RESUMO
BACKGROUND: To retain the spread of SARS-CoV-2, fast, sensitive and cost-effective testing is essential, particularly in resource limited settings (RLS). Current standard nucleic acid-based RT-PCR assays, although highly sensitive and specific, require transportation of samples to specialised laboratories, trained staff and expensive reagents. The latter are often not readily available in low- and middle-income countries and this may significantly impact on the successful disease management in these settings. Various studies have suggested a SARS-CoV-2 loop mediated isothermal amplification (LAMP) assay as an alternative method to RT-PCR. METHODS: Four previously published primer pairs were used for detection of SARS-CoV-2 in the LAMP assay. To determine optimal conditions, different temperatures, sample input and incubation times were tested. Ninety-three extracted RNA samples from St. George's Hospital, London, 10 non-extracted nasopharyngeal swab samples from Great Ormond Street Hospital for Children, London, and 92 non-extracted samples from Queen Elisabeth Central Hospital (QECH), Malawi, which have previously been tested for SARS-Cov-2 by quantitative reverse-transcription RealTime PCR (qRT-PCR), were analysed in the LAMP assay. RESULTS: In this study we report the optimisation of an extraction-free colourimetric SARS-CoV-2 LAMP assay and demonstrated that a lower limit of detection (LOD) between 10 and 100 copies/µL of SARS-CoV-2 could be readily detected by a colour change of the reaction within as little as 30 min. We further show that this assay could be quickly established in Malawi, as no expensive equipment is necessary. We tested 92 clinical samples from QECH and showed the sensitivity and specificity of the assay to be 86.7% and 98.4%, respectively. Some viral transport media, used routinely to stabilise RNA in clinical samples during transportation, caused a non-specific colour-change in the LAMP reaction and therefore we suggest collecting samples in phosphate buffered saline (which did not affect the colour) as the assay allows immediate sample analysis on-site. CONCLUSION: SARS-CoV-2 LAMP is a cheap and reliable assay that can be readily employed in RLS to improve disease monitoring and management.
Assuntos
COVID-19 , SARS-CoV-2 , COVID-19/diagnóstico , Criança , Humanos , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico/métodos , RNA , SARS-CoV-2/genéticaRESUMO
AIMS: This study evaluated detection methods for Salmonella Typhi (S. Typhi) in the environment, to establish a novel pathway from field sampling to isolation of viable organisms and molecular confirmation from complex environmental samples, thus enabling environmental surveillance of typhoid. METHODS AND RESULTS: Multiple media were assessed using clinical isolates from the Public Health England's (PHE) Culture collection. The culture pathway selected consisted of a primary 2% bile broth and secondary Selenite F broth, followed by modified Chromogenic Agar for Salmonella Esterase (mCASE). A qPCR assay was adapted from a validated S. Typhi PCR panel for confirmation of isolates, with comparison to biochemical and serological tests showing good specificity. Sampling locations in Blantyre, Malawi were used to compare sampling methods. Viable S. Typhi were isolated from a mixture of trap and grab river water samples on six occasions. CONCLUSIONS: Culture of viable S. Typhi from environmental samples was possible using effective capture and culture techniques. SIGNIFICANCE AND IMPACT OF STUDY: Whilst several studies have attempted to detect S. Typhi from the environment, this is the first successful attempt to isolate the organism from river water since the 1980s. Supplementing clinical data with environmental screening offers the potential for enhanced surveillance, which might inform interventions and assess vaccination programmes.
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Salmonella typhi , Febre Tifoide , Humanos , Reação em Cadeia da Polimerase em Tempo Real , Salmonella , Salmonella typhi/genética , Manejo de Espécimes , Febre Tifoide/diagnósticoRESUMO
BACKGROUND: Typhoid fever remains a major source of morbidity and mortality in low-income settings. Its most feared complication is intestinal perforation. However, due to the paucity of diagnostic facilities in typhoid-endemic settings, including microbiology, histopathology, and radiology, the etiology of intestinal perforation is frequently assumed but rarely confirmed. This poses a challenge for accurately estimating burden of disease. METHODS: We recruited a prospective cohort of patients with confirmed intestinal perforation in 2016 and performed enhanced microbiological investigations (blood and tissue culture, plus tissue polymerase chain reaction [PCR] for Salmonella Typhi). In addition, we used a Poisson generalized linear model to estimate excess perforations attributed to the typhoid epidemic, using temporal trends in S. Typhi bloodstream infection and perforated abdominal viscus at Queen Elizabeth Central Hospital from 2008-2017. RESULTS: We recruited 23 patients with intraoperative findings consistent with intestinal perforation. 50% (11/22) of patients recruited were culture or PCR positive for S. Typhi. Case fatality rate from typhoid-associated intestinal perforation was substantial at 18% (2/11). Our statistical model estimates that culture-confirmed cases of typhoid fever lead to an excess of 0.046 perforations per clinical typhoid fever case (95% CI, .03-.06). We therefore estimate that typhoid fever accounts for 43% of all bowel perforation during the period of enhanced surveillance. CONCLUSIONS: The morbidity and mortality associated with typhoid abdominal perforations are high. By placing clinical outcome data from a cohort in the context of longitudinal surgical registers and bacteremia data, we describe a valuable approach to adjusting estimates of the burden of typhoid fever.
