTIVES: For the purpose of blocking transmission of Plasmodium falciparum malaria from humans to mosquitoes, a single dose of primaquine is recommended by the WHO as an addition to artemisinin combination therapy. Primaquine clears gametocytes but causes dose-dependent haemolysis in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency. Evidence is needed to inform the optimal dosing of primaquine for malaria elimination programmes and for the purpose of interrupting the spread of artemisinin-resistant malaria. This study investigates the efficacy and safety of reducing doses of primaquine for clearance of gametocytes in participants with normal G6PD status. METHODS AND ANALYSIS: In this prospective, four-armed randomised placebo-controlled double-blinded trial, children aged 1-10 years, weighing over 10 kg, with haemoglobin >/=8 g/dl and uncomplicated P falciparum malaria are treated with artemether lumefantrine and randomised to receive a dose of primaquine (0.1, 0.4 or 0.75 mg base/kg) or placebo on the third day of treatment. Participants are followed up for 28 days. Gametocytaemia is measured by quantitative nucleic acid sequence-based analysis on days 0, 2, 3, 7, 10 and 14 with a primary endpoint of the number of days to gametocyte clearance in each treatment arm and secondarily the area under the curve of gametocyte density over time. Analysis is for non-inferiority of efficacy compared to the reference dose, 0.75 mg base/kg. Safety is assessed by pair-wise comparisons of the arithmetic mean (+/-SD) change in haemoglobin concentration per treatment arm and analysed for superiority to placebo and incidence of adverse events. Ethics and dissemination Approval was obtained from the ethical committees of Makerere University School of Medicine, the Ugandan National Council of Science and Technology and the London School of Hygiene and Tropical Medicine. RESULTS: These will be disseminated to inform malaria elimination policy, through peer-reviewed publication and academic presentations.

Contains fulltext : 117784.pdf (Publisher’s version ) (Open Access) BACKGROUND: Helminth infections are proposed to have immunomodulatory activities affecting health outcomes either detrimentally or beneficially. We evaluated the effects of albendazole treatment, every three months for 21 months, on STH, malarial parasitemia and allergy. METHODS AND FINDINGS: A household-based cluster-randomized, double-blind, placebo-controlled trial was conducted in an area in Indonesia endemic for STH. Using computer-aided block randomization, 481 households (2022 subjects) and 473 households (1982 subjects) were assigned to receive placebo and albendazole, respectively, every three months. The treatment code was concealed from trial investigators and participants. Malarial parasitemia and malaria-like symptoms were assessed in participants older than four years of age while skin prick test (SPT) to allergens as well as reported symptoms of allergy in children aged 5-15 years. The general impact of treatment on STH prevalence and body mass index (BMI) was evaluated. Primary outcomes were prevalence of malarial parasitemia and SPT to any allergen. Analysis was by intention to treat. At 9 and 21 months post-treatment 80.8% and 80.1% of the study subjects were retained, respectively. The intensive treatment regiment resulted in a reduction in the prevalence of STH by 48% in albendazole and 9% in placebo group. Albendazole treatment led to a transient increase in malarial parasitemia at 6 months post treatment (OR 4.16(1.35-12.80)) and no statistically significant increase in SPT reactivity (OR 1.18(0.74-1.86) at 9 months or 1.37 (0.93-2.01) 21 months). No effect of anthelminthic treatment was found on BMI, reported malaria-like- and allergy symptoms. No adverse effects were reported. CONCLUSIONS: The study indicates that intensive community treatment of 3 monthly albendazole administration for 21 months over two years leads to a reduction in STH. This degree of reduction appears safe without any increased risk of malaria or allergies. TRIAL REGISTRATION: Controlled-Trials.com ISRCTN83830814.

