Metabolomic Changes in Serum of Children with Different Clinical Diagnoses of Malnutrition.

BACKGROUND: Mortality in children with severe acute malnutrition (SAM) remains high despite standardized rehabilitation protocols. Two forms of SAM are classically distinguished: kwashiorkor and marasmus. Children with kwashiorkor have nutritional edema and metabolic disturbances, including hypoalbuminemia and hepatic steatosis, whereas marasmus is characterized by severe wasting. The metabolic changes underlying these phenotypes have been poorly characterized, and whether homeostasis is achieved during hospital stay is unclear. OBJECTIVES: We aimed to characterize metabolic differences between children with marasmus and kwashiorkor at hospital admission and after clinical stabilization and to compare them with stunted and nonstunted community controls. METHODS: We studied children aged 9-59 mo from Malawi who were hospitalized with SAM (n = 40; 21 with kwashiorkor and 19 with marasmus) or living in the community (n = 157; 78 stunted and 79 nonstunted). Serum from patients with SAM was obtained at hospital admission and 3 d after nutritional stabilization and from community controls. With the use of targeted metabolomics, 141 metabolites, including amino acids, biogenic amines, acylcarnitines, sphingomyelins, and phosphatidylcholines, were measured. RESULTS: At admission, most metabolites (128 of 141; 91%) were lower in children with kwashiorkor than in those with marasmus, with significant differences in several amino acids and biogenic amines, including those of the kynurenine-tryptophan pathway. Several phosphatidylcholines and some acylcarnitines also differed. Patients with SAM had profiles that were profoundly different from those of stunted and nonstunted controls, even after clinical stabilization. Amino acids and biogenic amines generally improved with nutritional rehabilitation, but most sphingomyelins and phosphatidylcholines did not. CONCLUSIONS: Children with kwashiorkor were metabolically distinct from those with marasmus, and were more prone to severe metabolic disruptions. Children with SAM showed metabolic profiles that were profoundly different from stunted and nonstunted controls, even after clinical stabilization. Therefore, metabolic recovery in children with SAM likely extends beyond discharge, which may explain the poor long-term outcomes in these children. This trial was registered at isrctn.org as ISRCTN13916953.

Zinc erythrocyte protoporphyrin as marker of malaria risk in pregnancy - a retrospective cross-sectional and longitudinal study.

BACKGROUND: The effects of iron interventions and host iron status on infection risk have been a recurrent clinical concern, although there has been little research on this interaction in pregnant women. METHODS: Cross-sectional and longitudinal analyses were undertaken to determine the association of whole blood zinc erythrocyte protoporphyrin (ZPP) with malaria parasitaemia in pregnant women attending antenatal and delivery care at Montfort and Chikwawa Hospitals, Shire Valley, Malawi. Prevalence of antenatal, delivery and placental malaria was assessed in relation to maternal ZPP levels. The main outcome measures were prevalence of peripheral and placental Plasmodium falciparum parasitaemia and odds ratios of malaria risk. RESULTS: A total of 4,103 women were evaluated at first antenatal visit, of whom at delivery 1327 were screened for peripheral and 1285 for placental parasitaemia. Risk of malaria at delivery (peripheral or placental) was higher in primigravidae (p < 0.001), and lower (peripheral) with use of intermittent preventive anti-malarials during pregnancy (p < 0.001). HIV infection was associated with increased malaria parasitaemia (p < 0.02, peripheral or placental). Parasitaemia prevalence was lower in women with normal ZPP levels compared to those with raised concentrations at both first antenatal visit (all gravidae, p = 0.048, and at delivery (all gravidae, p < 0.001; primigravidae, p = 0.056). Between first antenatal visit and delivery women who transitioned from raised (at first antenatal visit) to normal ZPP values (at delivery) had lower peripheral parasitaemia prevalence at delivery compared to those who maintained normal ZPP values at both these visits (all gravidae: 0.70, 95%CI 0.4-1.1; primigravidae: 0.3, 0.1-0.8). In regression analysis this difference was lost with inclusion of HIV infection in the model. CONCLUSIONS: Raised ZPP concentrations in pregnancy were positively associated with P. falciparum parasitaemia and were probably secondary to malaria inflammation, rather than indicating an increased malaria risk with iron deficiency. It was not possible from ZPP measurements alone to determine whether iron deficiency or repletion alters malaria susceptibility in pregnancy.

