Absence of Erythrocyte Sequestration and Lack of Multicopy Gene Family Expression in Plasmodium falciparum from a Splenectomized Malaria Patient

BACKGROUND:To avoid spleen-dependent killing mechanisms parasite-infected erythrocytes (IE) of Plasmodium falciparum malaria patients have the capacity to bind to endothelial receptors. This binding also known as sequestration, is mediated by parasite proteins, which are targeted to the erythrocyte surface. Candidate proteins are those encoded by P. falciparum multicopy gene families, such as var, rif, stevor or PfMC-2TM. However, a direct in vivo proof of IE sequestration and expression of multicopy gene families is still lacking. Here, we report on the analysis of IE from a black African immigrant, who received the diagnosis of a malignant lymphoproliferative disorder and subsequently underwent splenectomy. Three weeks after surgery, the patient experienced clinical falciparum malaria with high parasitemia and circulating developmental parasite stages usually sequestered to the vascular endothelium such as late trophozoites, schizonts or immature gametocytes. METHODOLOGY/PRINCIPAL FINDINGS:Initially, when isolated from the patient, the infected erythrocytes were incapable to bind to various endothelial receptors in vitro. Moreover, the parasites failed to express the multicopy gene families var, A-type rif and stevor but expression of B-type rif and PfMC-2TM genes were detected. In the course of in vitro cultivation, the parasites started to express all investigated multicopy gene families and concomitantly developed the ability to adhere to endothelial receptors such as CD36 and ICAM-1, respectively. CONCLUSION/SIGNIFICANCE:This case strongly supports the hypothesis that parasite surface proteins such as PfEMP1, A-type RIFIN or STEVOR are involved in interactions of infected erythrocytes with endothelial receptors mediating sequestration of mature asexual and immature sexual stages of P. falciparum. In contrast, multicopy gene families coding for B-type RIFIN and PfMC-2TM proteins may not be involved in sequestration, as these genes were transcribed in infected but not sequestered erythrocytes

Molecular Evidence for Flea-Borne Rickettsiosis in Febrile Patients from Madagascar.

Rickettsiae may cause febrile infections in humans in tropical and subtropical regions. From Madagascar, no molecular data on the role of rickettsioses in febrile patients are available. Blood samples from patients presenting with fever in the area of the capital Antananarivo were screened for the presence of rickettsial DNA. EDTA (ethylenediaminetetraacetic acid) blood from 1020 patients presenting with pyrexia > 38.5 °C was analyzed by gltA-specific qPCR. Positive samples were confirmed by ompB-specific qPCR. From confirmed samples, the gltA amplicons were sequenced and subjected to phylogenetic analysis. From five gltA-reactive samples, two were confirmed by ompB-specific qPCR. The gltA sequence in the sample taken from a 38-year-old female showed 100% homology with R. typhi. The other sample taken from a 1.5-year-old infant was 100% homologous to R. felis. Tick-borne rickettsiae were not identified. The overall rate of febrile patients with molecular evidence for a rickettsial infection from the Madagascan study site was 0.2% (2/1020 patients). Flea-borne rickettsiosis is a rare but neglected cause of infection in Madagascar. Accurate diagnosis may prompt adequate antimicrobial treatment

Are brucellosis, Q fever and melioidosis potential causes of febrile illness in Madagascar?

