Hepatoprotective effect of methanol fruit extract of Punica granatum L in highly active antiretroviral therapy-induced toxicity in Wistar rats.

Prolonged use of Highly Active Antiretroviral Therapy (HAART) has been linked to toxicity, particularly hepatotoxicity. There are few effective drugs for HAART patients that promote hepatic cell regeneration and prevent liver injury. Therefore, the purpose of this study was to investigate the hepato-protective activity of Methanol fruit extract of Punica granatum (MFEPG) in HAART-administered rats. Thirty rats weighing between 150-200 g were randomly divided into six groups and each group comprised of five rats. Distilled water was given to the rats in group one. Only HAART was given to the rats in group two. MFEPG at doses of 100 and 400 mg/kg was given to the rats in groups three and four. MFEPG dosages of 100 and 400 mg/kg along with HAART were given to the rats in groups five and six, respectively. All treatments were via oral gavage daily for 40 days. Under halothane anesthesia, all rats were sacrificed on day 41. Liver tissues were utilized for lipid peroxidation marker; Malondialdehyde (MDA), antioxidant enzymes; Superoxide dismutase (SOD) and Catalase (CAT) and histological evaluation, while blood samples were examined for biochemical parameters (AST, ALT, ALP, Total cholesterol, Total protein, and Albumin). The HAART-treated group exhibited a significantly higher amount of the lipid peroxidation end product; MDA, and significantly lower levels of antioxidant enzymes; SOD, and CAT. Liver enzymes and total cholesterol were significantly increased with a significant reduction in Total protein and Albumin levels in the HAART-treated group. Conversely, the liver function biomarkers were returned to normal levels in the HAART and MFEPG-treated groups. Histopathological studies revealed that when HAART-exposed rats were treated with MFEPG, both the biochemical and histological results significantly improved. Thus, the antioxidant activity of MFEPG provides protection against HAART-induced liver oxidative damage. More research is needed to determine the safety of using MFEPG in humans.

Plasma Neuron-Specific Enolase is not a reliable biomarker for staging Trypanosoma brucei rhodesiense sleeping sickness patients.

OBJECTIVE: Currently, the only available staging criterion for T. b. rhodesiense requires a lumber puncture to collect and later examine cerebrospinal fluid (CSF). This study examined the potential of plasma Neuron-Specific Enolase (NSE) in discriminating between early and late-stage patients. RESULTS: When median NSE levels were compared between early and late-stage patients, results showed a significant (P < 0.02) upregulation among late-stage patients (599.8 ng/mL). No significant differences (P > 0.9) in NSE levels were observed between early-stage patients (300 ng/mL) and controls (454 ng/mL). We used Receiver Operator Characteristic (ROC) curves to explore the likelihood of using plasma NSE as a potential stage biomarker in discriminating between early and late-stage HAT patients. Our results showed that NSE demonstrated an area under the curve (AUC) of 0.702 (95% CI 0.583-0.830). A high staging accuracy for NSE was obtained by using a cutoff of > 346.5 ng/mL with a sensitivity of 68.6% (95% CI 55-79.7%) and a specificity of 93.3% (95% CI 70.2-99.7%). Although our results demonstrate that plasma NSE is upregulated in T. b. rhodesiense sleeping sickness patients, its value in discriminating between late and early-stage patients is limited. However, future studies could consider improving its specificity by combining it with other identified plasma biomarkers.

Prevalence of pathogenic Klebsiella pneumoniae based on PCR capsular typing harbouring carbapenemases encoding genes in Uganda tertiary hospitals.

