Control of Schistosoma mekongi in Cambodia: results of eight years of control activities in the two endemic provinces.

In Cambodia, schistosomiasis is transmitted in the provinces of Kratie and Stung Treng where approximately 80000 individuals are estimated to be at risk of infection. The baseline prevalence of infection was estimated to be between 73% and 88%, and cases of severe morbidity (hepatosplenomegaly, puberty retardation) and mortality were very common. In 1994, the Ministry of Health of Cambodia started schistosomiasis control applying universal chemotherapy with praziquantel (40mg/kg). The coverage of the programme was between 62% and 86% for 8 years. This simple control measure resulted in the control of the disease: no cases were reported in 2004 and only three cases were reported in 2005. In addition, there are no longer reports of cases of severe morbidity due to schistosomiasis. Since the beginning of the control programme, a single dose of mebendazole (500mg) has been combined with praziquantel during the mass chemotherapy; as a result the prevalence of Ascaris lumbricoides and hookworms dropped from 74.5% to 10% and from 86% to 40% respectively. The experience in Cambodia demonstrates that, with political commitment, control of parasitic diseases is achievable even in a situation of minimal resources. The programme represents a successful model for other developing countries.

Counterfeit and substandard antimalarial drugs in Cambodia.

Counterfeit and substandard antimalarial drugs can cause death and contribute to the growing malaria drug resistance problem, particularly in Southeast Asia. Since 2003 in Cambodia the quality of antimalarial drugs both in the public and private health sector is regularly monitored in sentinel sites. We surveyed 34% of all 498 known facilities and drug outlets in four provinces. We collected 451 drug samples; 79% of these were not registered at the Cambodia Department of Drugs and Food (DDF). Twenty-seven percent of the samples failed the thin layer chromatography and disintegration tests; all of them were unregistered products. Immediate action against counterfeit drugs was taken by the National Malaria Control Program (NMCP) and the DDF. They communicated with the Provincial Health Department about the presence of counterfeit antimalarial drugs through alert letters, a manual, annual malaria conferencing and other training occasions. Television campaigns to alert the population about counterfeit drugs were conducted. Moreover, the NMCP has been promoting the use of good quality antimalarial drugs of a blister co-packaged combination of artesunate and mefloquine in public and private sectors. Appropriate strategies need to be developed and implemented by relevant government agencies and stakeholders to strengthen drug quality assurance and control systems in the country.

Investigation of bioavailability, pharmacokinetics and safety of new pediatric formulations of artesunate and mefloquine.

The bioavailability/pharmacokinetics of dihydroartemisinin and mefloquine following the oral doses of 4 mg/kg body weight artesunate (Cambodian Pharmaceutical Enterprise) given concurrently with 10 mg/kg body weight oral mefloquine artesunate (Cambodian Pharmaceutical Enterprise) were investigated in 15 healthy Cambodian male volunteers. Both formulations were generally well tolerated. Both produced satisfactory plasma/blood concentration-time profiles. Oral artesunate and mefloquine were rapidly absorbed from gastrointestinal tract with marked inter-individual variation. For the dihydroartemisinin, the median (95% Cl) Cmax of 748 (304-1,470) ng/ml was observed at 1.5 (0.3-3.0) hours (tmax) after drug administration. The median (95% CI) values for AUC0-infinity, lambda(z) and tl/2z were 1.673 (1.08-2.88) microg.h/ml, 0.54(0.24-1.1)/hour and 1.3 (0.6-2.9) hours, respectively. For mefloquine, a median (95% Cl) Cmax of 1,000 (591-1,500) ng/ml was observed at 4 (2-6) hours (tmax) after drug administration. The median (95% CI) value for AUC0-168h was 3.92 (2.88-7.02) microg.h/ml.

Positive control wells: a potential answer to remote-area quality assurance of malaria rapid diagnostic tests.

Malaria rapid diagnostic tests (RDTs) are a potential breakthrough in the provision of accurate diagnosis in remote areas, but widescale use is hampered by uncertainty over accuracy under field conditions. Positive control wells, which contain recombinant malaria parasite antigen, are a novel method for addressing this need for quality assurance. The potential of a commercially available positive control well, reconstituted with blood, was assessed for use in routine monitoring of RDT sensitivity in a remote malaria-endemic region. When maintained at 4 degrees C, the wells produced a consistent level of parasite lactate dehydrogenase (pLDH) antigen activity, as detected by pLDH-detecting RDTs, but activity reduced after cumulative exposure to temperatures likely to be encountered over a few months in a malaria-endemic area. This limitation was successfully overcome in the field through centralized, controlled storage. Monitoring of RDT sensitivity was successfully incorporated into routine supervisory visits to remote clinics. However, improved temperature stability of the wells would enhance their potential. The threshold at which the wells' signal reduced RDT sensitivity requires further investigation. The wells show potential to overcome an important obstacle to the wide implementation of accurate parasite-based diagnosis and appropriate treatment. Further assessment of their place in malaria management is warranted.