Impaired glucose absorption in children with severe malnutrition.

OBJECTIVE: To quantify intestinal glucose absorption in children with two types of severe malnutrition, kwashiorkor and marasmus, compared with healthy children. STUDY DESIGN: Children with kwashiorkor (n = 6) and marasmus (n = 9) and control subjects (n = 3) received a primed (13 mg/kg), constant infusion (0.15 mg/kg/min) of [6,6H2]glucose for 4.5 hours. Two hours after start of the infusion an oral bolus of glucose 1.75 g/kg labeled with [U-13C]glucose 10 mg/g was given and was followed by periodic blood sampling. Mathematical modeling was applied to determine oral glucose absorption. RESULTS: Median total glucose absorption was 5.9 mmol/kg, interquartile range (IQR) 4.5-6.7 mmol/kg and 4.4 (IQR 2.9-5.9) mmol/kg in children with kwashiorkor and marasmus compared with 7.7 (IQR 5.8-9.0) mmol/kg in control subjects; P = .03 compared with marasmus). Children with the lowest glucose absorption were found specifically in the kwashiorkor group and marasmic children with hypoalbuminemia. Severe impairment in absorption correlated with urinary 8-hydroxydeoxyguanosine secretion (r = -0.62, P = .01). CONCLUSIONS: Severe malnutrition is associated with an impaired glucose absorption and decreased glucose absorption correlates with oxidative stress in these children.

Reduced risk for placental malaria in iron deficient women.

BACKGROUND: Nutritional iron deficiency may limit iron availability to the malaria parasite reducing infection risk, and/or impair host immunity thereby increasing this risk. In pregnant women, there is evidence of an adverse effect with iron supplementation, but the few reported studies are strongly confounded. METHODS: A case control study in pregnant Malawian women was undertaken in Chikhwawa southern Malawi in order to describe iron status in relation to placental malaria controlling for several confounding factors. Pregnancy characteristics were obtained and a blood sample at delivery. A full blood count was performed and serum ferritin and transferrin receptor quantified by enzyme-linked immunoassay. DNA analysis was used to identify genetic polymorphisms for ABO phenotype, hemoglobin HbS, and glucose -6 phosphate dehydrogenase deficiency. Placental tissue was obtained and malaria histology classified as active, past or no malaria infection. RESULTS: 112 cases with placental malaria were identified and 110 women with no evidence of placental infection. Iron deficiency was less frequent in women with placental Plasmodium falciparum infection. In those with acute, chronic or past placental infections the odds ratio for iron deficiency was 0.4, 95% CI 0.2-0.8, p = 0.01; for acute and chronic infections 0.4, 0.2-0.8, p = 0.006; for acute infection 0.3, 0.1-0.7, p = 0.001. The association was greater in multigravidae. CONCLUSION: Women with either acute, or acute and chronic placental malaria were less likely to have iron deficiency than women without placental malaria infection There is a priority to establish if reversing iron deficiency through iron supplementation programs either prior to or during pregnancy enhances malaria risk.

Plasma cytokines quantification among Trypanosoma brucei rhodesiense sleeping sickness cases and controls in Rumphi, Malawi.

Introduction: Trypanosoma brucei (T.b.) rhodesiense is the cause of the acute form of human African trypanosomiasis (HAT) in eastern and southern African countries, including Malawi. For a long time, untreated HAT infections were believed to be 100% fatal. However, recent studies show that infection by T.b. rhodesiense can result in a wide range of clinical outcomes in its human host. Apart from other factors such as parasite diversity, cytokines have been strongly implicated to play a major role in the outcome of T.b. rhodesiense infections.In this study, we quantify the levels of three cytokines Interleukin-8 (IL-8), Tumor Necrotic Factor alpha (TNF-α) and Interleukin -10 (IL-10) in plasma amongst HAT cases (treated and untreated) and controls recruited during medical survey. Methods: Two-hundred and thirty-three plasma samples (HAT cases and controls) from Rumphi, one of the endemic areas in Malawi were used. Blood collected was centrifuged, plasma extracted and stored in cryovials at -80°C until processing. Plasma cytokine concentration was measured using ELISA. Results: Plasma samples for 233 individuals, 76 HAT cases and 157 controls were quantified. Among the cases, nine had their plasma collected before treatment (untreated) and the rest were treated before blood for plasma analysis was collected. Controls had significantly higher mean plasmatic levels of TNF-α (94.5 ±474.12 pg/ml) and IL-8 (2258.6 ±5227.4 pg/ml) than cases TNF-α (29.35±181.58 pg/ml) and IL-8 (1191.3±4236.09 pg/ml). Controls and cases had similar mean levels of IL-10 in plasma. Only IL-8 had statistically significant higher median levels in the untreated than treated HAT cases P=0.006. Conclusion: Our data suggest that cytokines could be considered as biomarkers of HAT infection and treatment. Further studies with a larger cohort of cases and additional cytokines which are known to be associated with HAT infection outcomes will be required to evaluate these cytokines further.

