The aim of this study was to examine whether the cumulative inflammatory burden is associated with an increase in arterial stiffness in a prospective cohort of psoriatic arthritis (PsA) patients.In total, 72 PsA patients were followed for a median of 6.5 years. Cumulative inflammatory burden was represented by the cumulative averages of repeated measures of erythrocyte sedimentation rate (ca-ESR) and C-reactive protein (ca-CRP). Brachial-ankle pulse wave velocity (PWV) was measured at the last visit. We also included 47 healthy controls for PWV assessment.P = 0.008). PsA patients were divided into two groups based on whether their PWV value is ≥1450 cm/s  or <1450 cm/s . The High PWV group had a significantly higher ca-ESR (29 (19 to 44) versus 18 (10 to 32) mm/1st hour, P = 0.005) and ca-CRP (0.7 (0.3 to 1.4) versus 0.4 (0.2 to 0.7) mg/dl, P = 0.029). Using regression analysis, high ca-ESR (defined as ≥75th percentile: 37 mm/1st hour) was associated with a higher likelihood of being in the High PWV group (odds ratio (OR): 9.455 (1.939 to 46.093), P = 0.005, adjusted for baseline clinical and cardiovascular risk factors; and 9.111 (1.875 to 44.275), P = 0.006, adjusted for last visit parameters).PWV was significantly higher in PsA patients compared with healthy controls after adjustment for age, gender and body weight (1466 ± 29 cm/s versus 1323 ± 38 cm/s, Cumulative inflammatory burden, as reflected by ca-ESR, was associated with increased arterial stiffness in PsA patients even after adjustment for cardiovascular risk factors, emphasizing the important role of chronic inflammation in accelerating the development of cardiovascular risks in PsA patients. Psoriatic arthritis (PsA) is a chronic inflammatory arthritis associated with an increased risk of subclinical ,2, cliniArterial stiffness is an independent predictor of CV events and mortality . Pulse wEighty-two PsA patients who participated in a prior subclinical study of atherosclerosis in PsA  were recWe recruited 47 healthy controls from a broad spectrum of hospital staff. None of the controls had a known history of hypertension, diabetes, hyperlipidemia, or overt CVD , or family history of CVD.Ethics approval was obtained from the Ethics Committee of The Chinese University of Hong Kong-New Territories East Cluster Hospitals, and written informed consent was obtained from all participants according to the Declaration of Helsinki.Pain and physicians’ and patients’ global assessments were evaluated using a 100-point visual analog scale, where 0 indicated excellent well-being and 100 indicated feeling extremely unwell. Physical examination included recording the number of tender and swollen joints using the 68 tender-joint/66 swollen-joint count, the presence of dactylitis, and the number of permanently deformed joints. The Health Assessment Questionnaire (HAQ) was used to evaluate physical function , and theComplete blood count, liver and renal function tests, ESR and CRP were checked at every visit. Fasting blood glucose, and lipid profile , low-density lipoprotein-cholesterol (LDL), high-density lipoprotein-cholesterol (HDL), and triglycerides) were checked at baseline and the last visit. Cumulative inflammation over time was represented by the cumulative averages of ESR (ca-ESR) and CRP (ca-CRP).Brachial-ankle PWV was assessed noninvasively in subjects in the supine position by a dedicated tonometry system  as described previously . All PWVt-test or Mann–Whitney U-test for continuous variables and the chi-square (χ2) test for categorical variables. Analysis of covariance (ANCOVA) was used to compare means of PWV between PsA and control subjects by adjusting for parameters that were distributed differently between groups. Spearman’s correlation was used to evaluate bivariate correlation. ca-ESR and ca-CRP were calculated from the AUC of all available measurements divided by the total number of months of follow-up. Univariate analysis was performed to ascertain the association between clinical parameters and PWV. The cutoff value for CV and atherosclerotic risk, which was derived from the large-scale Chinese population-based study [P <0.1 in the univariate analysis were included in the multivariate analysis. All statistical analyses were conducted using IBM SPSS Statistics Version 22 . A minimal level of significance of P <0.05 is used.Results are expressed as mean ± SD or median (interquartile range) as appropriate. Comparisons between two groups were assessed using the Student’s ed study , was useA total of 72  PsA patients were included in the analysis. At baseline, the mean ± SD age was 49.6 ± 11.7 years and the median (IQR) disease duration was 9.2 (2.4 to 14.1) years. The median follow-up duration from baseline to the time of PWV assessment (last visit) was 6.5 (range: 4.8 to 7.7) years. Table P <0.001) , had higher proportion of women  and a lower body weight  at PWV assessment compared with the PsA patients. However, after adjustment for age, gender and body weight, the adjusted mean PWV was still significantly greater in PsA patients compared with control subjects   .The associations between PWV and clinical features at both baseline and the last visit (PWV assessment visit) are shown in Table P <0.1). At baseline, the high-PWV group had significantly higher ESR levels compared with patients in the low-PWV group (P = 0.048). At PWV assessment, the high-PWV group had significantly higher damaged joint count, ESR and CRP levels.Disease-related variables were compared between the two PWV groups Table . The higP = 0.001) but not ca-CRP . The high-PWV group had significantly higher ca-ESR (31 (21 to 44) versus 17 (10 to 30) mm/1st h, P <0.001) and ca-CRP (0.7 (0.3 to 1.4) versus 0.4 (0.2 to 0.7) mg/dl, P = 0.029) compared with the low-PWV group  mm/1st h and the median ca-CRP was 0.6 (0.2 to 1.1) mg/dl in the PsA patients. There was a significant correlation between PWV and ca-ESR  or last visit , P = 0.006)  Table . In cont6) Table .Table 3P <0.001; adjusted for last-visit parameters: OR 1.135 , P = 0.001).Older age was the other independent risk factor associated with a higher likelihood of being in the high-PWV group , This is the first study to assess the association between cumulative inflammatory burden and arterial stiffness in PsA patients. High cumulative inflammatory burden as reflected by the ca-ESR was a predictor for higher PWV independently of traditional CV risk factors and other disease activity parameters. We also confirmed that PWV is increased in PsA patients compared with healthy control subjects in a larger cohort.P = 0.001) and a marginally significant correlation between ca-CRP and PWV (P = 0.061), suggesting that chronic inflammation may have a causative role in the development of arterial dysfunction in PsA patients. These results were consistent with previous findings that cumulative inflammatory burden (ca-ESR) was associated with increased aortic augmentation index in RA [We have demonstrated a significant correlation between ca-ESR and PWV  and impaired EPC function, leading to decreased release of nitric oxide (NO) . InflammThe strength of our study was the inclusion of a large prospective cohort with long-term follow up. Patients with CV risk factors were not excluded so that our results can be generalized to the usual PsA patient population. Our study also has a few limitations. First, our results may not be applicable to PsA patients from other ethnic backgrounds. Second, the outcome of this study (PWV) is only a surrogate of clinical CV events. In patients with PsA or RA, there are virtually no data to suggest whether PWV is a good surrogate of future CVD events. Third, arterial stiffness was assessed by brachial-ankle PWV but not the gold standard, carotid-femoral PWV. However, brachial-ankle PWV is highly correlated with carotid-femoral PWV and may provide qualitatively similar information . Fourth,In conclusion, PsA patients have increased arterial stiffness compared with healthy control subjects. Cumulative inflammatory burden contributes to the increased arterial stiffness independent of traditional CV risk factors, suggesting that increasing arterial stiffness may be one of the mechanisms linking inflammation and CVD in PsA.