Contains fulltext : 118124.pdf (Publisher’s version ) (Open Access)

Malaria caused an estimated 216 million cases and approximately 1 million deaths in 2010 [1], mainly in sub-Saharan Africa where most cases are caused by Plasmodium falciparum. Development of vaccines and new drugs and better understanding of immunological processes are essential to tackling this immense problem. Controlled human malaria infection (CHMI), in which healthy volunteers are exposed to bites of P. falciparum–infected mosquitoes, is a powerful tool to address questions regarding P. falciparum drug and vaccine efficacy, clinical signs and symptoms, parasite kinetics, and human immunology. Since the first CHMI by mosquitoes fed on cultures of P. falciparum, >1300 healthy volunteers have been exposed to CHMI with mainly the Nijmegen falciparum strain NF54 or its clone 3D7 [2]. Strain/parasite line NF54 stably produces sexual stages required for production of infectious mosquitoes. Parasites have been adapted to laboratory conditions by continuous in vitro culture for >3 decades. In the field, P. falciparum displays a wide genetic diversity, which is currently not represented by the available laboratory strains for CHMI. Other strains, including the South American 7G8 P. falciparum clone of the Brazilian strain IMTM22, have been sporadically used in limited number of volunteers [3–5]. We therefore aimed to identify, clone, and test an additional P. falciparum strain that can be used in CHMIs, and we developed several qualification criteria: The clone (a) must consistently produce gametocytes and sporozoites, (b) should be cloned to create a single genetically homogenous parasite population, (c) should be sensitive to commonly administered antimalarials, and (d) should be of non-African origin to be geographically and genetically distinct from the NF54 strain, an airport strain that probably originates from Africa [6]. Here we report the generation, characterization, and first CHMI for NF135.C10, a new Cambodian clone; findings include drug sensitivity, microsatellite profile, kinetics of parasitemia, and clinical and immunological properties in a direct comparison with NF54.

Although the genus Enterovirus contains many important human pathogens, there is no licensed drug for either the treatment or the prophylaxis of enterovirus infections. We report that fluoxetine (Prozac)—a selective serotonin reuptake inhibitor—inhibits the replication of human enterovirus B (HEV-B) and HEV-D but does not affect the replication of HEV-A and HEV-C or human rhinovirus A or B. We show that fluoxetine interferes with viral RNA replication, and we identified viral protein 2C as the target of this compound.

Item does not contain fulltext BACKGROUND: Molecular typing is a valuable tool for gaining insight into spread of Mycobacterium tuberculosis. Typing allows for clustering of cases whose isolates share an identical genotype, revealing epidemiologic relatedness. Observed distributions of genotypic cluster sizes of tuberculosis (TB) are highly skewed. A possible explanation for this skewness is the concept of "superspreading": a high heterogeneity in the number of secondary cases caused per infectious individual. Superspreading has been previously found for diseases such as severe acute respiratory syndrome and smallpox, where the entire transmission tree is known. So far, no method exists to relate superspreading to the distribution of genotypic cluster sizes. METHODS: We quantified heterogeneity in secondary infections per infectious individual by describing this number as a negative binomial distribution. The dispersion parameter k is a measure of superspreading; standard (homogeneous) models use values of k >/= 1, whereas small values of k imply superspreading. We estimated this negative binomial dispersion parameter for TB in the Netherlands, using the genotypic cluster size distribution for all 8330 cases of culture confirmed, pulmonary TB diagnosed between 1993 and 2007 in the Netherlands. RESULTS: The dispersion parameter k was estimated at 0.10 (95% confidence interval = 0.09-0.12), well in the range of values consistent with superspreading. Simulation studies showed the method reliably estimates the dispersion parameter across a range of scenarios and parameter values. CONCLUSION: Heterogeneity in the number of secondary cases caused per infectious individual is a plausible explanation for the observed skewness in genotypic cluster size distribution of TB. 01 mei 2013

Contains fulltext : 117716.pdf (Publisher’s version ) (Closed access) A key issue in the treatment of disease caused by slow-growing nontuberculous mycobacteria is the limited association between in vitro minimum inhibitory concentrations (MICs) of rifampicin and ethambutol alone and the in vivo outcome of treatment with these drugs. Combined susceptibility testing to rifampicin and ethambutol could provide a more realistic view of the efficacy of these drugs. In this study, Mycobacterium avium (n=5), Mycobacterium chimaera (n=6), Mycobacterium intracellulare (n=4), Mycobacterium xenopi (n=4), Mycobacterium malmoense (n=3) and Mycobacterium simiae (n=2) clinical isolates were selected and the MICs of rifampicin and ethambutol alone and in combination were measured using the Middlebrook 7H10 agar dilution method. Synergy was defined as a fractional inhibitory concentration index