Brucellosis, Q fever and melioidosis are zoonoses, which can lead to pyrexia. These diseases are often under-ascertained and underreported because of their unspecific clinical signs and symptoms, insufficient awareness by physicians and public health officers and limited diagnostic capabilities, especially in low-resource countries. Therefore, the presence of Brucella spp., Coxiella burnetii and Burkholderia pseudomallei was investigated in Malagasy patients exhibiting febrile illness. In addition, we analyzed zebu cattle and their ticks as potential reservoirs for Brucella and C. burnetii, respectively. Specific quantitative real-time PCR assays (qPCRs) were performed on 1020 blood samples drawn from febrile patients. In total, 15 samples (1.5%) were Brucella-positive, mainly originating from patients without travel history, while DNA from C. burnetii and Bu. pseudomallei was not detected. Anti-C. burnetii antibodies were found in four out of 201 zebu serum samples (2%), whereas anti-Brucella antibodies could not be detected. Brucella DNA was detected in a single zebu sample. Three out of 330 ticks analyzed (1%) were positively tested for C. burnetii DNA but with high Ct values in the qPCR assay. Our data suggest that zebus as well as Amblyomma and Boophilus ticks have to be considered as a natural reservoir or vector for C. burnetii, but the risk of cattle-to-human transmission is low. Since bovine brucellosis does not seem to contribute to human infections in Madagascar, other transmission routes have to be assumed

Determining the Best Immunization Strategy for Protecting African Children Against Invasive Salmonella Disease.

BACKGROUND: The World Health Organization recently prequalified a typhoid conjugate vaccine (TCV), recommending its use in persons ≥6 months to 45 years residing in typhoid fever (TF)-endemic areas. We now need to consider how TCVs can have the greatest impact in the most vulnerable populations. METHODS: The Typhoid Fever Surveillance in Africa Program (TSAP) was a blood culture-based surveillance of febrile patients from defined populations presenting at healthcare facilities in 10 African countries. TF and invasive non-typhoidal Salmonella (iNTS) disease incidences were estimated for 0-10 year-olds in one-year age increments. RESULTS: Salmonella Typhi and iNTS were the most frequently isolated pathogens; 135 and 94 cases were identified, respectively. Analysis from three countries was excluded (incomplete person-years of observation (PYO) data). Thirty-seven of 123 TF cases (30.1%) and 71/90 iNTS disease cases (78.9%) occurred in children aged <5 years. No TF and 8/90 iNTS infections (8.9%) were observed in infants aged <9 months. The TF incidences (/100 000 PYO) for children aged <1 year and 1 to <2 years were 5 and 39, respectively; the highest incidence was 304 per 100 000 PYO in 4 to <5 year-olds. The iNTS disease incidence in the defined age groups ranged between 81 and 233 per 100 000 PYO, highest in 1 to <2 year-olds. TF and iNTS disease incidences were higher in West Africa. CONCLUSIONS: High burden of TF detected in young children strengthens the need for TCV introduction. Given the concurrent iNTS disease burden, development of a trivalent vaccine against S. Typhi, S. Typhimurium, and S. Enteritidis may be timely in this region

Multicountry Distribution and Characterization of Extended-spectrum β-Lactamase-associated Gram-negative Bacteria From Bloodstream Infections in Sub-Saharan Africa.

BACKGROUND: Antimicrobial resistance (AMR) is a major global health concern, yet, there are noticeable gaps in AMR surveillance data in regions such as sub-Saharan Africa. We aimed to measure the prevalence of extended-spectrum β-lactamase (ESBL) producing Gram-negative bacteria in bloodstream infections from 12 sentinel sites in sub-Saharan Africa. METHODS: Data were generated during the Typhoid Fever Surveillance in Africa Program (TSAP), in which standardized blood cultures were performed on febrile patients attending 12 health facilities in 9 sub-Saharan African countries between 2010 and 2014. Pathogenic bloodstream isolates were identified at the sites and then subsequently confirmed at a central reference laboratory. Antimicrobial susceptibility testing, detection of ESBL production, and conventional multiplex polymerase chain reaction (PCR) testing for genes encoding for β-lactamase were performed on all pathogens. RESULTS: Five hundred and five pathogenic Gram-negative bloodstream isolates were isolated during the study period and available for further characterization. This included 423 Enterobacteriaceae. Phenotypically, 61 (12.1%) isolates exhibited ESBL activity, and genotypically, 47 (9.3%) yielded a PCR amplicon for at least one of the screened ESBL genes. Among specific Gram-negative isolates, 40 (45.5%) of 88 Klebsiella spp., 7 (5.7%) of 122 Escherichia coli, 6 (16.2%) of 37 Acinetobacter spp., and 2 (1.3%) of 159 of nontyphoidal Salmonella (NTS) showed phenotypic ESBL activity. CONCLUSIONS: Our findings confirm the presence of ESBL production among pathogens causing bloodstream infections in sub-Saharan Africa. With few alternatives for managing ESBL-producing pathogens in the African setting, measures to control the development and proliferation of AMR organisms are urgently needed