Most acute febrile illnesses (AFI) are usually not associated with a specific diagnosis because of limitations of available diagnostics. This study reports on the frequency of EBV viremia and viral load in children and adults presenting with febrile illness in hospitals in Kenya.A pathogen surveillance study was conducted on patients presenting with AFI (N = 796) at outpatient departments in 8 hospitals located in diverse regions of Kenya. Enrollment criterion to the study was fever without a readily diagnosable infection. All the patients had AFI not attributable to the common causes of fever in Kenyan hospitals, such as malaria or rickettsiae, leptospira, brucella and salmonella and they were hence categorized as having AFI of unknown etiology. EBV was detected in blood using quantitative TaqMan-based qPCR targeting a highly conserved BALF5 gene. The overall frequency of EBV viremia in this population was 29.2%, with significantly higher proportion in younger children of <5years  compared to patients aged ≥5 years (26.3% for 5–15 years or 18.8% for >15 years). With respect to geographical localities, the frequency of EBV viremia was higher in the Lake Victoria region (36.4%), compared to Kisii highland (24.6%), Coastal region (22.2%) and Semi-Arid region (25%). Furthermore, patients from the malaria endemic coastal region and the Lake Victoria region presented with significantly higher viremia than individuals from other regions of Kenya.This study provides profiles of EBV in patients with AFI from diverse eco-regions of Kenya. Of significant interest is the high frequency of EBV viremia in younger children. The observed high frequencies of EBV viremia and elevated viral loads in residents of high malaria transmission areas are probably related to malaria induced immune activation and resultant expansion of EBV infected B-cells. Acute febrile illnesses (AFI), defined as non-specific illnesses presenting with fever ≥ 38°C lasting for less than two weeks without a readily diagnosable source after routine clinical evaluation ,2, are tIn malaria endemic regions, AFI, especially in children are clinically managed as malaria even in the absence of parasites –9. This EBV is reported to account for a significant occurrence of AFI cases ,8. SymptEBV infection is common, with approximately 95% of the global adult population showing serological evidence of exposure ,15. In KThe current study investigated the frequency of EBV viremia and viral load in patients with non-malaria febrile illness residing in different eco-regions of Kenya.Eligible subjects were recruited under a study protocol that was approved by the Ethical Review Committee of the Kenya Medical Research Institute  and the Walter Reed Army Institute of Research Human Subject Protection Board in the United States of America (WRAIR HSPB # 1402).The study assessed patients presenting with febrile illness in 8 hospitals in Kenya for infections with EBV . RecruitWhole blood samples were collected from patients on the day of recruitment. Collected specimens were temporarily stored in liquid nitrogen dry shippers at the respective hospitals, before being transported to the KEMRI/WRP research laboratory in Kisumu for long-term storage at -80°C. During shipping, temperature in the liquid nitrogen dry shippers remained below -100°C.x g was performed for 3 minutes. Finally, the purified DNA was eluted from the column by addition of 100 μL of buffer AVE and stored at -20°C until analysis.DNA was isolated from whole blood using QIAamp MinElute Virus Spin Kit  in accordance with the manufacturer’s recommendations. Briefly, 200 μL of whole blood was added to 25 μL of protease. Cells were then lysed with 200 μL of buffer AL. This was followed with a 15-seconds pulse vortexing and 15 minutes incubation at 56°C. The deproteinised lysed DNA preparation was purified on QIAamp MinElute column by addition of 250 μL of absolute ethanol followed by centrifugation at 6000 x g for 1 minute. This was followed by a series of ‘wash and spin’ involving 500 μL of buffer AW1, buffer AW2, and absolute ethanol, in that order, at a centrifugation speed of 6000 x g for 1 minute for each wash. To completely dry the membrane inside the column, a full speed centrifugation at 20,000 The following target genes were used: Screening was by TaqMan-based PCR that targeted 18S rRNA for the plasmodia genus , 16S rRN5’-CGGAAGCCCTCTGGACTTC-3’, 5’-CCCTGTTTATCCGATGGAATG-3’ for the reverse primer and FAM-5’TGTACACGCACGAGAAATGCGCC-3-TAMRA for the probe. The PCR was performed in a 25 μL reaction mixture which contained 12.5 μL TaqMan PCR (1X) master mix, 0.5 μL of 0.2 μM of forward and reverse primers, 0.25 μL 0.1 μM of probe, 2 μL of DNA sample, and PCR grade water. Amplification was performed at 95°C for 10 minutes, and 45 cycles consisting of 95°C for 15 seconds followed by 60°C for 1 minute.Two microliters of DNA was used for the PCR. This amount corresponded to 4 μL of the original whole blood sample. BALF5 gene sequence of the EBV was amplified by a TaqMan-qPCR on the Applied Biosystems 7300  using primers and probe that have been described before . The priT at various dilutions and the negative controls (water for no template control and EBV negative human sample for non-target template) remained undetected.For each run, a standard curve was generated by serially diluting EBV plasmid DNA that contained the BALF5 gene sequence. The starting concentration for the serial dilution was 12,500 copies/μL of the plasmid DNA. EBV copy numbers of the unknown samples were extrapolated from the standard graph. For quality control, the amplification results were considered valid only when the positive control plasmid generated consistent CDNA concentrations in copies/mL of blood were then calculated according to the following equation: whole blood copies/mL = copy/rnx x 1/0.004 mL of sample /rxnNumerical data were expressed as proportions and compared using Pearson’s Chi-square or Fisher’s exact tests as appropriate. Viral load data were log transformed and expressed as geometric means with 95% confidence intervals. Comparison of geometric means in patient groups such as age, gender, and region of residence was carried out using One-way ANOVA or student T-test as appropriate. Multiple logistic regression was used to assess the association between clinical signs and EBV status or level of viremia. Significance levels were set at 0.05 at 95% confidence interval. Statistical analyses were performed using Stata version 12  or GraphPad Prism version 5 .A total of 796 patients presenting with non-malaria AFI were evaluated for presence of EBV. 5 copies/mL . Comparable viral loads were observed in patients under 5 years  and those over 15 years  and was lowest in patients 5–15 years   . Viral lWe next investigated whether the geographical locality of the patients determines the frequency of EBV viremia and the associated magnitude of viral load. As shown in EBV is a common viral infection, and its symptoms often resemble malaria and other febrile illnesses. The goal of this study was to determine the EBV profile in patients with febrile illnesses that could not be attributed to the malaria or the other commonly reported illnesses caused by rickettsiae, leptospira, brucella and salmonella. To achieve this goal, the frequency of EBV viremia and viral load levels in a patient cohort a priori determined not infected with these endemic infections was investigated. The patients were recruited from 8 surveillance hospitals in Kenya .Most studies of EBV in Kenya have focused on finding association of EBV and malaria in causation of Burkitt’s lymphoma –19,36. T5 copies/mL. Levels of viremia were age dependent  than the coastal region (22.2%), Kisii highland (24.6%) and semi arid areas 25.0%). Many factors could be responsible for the geographical trends, including human social behavior as well as economic activities . These f.0%. Manyantigens . Others antigens ,59,60 thThis study provides profiles of EBV in patients with AFI from diverse eco-regions of Kenya. Of significant interest is the high frequency of EBV viremia in younger children. It is important to note that the frequency reported is from a pool of febrile patients and therefore does not represent EBV prevalence in the community. The risk of EBV infection decreased with age. This is contrary to sero-prevalence data from other reports that have shown an increase in sero-prevalence with age. We think this discrepancy is due to the methods used. In our study, EBV detection by PCR indicates presence of the virus, while sero-prevalence indicates exposure. One limitation of the study is that the screening for the common endemic infections was not comprehensive enough, and therefore, it was not possible to rule out increased viremia from reactivation by other diseases. Finally, it is fair to state that our data on EBV viremia is inconclusive on the clinical importance of finding EBV in blood. We think this is because our tests for pathogens were not exhaustive and especially because we did not rule out other causes of upper respiratory tract infections. Nevertheless, this study serves as a nice platform for further studies on EBV.The opinions or assertions contained herein are the private views of the authors, and are not to be construed as official, or as reflecting true views of the Department of the Army or the Department of Defense. The U.S. Government has the right to retain a nonexclusive, royalty-free license and to any copyright covering this paper.