Contains fulltext : 125331.pdf (Publisher’s version ) (Open Access) BACKGROUND: Currently, migration has become one of the risk factors of high burden of tuberculosis in China. This study was to explore the influence of mass migration on the dynamics of Mycobacterium (M.) tuberculosis in Beijing, the capital and an urban area of China. METHODS: Three hundred and thirty-six M. tuberculosis strains from the Changping district, where the problem of urban migrants was more pronounced than in other Beijing regions, were genotyped by Spoligotyping, large sequence polymorphisms (LSPs 105 and 181), and variable number tandem repeat (VNTR) typing. Based on the genotype data, the phylogeny of the isolates was studied. RESULTS: In Changping district, the proportion of Beijing lineage M. tuberculosis isolates amounted to 89.0% (299/336), among which 86.6 % (252) belonged to the modern lineage. The frequency of modern Beijing lineage strains is so high (around 75% (252/336)) that associated risk factors affecting the tuberculosis epidemic cannot be determined. The time to the most recent common ancestor (TMRCA) of the Beijing lineage strains was estimated to be 5073 (95% CI: 4000-6200) years. There was no significant difference in the genetic variation of Beijing isolates from urban migrants and local residents. CONCLUSIONS: The clone of modern Beijing lineage M. tuberculosis, which is dominant in the Beijing area, most likely started to expand with the five thousand-year-old Chinese civilization. In the future, with the urbanization in the whole of China, modern Beijing lineage M. tuberculosis may gain the larger geographical spread. 01 november 2013

Contains fulltext : 117879.pdf (Publisher’s version ) (Open Access) A major challenge for strategies to combat the human malaria parasite Plasmodium vivax is the presence of hypnozoites in the liver. These dormant forms can cause renewed clinical disease after reactivation through unknown mechanisms. The closely related non-human primate malaria P. cynomolgi is a frequently used model for studying hypnozoite-induced relapses. Here we report the generation of the first transgenic P. cynomolgi parasites that stably express fluorescent markers in liver stages by transfection with novel DNA-constructs containing a P. cynomolgi centromere. Analysis of fluorescent liver stages in culture identified, in addition to developing liver-schizonts, uninucleate persisting parasites that were atovaquone resistant but primaquine sensitive, features associated with hypnozoites. We demonstrate that these hypnozoite-forms could be isolated by fluorescence-activated cell sorting. The fluorescently-tagged parasites in combination with FACS-purification open new avenues for a wide range of studies for analysing hypnozoite biology and reactivation.

Contains fulltext : 118774.pdf (Publisher’s version ) (Closed access) BACKGROUND: Artemisinin-based combination therapy (ACT) reduces the potential for malaria transmission, compared with non-ACTs. It is unclear whether this effect differs between ACTs. METHODS: A total of 298 children (age, 6 months to 10 years) with uncomplicated falciparum malaria were randomized to artemether-lumefantrine (AL; n = 153) or dihydroartemisinin-piperaquine (DP; n = 145) in Mbita, a community in western Kenya. Gametocyte carriage was determined by molecular methods on days 0, 1, 2, 3, 7, 14, 28, and 42 after treatment initiation. The gametocyte infectiousness to mosquitoes was determined by mosquito-feeding assays on day 7 after beginning therapy. RESULTS: The cumulative risk of recurrent parasitemia on day 42 after initiation of treatment, unadjusted by polymerase chain reaction findings, was 20.7% (95% confidence interval [CI], 14.4-28.2) for AL, compared with 3.7% (95% CI, 1.2-8.5) for DP (P < .001). The mean duration of gametocyte carriage was 5.5 days (95% CI, 3.6-8.5) for AL and 15.3 days (95% CI, 9.7-24.2) for DP (P = .001). The proportion of mosquitoes that became infected after feeding on blood from AL-treated children was 1.88% (43 of 2293), compared with 3.50% (83 of 2371) for those that fed on blood from DP-treated children (P = .06); the oocyst burden among mosquitoes was lower among those that fed on blood from AL-treated children (P = .005) CONCLUSIONS: While DP was associated with a longer prophylactic time after treatment, gametocyte carriage and malaria transmission to mosquitoes was lower after AL treatment. CLINICAL TRIALS REGISTRATION: NCT00868465.