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Epidemias , Perfuração Intestinal , Febre Tifoide , Humanos , Perfuração Intestinal/epidemiologia , Malaui , Estudos Prospectivos , Salmonella typhi , Febre Tifoide/complicações , Febre Tifoide/epidemiologiaRESUMO
OBJECTIVES: ESBL-producing Klebsiella pneumoniae (KPN) pose a major threat to human health globally. We carried out a WGS study to understand the genetic background of ESBL-producing KPN in Malawi and place them in the context of other global isolates. METHODS: We sequenced genomes of 72 invasive and carriage KPN isolates collected from patients admitted to Queen Elizabeth Central Hospital, Blantyre, Malawi. We performed phylogenetic and population structure analyses on these and previously published genomes from Kenya (nâ=â66) and from outside sub-Saharan Africa (nâ=â67). We screened for presence of antimicrobial resistance (AMR) genetic determinants and carried out association analyses by genomic sequence cluster, AMR phenotype and time. RESULTS: Malawian isolates fit within the global population structure of KPN, clustering into the major lineages of KpI, KpII and KpIII. KpI isolates from Malawi were more related to those from Kenya, with both collections exhibiting more clonality than isolates from the rest of the world. We identified multiple ESBL genes, including blaCTX-M-15, several blaSHV, blaTEM-63 and blaOXA-10, and other AMR genes, across diverse lineages of the KPN isolates from Malawi. No carbapenem resistance genes were detected; however, we detected IncFII and IncFIB plasmids that were similar to the carbapenem resistance-associated plasmid pNDM-mar. CONCLUSIONS: There are multiple ESBL genes across diverse KPN lineages in Malawi and plasmids in circulation that are capable of carrying carbapenem resistance. Unless appropriate interventions are rapidly put in place, these may lead to a high burden of locally untreatable infection in vulnerable populations.
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Genoma Bacteriano , Genômica , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/classificação , Klebsiella pneumoniae/genética , beta-Lactamases/genética , Antibacterianos/farmacologia , Biologia Computacional/métodos , Farmacorresistência Bacteriana Múltipla , Variação Genética , Genômica/métodos , Humanos , Klebsiella pneumoniae/isolamento & purificação , Malaui , Testes de Sensibilidade Microbiana , FilogeniaRESUMO
Objectives: Efforts to treat Escherichia coli infections are increasingly being compromised by the rapid, global spread of antimicrobial resistance (AMR). Whilst AMR in E. coli has been extensively investigated in resource-rich settings, in sub-Saharan Africa molecular patterns of AMR are not well described. In this study, we have begun to explore the population structure and molecular determinants of AMR amongst E. coli isolates from Malawi. Methods: Ninety-four E. coli isolates from patients admitted to Queen's Hospital, Malawi, were whole-genome sequenced. The isolates were selected on the basis of diversity of phenotypic resistance profiles and clinical source of isolation (blood, CSF and rectal swab). Sequence data were analysed using comparative genomics and phylogenetics. Results: Our results revealed the presence of five clades, which were strongly associated with E. coli phylogroups A, B1, B2, D and F. We identified 43 multilocus STs, of which ST131 (14.9%) and ST12 (9.6%) were the most common. We identified 25 AMR genes. The most common ESBL gene was bla CTX-M-15 and it was present in all five phylogroups and 11 STs, and most commonly detected in ST391 (4/4 isolates), ST648 (3/3 isolates) and ST131 [3/14 (21.4%) isolates]. Conclusions: This study has revealed a high diversity of lineages associated with AMR, including ESBL and fluoroquinolone resistance, in Malawi. The data highlight the value of longitudinal bacteraemia surveillance coupled with detailed molecular epidemiology in all settings, including low-income settings, in describing the global epidemiology of ESBL resistance.