BACKGROUND: Klebsiella pneumoniae is an opportunistic pathogen that has been implicated as one of commonest cause of hospital and community acquired infections. The K. pneumoniae infections have considerably contributed to morbidity and mortality in patients with protracted ailments. The capacity of K. pneumoniae to cause diseases depends on the presence of an array virulence factors. Coexistence and expression of virulence factors and genetic determinants of antibiotic resistance complicates treatment outcomes. Thus, emergence of pathogenic MDR K. pneumoniae poses a great threat to the healthcare system. However, the carriage of antibiotic resistance among pathogenic K. pneumoniae is yet to be investigated in Uganda. We sought to investigate the carbapenem resistance profiles and pathogenic potential based on capsular serotypes of K. pneumoniae clinical isolates. METHODS: This was a cross sectional study involving use of archived Klebsiella pneumoniae isolates collected between January and December, 2019 at four tertiary hospitals in Uganda. All isolates were subject to antimicrobial susceptibility assays to determine phenotypic antibiotic resistance, pentaplex PCR to detect carbapenemases encoding genes and heptaplex PCR to identify capsular serotypes K1, K2, K3, K5, K20, K54 and K57. RESULTS: The study found an overall phenotypic carbapenem resistance of 23.3% (53/227) and significantly higher genotypic resistance prevalence of 43.1% (98/227). Over all, the most prevalent gene was blaOXA-48-like (36.4%), followed by blaIMP-type (19.4%), blaVIM-type (17.1%), blaKPC-type (14.0%) and blaNDM-type (13.2%). blaVIM-type and blaOXA-48-like conferred phenotypic resistance in all isolates and 38.3% of isolates that harbored them respectively. Capsular multiplex PCR revealed that 46.7% (106/227) isolates were pathogenic and the predominantly prevalent pathotype was K5 (18.5%) followed by K20 (15.1%), K3 (7.1%), K2 (3.1%) and K1 (2.2%). Of the 106 capsular serotypes, 37 expressed phenotypic resistance; thus, 37 of the 53 carbapenem resistant K. pneumoniae were pathogenic. CONCLUSION: The high prevalence of virulent and antibiotic resistant K. pneumoniae among clinical isolates obtained from the four tertiary hospital as revealed by this study pose a great threat to healthcare. Our findings underline the epidemiological and public health risks and implications of this pathogen.

Methicillin-Resistant Staphylococcus aureus Nasal Colonization among Healthcare Workers at Kampala International University Teaching Hospital, Southwestern Uganda.

Whereas Staphylococcus aureus is a pathogen, it colonizes healthy people as normal flora without causing any symptoms or illness. Probably because of greater exposure, healthcare workers (HCWs) are more colonized, serving as reservoir for endogenous infections as well as dissemination. In developing countries including Uganda, there is scarcity of the literature on S. aureus carriage among HCWs, making infection control difficult. This study aimed at determining the nasal carriage rate and comparing the antimicrobial susceptibility profiles of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible Staphylococcus aureus (MSSA) isolates from HCWs of Kampala International University Teaching Hospital. Nasal swab specimens from HCWs were screened for MRSA using both phenotypic and genotypic methods. Antimicrobial susceptibility testing of the MRSA and MSSA isolates was performed using the Kirby-Bauer disc diffusion method. Out of the 97 participants, 28 (28.8%) participants were nasal carriers of S. aureus of which 13 (46.4%) were phenotypically MRSA (resistant to cefoxitin) and 8 (28.6%) were genotypically MRSA (had mecA gene). Only 6 isolates of the 13 isolates (46%) which showed resistance to cefoxitin had mecA gene detectable while 2 (13.3%) of the 15 cefoxitin susceptible isolates were found to carry mecA gene. The study thus shows that methicillin resistance in S. aureus may not only be determined by mecA gene.

Cytokine profiles among patients co-infected with Plasmodium falciparum malaria and soil borne helminths attending Kampala International University Teaching Hospital, in Uganda.

