Estimating the burden of disease attributable to unsafe water and lack of sanitation and hygiene in South Africa in 2000

Objectives To estimate the burden of disease attributable to unsafe water, sanitation and hygiene (WSH) by age group for South Africa in 2000. Design World Health Organization comparative risk assessment methodology was used to estimate the disease burden attributable to an exposure by comparing the observed risk factor distribution with a theoretical lowest possible population distribution. A scenario-based approach was applied for estimating diarrhoeal disease burden from unsafe WSH. Six exposure scenarios were defined based on the type of water and sanitation infrastructure and environmental faecal-oral pathogen load. For ‘intestinal parasites’ and schistosomiasis, the burden was assumed to be 100% attributable to exposure to unsafe WSH. Setting South Africa. Outcome measures Disease burden from diarrhoeal diseases, intestinal parasites and schistosomiasis, measured by deaths and disability-adjusted life years (DALYs). Results 13 434 deaths were attributable to unsafe WSH accounting for 2.6% (95% uncertainty interval 2.4 - 2.7%) of all deaths in South Africa in 2000. The burden was especially high in children under 5 years, accounting for 9.3% of total deaths in this age group and 7.4% of burden of disease. Overall, the burden due to unsafe WSH was equivalent to 2.6% (95% uncertainty interval 2.5 - 2.7%) of the total disease burden for South Africa, ranking this risk factor seventh for the country. Conclusions Unsafe WSH remains an important risk factor for disease in South Africa, especially in children under 5. High priority needs to be given to the provision of safe and sustainable sanitation and water facilities and to promoting safe hygiene behaviours, particularly among children.

Health education and the control of intestinal worm infections in China : a new vision

Background: The transmission of soil-transmitted helminths (STHs) is associated with poverty, poor hygiene behaviour, lack of clean water and inadequate waste disposal and sanitation. Periodic administration of benzimidazole drugs is the mainstay for global STH control but it does not prevent re-infection, and is unlikely to interrupt transmission as a stand-alone intervention. Findings: We reported recently on the development and successful testing in Hunan province, PR China, of a health education package to prevent STH infections in Han Chinese primary school students. We have recently commenced a new trial of the package in the ethnically diverse Xishuangbanna autonomous prefecture in Yunnan province and the approach is also being tested in West Africa, with further expansion into the Philippines in 2015. Conclusions: The work in China illustrates well the direct impact that health education can have in improving knowledge and awareness, and in changing hygiene behaviour. Further, it can provide insight into the public health outcomes of a multi-component integrated control program, where health education prevents re-infection and periodic drug treatment reduces prevalence and morbidity.

The p38 mitogen-activated protein kinase regulates interleukin-1beta-induced IL-8 expression via an effect on the IL-8 promoter in intestinal epithelial cells

Several lines of evidence implicate the p38 mitogen-activated protein kinase (p38 MAPK) in the proinflammatory response to bacterial agents and cytokines. Equally, the transcription factor, nuclear factor (NF)-kappaB, is recognized to be a critical determinant of the inflammatory response in intestinal epithelial cells (IECs). However, the precise inter-relationship between the activation of p38 MAPK and activation of the transcription factor NF-kappaB in the intestinal epithelial cell (IEC) system, remains unknown. Here we show that interleukin (IL)-1beta activates all three MAPKs in Caco-2 cells. The production of IL-8 and monocyte chemotactic protein 1 (MCP-1) was attenuated by 50% when these cells were preincubated with the p38 MAPK inhibitor, SB 203580. Further investigation of the NF-kappaB signalling system revealed that the inhibitory effect was independent of the phosphorylation and degradation of IkappaBalpha, the binding partner of NF-kappaB. This effect was also independent of the DNA binding of the p65 Rel A subunit, as well as transactivation, determined by an NF-kappaB luciferase construct, using both SB 203580 and dominant-negative p38 MAPK. Evaluation of IL-8 and MCP-1 RNA messages by reverse transcription-polymerase chain reaction (RT-PCR) revealed that the inhibitory effect of SB 203580 was associated with a reduction in this parameter. Using an IL-8-luciferase promoter construct, an effect of p38 upon its activation by both pharmacological and dominant-negative p38 construct co-transfection was demonstrated. It is concluded that p38 MAPK influences the expression of chemokines in intestinal epithelial cells, through an effect upon the activation of the chemokine promoter, and does not directly involve the activation of the transcription factor NF-kappaB