The acceptability of insecticide treated mosquito nets among community members in Zimbabwe.

OBJECTIVES: To assess the acceptability of insecticide treated mosquito nets (ITNs) among community members in Zimbabwe for the implementation of effective malaria intervention using ITNs. DESIGN: A cross sectional study and longitudinal study were carried out using questionnaires, which were administered by health workers from rural health centres. SETTING: Communities in rural areas from seven districts namely Bulilimamangwe, Chipinge, Gokwe, Hurungwe, Lupane, Mount Darwin and Uzumba Maramba Pfungwe (UMP) in Zimbabwe. SUBJECTS: 1,576 community members who bought ITNs in the project areas. MAIN OUTCOME MEASURE: Acceptance of ITNs and price, reasons for wanting or not wanting nets; ITNs usage and washing pattern among respondents. RESULTS: More than 90% of people said they bought the nets from the project in order to avoid mosquito bites and to prevent malaria. Quite a number of respondents (69%) gave the answer that they bought mosquito nets from the project because they were cheaper (Z$130 to Z$390) than the market price (Z$1,000) and more than half (58.3%) claimed that they could not afford to buy mosquito nets at the market price. The price was accepted by the majority as cheap or very cheap in Gokwe, Hurungwe, Mount Darwin, and UMP (92%, 82%, 98% and 90%, respectively). Other responses given for having bought nets wereat health staff's recommendation (71.3%), and they were suffering from malaria (40.9%). The reasons for not having bought mosquito nets were that they had not suffered from malaria (55.5%), that they did not like to use a mosquito net (31.1%), did not know the efficacy of the net (21.6%), or there were not many mosquitoes at home (28.9%). The percentage of ITN use was very high (90 to 100%) amongst those who bought nets. The percentage of children under five years and pregnant women who were ITN users ranged from six to 24% to 2.8 to 9.7%, respectively. Percentage of "others" which means those above five years and not pregnant was more than 70% in all the districts. The washing pattern of ITNs in Chipinge showed that most of the mosquito nets (74.5%) were washed every three to five months. CONCLUSION: Acceptability of ITNs was very high based on reported utilisation of ITNs. It is recommended that rural communities should be sensitised on the importance of treated nets for malaria prevention, and advised to purchase mosquito nets whenever cash is readily available in each community.

The knowledge and practice on malaria among community members in Zimbabwe.

OBJECTIVES: To assess the knowledge of the cause, preventive measures and symptoms of malaria, treatment seeking behaviour on malaria and source of information about malaria among community members for the planning of effective malaria intervention. DESIGN: A cross sectional study was carried out using a questionnaire, which was administered by health workers from rural health centres. SETTING: Households in rural areas from five districts namely Uzumba Maramba Pfungwe (UMP), Hurungwe, Mount Darwin, Bulilimamangwe and Chipinge in Zimbabwe. SUBJECTS: 2,531 community members who were in charge of households sampled systematically in the area. MAIN OUTCOME MEASURES: Percentage of knowledge of transmission, signs and symptoms, prevention methods such as residual house spraying and personal protection, and treatment seeking practice on malaria. RESULTS: Knowledge of malaria transmission was generally high ranging from 57.9% to 95.9%, although there were some misconceptions such as drinking dirty water and poor hygiene. Knowledge of malaria preventive methods was varied among districts ranging from 19.4% to 93.0% about residual house spraying, and low (0.2% to 22.7%), (8.1% to 24.5%) about mosquito nets and repellents, respectively. Knowledge of malaria signs was generally high although this also varied among districts. Treatment seeking practice on malaria shows that the majority of respondents (85.4%) visit the clinic first when seeking treatment although there are other options such as shops and traditional healers. Other sources of information about malaria mentioned were, health workers (72.9%), radio (44.0%), television (37.5%), teachers (25.0%) and village community workers (10.0%). CONCLUSION: Despite widespread knowledge about the cause of malaria and symptoms, and the fairly extensive sources of health information, understanding of the preventive measures was relatively low. Appropriate health education is essential to stimulate changes in both knowledge and behaviour in malaria control activities. Schools can be effective health education tools as well as health facilities.