Mechanisms behind decreased endogenous glucose production in malnourished children.

Severe malnutrition is a major health problem in developing countries and can present itself as kwashiorkor or marasmus. Although marasmus is characterized by clinical wasting, kwashiorkor is associated with peripheral edema, oxidative stress, hypoalbuminemia, and hypoglycemia. The etiology of the hypoglycemia is poorly understood. We determined endogenous glucose production (EGP) in children with severe malnutrition. Children with kwashiorkor, marasmus, and controls received a primed constant infusion of [6,6H2]glucose for 2 h. An i.v. bolus of 13C-ketoisocaproic acid (KIC) was given, and breath samples were obtained during 2 h. Isotope dilution was used to calculate EGP, and 13CO2/12CO2 production was determined. Mean EGP ± SEM was 5.5 ± 0.3 mg/kg/min in the kwashiorkor group and 6.9 ± 0.4 mg/kg/min and 7.6 ± 0.7 mg/kg/min in the marasmic and control group, respectively, (p < 0.05 kwashiorkor versus marasmus and controls). EGP correlated with serum albumin concentration (r = 0.67; p < 0.001) and urinary 8-hydroxydeoxyguanosine as a marker of oxidative stress (r = -0.62; p < 0.005). 13CO2 secretion as a marker of hepatic mitochondrial function was significantly higher in the marasmic group compared with kwashiorkor and controls. We conclude that decreased EGP in severely malnourished children is related to the degree of hypoalbuminemia and oxidative stress but is not associated with a clear defect in hepatic mitochondrial function.

The use of molecular technology to investigate trypanosome infections in tsetse flies at Liwonde Wild Life Reserve.

Background: Trypanosomes are protozoan flagellates that cause human African trypanosomiasis (HAT) and African animal trypanosomiasis (AAT). HAT is caused by Trypanosoma brucei rhodesiense in East and Central Africa and T.b. gambiense in West Africa, whereas AAT is caused by a number of trypanosome species, including T. brucei brucei, T. evansi, T. vivax, T. congolense, T. godfreyi and T. simiae. The aim of this study was to establish if tsetse flies at Liwonde Wild Life Reserve (LWLR) are infected with these trypanosomes and thus pose a risk to both humans and animals within and surrounding the LWLR. Methods: A total of 150 tsetse flies were caught. Of these, 82 remained alive after capture and were dissected such that the mid-gut could be examined microscopically for trypanosomes. DNA extractions were performed from both mid-guts and the 68 dead flies using a Qiagen Kit. Amplification techniques involved the Internal Transcriber Spacer 1 (ITS 1) conventional polymerase chain reaction (PCR) with primers designed to identify trypanosome species, and Repetitive Insertion Mobile Element - Loop Mediated Isothermal Amplification (RIME LAMP), a sequence specific to T. brucei. Results: Analysis showed that 79/82 (96.3%) of the mid-guts examined microscopically were positive for trypanosomes and that 75/150 (50%) of the DNA extracts (from the mid-gut, and tsetse fly carcasses) were positive for T. brucei, as determined by the RIME LAMP method. ITS1 PCR further showed that 87/150 (58.0%) flies were positive for trypanosomes, of which 56/87 (64.4%) were T. brucei, 9/87 (10.3%) were T. vivax; 7/87 (8.1%) were T. simiae; 6/87 (6.9%) were T. congolense, and 6/87 (6.9%) were T. godfreyi. Ten samples had a mixture of infections. Conclusion: Our analysis demonstrated a mixture of infections from trypanosome species in tsetse flies at LWLR, and that T. brucei, the species that causes HAT, was the most common. Our study successfully used molecular techniques to demonstrate the presence of T. b. rhodesiense at LWLR, a species that causes HAT in both East and Central Africa.