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Farmacorresistência Bacteriana Múltipla/genética , Infecções por Escherichia coli/microbiologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Resistência beta-Lactâmica/genética , beta-Lactamases/genética , Adolescente , Adulto , Criança , Pré-Escolar , Cloranfenicol/farmacologia , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/epidemiologia , Feminino , Genes Bacterianos , Variação Genética , Genômica , Humanos , Malaui/epidemiologia , Masculino , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Filogenia , População Urbana/estatística & dados numéricos , Adulto JovemRESUMO
BACKGROUND: Multiyear epidemics of Salmonella enterica serovar Typhi have been reported from countries across eastern and southern Africa in recent years. In Blantyre, Malawi, a dramatic increase in typhoid fever cases has recently occurred, and may be linked to the emergence of the H58 haplotype. Strains belonging to the H58 haplotype often exhibit multidrug resistance and may have a fitness advantage relative to other Salmonella Typhi strains. METHODS: To explore hypotheses for the increased number of typhoid fever cases in Blantyre, we fit a mathematical model to culture-confirmed cases of Salmonella enterica infections at Queen Elizabeth Central Hospital, Blantyre. We explored 4 hypotheses: (1) an increase in the basic reproductive number (R0) in response to increasing population density; (2) a decrease in the incidence of cross-immunizing infection with Salmonella Enteritidis; (3) an increase in the duration of infectiousness due to failure to respond to first-line antibiotics; and (4) an increase in the transmission rate following the emergence of the H58 haplotype. RESULTS: Increasing population density or decreasing cross-immunity could not fully explain the observed pattern of typhoid emergence in Blantyre, whereas models allowing for an increase in the duration of infectiousness and/or the transmission rate of typhoid following the emergence of the H58 haplotype provided a good fit to the data. CONCLUSIONS: Our results suggest that an increase in the transmissibility of typhoid due to the emergence of drug resistance associated with the H58 haplotype may help to explain recent outbreaks of typhoid in Malawi and similar settings in Africa.
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Salmonella typhi/genética , Febre Tifoide/epidemiologia , Febre Tifoide/transmissão , África , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Número Básico de Reprodução , Surtos de Doenças , Farmacorresistência Bacteriana Múltipla , Haplótipos , Humanos , Incidência , Malaui/epidemiologia , Modelos Teóricos , Filogenia , Densidade Demográfica , Salmonella enteritidis/genética , Salmonella enteritidis/imunologia , Salmonella typhi/efeitos dos fármacos , Salmonella typhi/imunologia , Febre Tifoide/imunologia , Febre Tifoide/microbiologiaRESUMO
BACKGROUND: The Malawi Liverpool Wellcome Trust Clinical Research Programme (MLW) has routinely collected specimens for blood culture from febrile patients, and cerebrospinal fluid from patients with suspected meningitis, presenting to Queen Elizabeth Central Hospital (QECH), Blantyre, Malawi, since 1998. METHODS: We present bloodstream infection (BSI) and meningitis surveillance data from 1998 to 2014. Automated blood culture, manual speciation, serotyping, and antimicrobial susceptibility testing were performed at MLW. Population data for minimum-incidence estimates in urban Blantyre were drawn from published estimates. RESULTS: Between 1998 and 2014, 167,028 blood cultures were taken from adult and pediatric medical patients presenting to QECH; Salmonella Typhi was isolated on 2054 occasions (1.2%) and nontyphoidal Salmonella (NTS) serovars were isolated 10,139 times (6.1%), of which 8017 (79.1%) were Salmonella Typhimurium and 1608 (15.8%) were Salmonella Enteritidis. There were 392 cases of NTS meningitis and 9 cases of Salmonella Typhi meningitis. There have been 3 epidemics of Salmonella BSI in Blantyre; Salmonella Enteritidis from 1999 to 2002, Salmonella Typhimurium from 2002 to 2008, and Salmonella Typhi, which began in 2011 and was ongoing in 2014. Multidrug resistance has emerged in all 3 serovars and is seen in the overwhelming majority of isolates, while resistance to third-generation cephalosporins and fluoroquinolones is currently uncommon but has been identified. CONCLUSIONS: Invasive Salmonella disease in Malawi is dynamic and not clearly attributable to a single risk factor, although all 3 epidemics were associated with multidrug resistance. To inform nonvaccine and vaccine interventions, reservoirs of disease and modes of transmission require further investigation.