Item does not contain fulltext So far, the best immunization strategies to achieve high levels of protection against malaria are based on whole parasites. Complete sterile protection can be obtained in rodent models after immunization with sporozoites and chemoprophylaxis, or with sporozoites attenuated either genetically or by radiation. These approaches target specific stages, with arrests occurring at different time-points of the parasite life cycle. Here, we review these different approaches in relation to their capacity to induce protection in both Plasmodium berghei and Plasmodium yoelii models. The combined data suggest that maximal liver-stage exposure without further development into blood stages may induce the most efficient protection in mice.

Contains fulltext : 126147.pdf (Publisher’s version ) (Open Access)

Contains fulltext : 125669.pdf (Publisher’s version ) (Open Access)

Item does not contain fulltext Data are scarce on demographical factors related to the population structure of Mycobacterium tuberculosis in Saudi Arabia. A study was conducted on 902 clinical isolates to explore current trends in the phylogeography and associated demographical factors of tuberculosis by using spoligotyping and 24 loci based MIRU-VNTR typing. Young male patients (aged 16-29 and 30-44) were predominant in this cohort. The phylogenetic diversity among M. tuberculosis isolates was found high, as almost all known genetic lineages were identified. Delhi/CAS (26.4%), EAI (13.7%) and Haarlem (11.3%) were the most common lineages observed, particularly among the low age groups (16-29 and 30-44 years), whereas elderly patients (>60 years) showed a predominance in the lineages S, Ghana, TUR and Uganda-I. A statistically significant association was observed between gender of the patients and lineages of EAI (p value 0.026) and LAM (p value 0.005). Overall, molecular strain cluster rate was 34.4% with an elevated rate among patients aged below 15 years (43.1%), while cases among the elderly (>60 years) showed the lowest degree of clustering (12.5%). The largest level of clustering was noticed among cases caused by strains of the lineages Haarlem (59.8%), Beijing (55.8%) and LAM (42.8%). The current population structure of M. tuberculosis in Saudi Arabia is highly diverse with significant associations to demography, transmission dynamics and origin of the patients. The difference in genotype distributions among low and high aged patients reflects the ongoing change in the strain population structure in the country. 01 juni 2013

Previous evaluations of the molecular GenoType tests have promoted their use to detect resistance to first- and second-line antituberculosis drugs in different geographical regions. However, there are known geographic variations in the mutations associated with drug resistance in Mycobacterium tuberculosis , and especially in South America, there is a paucity of information regarding the frequencies and types of mutations associated with resistance to first- and second-line antituberculosis drugs. We therefore evaluated the performance of the GenoType kits in this region by testing 228 M. tuberculosis isolates in Colombia, including 134 resistant and 94 pansusceptible strains. Overall, the sensitivity and specificity of the GenoType MTBDR plus test ranged from 92 to 96% and 97 to 100%, respectively; the agreement index was optimal (Cohen's kappa, >0.8). The sensitivity of the GenoType MTBDR sl test ranged from 84 to 100% and the specificity from 88 to 100%. The most common mutations were katG S315T1, rpoB S531L, embB M306V, gyrA D94G, and rrs A1401G. Our results reflect the utility of the GenoType tests in Colombia; however, as some discordance still exists between the conventional and molecular approaches in resistance testing, we adhere to the recommendation that the GenoType tests serve as early guides for therapy, followed by phenotypic drug susceptibility testing for all cases.