The genomic epidemiology of multi-drug resistant invasive non-typhoidal Salmonella in selected sub-Saharan African countries

Funder: Swedish International Development Cooperation Agency (SIDA)Funder: Government of Republic of KoreaFunder: US Centers for Disease Control and PreventionBackground: Invasive non-typhoidal Salmonella (iNTS) is one of the leading causes of bacteraemia in sub-Saharan Africa. We aimed to provide a better understanding of the genetic characteristics and transmission patterns associated with multi-drug resistant (MDR) iNTS serovars across the continent. Methods: A total of 166 iNTS isolates collected from a multi-centre surveillance in 10 African countries (2010–2014) and a fever study in Ghana (2007–2009) were genome sequenced to investigate the geographical distribution, antimicrobial genetic determinants and population structure of iNTS serotypes–genotypes. Phylogenetic analyses were conducted in the context of the existing genomic frameworks for various iNTS serovars. Population-based incidence of MDR-iNTS disease was estimated in each study site. Results: Salmonella Typhimurium sequence-type (ST) 313 and Salmonella Enteritidis ST11 were predominant, and both exhibited high frequencies of MDR; Salmonella Dublin ST10 was identified in West Africa only. Mutations in the gyrA gene (fluoroquinolone resistance) were identified in S. Enteritidis and S. Typhimurium in Ghana; an ST313 isolate carrying blaCTX-M-15 was found in Kenya. International transmission of MDR ST313 (lineage II) and MDR ST11 (West African clade) was observed between Ghana and neighbouring West African countries. The incidence of MDR-iNTS disease exceeded 100/100 000 person-years-of-observation in children aged <5 years in several West African countries. Conclusions: We identified the circulation of multiple MDR iNTS serovar STs in the sampled sub-Saharan African countries. Investment in the development and deployment of iNTS vaccines coupled with intensified antimicrobial resistance surveillance are essential to limit the impact of these pathogens in Africa

Incidence of invasive salmonella disease in sub-Saharan Africa: a multicentre population-based surveillance study.

BACKGROUND: Available incidence data for invasive salmonella disease in sub-Saharan Africa are scarce. Standardised, multicountry data are required to better understand the nature and burden of disease in Africa. We aimed to measure the adjusted incidence estimates of typhoid fever and invasive non-typhoidal salmonella (iNTS) disease in sub-Saharan Africa, and the antimicrobial susceptibility profiles of the causative agents. METHODS: We established a systematic, standardised surveillance of blood culture-based febrile illness in 13 African sentinel sites with previous reports of typhoid fever: Burkina Faso (two sites), Ethiopia, Ghana, Guinea-Bissau, Kenya, Madagascar (two sites), Senegal, South Africa, Sudan, and Tanzania (two sites). We used census data and health-care records to define study catchment areas and populations. Eligible participants were either inpatients or outpatients who resided within the catchment area and presented with tympanic (≥38·0°C) or axillary temperature (≥37·5°C). Inpatients with a reported history of fever for 72 h or longer were excluded. We also implemented a health-care utilisation survey in a sample of households randomly selected from each study area to investigate health-seeking behaviour in cases of self-reported fever lasting less than 3 days. Typhoid fever and iNTS disease incidences were corrected for health-care-seeking behaviour and recruitment. FINDINGS: Between March 1, 2010, and Jan 31, 2014, 135 Salmonella enterica serotype Typhi (S Typhi) and 94 iNTS isolates were cultured from the blood of 13 431 febrile patients. Salmonella spp accounted for 33% or more of all bacterial pathogens at nine sites. The adjusted incidence rate (AIR) of S Typhi per 100 000 person-years of observation ranged from 0 (95% CI 0-0) in Sudan to 383 (274-535) at one site in Burkina Faso; the AIR of iNTS ranged from 0 in Sudan, Ethiopia, Madagascar (Isotry site), and South Africa to 237 (178-316) at the second site in Burkina Faso. The AIR of iNTS and typhoid fever in individuals younger than 15 years old was typically higher than in those aged 15 years or older. Multidrug-resistant S Typhi was isolated in Ghana, Kenya, and Tanzania (both sites combined), and multidrug-resistant iNTS was isolated in Burkina Faso (both sites combined), Ghana, Kenya, and Guinea-Bissau. INTERPRETATION: Typhoid fever and iNTS disease are major causes of invasive bacterial febrile illness in the sampled locations, most commonly affecting children in both low and high population density settings. The development of iNTS vaccines and the introduction of S Typhi conjugate vaccines should be considered for high-incidence settings, such as those identified in this study. FUNDING: Bill & Melinda Gates Foundation