Urinary Organic Acids Increase After Clinical Stabilization of Hospitalized Children With Severe Acute Malnutrition.

BACKGROUND: Despite a reduction of child mortality in low-income countries, acutely ill undernourished children still have an elevated risk of death. Those at highest risk are children with severe acute malnutrition (SAM) who often show metabolic dysregulation that remains poorly understood. OBJECTIVE: We performed a pilot study to examine changes in urinary organic acids during nutritional rehabilitation of children with SAM, and to identify metabolites associated with the presence of edema or with mortality. METHODS: This study included 76 children aged between 6 and 60 months, hospitalized for SAM at the Moyo Nutritional Rehabilitation and Research Unit in Blantyre, Malawi. Urine was collected at admission and 3 days after clinical stabilization and metabolomics were performed using gas chromatography-mass spectrometry. Metabolite concentrations were evaluated with both uni- and multivariate approaches. RESULTS: Most metabolites increased 3 days after clinical stabilization, and total urinary concentration changed from 1.2 mM (interquartile range [IQR], 0.78-1.7) at admission to 3.8 mM (IQR, 2.1-6.6) after stabilization (P < .0001). In particular, 6 metabolites showed increases: 3-hydroxybutyric, 4-hydroxyhippuric, p-hydroxyphenylacetic, oxoglutaric, succinic, and lactic acids. Urinary creatinine was low at both time points, but levels did increase from 0.63 mM (IQR, 0.2-1.2) to 2.6 mM (IQR,1.6-4.4; P < .0001). No differences in urinary profiles were found between children who died versus those who survived, nor between children with severe wasting or edematous SAM. CONCLUSIONS: Total urinary metabolites and creatinine increase after stabilization and may reflect partial recovery of overall metabolism linked to refeeding. The use of urinary metabolites for risk assessment should be furthered explored. TRIAL REGISTRATION: TranSAM study (ISRCTN13916953).

Association of APOL1 renal disease risk alleles with Trypanosoma brucei rhodesiense infection outcomes in the northern part of Malawi.

Trypanosoma brucei (T.b.) rhodesiense is the cause of the acute form of human African trypanosomiasis (HAT) in eastern and southern African countries. There is some evidence that there is diversity in the disease progression of T.b. rhodesiense in different countries. HAT in Malawi is associated with a chronic haemo-lymphatic stage infection compared to other countries, such as Uganda, where the disease is acute with more marked neurological impairment. This has raised the question of the role of host genetic factors in infection outcomes. A candidate gene association study was conducted in the northern region of Malawi. This was a case-control study involving 202 subjects, 70 cases and 132 controls. All individuals were from one area; born in the area and had been exposed to the risk of infection since birth. Ninety-six markers were genotyped from 17 genes: IL10, IL8, IL4, HLA-G, TNFA, IL6, IFNG, MIF, APOL, HLA-A, IL1B, IL4R, IL12B, IL12R, HP, HPR, and CFH. There was a strong significant association with APOL1 G2 allele (p = 0.0000105, OR = 0.14, CI95 = [0.05-0.41], BONF = 0.00068) indicating that carriers of the G2 allele were protected against T.b. rhodesiense HAT. SNP rs2069845 in IL6 had raw p < 0.05, but did not remain significant after Bonferroni correction. There were no associations found with the other 15 candidate genes. Our finding confirms results from other studies that the G2 variant of APOL1 is associated with protection against T.b. rhodesiense HAT.

Comparative evaluation of dry and liquid RIME LAMP in detecting trypanosomes in dead tsetse flies.