Assuntos
Bacteriemia/epidemiologia , Bacteriemia/microbiologia , Epidemias , Meningites Bacterianas/epidemiologia , Infecções por Salmonella/epidemiologia , Infecções por Salmonella/microbiologia , Salmonella enterica/isolamento & purificação , Adolescente , Adulto , Idoso , Antibacterianos/farmacologia , Cefalosporinas/farmacologia , Criança , Pré-Escolar , Farmacorresistência Bacteriana Múltipla , Monitoramento Epidemiológico , Feminino , Fluoroquinolonas/farmacologia , Humanos , Incidência , Lactente , Recém-Nascido , Malaui/epidemiologia , Masculino , Meningites Bacterianas/microbiologia , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Fatores de Risco , Infecções por Salmonella/etiologia , Salmonella enterica/classificação , Salmonella enterica/efeitos dos fármacos , Salmonella enteritidis/efeitos dos fármacos , Salmonella enteritidis/isolamento & purificação , Salmonella typhi/efeitos dos fármacos , Salmonella typhi/isolamento & purificação , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/isolamento & purificação , Sorotipagem , Adulto JovemRESUMO
BACKGROUND: Carriage of either single or multiple pneumococcal serotypes (multiple carriage) is a prerequisite for developing invasive pneumococcal disease. However, despite the reported high rates of pneumococcal carriage in Malawi, no data on carriage of multiple serotypes has been reported previously. Our study provides the first description of the prevalence of multiple pneumococcal carriage in Malawi. METHODS: The study was conducted in Blantyre and Karonga districts in Malawi, from 2008 to 2012. We recruited 116 children aged 0-13 years. These children were either HIV-infected (N = 44) or uninfected (N = 72). Nasopharyngeal samples were collected using sterile swabs. Pneumococcal serotypes in the samples were identified by microarray. Strains that could not be typed by microarray were sequenced to characterise possible genetic alterations within the capsular polysaccharide (CPS) locus. RESULTS: The microarray identified 179 pneumococcal strains (from 116 subjects), encompassing 43 distinct serotypes and non-typeable (NT) strains. Forty per cent (46/116) of children carried multiple serotypes. Carriage of vaccine type (VT) strains was higher (p = 0.028) in younger (0-2 years) children (71 %, 40/56) compared to older (3-13 years) children (50 %, 30/60). Genetic variations within the CPS locus of known serotypes were observed in 19 % (34/179) of the strains identified. The variants included 13-valent pneumococcal conjugate vaccine (PCV13) serotypes 6B and 19A, and the polysaccharide vaccine serotype 20. Serotype 6B variants were the most frequently isolated (47 %, 16/34). Unlike the wild type, the CPS locus of the 6B variants contained an insertion of the licD-family phosphotransferase gene. The CPS locus of 19A- and 20-variants contained an inversion in the sugar-biosynthesis (rmlD) gene and a 717 bp deletion within the transferase (whaF) gene, respectively. CONCLUSIONS: The high multiple carriage in Malawian children provides opportunities for genetic exchange through horizontal gene transfer. This may potentially lead to CPS locus variants and vaccine escape. Variants reported here occurred naturally, however, PCV13 introduction could exacerbate the CPS genetic variations. Further studies are therefore recommended to assess the invasive potential of these variants and establish whether PCV13 would offer cross-protection. We have shown that younger children (0-2 years) are a reservoir of VT serotypes, which makes them an ideal target for vaccination.
Assuntos
Infecções Pneumocócicas/microbiologia , Streptococcus pneumoniae/genética , Adolescente , Cápsulas Bacterianas/genética , Técnicas Bacteriológicas , Criança , Pré-Escolar , Proteção Cruzada/imunologia , DNA Bacteriano/análise , DNA Bacteriano/isolamento & purificação , Feminino , Variação Genética , Infecções por HIV/complicações , Infecções por HIV/diagnóstico , Humanos , Lactente , Recém-Nascido , Malaui , Masculino , Nasofaringe/microbiologia , Análise de Sequência com Séries de Oligonucleotídeos , Filogenia , Infecções Pneumocócicas/complicações , Infecções Pneumocócicas/prevenção & controle , Vacinas Pneumocócicas/imunologia , Análise de Sequência de DNA , Sorogrupo , Streptococcus pneumoniae/classificação , Streptococcus pneumoniae/isolamento & purificaçãoRESUMO
Despite the importance of Salmonella infections in human and animal health, the target antigens of Salmonella-specific immunity remain poorly defined. We have previously shown evidence for antibody-mediating protection against invasive Salmonellosis in mice and African children. To generate an overview of antibody targeting in systemic Salmonellosis, a Salmonella proteomic array containing over 2,700 proteins was constructed and probed with immune sera from Salmonella-infected mice and humans. Analysis of multiple inbred mouse strains identified 117 antigens recognized by systemic antibody responses in murine Salmonellosis. Importantly, many of these antigens were independently identified as target antigens using sera from Malawian children with Salmonella bacteremia, validating the study of the murine model. Furthermore, vaccination with SseB, the most prominent antigenic target in Malawian children, provided mice with significant protection against Salmonella infection. Together, these data uncover an overlapping immune signature of disseminated Salmonellosis in mice and humans and provide a foundation for the generation of a protective subunit vaccine.