Plasmodium parasites and a leading cause of malaria mortality. Currently available antimalarial therapy has proven insufficient to prevent neurological complications and death in all cases of cerebral malaria. Souza and colleagues observed that transplantation of bone marrow-derived mesenchymal stromal cells (BM-MSCs) increased survival, reduced parasitemia, decreased malaria pigment accumulation in the spleen, liver and kidney, elevated Kupffer cell count in liver, alleviated renal injury and lung inflammation, and improved lung mechanics in an experimental mouse model of cerebral malaria. Although plenty of challenges lie ahead, their findings show the promise of BM-MSC therapy for the treatment of cerebral malaria.Cerebral malaria, a severe form of the disease, is one of the most severe complications of infection with  Plasmodium parasites, cerebral malaria may cause neurological disorders that manifest as severe headache, drowsiness, confusion, coma, and convulsion, and affect other vital organs such as lung, kidney, heart, spleen and liver. Worldwide, cerebral malaria is a leading cause of malaria mortality, responsible for almost 20 % of adult deaths and 15 % of childhood deaths. To date, there is no effective and safe treatment available for cerebral malaria, notably for use in children, who represent the majority of cases. Even patients that are given standard antimalarial therapy at an early phase still face a high risk of dying despite clearance of the parasite, while approximately 25 % of survivors may develop neurological complications and cognitive impairment. In a recent issue of Stem Cell Research & Therapy, Souza and colleagues [As one of the most severe complications of infection with Trypanosoma cruzi that may cause fatal heart diseases [Plasmodium berghei infection, and increased the survival of infected mice [+ c-Kithi progenitors, which were isolated from mice experimentally infected with P. chabaudi, was found to result in clearance of infected erythrocytes in infected mice, and promote recovery from the infection [in vivo efficacy of MSC treatment against malaria was evaluated in a mouse model of P. berghei infection; administration of MSCs was found to confer host resistance against malaria through increasing IL-12 production, and suppressing IL-10 and regulatory T cell production [To date, stem cell therapy has shown satisfactory outcomes in the treatment of a wide variety of human diseases, mainly tissue injury and immune disorders . With thdiseases . The valted mice . Subsequnfection . The advnfection . As a cooduction . Indeed,P. berghei. Surprisingly, however, these alterations seem not be associated with changes in the expression of pro-inflammatory cytokines, including interferon-gamma, tumor necrosis factor alpha and chemokine (C-X-C motif) ligand 1 (CXCL1) as detected using enzyme-linked immunosorbent assays 5 days post-infection, but rather to reduced peripheral parasitemia. As stated in the discussion, the duration of observation seems relatively short. In the future, the time-course effect of MSC therapy on Plasmodium infection should be evaluated over an extended period of time. Since multiple cell tracking strategies have been developed to track the migration, distribution and target tissues of intravenously injected stem cells [The first insight from this study is that intravenous injection of BM-MSCs results in reduced mortality and parasitemia, decreased malaria pigment accumulation in the spleen, liver and kidney, elevated Kupffer cell count in liver, alleviated renal injury and lung inflammation, and improved lung mechanics in mouse cerebral malaria models experimentally infected with em cells , trackinP. berghei-infected mice, which implicates tissue repair. Interestingly, assessment of brain injury with a five-point, semi-quantitative, severity-based scoring system revealed no significant difference in tissue inflammation or severity of histoarchitectural damage between the treated and untreated infected mice, indicating that BM-MSC therapy does not affect the inflammatory process of the brain tissues. In addition, BM-MSC therapy was found to mitigate renal injury caused by P. berghei, while it did not improve renal function, as reflected by no significant difference in serum creatinine, blood urea nitrogen, blood urea nitrogen/serum creatinine ratio or urinary creatinine between the treated and untreated groups. Further studies to investigate the exact mechanisms underlying these differences seem justified.Second, this study demonstrates that administration of BM-MSCs results in elevated counts of astrocytes and oligodendrocytes in P. berghei ANKA models is thought to be the extent of parasite sequestration in the brain [However, the use of rodent malaria parasite for modeling experimental cerebral malaria remains controversial, since the major difference between human cerebral malaria and the he brain . TherefoSouza and colleagues  evaluate


Malarial retinopathy allows detailed study of central nervous system vascular pathology in living patients with severe malaria. An adult with cerebral malaria is described who had prominent retinal whitening with corresponding retinal microvascular obstruction, vessel dilatation, increased vascular tortuosity, and blood retinal barrier leakage with decreased visual acuity, all of which resolved on recovery. Additional study of these features and their potential role in elucidating the pathogenesis of cerebral malaria is warranted. The pathogenesis of coma in falciparum malaria and its rapid reversibility are potential targets for adjunctive therapies, but they are not well-understood. Microvascular obstruction is probably an important contributor. The brain microvasculature is relatively inaccessible; it can be studied in detail only at post-mortem. Similarly, microvascular obstruction in the retina is thought to be a major contributor to the unique retinopathy of severe falciparum malaria, and, because it is easily visualized in living subjects, in-depth study is providing new and valuable insights. We describe an adult patient with cerebral malaria who had prominent retinal changes with some previously unrecognized features that resolved on recovery.Plasmodium falciparum malaria (parasitemia = 0.3%) with coma, generalized convulsions, hyperlactatemia, renal failure, and black urine. He had no prior medical history. Retinal photography showed bilateral patchy macular whitening with corresponding capillary non-perfusion and leakage of fluorescein caused by blood retinal barrier breakdown on fluorescein angiography , which are distributed particularly around the optic disk and typically more opaque. Malarial retinopathy is considered reversible,The angiogram in this patient showed that the whitening corresponds closely to capillary non-perfusion. This finding has not been described previously in adults but is common in Malawian children with cerebral malaria.This patient had mildly increased tortuosity of retinal blood vessels that decreased on recovery. Although increased vascular tortuosity has not been well-described in malaria, it is a recognized feature of other vascular occlusive diseases of the retina. Vessel tortuosity is caused by a combination of vessel dilation from radial stretching and the vessel taking a more serpentine path because of longitudinal stretching.1Increased vascular tortuosity has not been well-described previously in severe falciparum malaria, possibly because the normal appearance of retinal vessels varies significantly between individuals and subtle changes are difficult to identify. Ophthalmoscopy revealed engorgement and tortuosity of retinal veins in 26% of children with cerebral malaria in Ghana, which mostly resolved by 1 week.The angiogram in this patient showed focal leakage of fluorescein across the blood–retinal barrier (BRB) in areas of non-perfusion, suggesting a common etiology. The BRB is analogous to the blood–brain barrier, which is also mildly disrupted in cerebral malaria. Leakage from larger retinal vessels crossing ischemic areas is a well-known phenomenon in retinal ischemia. The significance of this finding as a contributor to the pathogenesis of malarial coma is not known. More angiographic studies are needed.This patient had decreased visual acuity, which had resolved at follow-up. Although it is not possible to give a cause, it is the first report of an association between macular retinal whitening and decreased visual acuity with subsequent recovery.Additional studies of malarial retinopathy have great potential to enhance our understanding of vascular changes in severe malaria. To maximize their impact, studies should use retinal photography, where possible, to allow detailed examination of the full range of fundus signs by multiple blinded observers. This examination should be done both acutely and at follow-up. Fluorescein angiography provides a highly detailed map of CNS retinal perfusion. There is a need for additional detailed studies to include assessment of vascular tortuosity to investigate its role as a potential early and sensitive marker in studies of severe malaria.The rate of reversibility of malarial retinopathy has potential as an end point in intervention studies of severe malaria, particularly for adjunctive therapies that directly target the pathogenesis. Additional information on the speed of reversibility of the various components of malarial retinopathy is needed, and studies are underway to investigate this.


Following enhanced malaria control measures, such as nationwide free distribution of insecticide-treated bed nets (ITN) by the government of Cameroon, its impact on malariometric and red cell indices in children ≤14 years in Muea, in the Mount Cameroon area was evaluated.Two cross-sectional studies were conducted during the malaria transmission season (March-July) in 2006 (baseline) and 2013 (follow-up), respectively. The investigative methods included the use of a questionnaire to assess ITN use and coverage, clinical evaluation and laboratory investigations. Blood sample collected from each child was used for the preparation of blood films for detection of malaria parasites and density as well as full blood count determination using standard procedures and also an automated haematology analyzer.χ2 = 18.3, P <0.001) in the youngest group of children while in the follow-up study, it was highest in the oldest . The overall prevalence of mild, moderate and severe anaemia in the study population at baseline  decreased significantly (P <0.001) to 24.4, 2.7 and 1.3%, respectively during the follow-up with the highest relative risk reduction in prevalence occurring in moderate anaemia . Microcytic anaemia also decreased significantly (P <0.001) from 56 to 7.7% during the follow-up survey.The majority of children (81.5%) possessed an ITN in 2013. The proportion of effective users of ITN increased significantly from 20.9% (CI = 17.3-25%) in 2006 to 35.2% (CI = 31–39.7%) in 2013. The highest relative risk reduction in prevalence during the follow-up study was observed in malaria anaemia , followed by gametocytaemia , anaemia , and malaria parasitaemia . In the baseline survey, the prevalence of splenomegaly was significantly highest  was a more sensitive measure to changes in malaria exposure and children between 11–14 years of age experienced a significant increase in malaria-related morbidity. Plasmodium induces haematological alterations. It is essential to monitor these alterations in order to assess the impact of malaria control interventions.The estimates of malaria burden are based on malariometric indices, such as prevalence of malaria parasitaemia, spleen rate and anaemia in defined risk groups . ChildhoThere are many preventive measures of malaria both at community and individual level, with the most promising being insecticide-treated bed nets (ITNs) –5. The uPlasmodium falciparum and Plasmodium vivax infection by 50% on average (range 39-62%), as well as reducing the prevalence of high density parasitaemia [A strong association between interventions and their impact on malaria morbidity and mortality has been reported in Sao Tome and Principe . When fuge range 9-62%, aset al.