Cytokine synthesis by intestinal intraepithelial lymphocytes. Both gamma/delta T cell receptor-positive and alpha/beta T cell receptor-positive T cells in the G1 phase of cell cycle produce IFN-gamma and IL-5

Murine intestinal intraepithelial lymphocytes (IEL) have been shown to contain subsets of alpha/beta TCR+ and gamma/delta TCR+ T cells that spontaneously produce cytokines such as IFN-gamma and IL-5. We have now determined the nature and cell cycle stage of these cytokine-producing T lymphocytes in EIL by using IFN-gamma- and IL-5-specific ELISPOT assay, cytokine-specific mRNA-cDNA dot-blot hybridization and polymerase chain reaction, and flow cytometry (FACS) for DNA analysis. When CD3+ T cells from IEL of normal C3H/HeN mice were separated into low and high density fractions by discontinuous Percoll gradients, IFN-gamma and IL-5 spot-forming cells were only found in the former population. Analysis of mRNA for these cytokines by both IFN-gamma- and IL-5-specific dot-blot hybridization and polymerase chain reaction revealed that higher levels of message for IFN-gamma and IL-5 were also seen in the low density fraction. However, cell cycle analysis of these two fractions by FACS using propidium iodide showed a similar pattern of cell cycle stages in both low and high density populations (G0 + G1 approximately 96 to 98% and S/G2 + M approximately 2 to 4%). Finally, mRNA from gamma/delta TCR+ and alpha/beta TCR+ T cells in both low and high density fractions of IEL were analyzed for IFN-gamma and IL-5 message by polymerase chain reaction. After 35 cycles of amplification, both gamma/delta TCR+ and alpha/beta TCR+ T cells in the low density fraction expressed higher levels of message for these two cytokines when compared with the high density population. These results have now shown that both gamma/delta and alpha/beta TCR+ IEL can be separated into low and high density subsets and both fractions possess a similar stage of cell cycle. However, only the low density cells (in G1 phase) of both gamma/delta and alpha/beta TCR types possess increased cytokine-specific mRNA and produce the cytokines IFN-gamma and IL-5. Our results suggest that alpha/beta TCR+ and gamma/delta TCR+ IEL can produce cytokines without cell proliferation.

Hepatocyte growth factor stimulates invasion across reconstituted basement membranes by a new human small intestinal cell line

Hepatocyte growth factor/scatter factor (HGF/SF) is a protein growth factor whose pleiotropic effects on epithelial cells include the stimulation of motility, mitosis and tubulogenesis. These responses are mediated by the cell surface tyrosine kinase receptor c-met. Because both the cytokine and receptor are found in the gastrointestinal tract, we have studied the effects of HGF/SF on transformed gut epithelial cells which express c-met. Here we describe the response of a new transformed human jejunal epithelioid cell line (HIE-7) to HGF/SF. Morphologically HIE-7 cells are immature. Their epithelial lineage was confirmed by reactivity with the epithelial specific antibodies AE1/AE3, Cam 5.2, Ber-EP4 and anti-EMA and is consistent with their expression of c-met mRNA and protein. In addition, electron microscopic analysis revealed the presence of primitive junctions and rudimentary microvilli, but features of polarization were absent. When grown on reconstituted basement membranes, HIE-7 cells formed closely associated multicellular cord-like structures adjacent to acellular spaces. However, the cells did not mature structurally, form lumen-like structures or express disaccharidase mRNA, even in the presence of recombinant HGF (rHGF). On the other hand, rHGF induced HIE-7 cells to scatter and stimulated their rapid migration in a modified wound assay. To determine whether the motogenic effect caused by rHGF is associated with HIE-7 cell invasiveness across reconstituted basement membranes, a Boyden chamber chemoinvasion assay was performed. rHGF stimulated a 10-fold increase in the number of HIE-7 cells that crossed the basement membrane barrier, while only stimulating a small increase in chemotaxis across a collagen IV matrix, suggesting that the cytokine activates matrix penetration by these cells. rHGF also stimulated the invasion of basement membranes by an undifferentiated rat intestinal cell line (IEC-6) and by two human colon cancer cell lines which are poorly differentiated (DLD-1 and SW 948). In contrast, two moderately well differentiated colon cancer cell lines (Caco-2 and HT-29) did not manifest an invasive response when exposed to rHGF. These results suggest that HGF/SF may play a significant role in the invasive behavior of anaplastic and poorly differentiated gut epithelial tumors.