Contains fulltext : 118709.pdf (Publisher’s version ) (Open Access) BACKGROUND: The heterogeneity of malaria transmission makes widespread elimination a difficult goal to achieve. Most of the current vector control measures insufficiently target outdoor transmission. Also, insecticide resistance threatens to diminish the efficacy of the most prevalent measures, indoor residual spray and insecticide treated nets. Innovative approaches are needed. The use of endectocides, such as ivermectin, could be an important new addition to the toolbox of anti-malarial measures. Ivermectin effectively targets outdoor transmission, has a novel mechanism of action that could circumvent resistance and might be distributed over the channels already in place for the control of onchocerciasis and lymphatic filariasis. METHODS: The previous works involving ivermectin and Anopheles vectors are reviewed and summarized. A review of ivermectin's safety profile is also provided. Finally three definitive clinical trials are described in detail and proposed as the evidence needed for implementation. Several smaller and specific supportive studies are also proposed. CONCLUSIONS: The use of ivermectin solves many challenges identified for future vector control strategies. It is an effective and safe endectocide that was approved for human use more than 25 years ago. Recent studies suggest it might become an effective and complementary strategy in malaria elimination and eradication efforts; however, intensive research will be needed to make this a reality.

Background Informing and evaluating malaria control efforts relies on knowledge of local transmission dynamics. Serological and molecular tools have demonstrated great sensitivity to quantify transmission intensity in low endemic settings where the sensitivity of traditional methods is limited. Filter paper blood spots are commonly used a source of both DNA and antibodies. To enhance the operational practicability of malaria surveys, a method is presented for combined DNA extraction and antibody elution. Methods Filter paper blood spots were collected as part of a large cross-sectional survey in the Kenyan highlands. DNA was extracted using a saponin/chelex method. The eluate of the first wash during the DNA extraction process was used for antibody detection and compared with previously validated antibody elution procedures. Antibody elution efficiency was assessed by total IgG ELISA for malaria antigens apical membrane antigen-1 (AMA-1) and merozoite-surface protein-1 (MSP-142). The sensitivity of nested 18S rRNA and cytochrome b PCR assays and the impact of doubling filter paper material for PCR sensitivity were determined. The distribution of cell material and antibodies throughout filter paper blood spots were examined using luminescent and fluorescent reporter assays. Results Antibody levels measured after the combined antibody/DNA extraction technique were strongly correlated to those measured after standard antibody elution (p 42 were generally slightly lower (11.3-21.4%) but age-seroprevalence patterns were indistinguishable. The proportion of parasite positive samples ranged from 12.9% to 19.2% in the different PCR assays. Despite strong agreement between outcomes of different PCR assays, none of the assays detected all parasite-positive individuals. For all assays doubling filter paper material for DNA extraction increased sensitivity. The concentration of cell and antibody material was not homogenously distributed throughout blood spots. Conclusion Combined DNA extraction and antibody elution is an operationally attractive approach for high throughput assessment of cumulative malaria exposure and current infection prevalence in endemic settings. Estimates of antibody prevalence are unaffected by the combined extraction and elution procedure. The choice of target gene and the amount and source of filter paper material for DNA extraction can have a marked impact on PCR sensitivity.

Organosilicon compounds are efflux pump inhibitors with potency as an antituberculosis drug. Of the organisilicon compounds tested, SILA 421 has been shown to have a highest potency as an antituberculosis drug (1). It shares the common pathways for antimycobacterial killing with other efflux pump inhibitors: it revealed direct in vitro activity against M. tuberculosis (1), it has been shown to modify resistance by inhibiting mdr-1 efflux pumps and has shown to enhance killing of M. tuberculosis by macrophages (1).

Malaria Cloak and Dagger Mosquitoes have a complex immune system capable of effective antiparasite responses; however, malaria transmission is still highly efficient. Molina-Cruz et al. (p. 984 , published online 9 May; see the Perspective by Philip and Waters ) show that Plasmodium falciparum has a gene product, Pfs47 , that makes the parasite's ookinetes “invisible” to the mosquito immune system. Disruption of this mechanism could potentially be used to block malaria transmission.

Contains fulltext : 112643.pdf (Publisher’s version ) (Open Access) Radboud Universiteit Nijmegen, 10 juni 2013 Promotor : Sauerwein, R.W. Co-promotor : Janse, C.J.