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Salmonelose Animal/imunologia , Infecções por Salmonella/imunologia , Animais , Formação de Anticorpos/imunologia , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Atividade Bactericida do Sangue , Criança , Pré-Escolar , Convalescença , Feminino , Humanos , Lactente , Recém-Nascido , Malaui , Masculino , Camundongos , Camundongos Endogâmicos , Análise Serial de Proteínas , Reprodutibilidade dos Testes , Infecções por Salmonella/sangue , Vacinação , Vacinas Atenuadas/imunologia , Vacinas de Subunidades Antigênicas/imunologiaRESUMO
Reducing the inappropriate use of antibiotics in food animals is a global priority to address antimicrobial resistance (AMR). We investigated practices and factors associated with antibiotic use in small-scale commercial broiler farms in Lilongwe district, Malawi. We used structured questionnaires to collect data on recent antibiotic use practices among 128 broiler farmers, who kept between 50 and 1 000 birds, from December 2022 to March 2023. Logistic regression analysis was used to identify risk factors associated with antibiotic use. Over half (53.1â¯%, n=68) of the farms reported using antibiotics at least once in the previous production cycle. Overall, 11 different types of antibiotics were used either for treatment and/or preventive purposes, with oxytetracycline (88.2â¯%), erythromycin (29.4â¯%), and enrofloxacin (26.5â¯%) reported as the frequently used. One-third of all antibiotic formulations contained multiple active antibiotic ingredients, with 12â¯% containing four antibiotics. Covariates associated with an increased likelihood of antibiotic use include disease incidence (OR=13.8, 95â¯% CI 5.27-42.50, p<0.001) and entry of wild birds into poultry houses (OR=3.56, 95â¯% CI =1.44-9.61, p=0.008). Our study highlights inappropriate usage of antibiotics, largely associated with reduced biosecurity and disease incidence. These findings underscore the need to strengthen veterinary services, reinforce regulations on antibiotic access and use, and farmer education programs promoting proper husbandry, biosecurity, and responsible antibiotic use.
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Criação de Animais Domésticos , Antibacterianos , Galinhas , Animais , Malaui , Antibacterianos/uso terapêutico , Criação de Animais Domésticos/métodos , Criação de Animais Domésticos/estatística & dados numéricos , Doenças das Aves Domésticas/tratamento farmacológico , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/prevenção & controle , Inquéritos e Questionários , Fazendeiros/psicologiaRESUMO
The World Health Organization End TB strategy aims for a 90% reduction of tuberculosis (TB) incidence by 2035. Systematic testing and treatment of latent TB infection (LTBI) among contacts of active TB patients is recommended as one of the ways to curtail TB incidence. However, there is a shortage of tools to accurately diagnose LTBI. We assessed the appropriateness of whole blood host transcriptomic markers (TM) to diagnose LTBI among household contacts of bacteriologically confirmed index cases compared to HIV negative healthy controls (HC). QuantiFERON-TB Gold Plus Interferon gamma release assay (IGRA) and reverse-transcriptase quantitative PCR were used to determine LTBI and quantify TM expression respectively. Association between TM expression and LTBI was evaluated by logistic regression modelling. A total of 100 participants, 49 TB exposed (TBEx) household contacts and 51 HC, were enrolled. Twenty-five (51%) TBEx individuals tested positive by IGRA, and were denoted as LTBI individuals, and 37 (72.5%) HC were IGRA-negative. Expression of 11 evaluated TM was significantly suppressed among LTBI compared to HC. Out of the 11 TM, ZNF296 and KLF2 expression were strongly associated with LTBI and successfully differentiated LTBI from HC. Paradoxically, 21 (49%) TBEx participants who tested IGRA negative exhibited the same pattern of suppressed TM expression as IGRA positive (LTBI-confirmed individuals). Results suggest that suppression of gene expression underlies LTBI and may be a more sensitive diagnostic biomarker than standard-of-care IGRA.
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Biomarcadores , Tuberculose Latente , Humanos , Tuberculose Latente/diagnóstico , Tuberculose Latente/sangue , Tuberculose Latente/genética , Masculino , Feminino , Adulto , Biomarcadores/sangue , Pessoa de Meia-Idade , Testes de Liberação de Interferon-gama/métodos , Adulto Jovem , Transcriptoma , Estudos de Casos e Controles , AdolescenteRESUMO
BACKGROUND: Universal antiretroviral therapy (ART) has led to improved treatment outcomes in persons living with HIV. Adherence to ART is required to achieve viral suppression. Real-time medication monitoring (RTMM)-based digital adherence tools (DATs) could be effective in improving ART adherence and viral suppression in persons living with HIV. OBJECTIVES: The primary and secondary objectives of this review were to assess the effect of RTMM-based DATs on improving ART adherence and viral load suppression. METHODS: We searched MEDLINE, Embase, and Global Health for publications published through October 11, 2022. Narrative synthesis and random effects meta-analyses were conducted to synthesize the results. RESULTS: Of 638 papers identified, 8 were included. Six studies were randomized controlled trials (RCTs), and 2 were cohort studies. Two studies, an RCT in China (mean adherence: 96.2% vs 89.1%) and a crossover cohort study in Uganda (mean adherence: 84% vs 93%), demonstrated improved ART adherence. No studies demonstrated improved viral suppression. In the meta-analyses, we estimated that RTMM-based digital adherence tools had a statistically insignificant small positive effect on ART adherence and viral suppression with a standardized mean difference of 0.1922 [95% CI: -0.0268 to 0.4112, P-value: 0.0854] and viral suppression with an odds ratio of 1.3148 [95% CI: 0.9199 to 1.8791, P-value: 0.1331]. CONCLUSIONS: Our meta-analyses found that RTMM-based DATs did not have a significant effect on ART adherence and viral suppression. However, due to few published studies available, heterogeneity of target populations, intervention designs, and adherence measurement instruments, more data are required to provide conclusive evidence.