Plasmodium falciparum parasites lacking histidine-rich protein 2 and 3 : a review and recommendations for accurate reporting

Malaria rapid diagnostic tests (RDTs) play a critical role in malaria case management, surveillance and case investigations. Test performance is largely determined by design and quality characteristics, such as detection sensitivity, specificity, and thermal stability. However, parasite characteristics such as variable or absent expression of antigens targeted by RDTs can also affect RDT performance. Plasmodium falciparum parasites lacking the PfHRP2 protein, the most common target antigen for detection of P. falciparum, have been reported in some regions. Therefore, accurately mapping the presence and prevalence of P. falciparum parasites lacking pfhrp2 would be an important step so that RDTs targeting alternative antigens, or microscopy, can be preferentially selected for use in such regions. Herein the available evidence and molecular basis for identifying malaria parasites lacking PfHRP2 is reviewed, and a set of recommended procedures to apply for future investigations for parasites lacking PfHRP2, is proposed.

An improved method for undertaking limiting dilution assays for in vitro cloning of Plasmodium falciparum parasites

Background Obtaining single parasite clones is required for many techniques in malaria research. Cloning by limiting dilution using microscopy-based assessment for parasite growth is an arduous and labor-intensive process. An alternative method for the detection of parasite growth in limiting dilution assays is using a commercial ELISA histidine-rich protein II (HRP2) detection kit. Methods Detection of parasite growth was undertaken using HRP2 ELISA and compared to thick film microscopy. An HRP2 protein standard was used to determine the detection threshold of the HRP2 ELISA assay, and a HRP2 release model was used to extrapolate the amount of parasite growth required for a positive result. Results The HRP2 ELISA was more sensitive than microscopy for detecting parasite growth. The minimum level of HRP2 protein detection of the ELISA was 0.11ng/ml. Modeling of HRP2 release determined that 2,116 parasites are required to complete a full erythrocytic cycle to produce sufficient HRP2 to be detected by the ELISA. Under standard culture conditions this number of parasites is likely to be reached between 8 to 14 days of culture. Conclusions This method provides an accurate and simple way for the detection of parasite growth in limiting dilution assays, reducing time and resources required in traditional methods. Furthermore the method uses spent culture media instead of the parasite-infected red blood cells, enabling culture to continue.

Slaving and release in co-infection control

Background Animal and human infection with multiple parasite species is the norm rather than the exception, and empirical studies and animal models have provided evidence for a diverse range of interactions among parasites. We demonstrate how an optimal control strategy should be tailored to the pathogen community and tempered by species-level knowledge of drug sensitivity with use of a simple epidemiological model of gastro-intestinal nematodes. Methods We construct a fully mechanistic model of macroparasite co-infection and use it to explore a range of control scenarios involving chemotherapy as well as improvements to sanitation. Results Scenarios are presented whereby control not only releases a more resistant parasite from antagonistic interactions, but risks increasing co-infection rates, exacerbating the burden of disease. In contrast, synergisms between species result in their becoming epidemiologically slaved within hosts, presenting a novel opportunity for controlling drug resistant parasites by targeting co-circulating species. Conclusions Understanding the effects on control of multi-parasite species interactions, and vice versa, is of increasing urgency in the advent of integrated mass intervention programmes.