[The study was carried out in Muea village, a semi-rural setting in the Mount Cameroon area. The coordinates of the study area ranged from altitude 540 m, latitude 04°10.464′N, longitude 009°18.168′E to 556 m, 04°10.015′N and 009°18.009′E. The study area has been described in detail by Sumbele Following several control programmes, including the free distribution of ITNs Figure  by the gAt the start of each study, the parents, guardians and children were educated at their various neighbourhoods. During education, the study protocol was explained and the benefits of participating in the study highlighted. Children were enrolled into the study only when parent or guardian signed the assent form or gave a verbal consent. Thereafter the children were clinically evaluated and their parent/caregivers interviewed using a semi-structured questionnaire to obtain information on ITN possession and use. Blood samples were collected for thick and thin smear microscopy as well as a standard full blood count. Labelled blood samples were transported on ice in a cool box of temperature between 8-10°C to the University of Buea Malaria Research Laboratory for further analyses.A semi-structured questionnaire was administered to the participant/parent/guardians/caregiver to obtain information about ITN/LLINs use, its state, various malaria control measures and occurrence of malaria since they started using the nets. Children, who had good nets  that were used regularly, were considered effective users while children who had good nets but used them irregularly or had torn nets were considered as ineffective users.The clinical evaluation carried out by trained medical personnel in their homes consisted in part by the measurement of body temperature and examination of the spleen. Axillary body temperature was measured using a digital thermometer. A child with a body temperature ≥37.5°C was considered febrile. Ages of the children were obtained from their mothers and verified from their birth certificates. The tip of the spleen was felt by pressing the abdomen under the left coastal border and splenomegaly was graded according to the classification of Hackett .R Test Kit  following the manufacturer’s instructions. Also WBC and red blood cell counts (RBC) were determined using the improved Neubauer haemocytometer as described by Cheesbrough [Approximately 4–5 ml of blood sample was collected from the children by venipuncture into 5 ml sterile disposable syringes and dispensed into micro-containers or vacutainers containing ethylenediaminetetraacetate (EDTA) solution. Drops of whole blood were dispensed immediately on slides to prepare blood films. The thick and thin blood smears prepared on glass slides at the time of blood sampling were stained with Giemsa and examined following standard protocols . Parasitesbrough  and Daciesbrough . In 2013 Fever was defined as temperature ≥37.5°C;Plasmodium with an axillary temperature of <37.5°C; Asymptomatic malaria (AM) was defined as the presence of Plasmodium, with an axillary temperature of ≥37.5°C or reported fever in previous 48 hours; Clinical malaria (CM) was defined as the presence of any species of  Parasitaemia was categorized as low , moderate  and high ; A Hb level of <11 g/dL was classified as anaemic and further categorized as mild (Hb between 10.1 and 10.9 g/dL), moderate (Hb between 7.0 and 10.0 g/dL) and severe (Hb <7 g/dL) anaemia , 15; Malarial anaemia (MA) was defined as children with Hb <11 g/dL and malaria parasite positive ; Microcytosis was defined as MCV of less than 73 fl ; Hypochromasia was defined as a MCHC of less than 320 g/L .2). Parasitaemia was log transformed before analysis. Means were compared using independent sample t-test and analysis of variance (ANOVA) where appropriate. The 95% confidence interval (CI) for the difference between two proportions and relative risk reduction were calculated using a Microsoft Excel confidence interval calculator as described by the Newcombe-Wilson method [Data were entered into spreadsheets using Microsoft Excel and analysed with the IBM Statistical package for Social Sciences (SPSS) version 19 . Data were summarized into means and standard deviations (SD), and percentages were used in the evaluation of the descriptive statistics. Proportions were compared using the Chi-square test  ≤14 years with a mean age of 6.4 (7.4) years were examined for the presence of malaria parasites and some malaria-related indices. Two-hundred and twenty (53.5%) of the children were age ≤ five years while 134 (32.6%) and 57 (13.9%) belonged to the six to ten and 11–14 years age groups, respectively. A total of 129  children possessed or used an ITN. Only 86  of them used them effectively. The proportion of effective users in the different age groups was comparable (P = 0.82).In the follow-up study in 2013, 454 children  ≤14 years with a mean age of 6.7 (3.4) years were examined. Of these, 177 (39.0%), 205 (45.2%) and 72 (15.9%) of the children were in the ≤ five, six to ten and 11–14 years age groups, respectively. The majority of the children  in the community possessed or used an ITN. The proportion of effective users of ITN in 2013 increased significantly from 20.9%  in 2006 to 35.2% . The proportion of effective users was significantly highest  in the < five years age group  when compared with the six to ten  and the 11–14 years  age groups.The prevalence of fever, malaria parasitaemia, gametocytaemia, splenomegaly, CM, AM, anaemia, and MA decreased significantly (P <0.05) in 2013 when compared with the baseline data in 2006 , while in the baseline study children in the six to ten years age group had the highest prevalence  as indicated in Table 2 = 18.3, P <0.001) in the youngest group of children while in the follow-up study, the highest prevalence of splenomegaly that approached significance  was observed in the oldest group of children  in the follow-up study  than at the baseline  with an odds ratio of 2.63 (CI = 0.96-7.18).In the baseline survey, the prevalence of splenomegaly was significantly highest  with increased age Table . The higAt baseline a comparison of MA prevalence within the age groups showed no significant difference (P >0.05) while in 2013, the prevalence decreased significantly (P <0.001) with an increase in the age group with children in the 11–14 years age group having a 97.7% (CI = 83.5-99.7%) relative risk reduction in prevalence /μL of blood in 2013 was higher (834.8) than the baseline value (789.9). As shown in Table The disparity in prevalence of low, moderate and high asexual malaria parasite at baseline  and follow-up  was not significant (P = 0.74).Between the baseline survey and the follow-up study, the overall mean Hb concentration significantly increased  from 9.5 (1.7) g/dL (CI = 9.3-9.7 g/dL) to 11.6 (1.9) g/dL (CI = 11.4-11.8 g/dL) with a mean difference of 2.1 g/dL (CI = 1.9-2.4 g/dL). Children who were malaria parasite positive in the follow-up study had significantly higher (P <0.001) mean Hb, Hct, MCH, MCHC, and MCV when compared with their baseline counterpart. On the other hand, the mean RBC was significantly lower in malaria parasite positive children in the follow-up study than those at baseline  decreased significantly (P <0.001) to 24.4, 2.7 and 1.3%, respectively during follow-up as shown in Table The prevalence of microcytosis, hypochromasia and microcytic anaemia (MCV <73 fl and Hb <11 g/dL) decreased significantly (P <0.001) from >55% in 2006 to <29% in 2013  household ITN coverage of at least 60% as reported by previous studies . Howeveret al. [Parasitaemia is one of the indices used in estimating the burden of malaria and the impact of interventions. The prevalence of malaria parasitaemia is on the decline in the Mount Cameroon area as revealed by the study. Overall, the prevalence of malaria parasite obtained in the follow-up study (36.6%) was lower than those obtained at baseline (85.4%) and a similar study conducted in 2012 (44.3%) in this part of the Mount Cameroon area by Kimbi et al. . The achper se but may have acquired immunity that prevented the development of febrile disease as the study revealed an increase in asymptomatic malaria in the age group  than the baseline value (789.9). Furthermore, in the follow-up study, children in the 11–14 years age group had a significantly higher (P = 0.04) GMPD/μL of blood than their baseline equivalent. The highest prevalence of malaria parasite and the significant increase in parasite density observed in children 11–14 years old, unlike in the baseline study, probably reflects the manner of usage of ITNs by children in the study area. Results from the study indicate that while the effective use of ITNs in 2006 was comparable in the different age groups, in 2013 children in the 11–14 years age group registered a significant decrease in usage. Consequently, they were more exposed to infective mosquito bites than the other age groups. This may have resulted in the high prevalence and density of malaria parasite observed in this age group. Hence, children in the 11–14 years age group most likely did not acquire immunity to parasitization e et al.  who repoe et al. , althouge et al. .It is not surprising that children under five years recorded the highest parasitaemia when compared with the older children in spite of their effective use of ITNs. Children under five years are more vulnerable to the disease in areas of high transmission , 25 as nPlasmodium gametocytes, the sexual, non-proliferating blood stage in the life cycle, significantly reduced in the follow-up study. It is plausible that the implementation of a highly gametocytocidal, artemisinin-based combination therapy (ACT) [Anopheles mosquitoes.The prevalence of py (ACT) , namely P. falciparum transmission areas [meso-endemic [P. falciparum[Splenomegaly is the main clinical marker of endemicity in on areas . The min-endemic  in the s-endemic  and a pr-endemic  are knowlciparum, thus prlciparum.The increase in mean Hb concentration from 9.4 (1.8) in the baseline study to 11.3 (1.9) g/dL in the follow-up study, regardless of the status, signifies a great improvement in the health of children in this study area. WHO and RBM recommend that anaemia be used as an additional indicator to monitor malaria burden at community level as interventions are nationally scaled-up . The sigEven though both studies had as a limitation a small number of children in the 11–14 years age group, it is unlikely that the difference in numbers may have introduced a bias as the mean difference in age was of no consequence. The higher prevalence of anaemia in the ≤ five years group with decreasing severity with age is in line with previous studies that anaemia due to malaria is more severe in younger children in areas of intense transmission . Childreet al. [Studies by Korenromp et al.  reportedP falciparum malaria is typically normocytic and normochromic, with a notable absence of reticulocytes, although microcytosis and hypochromia may be present due to the very high frequency of alpha- and beta-thalassaemia traits and/or iron deficiency in many endemic areas [et al. [P. falciparum to be iron deficient. In a related study in the same area, 28% of the children with P. falciparum infections were iron deficient albeit the majority had microcytic anaemia [Results from the baseline survey indicated a prevalence of 56% for microcytic anaemia to be a common finding in the children examined when compared with 7.7% at follow-up. Microcytic anaemia, a surrogate marker of iron deficiency, is also a characteristic feature of anaemia due to malaria . The anaic areas . However [et al.  reported anaemia . It is tThe primary limitation of the study is the lack of information on the variation in consultations with children under five years in health facilities following the commencement of free treatment of malaria as inferences have been made. However, irrespective of the age group, all the children benefitted in the same way from the environmental management instituted to control the breeding of the mosquito vector and the distribution of LLINs.The results from the studies suggest that even though a greater majority of the population is in possession of an ITN, less than half of the population use them effectively. Although malaria is still meso-endemic, most malariometric indices declined and the red cell indices markedly improved. Following interventions, anaemia (moderate to severe) was a more sensitive measure to detect changes in malaria exposure. Children in the 11–14 years age group experienced a significant increase in malaria-related morbidity, and microcytic anaemia was no longer a common finding in children in the area.