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Fármacos Anti-HIV , Infecções por HIV , Adesão à Medicação , Carga Viral , Humanos , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Fármacos Anti-HIV/uso terapêutico , Monitoramento de Medicamentos/métodos , Ensaios Clínicos Controlados Aleatórios como Assunto , Antirretrovirais/uso terapêuticoRESUMO
BACKGROUND: Infections caused by multidrug-resistant gram-negative bacteria present a severe threat to global public health. The WHO defines drug-resistant Klebsiella pneumoniae as a priority pathogen for which alternative treatments are needed given the limited treatment options and the rapid acquisition of novel resistance mechanisms by this species. Longitudinal descriptions of genomic epidemiology of Klebsiella pneumoniae can inform management strategies but data from sub-Saharan Africa are lacking. METHODS: We present a longitudinal analysis of all invasive K. pneumoniae isolates from a single hospital in Blantyre, Malawi, southern Africa, from 1998 to 2020, combining clinical data with genome sequence analysis of the isolates. RESULTS: We show that after a dramatic increase in the number of infections from 2016 K. pneumoniae becomes hyperendemic, driven by an increase in neonatal infections. Genomic data show repeated waves of clonal expansion of different, often ward-restricted, lineages, suggestive of hospital-associated transmission. We describe temporal trends in resistance and surface antigens, of relevance for vaccine development. CONCLUSIONS: Our data highlight a clear need for new interventions to prevent rather than treat K. pneumoniae infections in our setting. Whilst one option may be a vaccine, the majority of cases could be avoided by an increased focus on and investment in infection prevention and control measures, which would reduce all healthcare-associated infections and not just one.
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Infecções por Klebsiella , Klebsiella pneumoniae , Klebsiella pneumoniae/genética , Humanos , Infecções por Klebsiella/epidemiologia , Infecções por Klebsiella/microbiologia , Estudos Longitudinais , Vacinas Bacterianas/imunologia , Adulto , Feminino , Hospitais , Criança , Masculino , Pré-Escolar , Lactente , Pessoa de Meia-Idade , África Subsaariana/epidemiologia , Infecção Hospitalar/microbiologia , Adolescente , Genoma Bacteriano , Farmacorresistência Bacteriana Múltipla/genética , Recém-Nascido , Malaui/epidemiologia , Adulto JovemRESUMO
Invasive non-typhoidal Salmonella (iNTS) disease is a serious bloodstream infection that targets immune-compromised individuals, and causes significant mortality in sub-Saharan Africa. Salmonella enterica serovar Typhimurium ST313 causes the majority of iNTS in Malawi. We performed an intensive comparative genomic analysis of 608 S. Typhimurium ST313 isolates dating between 1996 and 2018 from Blantyre, Malawi. We discovered that following the arrival of the well-characterized S. Typhimurium ST313 lineage 2 in 1999, two multidrug-resistant variants emerged in Malawi in 2006 and 2008, designated sublineages 2.2 and 2.3, respectively. The majority of S. Typhimurium isolates from human bloodstream infections in Malawi now belong to sublineages 2.2 or 2.3. To understand the emergence of the prevalent ST313 sublineage 2.2, we studied two representative strains, D23580 (lineage 2) and D37712 (sublineage 2.2). The chromosome of ST313 lineage 2 and sublineage 2.2 only differed by 29 SNPs/small indels and a 3 kb deletion of a Gifsy-2 prophage region including the sseI pseudogene. Lineage 2 and sublineage 2.2 had distinctive plasmid profiles. The transcriptome was investigated in 15 infection-relevant in vitro conditions and within macrophages. During growth in physiological conditions that do not usually trigger S. Typhimurium SPI2 gene expression, the SPI2 genes of D37712 were transcriptionally active. We identified down-regulation of flagellar genes in D37712 compared with D23580. Following phenotypic confirmation of transcriptomic differences, we discovered that sublineage 2.2 had increased fitness compared with lineage 2 during mixed growth in minimal media. We speculate that this competitive advantage is contributing to the emergence of sublineage 2.2 in Malawi.