Anemia and intestinal parasitic infections in primary school students in Aracaju, Sergipe, Brazil.

Anemia is estimated to affect half the school-age children and adolescents in developing countries. The main causes are parasitic infections, malaria, and low iron intake. This study aimed to describe the prevalence of anemia, parasitic infections, and nutritional status of children attending public primary schools in Aracaju, Northeast Brazil. Of 360 students, 26.7% were anemic, and prevalence was higher in children under 8 and over 15 years of age. Overall prevalence of intestinal parasites was 42%, with Ascaris lumbricoides (28.7%), Trichuris trichiura (15.6%), and hookworm (1. 7%) most frequently found. There was an association between parasitic infections and poor sanitary conditions, but there was no association between anemia and presence of intestinal parasites. Height-for-age Z scores were lower than the NCHS standard, and prevalence of stunting was 5.4%. Although intestinal parasites were not associated with anemia, children with parasites had lower nutritional indices (weight- and height-for-age Z scores) than those without parasites.

1,25-dihydroxyvitamin D3 differentiates normal neutrophilic promyelocytes to monocytes/macrophages in vitro.

Although it is well established that the addition of 1,25-dihydroxyvitamin D3 (D3) to the culture of normal human granulocyte/macrophage progenitors induces monocyte/macrophage (Mo/M phi) colonies, the target cells of D3 in the Mo/M phi differentiation have not been identified. We examined whether neutrophilic promyelocytes are the target cells. As a source of the promyelocyte fraction, we used colonies after 5 days of culture (5-day colonies) of colony-forming unit-granulocyte. The culture contained granulocyte colony-stimulating factor (G-CSF) as the growth factor and generated only neutrophilic colonies. The promyelocytic nature of the 5-day colonies was confirmed morphologically, cytochemically, and ultrastructurally. After morphological evaluation on part of the individual colonies, they were transferred into new semisolid cultures with or without D3 (10(-7) mol/L) in the presence of G-CSF, then incubated for the subsequent 7 days. With D3, the colonies were loose, and all the constituent cells were morphologically small macrophages, which were positive for alpha-naphthyl butyrate (alpha NB) esterase, strongly positive for CD14 antigen, and plastic-adherent. While without D3, the colonies were rather compact, and all the constituent cells were morphologically mature neutrophils, which were positive for naphthol ASD-chloroacetate esterase and weakly positive for CD14 antigen. Secondary culture of the 8- or 10-day colonies with D3 induced a lower number of alpha NB-positive cells, in proportion to the percentage of promyelocytes at the time of transfer in each colony. Four days of secondary culture with D3 was sufficient to induce alpha NB-positive cells. G-CSF was not an essential factor to induce alpha NB-positive cells. These findings indicate that D3 differentiates normal human neutrophilic promyelocytes into the Mo/M phi lineage in vitro.

Intestinal perforation in temporal arteritis, associated with paroxysmal nocturnal hemoglobinuria.

Temporal arteritis (TA) is an adult-onset, focal granulomatous inflammatory disorder of the small and medium sized arteries. Intestinal perforation is a rare complication of TA. Regarding its etiology, steroid-induced or arteritis-induced ulceration have been proposed. We describe a patient who developed TA in addition to preceding paroxysmal nocturnal hemoglobinuria. During steroid therapy for TA, intestinal perforation manifested, and it was proven to be arteritis induced perforation on histological examinations. The patient may be the 5th reported case of TA complicated with arteritis-induced intestinal perforation. The possibility of polyangitis overlap syndrome of TA and polyarteritis nodosa is discussed.

Long term complete remission in primary plasma cell leukemia.

A 76-year-old female was diagnosed as having primary plasma cell leukemia (PCL), based on abundant atypical plasma cells in the circulation and bone marrow, monoclonal kappa light chain in the serum and urine, the immunophenotype of the plasma cells and the lack of preceding multiple myeloma. The patient was treated with melphalan and prednisolone (MP), and complete remission (CR) was achieved; this was maintained for 28 months. The duration of CR in this patient appears to be the longest of those reported in well-documented primary PCL patients who have been treated with MP chemotherapy.

A newly established megakaryoblastic/erythroid cell line that differentiates to red cells in the presence of erythropoietin and produces platelet-like particles.