Plasmodium chaubaudi infection reduced pre-existing influenza-specific antibodies—raising concerns that malaria may compromise pre-existing vaccine responses. We extended these findings to P. yoelii infection, observing decreases in antibodies to model antigens in inbred mice and to influenza in outbred mice, associated with LLPC depletion and increased susceptibility to influenza rechallenge. We investigated the implications of these findings in Malian children by measuring vaccine-specific IgG  before and after the malaria-free 6-month dry season, 10 days after the first malaria episode of the malaria season, and after the subsequent dry season. On average, vaccine-specific IgG did not decrease following acute malaria. However, in some children malaria was associated with an accelerated decline in vaccine-specific IgG, underscoring the need to further investigate the impact of malaria on pre-existing vaccine-specific antibodies.Vaccine-induced immunity depends on long-lived plasma cells (LLPCs) that maintain antibody levels. A recent mouse study showed that  Plasmodium falciparum infection reduces vaccine efficacy in children [Since the Expanded Program on Immunization was implemented in the mid-1970s , yet ~20children .Most licensed vaccines confer protection through antibodies . Long-liP. falciparum infection are relatively short-lived, particularly in children [Plasmodium blood-stage infection is associated with dysregulated B cell and CD4+ helper T cell function and that this contributes to short-lived antibody responses to malaria [P. falciparum infections—a scenario that would dictate re-evaluation of policies regarding re-immunization frequency in malaria-endemic areas.In contrast, antibody responses to children , which lchildren . Mountin malaria –14. WhatPlasmodium blood-stage infection is deleterious to pre-existing levels of heterologous antibodies. Specifically, Ng et al. showed that Plasmodium chaubaudi (Pcc) blood-stage infection of influenza-immune B6 mice resulted in a transient drop in influenza-specific antibodies and ASCs in bone marrow that, paradoxically, recovered ~21 days post Pcc infection [Pcc model are generalizable to other Plasmodium species in mice of diverse genetic backgrounds and, of clinical relevance, whether acute malaria in children is associated with a decrease in levels of pre-existing antibodies to vaccines.Interestingly, a recent study in mice suggested that nfection . Given tBALB/c and C57BL/6 mice  and Swiss Webster mice (Harlan Laboratories) were housed at the University of Iowa. Female mice were used to initiate experiments at 8–12 weeks of age. Euthanasia was carried out by cervical dislocation. Mouse experiments were approved by The University of Iowa Animal Care and Use Committee.11 /ml were diluted with PBS to 1:200,000 and 50 μl of viral suspension was given intranasally to anesthetized mice.SRBC immunization: 5 ml of Sheep blood  was washed twice with Gibco PBS (Life Technologies). Red blood cells (RBCs) were pelleted at 1000 rpm for 10 min and the volume of the cell pellet measured. One volume of pellet was resuspended in 9 volumes PBS to prepare 10% SRBC suspension. 200 μl of the 10% SRBC suspension was injected intraperitoneally. Influenza virus immunization: Stock A/37/PR/8 (PR8) with TCID titers of 10P. yoelii (Py) 17XNL infected RBCs were diluted with saline such that mice were intravenously injected with 100,000 P. yoelii (Py)-infected RBCs at the time indicated relative to immunization with SRBC or IAV. Parasitemia was monitored by Giemsa stained blood smears. In the indicated experiments, mice were treated on day 4 post infection and every second day thereafter by intraperitoneal injection with 50 mg/kg of chloroquine to prevent high parasite burden. Parasitemia in chlorquine-treated mice did not exceed 4% at any time point during the infection. Listeria monocytogenes (DPL-1942) was appropriately diluted and 5 x 105 cfu were injected intravenously. LCMV Armstrong was appropriately diluted and 2 x 105 pfu injected intraperitoneally. For challenge studies, PR8 stock virus was diluted 1:20,000 and mice were infected intranasally.Cryopreserved 2).Serum samples obtained at the indicated days relative to immunization or infection were serially diluted in PBS and mixed 1:1 with a 1% suspension of SRBC in triplicate in round-bottom 96-well plates. Plates were mixed and incubated at 4°C for 60 minutes then observed for hemagglutination. Hemagglutinin antibody titers are the highest dilution of serum with a positive result. Titers are expressed as logarithms to base 2 .Sera obtained at indicated days relative to immunization and infection were serially diluted in DMEM and 60 μl of each dilution (in triplicate) was mixed with equal volume containing 1000 TCIDOn day three post-challenge infection with PR8, lungs were homogenized in DMEM and stored at −80°C until use. Samples were thawed and virus levels determined as above.+ (104), B220+ (6B2), CD19+ (6D5), IgD- (11-26c.2a), CD138+ (281–2). Apoptosis in plasma cells was determined by detection of active Caspase-3/7 using the VybrantFAMCaspase-3 and Caspase-7 Assay Kit (Invitrogen) according to manufacturer’s protocol. BAFF receptor (R) expression was detected on CD45.2+B220+CD19+IgD-CD138+ cells by staining with anti-BAFF-R mAb (11C1). Serum BAFF levels were quantified by ELISA according to manufacturer’s protocol (R&D Systems).Plasma cells in the spleen and bone marrow were identified with the indicated antibodies as CD45.2The Ethics Committee of the Faculty of Medicine, Pharmacy and Dentistry at the University of Sciences, Technique and Technology of Bamako, and the NIAID/NIH IRB approved this study. Written informed consent was obtained from parents or guardians of participating children.P. falciparum transmission occurs annually (June-December). The cohort was described in detail elsewhere [P. falciparum (by PCR) in May 2012 who had plasma available before (January 2012) and after (May 2012) the 6-month dry season, 10 days after the first malaria episode of the ensuing 6-month malaria season (variable date) and after the subsequent dry season (May 2013). Malaria was defined as ≥2,500 asexual parasites/μL, axillary temperature ≥37.5°C and no other cause of fever by exam. Malaria was detected by passive and weekly active surveillance. Individuals with malaria symptoms and any parasitemia were treated according to Malian National Malaria Control Program guidelines. Blood was drawn by venipuncture into sodium citrate–containing Vacutainer tubes (BD) and plasma separated and cryopreserved.The field study was conducted in Kalifabougou, Mali where lsewhere . Here, wP. falciparum PCR methods were described previously [Thick blood smears stained with Giemsa were counted against 300 leukocytes. Densities were recorded as the number of asexual parasites/microliter blood based on a mean leukocyte count of 7500 cells/μL. eviously .IgG specific for tetanus toxoid, measles and Hepatitis B surface antigen was measured using ELISA kits according to manufacturer’s instructions . Plasma diluted 1:100 was analyzed in duplicate on single plates for each vaccine-specific ELISA. IgG concentrations were calculated by fitting to standard curves generated from standards in each kit and expressed as Units/milliliter.10 (1/2) and fixed-effect slope of age; 95% CIs were obtained by similar calculations using endpoints of the 95% CI of fixed-effect slopes. Statistical significance was defined as 2-tailed P value<0.05. Analyses were performed in R v.2.15.1 (http://www.R-project.org) or Prism 5.0d (GraphPad).Mouse data were analyzed with two-tailed unpaired Student’s t-test using GraphPad Prism software. Humans ELISA data were log-transformed (with base 10). The Wilcoxon rank-sum test was used to compare slopes. Linear mixed-effects models compared rates of change during periods of malaria exposure and non-exposure, where the random intercept accounted for correlations among data points  and fixed effects included time from enrollment and the minimum of 0 and time since end of dry season. Half-lives of IgG levels were estimated by fitting a linear mixed-effects model with three data points: before and after first dry season, and after second dry season, where random intercept accounted for within-subject correlation, and age at visit as fixed effect. IgG half-lives were estimated by the ratio of logPlasmodium infection and whether they reflect malaria-specific or generalized immune system abnormalities. To address this, we immunized BALB/c mice with Sheep Red Blood Cells (SRBC), then infected some mice in each group two days later with 105P. yoelii (Py) 17XNL parasitized red blood cells (pRBC) (Listeria monocytogenes) or viral (lymphocytic choriomeningitis virus) infections did not compromise anti-SRBC antibody responses  . This exs (pRBC) . Importaesponses . Thus, tesponses . Of notePcc blood-stage infection of influenza virus-immune B6 mice resulted in a transient drop in circulating antibody levels and ASCs in bone marrow that, paradoxically, recovered by ~21 days post-infection [Pcc in B6 mice, outbred Swiss Webster mice were sublethally infected with the mouse adapted H1N1 influenza A virus (IAV) A/PR/8 (PR8). At day 40 post-infection, when PR8-specific neutralizing antibody titers had been stable for several weeks, some mice were infected with Py pRBC   . BAFF-ree marrow  of Py ine marrow . The losAlthough antibody titers were decreased by Py infection, they were not eliminated. To determine if compromised antibody response also eliminated MBCs, SRBC-immune mice that were infected or not with Py were subjected to booster immunization with SRBC . BoostinTogether, the observations in mouse models prompted us to investigate the impact of acute malaria on pre-existing antibodies to common vaccines in children. In a longitudinal study in Mali of 54 children aged 2–5 years (mean 3.5 years), we took advantage of the sharply demarcated 6-month rainy season  and 6-month dry season   Fig 5A)Fig 5A) tP. falciparum PCR) at the end of the first dry season. The median time between the end of the first dry season and the first malaria episode was 107 days (IQR 65 days). At the first malaria episode children had an average temperature of 38.4°C (range 37.5–40.3), were infected with P. falciparum  and had no other cause of fever on examination. Malaria was treated with a standard course of artemether/lumefantrine. Demographic/clinical data of subjects are shown in Subjects were asymptomatic and uninfected  indicated that 6.5% of hospital