RESUMO
BACKGROUND: Enteric fever is a serious public health concern. The causative agents, Salmonella enterica serovars Typhi and Paratyphi A, frequently have antimicrobial resistance (AMR), leading to limited treatment options and poorer clinical outcomes. We investigated the genomic epidemiology, resistance mechanisms, and transmission dynamics of these pathogens at three urban sites in Africa and Asia. METHODS: S Typhi and S Paratyphi A bacteria isolated from blood cultures of febrile children and adults at study sites in Dhaka (Bangladesh), Kathmandu (Nepal), and Blantyre (Malawi) during STRATAA surveillance were sequenced. Isolates were charactered in terms of their serotypes, genotypes (according to GenoTyphi and Paratype), molecular determinants of AMR, and population structure. We used phylogenomic analyses incorporating globally representative genomic data from previously published surveillance studies and ancestral state reconstruction to differentiate locally circulating from imported pathogen AMR variants. Clusters of sequences without any single-nucleotide variants in their core genome were identified and used to explore spatiotemporal patterns and transmission dynamics. FINDINGS: We sequenced 731 genomes from isolates obtained during surveillance across the three sites between Oct 1, 2016, and Aug 31, 2019 (24 months in Dhaka and Kathmandu and 34 months in Blantyre). S Paratyphi A was present in Dhaka and Kathmandu but not Blantyre. S Typhi genotype 4.3.1 (H58) was common in all sites, but with different dominant variants (4.3.1.1.EA1 in Blantyre, 4.3.1.1 in Dhaka, and 4.3.1.2 in Kathmandu). Multidrug resistance (ie, resistance to chloramphenicol, co-trimoxazole, and ampicillin) was common in Blantyre (138 [98%] of 141 cases) and Dhaka (143 [32%] of 452), but absent from Kathmandu. Quinolone-resistance mutations were common in Dhaka (451 [>99%] of 452) and Kathmandu (123 [89%] of 138), but not in Blantyre (three [2%] of 141). Azithromycin-resistance mutations in acrB were rare, appearing only in Dhaka (five [1%] of 452). Phylogenetic analyses showed that most cases derived from pre-existing, locally established pathogen variants; 702 (98%) of 713 drug-resistant infections resulted from local circulation of AMR variants, not imported variants or recent de novo emergence; and pathogen variants circulated across age groups. 479 (66%) of 731 cases clustered with others that were indistinguishable by point mutations; individual clusters included multiple age groups and persisted for up to 2·3 years, and AMR determinants were invariant within clusters. INTERPRETATION: Enteric fever was associated with locally established pathogen variants that circulate across age groups. AMR infections resulted from local transmission of resistant strains. These results form a baseline against which to monitor the impacts of control measures. FUNDING: Wellcome Trust, Bill & Melinda Gates Foundation, EU Horizon 2020, and UK National Institute for Health and Care Research.
Assuntos
Antibacterianos , Filogenia , Salmonella paratyphi A , Salmonella typhi , Febre Tifoide , Humanos , Bangladesh/epidemiologia , Nepal/epidemiologia , Febre Tifoide/epidemiologia , Febre Tifoide/microbiologia , Febre Tifoide/transmissão , Febre Tifoide/tratamento farmacológico , Salmonella typhi/genética , Salmonella typhi/efeitos dos fármacos , Criança , Antibacterianos/farmacologia , Adulto , Pré-Escolar , Malaui/epidemiologia , Salmonella paratyphi A/genética , Salmonella paratyphi A/efeitos dos fármacos , Masculino , Adolescente , Farmacorresistência Bacteriana/genética , Feminino , Lactente , Febre Paratifoide/epidemiologia , Febre Paratifoide/microbiologia , Febre Paratifoide/transmissão , Febre Paratifoide/tratamento farmacológico , Adulto Jovem , Genótipo , Genoma Bacteriano/genética , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , GenômicaRESUMO
Malawi experienced its deadliest Vibrio cholerae (Vc) outbreak following devastating cyclones, with >58,000 cases and >1700 deaths reported between March 2022 and May 2023. Here, we use population genomics to investigate the attributes and origin of the Malawi 2022-2023 Vc outbreak isolates. Our results demonstrate the predominance of ST69 clone, also known as the seventh cholera pandemic El Tor (7PET) lineage, expressing O1 Ogawa (~ 80%) serotype followed by Inaba (~ 16%) and sporadic non-O1/non-7PET serogroups (~ 4%). Phylogenetic reconstruction revealed that the Malawi outbreak strains correspond to a recent importation from Asia into Africa (sublineage AFR15). These isolates harboured known antimicrobial resistance and virulence elements, notably the ICEGEN/ICEVchHai1/ICEVchind5 SXT/R391-like integrative conjugative elements and a CTXφ prophage with the ctxB7 genotype compared to historical Malawian Vc isolates. These data suggest that the devastating cyclones coupled with the recent importation of 7PET serogroup O1 strains, may explain the magnitude of the 2022-2023 cholera outbreak in Malawi.