Xenomonitoring is an important approach in assessing the progress of trypanosomiasis control as well as in estimating the endemicity of trypanosomes in affected areas. One of the major challenges in this approach is the unavailability of sensitive and easy to use xenomonitoring tools that can be used in the remote areas where the disease occurs. One tool that has been used successfully in detecting the parasites in tsetse flies is the repetitive insertion mobile element loop-mediated isothermal amplification (RIME LAMP). This tool has recently been modified from the liquid form to dry form for use in remote areas; however, uptake for use in the field has been slow. Field-collected tsetse flies were used to evaluate the performance of dry RIME LAMP over the conventional liquid RIME LAMP. All the samples were also subjected to internal transcribed spacer 1 (ITS1) ribosomal deoxyribonucleic acid (DNA) polymerase chain reaction (PCR) as a standard. ITS1-PCR-positive samples were further sequenced for confirmation of the species. A total of 86 wild tsetse flies were left to dry at room temperature for 3 months and DNA was extracted subsequently. All 86 flies were Glossina morsitans morsitans. From these, dry RIME LAMP detected 16.3% while liquid RIME LAMP detected 11.6% as infected with trypanosomes. Ten positive samples on ITS1-PCR were sequenced and all were shown to be trypanosomes. The use of dry RIME LAMP in the field for xenomonitoring of trypanosomes in tsetse flies will greatly contribute towards control of this neglected tropical disease as it provides the cheapest, fastest and simplest way to estimate possible human infective trypanosome infection rates in the tsetse fly vectors.

ABO blood group phenotypes influence parity specific immunity to Plasmodium falciparum malaria in Malawian women.

BACKGROUND: Blood group O has been significantly associated with increased placental malaria infection in primiparae and reduced risk of infection in multiparae in the Gambia, an area with markedly seasonal malaria transmission. This study analyses the association between ABO blood group phenotypes in relation to placental malaria pathology and birth outcomes in southern Malawi, an area with perennial malaria transmission. METHODS: A cross-sectional study of 647 mother/child pairs delivering in Montfort Hospital, Chikwawa District between February-June 2004 and January-July 2005 was undertaken. Maternal peripheral and cord blood samples were obtained at delivery. Placental tissue was obtained and malaria histology classified as active, past or no malaria infection. Birth anthropometry was recorded. ABO blood group was measured by agglutination. RESULTS: In primiparae, blood group O was significantly associated with increased risk of active placental infection (OR 2.18, 95% CI 1.15-4.6, p = 0.02) and an increased foetal-placental weight ratio compared to non-O phenotypes (5.68 versus 5.45, p = 0.03) In multiparae blood group O was significantly associated with less frequent active placental infection (OR 0.59, 95% CI 0.36-0.98, p = 0.04), and a higher newborn ponderal index compared to non-O phenotypes (2.65 versus 2.55, p = 0.007). In multivariate regression parity was independently associated with increased risk of placental malaria (active andpast infection) in primiparae with blood group O (p = 0.034) and reduced risk in multiparae with the same phenotype (p = 0.015). CONCLUSION: Parity related susceptibility to placental malaria is associated with the mothers ABO phenotype. This interaction influences foetal and placental growth and could be an important modifying factor for pregnancy outcomes. The biological explanation could relate to sialic acid dependent placental membrane differences which vary with ABO blood group.

AcSDKP is down-regulated in anaemia induced by Trypanosoma brucei infection in mice.