admissions were for tetanus with a mortality rate of 46.2% [Here we provide evidence in mice and some children that cc model  and descof 46.2% . Moreoveof 46.2% , furtherof 46.2% , 26. HowP. falciparum infections earlier in life would accelerate the loss of vaccine-specific IgG in more children and to a greater degree than observed here. Controlled human malaria infections [P. falciparum infections on pre-existing levels of vaccine-specific antibodies.Mice used in this study were malaria-naïve, whereas children had already experienced 2–5 malaria seasons; therefore, it remains possible that fections  could dePlasmodium rather than a general feature of infection. Accordingly, we did not detect a decline in IgG to unrelated antigens in mice after infection with Listeria monocytogenes or LCMV  in humans did not significantly alter levels of pre-existing IgG to unrelated antigens, including tetanus and measles , suggest or LCMV .Plasmodium accelerates the decline of IgG to unrelated antigens remains unclear. It has been suggested that binding of Plasmodium-infected erythrocytes to bone marrow stromal cells may disrupt LLPC survival signals [Py-infected mice . Consistent with this, we observed substantial increases in apoptotic plasma cells in the bone marrow of mice after Py infection  as a T cell–independent polyclonal B cell activator [We observed that acute malaria is associated with a transient increase in vaccine-specific IgG in some children. It is possible that bystander proliferation and differentiation of vaccine-specific MBCs into ASCs  masked tctivator . The facApproximately 3.4 billion people in 97 countries are at risk of malaria . Here we


Human co-infection with malaria and helmimths is ubiquitous throughout Africa. Nevertheless, its public health significance on malaria severity remains poorly understood.Plasmodium species by PCR. Fresh stool specimens were also collected for the identification and quantification of geohelminths by the Kato-Katz method. The prevalence of malaria, geohelminths, and co-infections were 77.2%, 28.6%, and 22.1%, respectively. Plasmodium falciparum was the only malaria parasite species identified with mean parasite density of 111  parasites/µl of blood. The geohelminths found were Ascaris lumbricoides (21.6%) and Trichuris trichiura (10.8%), with mean parasite densities of 243  and 36  eggs/gram of faeces, respectively. Co-infections of A. lumbricoides and P. falciparum were the most frequent and correlated positively. While no significant difference was observed on the prevalences of single and co-infections between the two localities, there was a significant difference in the density of A. lumbricoides infection between the two localities. The overall prevalence of anaemia was 42%, with individuals co-infected with T. trichiura and P. falciparum (60%) being the most at risk. While the prevalence of malaria and anaemia were inversely related to age, children aged 5–14 years were more susceptible to geohelminthiasis and their co-infections with malaria.To contribute to a better understanding of epidemiology and control of this co-infection in Cameroon, a cross-sectional study was carried out to assess the prevalence of concomitant intestinal geohelminthiasis and malaria, and to evaluate its association with malaria and anaemia in Nkassomo and Vian. Finger prick blood specimens from a total of 263 participants aged 1–95 years were collected for malaria microscopy, assessment of haemoglobin levels, and molecular identification of Co-existence of geohelminths and malaria parasites in Nkassomo and Vian enhances the occurrence of co-infections, and consequently, increases the risk for anaemia. Plasmodium falciparum (77.2%) was the only malaria parasite species found in the study population, Ascaris lumbricoides (21.6%) and Trichuris trichiura (10.8%) were the only geohelminths detected. The prevalence of malaria and geohelminth co-infections was 22.1% with co-infections of A. lumbricoides and P. falciparum being the most frequently encountered and associating positively. The prevalence of anaemia was 42%, with the most affected being those with co-infections of T. trichiura and P. falciparum. Schoolchildren had significantly higher risk of geohelminthiasis, and their co-infection is a cause for concern and should strategically be considered in the designing and implementation of their control.Co-infection of malaria and intestinal helminths causes significant and additive problems against the host. In order to contribute to a better understanding of the epidemiology and control of concomitant intestinal geohelminthiasis and malaria infections in Cameroon, a cross-sectional study to assess its prevalence and evaluate its influence on malaria and anaemia was carried out in two rural communities (Nkassomo and Vian) of the Mfou health district. Fresh stool and finger prick blood samples from 263 participants aged 1–95 years were analysed to identify and quantify the geohelminths and malaria parasites respectively. Anaemia was assessed by measuring the haemoglobin levels. Whereas  Plasmodium falciparum is the most prevalent and virulent of the malaria parasites Ascaris lumbricoides, Trichuris trichiura, and Hookworm ssp.) and Schistosoma mansoni are the major helminthic parasites In many afro-tropical countries, parasitic co-existence is common with increased potential for co-infection, which may adversely impact the outcome of the diseases they cause. This study received ethical and administrative authorizations respectively from the IMPM institutional ethics review committee and the competent authorities of the Mfou health district. Participation was strictly voluntary and was dependent on informed written consent by the participant or assent by the parent/guardian for children. All positive cases for malaria and/or intestinal worms were treated for free according to the Cameroonian anti-malarial and anti-helminth treatment guidelines and if necessary referred to the Mfou district hospital for appropriate management. A single dose of Paracetamol and iron tablets was respectively given to participants with fever or anaemia.2. Mfou is a multiethnic community made up of the Ewondo, Bané and Tsinga, with farming being the main economic pursuit. Commonly grown crops include cassava, maize and oil palm The study was carried out in two rural communities (Nkassomo and Vian) of the Mfou health district (4°27′N and 11°38′E), a forest area located in the Mefou-Afamba division, in the Centre region of Cameroon. The climate is typically equatorial with two discontinuous dry and wet seasons. The annual average rainfall measures 2,000 mm with an annual average temperature of 24°C. The population is made up of 71,373 inhabitants  with about 82.7 inhabitants/kmThe study was conducted from February to March 2011 during the long dry season in the Mfou health district. Samples were collected from each household and only from individuals who willingly accepted to participate and provided written and signed informed consent and have lived in the village for at least the past six months. A total of 263 participants from the two rural communities (131 from Nkassomo and 132 from Vian) aged 1 to 95 years old took part in the survey and were divided into 6 groups as follows: pre-school children (<5 yrs), young school children (5–9 yrs), old school children (10–14 yrs), adolescents and young adults (15–24 yrs), adults (25–49 yrs), and aged people (≥50 yrs). Each participant was clinically examined by a medical doctor for febrile symptoms  or any other clinical conditions , and information on age and sex taken prior to specimen collection.Plasmodium DNA isolation and PCR amplification for species determination, and measure the haemoglobin level.Finger prick blood samples collected in heparinised micro-capillary tubes were used to prepare thick and thin smears for malaria microscopy, blot filter papers (Whatmann #3) for The thin and thick blood smears air-dried were fixed in 75% methanol and 50% May-Grunwald (thin film only) and stained using 10% Giemsa solution for both thin and thick films. Slides were then microscopically examined for the presence of malaria parasites by two independent microscopists. Where parasites were found the parasite density was determined by counting the number of parasites against 200 leucocytes. The counts were expressed as the number of parasites per micro litre of blood (p/µl), assuming an average leukocyte count of 8,000 cells/µl of blood Plasmodium DNA and subsequent PCR amplification for the identification of Plasmodium species. Fresh blood specimens from heparinised micro-hematocit tubes were transferred onto Whatmann #3 filter paper and used for the extraction of A drop of capillary blood was set down on a strip of a portable haemoglobinometer  to determine the total haemoglobin level according to the product instructions. Anaemia was considered as total haemoglobin level <11 g/dl of blood and classified according to the WHO definition as either mild, moderate or severe A. lumbricoides, T. trichiura, and Hookworms spp., or larval stage for S. stercoralisPrior to stool sampling, clear instructions and clean well labelled stool collection vials were given to each participant for proper stool collection. Stool smears were prepared using the Kato Katz technique for the morphological identification of geohelminth eggs for Plasmodium densities, geohelminth densities, fever, and anaemia). Threshold for statistical significance was set at P<0.05. Prevalence was defined as the proportion of individuals found harbouring the parasite in question to the sum total of the study population. Co-infection was defined as the simultaneous presence of at least one helminth parasite species plus malaria parasite in the same host, and was classified as either single or mixed, depending on the number of parasites found. Qualitative values were expressed in frequency or percentage, whilst quantitative values were expressed as geometric mean values  or arithmetic mean value ± Standard Deviation (SD).Statistical analysis of the data was performed using SPSS 18.0 software . Chi-square test and One Way ANOVA were used to set the difference in proportions and means respectively, whereas the Pearson logistic regression test was used to establish the correlation between variables .Of the 263 study participants 131 (49.8%) were from Nkassomo and 132 (50.2%) from Vian. Of these, there were 123 (49.6%) males and 139 (53.1%) females. The median age in the study population was 26.1±22.4 yrs . Based on the WHO criteria, the median axial temperature was 37.3±0.5°C  and 91 (34.6%) participants had fever. Although children in the age group of 5–9 yrs had the most cases of fever no difference was observed based on age, sex and locality. A total of two hundred and eighteen participants (82.9%) was found to carry at least one of the identified parasites  parasites per micro litre of blood (P/µl). Based on the study locality, the prevalence of malaria in Nkassomo was 78.6% (n = 103) and 75.8% (n = 100) in Vian. This did not vary significantly in the prevalence. A similar trend was observed for the parasite density with an average parasite load of 125  P/µl in Nkassomo, and 98  P/µl in Vian. Sex did not influence the prevalence and intensity of P. falciparum in both localities. Although all age groups were affected, the peak of malaria prevalence was observed in children less than 9 years old. Only 32.5% (n = 66) of malaria positive cases had fever and with the highest parasite densities  with ensities  and 2. NAscaris lumbricoides, Trichuris trichiura were the only geohelminth parasites detected. Of the 66 (28.6%) participants positive for geohelminth infection, there were 50 (21.6%) A. lumbricoides, 25 (10.8%) T. trichiura. However, there were 9 (3.9%) cases with both infections. The geometric mean of the parasite density for each species was 243  eggs per gram of faeces (eps) for A. lumbricoides, and 36  eps for T. trichiura. There was no observed significant difference in the prevalence of the two parasites between the two localities. However, the parasite densities were higher in Vian with a significant difference observed for A. lumbricoides . With regards to sex and age, the prevalence of A. lumbricoides was similarly distributed and was higher in children between the ages of 5 years and 14 years (p = 0.002), whilst no difference was observed for T. trichiura infection between the males and the females  and by age groups. Overall, school-aged children (5–14 yrs) had the highest prevalence and parasite density of geohelminth infections. However, only the prevalence was significantly different when compared to other age groups (p = 0.008) (= 0.008) .P. falciparum. Two cases of co-infections were observed and classified as either single co-infections of A. lumbricoides and P. falciparum (38 [16.5%]), and of T. trichiura and P. falciparum (20 [8.7%]); or mixed co-infections of the three parasites (7 [3.0%]). As observed with single infections of individual parasites, there was significant difference in the prevalence of co-infections between the two study localities. Co-infections with T. trichiura and P. falciparum was observably higher in males than females (p = 0.009). Similarly the overall co-infection was also higher in males than females (p = 0.025). The highest prevalence of co-infection was observed in children between 5 and 14 years old (p = 0.010). With regards to the parasite densities, single infections with only A. lumbricoides  and only P. falciparum  had generally lower parasite densities compared to when the two occurred as co-infections of A. lumbricoides  and P. falciparum . Although there was a similar observation for mixed co-infections for A. lumbricoides, T. trichiura was associated with low densities of P. falciparum, but the difference was not significant (A. lumbricoides (independent variable) and P. falciparum (dependant variable) in co-infected individuals .With regards to malaria and geohelminth co-infection, a total of 51 (22.1%) participants were infected with at least one geohelminth parasite and nificant . On the nificant  showed aAccording to the WHO classification, out of the 263 participants in the study, 110 had haemoglobin levels less than 11 g/dl of blood, giving an overall anaemia prevalence of 42%. However, most the anaemic cases were classified as moderate anaemia with a mean haemoglobin level of 9.8±0.8 g/dl. Only one case of severe anaemia (0.9%) was observed with a total haemoglobin level of 6.7 g/dl. Fifty three (40.5%) and 57 (43.5%) participants were found to be anaemic in Nkassomo and Vian, respectively. Although anaemia did not differ by locality and sex, the prevalence of anaemia decreased significantly with the age groups (p = 0.003) .T. trichiura, P. falciparum, and A. Lumbricoides, respectively. The prevalence however was observed to increase by up to 71% in co-infection cases  considered as the independent variables with anaemia and fever, considered as the dependent variables. However, these infections were not significant predictors of anaemia and fever in the area.Overall, all anaemic cases were found to carry at least one of the detected parasitic infections. There were 13 (52%), 95 (46.8%), and 19 (38%) cases of anaemia amongst those infected with on cases . AlthougPlasmodium falciparum. The overall observed prevalence of 77.2% is higher than the 40.6% recorded in 2009 amongst schoolchildren in the same health district P. falciparum malaria can reach the severe level  in most regions where malaria is endemic et al.et al.Anopheles mosquitoes and consequently to the development and propagation of Plasmodium parasite for which clinical diagnosis and non-compliance to confirmed diagnosis remain a subject of case study. In fact, this study revealed that only 33% of participants positive to Plasmodium falciparum were symptomatic to malaria. This low symptomatology in malaria could be attributed to low parasitic intensities of P. falciparum and probably the carriage of Ascaris lumbricoides, as most participants infected with both P. falciparum and A. lumbricoides were significantly asymptomatic to malaria  was found. However, as hookworm ova is likely deform and may clear within 24 hours following Kato Katz smear preparation spp. could stand as a possible bias for this study due to the time spent (7–9 hours) between the preparation and examining of the Kato Katz smears. Knowing that A. lumbricoides infection is usually associated with higher parasite intensity in contrast to T. trichiura infection, and according to the WHO classification of helminth parasite densities A. lumbricoides were significantly higher compared to Nkassomo. The proximity of Nkassomo to the lone health facility in the health area could as well be central to this observed difference as there is decreased access of the population of Vian to appropriate drugs in case of infection. Similar results have been observed in two connected villages in previous studies by Nkengazong et al. T. trichiura infection, A. lumbricoides infection was significantly different amongst age groups with school children (5–14 yrs) having the highest prevalence of Ascariasis (p = 0.002) and total geohelminth infection (p = 0.008). This shows that geophagia or soil ingestion is more important in males and school children who are usually in contact with the soil, the main source of contamination for soil-transmitted helminths or geohelminths. Similar results were observed by Hamit et al. Helminth parasites are highly prevalent in rural communities due to poor sanitary conditions prevailing in most of these areas P. falciparum and A. lumbricoides were the main parasite species for malaria and geohelminthiasis and constituted the main co-infection. This was followed by T. trichiura-P. falciparum co-infection and mixed co-infection. Our data showed that co-infection had the same distribution with single infections between localities, and that increasing endemicity of single infections is likely to enhance of the establishment of co-infections. In fact, schoolchildren aged 5–14 years had significantly higher prevalence of co-infection (p = 0.001), due of increased susceptibility of this group to A. lumbricoides as reported elsewhere T. trichiura and P. falciparum was significantly higher in males due to increased susceptibility of men to trichinosisSo far, very little is known about the distribution and impact of intestinal worm co-infections across Cameroon. This study revealed that many malarial patients were infected by at least a geohelminth parasite with overall prevalence of 22.1%. Observably, P. falciparum and A. lumbricoides in their co-infection was significant. However, akin to earlier studies, the observed influence of A. lumbricoides on P. falciparum infection in this study was important though not statistically significant due to a decrease in anti-malarial immunity by concomitant helminth infections Ascariasis due to an increase in malaria intensity A. lumbricoides, T. trichiura parasite was associated with a decrease in intensity of P. falciparum at a non significant level. Nevertheless, interactions between T. trichiura and P. falciparum seemed be higher than those between A. lumbricoides and P. falciparum in the mixed co-infection when compared to single co-infections , nutritional status, socio-economic level, etc. Anaemia did not significantly differ by locality and sex, but total haemoglobin level increased with age, with children less than five years having the highest prevalence of anaemia probably due to their high infection rate for P. falciparum and T. trichiura parasites. However, age was not a predictor of anaemia in our study area in contrast to Nkuo-Akenji et al. in South-West Cameroon T. trichiura and P. falciparum; because A. lumbricoides does not seem to have any impact on haemoglobin level in co-infections as reported by Kouontchou at et al. T. trichiura is associated with chronic loss of blood and iron in gastrointestinal tube, while P. falciparum is associated with bursting and loss of red blood cells during its blood stage, all contributing to haemoglobin loss and anaemia. However, unlike in the southwest region of the country, the intensities of these parasites were not predictors of anaemia in Nkassomo and Vian, Overall, all cases of anaemia were carriers of at least one infection with co-infection being the main parasitic factor of anaemia, followed by single infections of The findings suggest that malaria is hyperendemic in the study localities, and co-exists with geohelminths with their co-infections common amongst schoolchildren. This co-infection constitutes an important risk to anaemia while exacerbating malaria intensity. Though further epidemiological studies are needed to support these observations and assess the mechanisms involved in such interactions, the results provide useful information necessary to design control management strategies for malaria in the context of co-infection.Checklist S1STROBE checklist.(DOC)Click here for additional data file.