In August, 1992, we established a leukemic cell line (NS-Meg) from a patient in megakaryoblastic transformation of Philadelphia chromosome-positive chronic myeloid leukemia. The NS-Meg cells were positive for alpha-naphthyl acetate esterase and periodic acid-Schiff (PAS) staining and for surface CD4, CD7, CD13, CD34, CD41a, and glycophorin A antigens. Ultrastructurally, the cells had alpha-granules, demarcation membranes, and platelet peroxidase activity. The NS-Meg cells spontaneously produced platelet-like particles which contained alpha-granules, mitochondria and dense bodies, strongly suggesting platelet production. Erythropoietin (Epo), granulocyte/macrophage colony stimulating factor(GM-CSF), and interleukin 3 (IL-3) promoted the growth of NS-Meg cells. Phorbol-12-myristate-13-acetate increased the expression of both CD41a and CD61 antigens. Ten-day exposure to Epo induced mature erythroblasts and red cells. These benzidine-positive cells were positive for hemoglobin F staining. Untreated NS-Meg cells expressed mRNA for the Epo receptor (EpoR), for GATA-1, and for alpha 1, alpha 2 and gamma globin genes. These results indicate that NS-Meg cells undergo terminal differentiation of both megakaryocytic and erythroid lineages. This cell line should be a very useful tool for the investigation of both megakaryocytic and erythroid maturation.

Colony-stimulating factor-producing tumours: production of granulocyte colony-stimulating factor and interleukin-6 is secondary to interleukin-1 production.

We have reported that tumours producing colony-stimulating factor (CSF) also secrete interleukin-1 (IL-1) and IL-6. In the present study, we characterised the role played by IL-1 in CSF production. IL-1 receptor antagonist inhibited the production of granulocyte (G)-CSF and IL-6 by approximately 90% in CSF-producing human lung carcinoma cell lines. Similar results were obtained with hydrocortisone, which suppresses IL-1 gene expression. In contrast, 15 non-CSF-producing human lung carcinoma cell lines did not show detectable IL-1 production, although seven of them were induced to produce G-CSF and IL-6 by exogenous IL-1 alpha. Cell lines that responded to IL-1 alpha, including the CSF-producing lines, expressed receptors for IL-1 alpha. These results indicate that CSF-producing tumours can be characterised by their constitutive IL-1 production, IL-1 receptor expression and IL-1-dependent excess production of G-CSF and IL-6. Furthermore, transcription factor(s) may be involved in the abnormal IL-1 alpha production.

Production of parathyroid hormone-related protein by cultured human myeloma cells.

We examined whether or not parathyroid hormone-related protein (PTH-rP) is produced in cultured human myeloma cells. PTH-rP protein was detected in the cell extract of one (U266) of 6 myeloma cell lines examined by radioimmunoassay. Also we demonstrated the expression of PTH-rP mRNA in all of the 6 lines. These findings indicate that PTH-rP may be one of the factors responsible for the bone destruction or hypercalcemia in multiple myeloma.

Thrombocytosis in patients with tumors producing colony-stimulating factor.

We investigated the cause of thrombocytosis in 14 patients with tumors producing colony-stimulating factor (CSF). Of the 14 patients, 10 had tumors producing granulocyte-CSF (G-CSF) and 4 had tumors producing granulocyte-macrophage--CSF (GM-CSF). Thrombocytosis of greater than 400 x 10(9)/L was noted in 8 of 10 patients with G-CSF-producing tumors and all 4 patients with GM-CSF-producing tumors. Median peak platelet counts were, respectively, 511 x 10(9)/L (range, 384 to 694 x 10(9)/L) and 579 x 10(9)/L (range, 526 to 910 x 10(9)/L) in patients with tumors producing G-CSF and GM-CSF. In most patients, thrombocytosis declined towards the terminal stage. High interleukin-1 (IL-1) and IL-6 levels were found in addition to CSFs in the plasma or culture supernatants of tumor cells obtained from most patients. In patients with GM-CSF-producing tumors, these specimens had megakaryocyte-CSF (Meg-CSF) activity, which was abolished by anti-GM-CSF antibody. These specimens also had megakaryocyte potentiating (Meg-Pot) activity attributable to both GM-CSF and IL-6. In patients with G-CSF-producing tumors, only Meg-Pot activity due to IL-6 was detected. These results indicate that the thrombocytosis in GM-CSF-producing tumors was caused by both the Meg-CSF activity of GM-CSF and the Meg-Pot activity of IL-6 plus GM-CSF, while that in G-CSF-producing tumors was due to the Meg-Pot activity of IL-6.

Interleukin-5 in eosinophilic gastroenteritis.