The phylogeography and incidence of multi-drug resistant typhoid fever in sub-Saharan Africa.

There is paucity of data regarding the geographical distribution, incidence, and phylogenetics of multi-drug resistant (MDR) Salmonella Typhi in sub-Saharan Africa. Here we present a phylogenetic reconstruction of whole genome sequenced 249 contemporaneous S. Typhi isolated between 2008-2015 in 11 sub-Saharan African countries, in context of the 2,057 global S. Typhi genomic framework. Despite the broad genetic diversity, the majority of organisms (225/249; 90%) belong to only three genotypes, 4.3.1 (H58) (99/249; 40%), 3.1.1 (97/249; 39%), and 2.3.2 (29/249; 12%). Genotypes 4.3.1 and 3.1.1 are confined within East and West Africa, respectively. MDR phenotype is found in over 50% of organisms restricted within these dominant genotypes. High incidences of MDR S. Typhi are calculated in locations with a high burden of typhoid, specifically in children aged <15 years. Antimicrobial stewardship, MDR surveillance, and the introduction of typhoid conjugate vaccines will be critical for the control of MDR typhoid in Africa

A Qualitative Study Investigating Experiences, Perceptions, and Healthcare System Performance in Relation to the Surveillance of Typhoid Fever in Madagascar.

BACKGROUND: The burden of typhoid fever (TF) in sub-Saharan Africa is largely unknown but is increasingly thought to be high, given that water and sanitary conditions remain unimproved in many countries. To address this gap in information, the Typhoid Fever Surveillance in Africa Program (TSAP) founded a surveillance system for TF in 10 African countries. This study was a component of the TSAP surveillance project in Madagascar. METHODS: The study entailed a qualitative assessment of patients' experiences and perceptions of services for febrile symptoms at the studies' rural and urban sentinel public health clinics. The study examined influences on the use of these facilities, alternative sources of care, and providers' descriptions of medical consultations and challenges in providing services. Data were collected through semistructured and open-ended individual interviews and a focus group with patients, caregivers, and medical personnel. RESULTS: Thirty-three patients and 12 healthcare providers participated in the data collection across the 2 healthcare facilities. The quality of services, cost, and travel distance were key factors that enabled access to and use of these clinics. Divergent healthcare-seeking patterns were related to variability in the care utilized, socioeconomic status, and potential distance from the facilities : These factors influenced delivery of care, patient access, and the health facilities' capacity to identify cases of febrile illness such as TF. CONCLUSIONS: This approach provided an in-depth investigation and understanding of healthcare-seeking behavior at the study facilities, and factors that facilitated or acted as barriers to their use. Our findings demonstrate the relevance of these public health clinics as sites for the surveillance of TF in their role as central healthcare sources for families and communities within these rural and urban areas of Madagascar