Plasmodium and 18% coinfected with intestinal parasites. Protozoan parasites comprised the bulk of the intestinal parasites infections and subjects infected with intestinal parasites were more likely to have malaria. The use of principal component analysis and cluster analysis associated increased levels of IL-6, TNF-α, IL-10, and CRP and low levels of IL-17A predominantly with individuals with malaria alone and coinfected individuals. In contrast, low levels of almost all inflammatory mediators were associated predominantly with individuals uninfected while increased levels of IL-17A were associated predominantly with individuals with intestinal parasites only. In conclusion, our data suggest that, in our population, the infection with intestinal parasites (mainly protozoan) does not modify the pattern of cytokine production in individuals infected with P. falciparum and P. vivax.In Brazil, malaria is prevalent in the Amazon region and these regions coincide with high prevalence of intestinal parasites but few studies explore the interaction between malaria and other parasites. Therefore, the present study evaluates changes in cytokine, chemokine, C-reactive protein, and nitric oxide (NO) concentrations in 264 individuals, comparing plasma from infected individuals with concurrent malaria and intestinal parasites to individuals with either malaria infection alone and uninfected. In the studied population 24% of the individuals were infected with  Plasmodium and intestinal parasites are overlapped over the world; therefore malaria coinfection with intestinal parasites is common in tropical regions of the planet  was added and proteins were subsequently precipitated by trichloroacetic acid . The tube contents were mixed and centrifuged ; two samples of each supernatant were transferred to a flat-bottomed microplate and their absorbencies were read at 520 nm using a microplate reader . NO values were calculated from standard calibration plots [A modified Griess reaction was used to detect nitrite and nitrate  ] and three chemokines  were analyzed using a BioPlex Kit assay  following the manufacturer instructions as described in . Cluster analyses were performed from Z-scores using Euclidean distance metrics and Ward as the linkage algorithm. All analyses were performed using R statistical environment [ lmPerm package [ vegan [ gplots package [To further investigate the relationship between infection groups and cytokine profile, we elaborated a heatmap that relates two hierarchical cluster analyses in a bidimensional plot.ironment . Permuta package  and PCA  [ vegan  and gplo package  was used P. vivax and/or P. falciparum at the time of blood collection were subsequently treated using the chemotherapeutic regimen recommended by the Brazilian Ministry of Health.The study was conducted after obtaining ethical clearance from Fundação Oswaldo Cruz Ethical Committee . Individual oral and written consents were obtained from all participants. Donors positive for P. vivax was more prevalent in both malaria infected only (81.2%) and coinfected with intestinal parasites (75%). Protozoan parasites comprised the bulk of the infections in subjects infected with intestinal parasites only (70.4%) or coinfected with malaria and intestinal parasites (81.2%). The prevalence of intestinal parasites was significantly higher in individuals infected with malaria than with those who were not infected . The most prevalent protozoan was Giardia intestinalis and helminths were Ancylostoma duodenale and Strongyloides stercoralis. Multiple protozoa species were common in both intestinal parasites only (14.5%) and coinfected with malaria (33.3%) while only one subject presented multiple helminthes species infection. Dual, triple, and quadruple protozoan and helminthes infections were observed. Data in  P. vivax malaria was 75% (36) and P. falciparum was 23% (11) whereas 2% (1) had mixed species infection. G. intestinalis was the most prevalent intestinal parasite in both P. falcilparum and P. vivax coinfections. However, in P. vivax coinfected subjects the species of protozoans and helminthes were more diverse  were infected with malaria only, 48 (18.2%) were coinfected with malaria and intestinal parasites, 98 (37%) were infected with intestinal parasites only, and 102 (38.7%) were uninfected with either malaria or intestinal parasites. diverse .P < 0.0001 and uninfected P < 0.05). The analysis of some hematological parameters revealed that malaria and coinfected groups were similar and no differences were observed in the median hemoglobin, platelets, lymphocytes, band cells values, and Plasmodium parasites counts. However, they differ from intestinal parasites and uninfected groups presenting lower mean lymphocytes and platelets counts while band cells counts were higher. Hemoglobin levels in malaria and coinfected groups were slightly higher when compared with the uninfected group.As shown in α, IL-2, IL-10, IL1-β, MCP-1, and IL-6 were observed to be mostly significantly increased compared with those intestinal parasites and uninfected groups . The median levels of IFN-γ and IL-8 were increased in all groups compared to uninfected group. Changes in cytokine and chemokine levels were very similar in malaria and coinfected groups and with this analysis we could not associate changes in cytokine profile that could be due to coinfection.Firstly, we analyzed the median plasma levels of individual cytokines, chemokines, NO, and CRP comparing the concentration between groups . Cytokinβ, IL-6, TNF-α IL-10, and CRP and decreased levels of IL-17A and NO while IP and UN presented higher levels of IL-17A and NO and decreased levels of IL-10 and CRP . This method allows coupled clustering of both the subject and the measured parameters without taking in account the groups. Interestingly, the clustering algorithm could discriminate two major clusters: one cluster included increased levels of IL-6, TNF-α, IL-10, and CRP and low levels of IL-17A predominantly in individuals from M and CI groups and the second cluster included low levels of almost all inflammatory mediators predominantly in individuals from UN group and increased levels of IL-17A predominantly in individuals from IP group.In order to assess differences in the whole multivariate set of cytokines, chemokines, NO, and CRP data in individuals between the groups, we performed an exploratory principal component analysis (PCA), a multivariate technique to identify whether inflammatory mediators could indicate coinfection.  and CRP . In orde and CRP  using th Plasmodium, 55% with intestinal parasites, and 18% with malaria and intestinal parasites. In the studied area, the likelihood of being infected with malaria was significantly higher in individuals infected with intestinal parasites. Several studies from human populations conducted in Africa reveal that helminthic infection can have a negative effect on host response to malaria, including increased susceptibility to Plasmodium infection and increased severity of disease [Overlapping distribution of intestinal parasites and malaria might result in high rate of coinfection. In the studied population 24% of the individuals were infected with disease –14. In o disease , 46–50.Differences in results obtained in different studies might depend on the species of the intestinal parasites and the age of studied population. While most of the coinfection studies are with helminthes and in children , 10, in  Plasmodium coinfection has gained interest because it has been hypothesized that Th2 polarized immune response elicited by helminthes could alter the natural immune response of the host to Plasmodium parasites [β, IL-6, TNF-α IL-10, and CRP and decreased levels of IL-17A while for malaria negative individuals (IP and UN) the profile was high levels of IL-17A, NO and decreased levels of IL-10 and CRP.The influence of intestinal parasites (mainly helminthes) inarasites , 34, 36.arasites , 51. Fewarasites –55. To oα, and IL-6 observed in our analysis in M and CI groups is reported in several studies in acute malaria infections [ P. vivax [The high production of CRP, IL-10, TNF-fections , 56, 57.P. vivax , 50, 56. P. falciparum and Schistosoma haematobium. Higher IFN-γ and similar TNF-α, TGF-β, and IL-10 levels were found when comparing coinfected and single P. falciparum infected children [γ, TNF-α, and TGF-β levels were detected [ S. haematobium when compared to children infected with S. haematobium alone [The role of cytokines production in acute malaria is far from being understood and little is known about their effect in coinfection with other parasites. In our study, the levels of inflammatory mediators in individuals with acute malaria did not differ from individuals coinfected with intestinal parasites when compared with individuals single infected with malaria. However, few reports demonstrated altered cytokines levels in children and adults coinfected withchildren  while, idetected . Elevateum alone . In thes P. falciparum in vitro, and low levels of this marker have been associated with suppression of NO synthesis depending of the gravity in malaria [ Ascaris lumbricoides and Schistosoma mansoni.It is worth it to highlight that, specifically using PCA analysis, we observed the importance of the contribution of IL-17A to separate the M and CI from IP and UN, although IL-17A seems to be higher in IP individuals. It has been observed that IL-17 is an important marker of intestinal inflammation contributing to enhance innate barrier defenses at mucosal surfaces , 59. Int malaria . The con malaria , 60–62.  malaria  detected Plasmodium.In conclusion, our data suggest that, in our population, the infection with intestinal parasites (mainly protozoan) does not modify the pattern of cytokine production in individuals infected with