BACKGROUND: Malaria and helminths share the same geographical distribution in tropical Africa. Studies of the interaction of helminth and malaria co-infection in humans have been few and are mainly epidemiological, with little information on cellular immune responses. This study aimed to determine Cytokine profiles among patients co-infected with Plasmodium falciparum malaria and soil borne helminth attending Kampala International University Teaching Hospital (KIU). METHODS: A case control study of 240 patients were recruited at KIU teaching hospital. Patients with Plasmodium falciparum malaria were 55 (22.9%) and those with soil-borne helminths were 63 (26.3%). The controls were 89 (37.1%), while those co-infected with Plasmodium falciparum malaria and soil-borne helminths were 33 (13.8%). Cases were defined as having a positive blood smear for P. falciparum malaria, those with helminths or co-infections of the two. Negative controls were those with a negative blood smear for P. falciparum malaria and those with no stool parasitic infections. Patients presenting with signs and symptoms of malaria or those suspected of having helminths were recruited for the study. A panel of five cytokines (IFN-γ, TNF-α, IL-6, TGF-ß and IL-10) were assayed from plasma samples in patients with and without Plasmodium falciparum malaria, patients with and without helminth, and then those co-infected with the two diseases diagnosis was done using thick blood smears stained with 10% Giemsa and stool examination was done following the Kato Katz technique following standard procedures. RESULTS: The prevalence of Plasmodium falciparum malaria by sex was 28 (11.7%) and 27 (11.3%) in male and female respectively. The overall prevalence of soil borne helminth was 26.3%, and among those harbouring helminths, 13.8% were co-infected with Plasmodium falciparum. Cytokine levels significantly differed across Plasmodium falciparum malaria, soil borne helminth infected patients and health controls for IFN-γ (P = 0.023), IL-10 (P = 0.008) and TGF-ß (P = 0.0001). Cytokine levels significantly differed across Plasmodium falciparum malaria, soil borne helminth infected patients and patients co-infected with Plasmodium falciparum malaria and soil borne helminth for IL-10 (P = 0.004), IL-6 (P = 0.011) and TGF-ß (P = 0.003). CONCLUSION: An up-regulation of IFN-γ during Plasmodium falciparum malaria and an up-regulation of IL-10 and TGF-ß in soil borne helminth infections was demonstrated. We demonstrate that co-infections of Plasmodium falciparum and soil borne helminth lead to an up-regulation of IL-10 and IL-6 and a down-regulation of TGF-ß.Trial registration No17/10-16.

Relationship between Trypanosoma brucei rhodesiense genetic diversity and clinical spectrum among sleeping sickness patients in Uganda.

OBJECTIVE: Human African trypanosomiasis (HAT) due to Trypanosoma brucei rhodesiense in East and southern Africa is reported to be clinically diverse. We tested the hypothesis that this clinical diversity is associated with a variation in trypanosome genotypes. RESULTS: Trypanosome DNA isolated from HAT patients was genotyped using 7 microsatellite markers directly from blood spotted FTA cards following a whole genome amplification. All markers were polymorphic and identified 17 multi-locus genotypes with 56% of the isolates having replicate genotypes. We did not observe any significant clustering between isolates and bootstrap values across major tree nodes were insignificant. When genotypes were compared among patients with varying clinical presentation or outcome, replicate genotypes were observed at both extremes showing no significant association between genetic diversity and clinical outcome. Our study shows that T. b. rhodesiense isolates are homogeneous within a focus and that observed clinical diversity may not be associated with parasite genetic diversity. Other factors like host genetics and environmental factors might be involved in determining clinical diversity. Our study may be important in designing appropriate control measures that target the parasite.

Population genetic structure and temporal stability among Trypanosoma brucei rhodesiense isolates in Uganda.

BACKGROUND: The population structure and role of genetic exchange in African trypanosomes have been previously analyzed albeit with contradictory findings. To further investigate the role of genetic polymorphism on the population genetic structure of Trypanosoma b. rhodesiense, we hypothesized that parasite genotypes are clonal and stable over time. METHODS: We have undertaken a microsatellite marker analysis of T. b. rhodesiense isolates in a relatively new active HAT focus in Uganda (Kaberamaido-Dokolo-Amolatar) over a six-year period (2006-2012). We amplified six microsatellite markers by PCR directly from blood spotted FTA cards following whole genome amplification. RESULTS: The majority of loci demonstrated an excess of heterozygosity (Ho > He, F(IS) < 0). We identified 26 unique genotypes among the 57 isolates, accounting for 45.6% genotypic polymorphism. The presence of a high proportion of samples with repeated genotypes (54.4%, 31/57), disagreement with Hardy-Weinberg equilibrium, and significant linkage disequilibrium between loci pairs, provide evidence that T. b. rhodesiense isolates from this focus are clonal. Our results show low values of F(ST)' (0-0.115) indicating negligible genetic differentiation across temporal isolates. Furthermore, predominant genotypes isolated in 2006 were still detectable in 2012. CONCLUSIONS: Our findings confirm the notion that endemicity is maintained by stable genotypes rather than an influx of new genotypes. Our results have considerable importance in understanding and tracking the spread of sleeping sickness with significant implication to disease control.

The role of cytokines in the pathogenesis and staging of Trypanosoma brucei rhodesiense sleeping sickness.