Exploring diarrhoea, enteral nutrition and intestinal microbial flora relationships in critically ill patients

Diarrhoea is a common complication observed in critically ill patients. Relationships between diarrhoea, enteral nutrition and aerobic intestinal microflora have been disconnectedly examined in this patient cohort. This research used a two-study, observational design to examine these associations. Higher diarrhoea incidence rates were observed when patients received enteral tube feeding, had abnormal serum blood results, received multiple medications and had aerobic microflora dysbiosis. Further, significant aerobic intestinal microflora changes were observed over time in patients who experienced diarrhoea. These results establish a platform for further work to improve the intestinal health of critically ill patients.

Mitochondrial membrane potential in a small subset of artemisinin-induced dormant plasmodium falciparum parasites in vitro

Artemisinin induced dormancy is a proposed mechanism for failures of mono-therapy and is linked with artemisinin resistance in Plasmodium falciparum. The biological characterization and dynamics of dormant parasites are not well understood. Here we report that following dihydroartemisinin (DHA) treatment in vitro, a small subset of morphologically dormant parasites was stained with rhodamine 123 (RH), a mitochondrial membrane potential (MMP) marker, and persisted to recovery. FACS sorted RH-positive parasites resumed growth at 10,000/well while RH-negative parasites failed to recover at 5 million/well. Furthermore, transcriptional activity for mitochondrial enzymes was only detected in RH-positive dormant parasites. Importantly, after treating dormant parasites with different concentrations of atovaquone, a mitochondrial inhibitor, the recovery of dormant parasites was delayed or stopped. This demonstrates that mitochondrial activity is critical for survival and regrowth of dormant parasites and that RH staining provides a means of identifying these parasites. These findings provide novel paths for studying and eradicating this dormant stage.

Combination chemotherapy for primary small intestinal lymphoma in the Middle East

Twelve patients with primary small intestinal lymphoma were followed prospectively for 3 years. Endoscopic abnormalities were diagnostic of lymphoma in all cases where the duodenum was involved (83%). In three cases (25%) the disease extended to the stomach. One patient (8%) had diffuse small cell cleaved and 11 (92%) diffuse large cell lymphoma stages I (8%), II (25%), III (58%) and IV (8%). Nine of them were unresectable and primarily treated with combination chemotherapy; 67% achieved complete remission, 22% partial response and 11% no response. Only one patient relapsed and achieved a second remission. All complete remission patients are currently alive and free of disease at a median follow-up of 36 months. Overall survival for all patients is 58%, and disease-free survival is 50%. No instance of chemotherapy-related bleeding or perforation was seen. Tetracycline was necessary for the treatment of IPSID-associated diarrhea and malabsorption in spite of cytotoxic chemotherapy.

Microscopy for the detection, identification and quantification of malaria parasites on stained thick and thin blood films in research settings

Summary This manual was developed to guide a move towards common standards for undertaking and reporting research microscopy for malaria parasite detection, identification and quantification. It contains procedures based on agreed quality assurance standards for research malaria microscopy defined at a consultation of: TDR, the Special Programme for Research and Training in Tropical Diseases; the Worldwide Antimalarial Resistance Network (WWARN), United Kingdom; the Foundation for Innovative New Diagnostics (FIND), Switzerland; the Centers for Disease Control and Prevention (CDC), USA; the Kenya Medical Research Institute (KEMRI) and later expanded to include Amref Health Africa (Kenya); the Eijkman-Oxford Clinical Research Unit (EOCRU), Indonesia; Institut Pasteur du Cambodge (IPC); Institut de recherche pour le Développement (IRD), Senegal; the Global Good and Intellectual Ventures Laboratory (GG-IVL), USA; the Mahidol-Oxford Tropical Medicine Research Unit (MORU), Thailand; Queensland University of Technology (QUT), Australia, and the Shoklo Malaria Research Unit (SMRU), Thailand. These collaborating institutions commit to adhering to these standards in published research studies. It is hoped that they will form a solid basis for the wider adoption of standardized reference microscopy protocols for malaria research.

Understanding the function and mechanisms of intestinal cell kinase in the growth and survival of prostate cancer cells

This project was a step forward in discovering the potential role of intestinal cell kinase in prostate cancer development. Intestinal cell kinase was shown to be upregulated in prostate cancer cells and altered expression led to changes in key cell survival proteins. This study used in vitro experiments to monitor changes in cell growth, protein and RNA expression.