Background: Anaemia commonly results from destruction of erythrocytes in the peripheral blood and failure of the bone marrow haematopoietic cells to replenish the erythrocytes. The mechanisms involved in trypanosoma-induced anaemia, including the role of the bone marrow haematopoietic cells are incompletely understood. We studied the responses of a tetrapeptide, AcSDKP, and IL-10, and their association with bone marrow nucleated cells in a Trypanosoma brucei brucei GVR35 experimental infection model. Methods: Mouse infection was done intraperitoneally with 1 × 103 trypanosomes/mL. Mice were either infected or left uninfected (N = 100). At days 0, 9, 16, 23, 30, 37, and 44 post-infection, mice were euthanised and blood was collected by cardiac puncture to examine for parasitaemia and packed cell volume (PCV) and then centrifuged for plasma, which was used for cytokine ELISA. The mice's femurs were also dissected and bone marrow was collected for femur cellularity. Results: PCV dropped from 39.6% to 27% in infected animals by day 9 and remained low (relative to uninfected mice) for the duration of the experiment. AcSDKP levels decreased from day 0 (11.5 × 104 pg/mL) to day 16 (10 × 104), and increased by day 30 (12.6 × 104). There was a significant difference at day 16 (P = 0.023) between the infected and uninfected groups. By contrast, expression of IL-10 markedly increased between day 0 (18.6 pg/mL) and day 16 (145 pg/mL) and decreased by day 30 (42.8 pg/mL). There was also a significant difference in IL-10 expression between infected and uninfected mice at day 16 (P < 0.001). Bone marrow nucleated cells were significantly reduced during periods of low plasma AcSDKP and high plasma IL-10 concentrations (5.4 × 106 infected vs 6.2 × 106 on day 0 and 4.9 × 106 infected vs 10 × 106 uninfected on day 16). Conclusions: These data unravel a possible negative feedback interaction between AcSDKP and IL-10 in trypanosome infection. More importantly, this study implicates an IL-10/AcSDKP cytokine network in the regulation of bone marrow nucleated cells and provides a new potential mechanism in the pathogenesis of trypanosoma-induced anaemia. Further mechanistic blocking experiments on AcSDKP and IL-10 are recommended to further clarify understanding of the interaction.

Polymerase chain reaction identification of Trypanosoma brucei rhodesiense in wild tsetse flies from Nkhotakota Wildlife Reserve, Malawi.

BACKGROUND: Trypanosoma brucei rhodesiense is the causative agent of acute human African trypanosomiasis. Identification of T. b. rhodesiense in tsetse populations is essential for understanding transmission dynamics, assessng human disease risk, and monitoring spatiotemporal trends and impact of control interventions. Accurate detection and characterisation of trypanosomes in vectors relies on molecular techniques. For the first time in Malawi, a molecular technique has been used to detect trypanosomes in tsetse flies in Nkhotakota Wildlife Reserve. METHODS: A polymerase chain reaction (PCR) technique was used to identify the serum resistance associated (SRA) gene of T. b. rhodesiense in tsetse flies. Of 257 tsetse flies that were randomly caught, 42 flies were dissected for microscopic examination. The midguts of 206 flies were positive and were individually put in eppendorf tubes containing phosphate-buffered saline (PBS buffer) for DNA extraction. Internal transcribed spacer (ITS)-PCR was first used to isolate all trypanosome species from the flies. TBR PCR was then used to isolate the Trypanozoon group. T. brucei-positive samples were further evaluated by SRA PCR for the presence of the SRA gene. RESULTS: Of 257 flies caught, 185 (72%) were Glossina morsitans morsitans and 72 (28%) were Glossina pallidipes. Three were tenerals and 242 were mature live flies. Of the 242 flies dissected, 206 were positive, representing an 85.1% infection rate. From 206 infected flies, 106 (51.5%) were positive using ITS-PCR, 68 (33.0%) being mixed infections, 18 (8.7%) T. brucei, 9 (4.4%) Trypanosoma vivax, 4 (1.9%) Trypanosoma godfrey, 3 (1.5%) Trypanosoma congolense savanna, 3 (1.5%) Trypanosoma simae, and 1 (0.4%) Trypanosoma simaetsavo. When subjected to TBR PCR, 107(51.9%) were positive for T. brucei. Of the 107 T. brucei-positive samples, 5 (4.7%) were found to have the SRA gene. CONCLUSIONS: These results suggest that wild tsetse flies in Malawi are infected with human-infective trypanosomes that put communities around wildlife reserves at risk of human African trypanosomiasis outbreaks. Further studies need to be done to identify sources of blood meals for the flies and for surveillance of communities around wildlife reserves.

Kwashiorkor and marasmus are both associated with impaired glucose clearance related to pancreatic ß-cell dysfunction.