During a 4-year period a 28-year-old female had 4 episodes of eosinophilia of over 10,000/mu 1; these episodes were associated with nausea, vomiting, diarrhea, and abdominal pain. On one occasion, she had ascites and pleural effusion which contained numerous mature eosinophils. On each occasion, these attacks disappeared within several weeks without any specific treatment. A diagnosis of eosinophilic gastroenteritis was made. A plasma sample obtained during the eosinophilia generated in vitro eosinophilic colonies when added to granulocyte/macrophage-progenitor (CFU-GM) cultures without exogenous growth factors. Colony formation was inhibited by anti-interleukin-5 (IL-5) antibody but not by antibodies toward IL-3, granulocyte colony-stimulating factor (G-CSF) or GM-CSF. A high plasma interleukin-5 (IL-5) level was noted when measured by enzyme-linked immunosorbent assay, while IL-3, G-CSF, and GM-CSF were undetectable. During remission the plasma gave negative results both for colony formation and IL-5 level. These results indicate that the eosinophilia of this disease is mediated by IL-5.

IL-1 production as a regulator of G-CSF and IL-6 production in CSF-producing cell lines.

We previously demonstrated that colony stimulating factor (CSF)-producing cell lines co-produce interleukin-1 (IL-1) and IL-6 in addition to CSFs. In the present study, we examined the role of IL-1 production in three human tumour cell lines producing granulocyte (G)-CSF, IL-1 and IL-6. Addition of anti-human IL-1 alpha antiserum to the culture caused a 90-62% reduction of G-CSF and a 85-44% reduction of IL-6 production, respectively, as evaluated by enzyme immunoassay in all three cell lines. The decrease of G-CSF and IL-6 production by the anti-IL-1 alpha antiserum was also confirmed at the level of mRNA expression. The anti-IL-1 alpha antiserum did not affect the growth of these cell lines. Excess recombinant IL-1 alpha exogenously added to the culture enhanced G-CSF and IL-6 production in all three cell lines. However, IL-1 alpha had little effect on the growth of these three cell lines. Neither anti-IL-6 nor anti-G-CSF antibodies affected the production of the other cytokines. These results indicate that IL-1 alpha regulates G-CSF and IL-6 production in these tumour cell lines, and suggest that the IL-1 production plays an important role in CSF-producing tumours.

Identification of colony-stimulating factor activity in patients with malignant tumors associated with excessive leukocytosis.

We have tried to demonstrate and identify colony-stimulating factor (CSF) activity in the plasma, pleural fluid, ascites or culture supernatant of tumor cells in 11 patients with malignant tumors associated with unexplained persistent leukocytosis. The specimens were treated with anti-granulocyte (G)-CSF or anti-granulocyte/macrophage (GM)-CSF monoclonal antibodies, then added to GM-progenitor (CFU-GM) cultures without exogenous CSFs. In all patients, untreated specimens generated CFU-GM-derived colonies, and colony formation was clearly inhibited by only one of the two antibodies, indicating the presence of either G-CSF or GM-CSF in the specimens. Furthermore, we measured the concentrations of G-CSF or GM-CSF in the specimens using an enzyme-linked immunosorbent assay, and confirmed the results by CFU-GM assay. Two patients were shown to have GM-CSF-producing tumors, while the other patients were G-CSF-producing. These assays are useful in identifying CSF activity in patients with CSF-producing tumors.

Megakaryocyte potentiating activity of IL-1, IL-6 and GM-CSF as evaluated by their action on in vitro human megakaryocytic colonies.

We examined whether recombinant cytokines enhance the in vitro platelet production of interleukin-3 (IL-3)-induced human megakaryocytic colonies (Meg-colony). We classified Meg-colonies into four categories based on platelet production during in situ observation on day 14: type 0, absence of cytoplasmic processes in a colony; type 1, one to three processes in at least one megakaryocyte in a colony; type 2, four to eight processes; type 3, more than nine processes or division of cytoplasm. Type 3 colonies were considered to be platelet-producing. In control cultures, type 1 Meg-colonies were dominant, followed by type 2, type 3 and type 0. Of the cytokines added at the initiation of culture, interleukin-1 alpha (IL-1 alpha), interleukin-6 (IL-6), and granulocyte/macrophage colony stimulating factor (GM-CSF) significantly increased the number of colonies. Furthermore, these three cytokines significantly elevated the proportion of type 3 colonies. Interleukin-4 (IL-4), granulocyte-CSF, macrophage-CSF and erythropoietin did not affect the colony count or distribution of colony type. IL-1 alpha, IL-6 and GM-CSF also significantly elevated the proportion of type 3 colonies, even when added to the culture on days 8 or 11. These results indicate that IL-1 alpha, IL-6 and GM-CSF promote platelet production of in vitro Meg-colonies.