Assuntos
Cólera , Surtos de Doenças , Filogenia , Vibrio cholerae , Malaui/epidemiologia , Cólera/epidemiologia , Cólera/microbiologia , Humanos , Vibrio cholerae/genética , Vibrio cholerae/classificação , Genômica , Genoma Bacteriano/genética , Prófagos/genética , Genótipo , SorogrupoRESUMO
BACKGROUND: TB is a leading cause of morbidity among HIV positive individuals. Accurate algorithms are needed to achieve early TB diagnosis and treatment. We investigated the use of Xpert MTB/RIF Ultra in combination with chest radiography for TB diagnosis in ambulatory HIV positive individuals. METHODS: This was a randomised controlled trial with a 2-by-2 factorial design. Outpatient HIV clinic attendees with cough were randomised to four arms: Arm 1-Standard Xpert/no chest radiography (CXR); Arm 2-Standard Xpert/CXR; Arm 3-Xpert Ultra/no CXR; and Arm 4-Xpert Ultra/CXR. Participants were followed up at days 28 and 56 to assess for TB treatment initiation. RESULTS: We randomised 640 participants. Bacteriologically confirmed TB treatment initiation at day 28 were: Arm 1 (8.4% [14/162]), Arm 2 (6.9% [11/159]), Arm 3 (8.2% [13/159]) and Arm 4 (5.6% [9/160]) and between Xpert Ultra group (Arms 3 and 4) (6.9% [22/319]) vs Standard Xpert group (Arms 1 and 2) (7.8% [25/321]), risk ratio 0.89 (95% CI 0.51 to 1.54). By day 56, there were also similar all-TB treatment initiations in the x-ray group (Arms 2 and 4) (16.0% [51/319]) compared with the no x-ray group (Arms 1 and 3) (13.1% [42/321]), risk ratio 1.22 (95% CI 0.84 to 1.78); however, the contribution of clinically diagnosed treatment initiations were higher in x-ray groups (50.9% vs 19.0%). CONCLUSIONS: Xpert Ultra performed similarly to Xpert MTB/RIF. X-rays are useful for TB screening but further research should investigate how to mitigate false-positive treatment initiations.
Assuntos
Infecções por HIV , Mycobacterium tuberculosis , Tuberculose Pulmonar , Humanos , Tuberculose Pulmonar/diagnóstico por imagem , Tuberculose Pulmonar/tratamento farmacológico , Radiografia , Instituições de Assistência Ambulatorial , Infecções por HIV/complicações , Sensibilidade e Especificidade , EscarroRESUMO
Early infant diagnosis of HIV (EID-HIV) is key to reducing paediatric HIV mortality. Traditional approaches for diagnosing HIV in exposed infants are usually unable to optimally contribute to EID. Point-of-care testing such as Cepheid Xpert HIV-1 Qual assay-1 (XPertHIV) are available and could improve EID-HIV in resource constrained and high HIV burden contexts. We investigated the acceptability and perceived appropriateness of XpertHIV for EID-HIV in Mulanje Hospital, Malawi. Qualitative cross-sectional study using semi-structured interviews (SSI) among caregivers and health care workers at Mulanje District Hospital. The qualitative study was nested within a larger diagnostic study that evaluated the performance of XpertHIV using whole-blood-sample in a resource limited and high burden setting. A total of 65 SSIs were conducted among caregivers (n = 60) and health care providers (n = 5). Data were coded using deductive and inductive approaches while thematic approach was used to analyse data. Point-of-care XPertHIV was perceived to be acceptable among caregivers and health care providers. Caregivers' motivations for accepting XPertHIV HIV-testing for their infants included perceived risk of HIV emanating from child's exposure and validation of caregiver's own HIV sero-status. Although concerns about pain of testing and blood sample volumes taken from an infant remained amplified, overall, both caregivers and health care providers felt XpertHIV was appropriate because of its quick result turn-around-time which decreased anxiety and stress, the prospect of early treatment initiation and reduction in hospital visits and related costs. Implementation of XpertHIV has a great potential to improve EID-HIV in Malawi because of its quick turn-around-time and associated benefits including overcoming access-related barriers. Scaled implementation of this diagnostic technology require a robust community engagement strategy for managing caregivers and community myths and misconceptions towards the amount of blood sample collected from infants.
RESUMO
Aeromonas caviae is an increasingly recognized etiological agent of acute gastroenteritis. Here, we report five draft genomes of A. caviae isolated from suspected cholera cases during the 2022-2023 cholera outbreak in Malawi.
RESUMO
Invasive non-typhoidal Salmonella (iNTS) disease manifesting as bloodstream infection with high mortality is responsible for a huge public health burden in sub-Saharan Africa. Salmonella enterica serovar Typhimurium (S. Typhimurium) is the main cause of iNTS disease in Africa. By analysing whole genome sequence data from 1303 S. Typhimurium isolates originating from 19 African countries and isolated between 1979 and 2017, here we show a thorough scaled appraisal of the population structure of iNTS disease caused by S. Typhimurium across many of Africa's most impacted countries. At least six invasive S. Typhimurium clades have already emerged, with ST313 lineage 2 or ST313-L2 driving the current pandemic. ST313-L2 likely emerged in the Democratic Republic of Congo around 1980 and further spread in the mid 1990s. We observed plasmid-borne as well as chromosomally encoded fluoroquinolone resistance underlying emergences of extensive-drug and pan-drug resistance. Our work provides an overview of the evolution of invasive S. Typhimurium disease, and can be exploited to target control measures.