Severe malnutrition is a major health problem in developing countries and can present as kwashiorkor or marasmus. Kwashiorkor is associated with septicaemia, profound metabolic changes including hepatic steatosis, altered protein metabolism and increased oxidative stress. Limited data suggest that children with kwashiorkor have an impaired glucose tolerance and insulin secretion. Our objective was to determine glucose tolerance in children with kwashiorkor compared to marasmus and its relation to insulin secretion and sensitivity. Six children with kwashiorkor and 8 children with marasmus were studied. We were also able to include 3 healthy children for comparison. They received a primed (13 mg/kg), constant infusion (0.15 mg/kg/min) of [6,6-(2)H(2)]glucose for 4 h with serial blood sampling. In addition, an oral glucose tolerance test (OGTT) was performed with labeled 10 mg/g [U-(13)C]glucose. Glucose clearance was determined using mathematical modeling. Glucose clearance rates during the OGTT were -392 (range 309) mL/kg in children with kwashiorkor, -156 (426) mL/kg in marasmus and 279 (345) mL/kg in the control group. Glucose clearance rates correlated with plasma albumin concentrations (r=0.67, P=.001). Insulin responses were strongly impaired in both kwashiorkor and marasmus. There was no indication of peripheral or hepatic insulin resistance in the malnourished groups. We show that glucose clearance rates are affected in both children with marasmus as well as kwashiorkor, which correlate with plasma albumin concentrations. The disturbed glucose clearance in malnutrition is related to an impairment in insulin availability.

Cytosine arabinoside reduces the numbers of granulocyte macrophage colony forming cells (GM-CFC) and high proliferative potential colony forming cells (HPP-CFC) in vivo in mice.

BACKGROUND: Cytosine arabinoside (Ara-C) is an S-phase specific cytotoxic drug used in the treatment of malignancies. It is converted to Cytosine Arabinoside triphosphate (Ara-CTP) in the cell. Cytosine Arabinoside triphosphate, reversibly displaces deoxy cytidine triphosphate from DNA polymerase for incorporation into DNA. This process leads to cell death. OBJECTIVE: To investigate the in vivo effects of Ara-C on the Granulocyte Macrophage Colony Forming Cells (GM-CFC) and High Proliferative Potential Colony Forming Cells (HPP-CFC) respectively in mice. METHODOLOGY: Ara-C (150mg/kg) was administered intraperitoneally (i.p) once to mice and bone marrow cells sampled on days 1, 3 and 6. RESULTS: Ara-C reduced the numbers of both GM-CFC and HPP-CFC in the bone marrow. HPP-CFCs were initially more sensitive to Ara-C treatment than GM-CFCs. In the six days after treatment the effect on GM-CFC persisted, while there was a partial recovery in the number of HPP-CFCs. CONCLUSION: It is possible that Ara-C disturbs the stem cells niche by damaging the stromal cells of the bone marrow microenvironment. This would result in derangement of HPP-CFC proliferation.

Affordable pediatric CD4 counting by flow cytometry in Malawi.

BACKGROUND: Rapid expansion of antiretroviral therapy in Malawi has occurred in the relative absence of suitable pediatric CD4 counting facilities. We have recently validated in adults a simplified affordable flow cytometric CD4 counting method, the Blantyre count. There is a need for this technology to transfer to government laboratories run by local staff, and to be validated in children, where %CD4/lymphocyte values are required. METHODS: We assessed agreement of %CD4/lymphocyte values determined by the Blantyre count and Panleucogate methods on an EPICS XL-MCL flow cytometer on 113 venous blood samples from HIV-seropositive children in Blantyre, Malawi. All assays were performed by two Malawian laboratory technicians. RESULTS: Overall bias between the two methods was -0.13% (95% CI -0.37 to 0.11) and limits of agreement were -2.69 to 2.43% (95% CI -3.11 to -2.27 and 2.01 to 2.85). Limits of agreement were within -3.00 and 3.00 for each laboratory technician. Coefficient of variation for the Blantyre count assay was 2.0% and samples showed good stability over 5 days. CONCLUSIONS: The Blantyre count method can accurately determine %CD4/lymphocyte values in blood of HIV-seropositive children on an EPIC XL-MCL flow cytometer at a reagent cost of US $0.21 per test or less. The assay can be competently carried out by local laboratory technicians.

Effects of cooking additives on the content of vitamin C in some vegetables in southern Malawi.

We report a study aimed to determine the effects of using soda sodium bicarbonate), chidulo (ashes obtained from certain plant materials), groundnut flour and cooking oil on the content of Vitamin C (ascorbic acid) in some common vegetables found in the south of Malawi. Soda and chidulo reduced significantly the content of ascorbic acid while groundnut flour and cooking oil increased the availability of ascorbic acid.