Human African trypanosomiasis due to Trypanosoma brucei rhodesiense is invariably fatal if untreated with up to 12.3 million people at a risk of developing the disease in Sub-Saharan Africa. The disease is characterized by a wide spectrum of clinical presentation coupled with differences in disease progression and severity. While the factors determining this varied response have not been clearly characterized, inflammatory cytokines have been partially implicated as key players. In this review, we consolidate available literature on the role of specific cytokines in the pathogenesis of T. b. rhodesiense sleeping sickness and further discuss their potential as stage biomarkers. Such information would guide upcoming research on the immunology of sleeping sickness and further assist in the selection and evaluation of cytokines as disease stage or diagnostic biomarkers.

Interleukin (IL)-6 and IL-10 Are Up Regulated in Late Stage Trypanosoma brucei rhodesiense Sleeping Sickness.

BACKGROUND: Sleeping sickness due to Trypanosoma brucei rhodesiense has a wide spectrum of clinical presentations coupled with differences in disease progression and severity across East and Southern Africa. The disease progresses from an early (hemo-lymphatic) stage to the late (meningoencephalitic) stage characterized by presence of parasites in the central nervous system. We hypothesized that disease progression and severity of the neurological response is modulated by cytokines. METHODS: A total of 55 sleeping sickness cases and 41 healthy controls were recruited passively at Lwala hospital, in Northern Uganda. A panel of six cytokines (IFN-γ, IL1-ß, TNF-α, IL-6, TGF-ß and IL-10) were assayed from paired plasma and cerebrospinal fluid (CSF) samples. Cytokine concentrations were analyzed in relation to disease progression, clinical presentation and severity of neurological responses. RESULTS: Median plasma levels (pg/ml) of IFN-γ (46.3), IL-6 (61.7), TGF-ß (8755) and IL-10 (256.6) were significantly higher in cases compared to controls (p< 0.0001). When early stage and late stage CSF cytokines were compared, IL-10 and IL-6 were up regulated in late stage patients and were associated with a reduction in tremors and cranioneuropathy. IL-10 had a higher staging accuracy with a sensitivity of 85.7% (95% CI, 63.7%-97%) and a specificity of 100% (95% CI, 39.8%-100%) while for IL-6, a specificity of 100% (95% CI, 47.8%-100%) gave a sensitivity of 83.3% (95% CI, 62.2%-95.3%). CONCLUSION: Our study demonstrates the role of host inflammatory cytokines in modulating the progression and severity of neurological responses in sleeping sickness. We demonstrate here an up-regulation of IL-6 and IL-10 during the late stage with a potential as adjunct stage biomarkers. Given that both cytokines could potentially be elevated by other CNS infections, our findings should be further validated in a large cohort of patients including those with other inflammatory diseases such as cerebral malaria.

Clinical profiles, disease outcome and co-morbidities among T. b. rhodesiense sleeping sickness patients in Uganda.

BACKGROUND: The acute form of Human African Trypanosomiasis (HAT, also known as Sleeping sickness) caused by Trypanosoma brucei rhodesiense has been shown to have a wide spectrum of focus specific clinical presentation and severity in East and Southern Africa. Indeed HAT occurs in regions endemic for other tropical diseases, however data on how these co-morbidities might complicate the clinical picture and affect disease outcome remains largely scanty. We here describe the clinical presentation, presence of co-infections, and how the latter impact on HAT prognosis. METHODS AND FINDINGS: We carried out a retrospective analysis of clinical data from 258 sleeping sickness patients reporting to Lwala hospital between 2005 and 2012. The mean patient age was 28.6 years with a significant number of cases below 18 years (p< 0.0001). About 93.4% of the cases were diagnosed as late stage (p< 0.0001). The case fatality rate was 10.5% with post treatment reactive encephalopathys reported in 7.9% of the cases, of whom 36.8% eventually died. Fever was significantly (p = 0.045) higher in patients under 18 years. Of the early stage patients, 26.7% and 6.7% presented with late stage signs of sleep disorder and mental confusion respectively. Among the co-infections, malaria was significantly more prevalent (28.9%; p< 0.0001) followed by urinary tract infections (4.2%). Co-infections were present in 14.3% of in-hospital deaths, 38.5% of which were recorded as Malaria. Malaria was significantly more common in patients under 18 years (45.5%; p< 0.02), and was reported in 60% of the fatal cases in this age group. CONCLUSIONS: We show a wide spectrum of sleeping sickness clinical presentation and disease outcome that was apparently not significantly influenced by concurrent infections. It would thus be interesting to determine the host and/or parasite factors that might be responsible for the observed diverse clinical presentation.