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The nasopharynx is an important reservoir of disease-associated and antimicrobial-resistant bacterial species. This proof-of-concept study assessed the utility of a combined culture, matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS), and targeted metagenomic sequencing workflow for the study of the pediatric nasopharyngeal bacterial microbiota. Nasopharyngeal swabs and clinical metadata were collected from Cambodian children during a hospital outpatient visit and then biweekly for 12 weeks. Swabs were cultured on chocolate and blood-gentamicin agar, and all colony morphotypes were identified by MALDI-TOF MS. Metagenomic sequencing was done on a scrape of all colonies from a chocolate agar culture and processed using the mSWEEP pipeline. One hundred one children were enrolled, yielding 620 swabs. MALDI-TOF MS identified 106 bacterial species/40 genera: 20 species accounted for 88.5% (2,190/2,474) of isolates. Colonization by Moraxella catarrhalis (92.1% of children on ≥1 swab), Haemophilus influenzae (87.1%), and Streptococcus pneumoniae (83.2%) was particularly common. In S. pneumoniae-colonized children, a median of two serotypes [inter-quartile range (IQR) 1-2, range 1-4] was detected. For the 21 bacterial species included in the mSWEEP database and identifiable by MALDI-TOF, detection by culture + MALDI-TOF MS and culture + mSWEEP was highly concordant with a median species-level agreement of 96.9% (IQR 86.8%-98.8%). mSWEEP revealed highly dynamic lineage-level colonization patterns for S. pneumoniae which were quite different to those for S. aureus. A combined culture, MALDI-TOF MS, targeted metagenomic sequencing approach for the exploration of the young child nasopharyngeal microbiome was technically feasible, and each component yielded complementary data. IMPORTANCE: The human upper respiratory tract is an important source of disease-causing and antibiotic-resistant bacteria. However, understanding the interactions and stability of these bacterial populations is technically challenging. We used a combination of approaches to determine colonization patterns over a 3-month period in 101 Cambodian children. The combined approach was feasible to implement, and each component gave complementary data to enable a better understanding of the complex patterns of bacterial colonization.
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Bactérias , Metagenômica , Microbiota , Nasofaringe , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Humanos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Nasofaringe/microbiologia , Microbiota/genética , Pré-Escolar , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Feminino , Metagenômica/métodos , Criança , Lactente , Masculino , Camboja , Haemophilus influenzae/genética , Haemophilus influenzae/isolamento & purificação , Haemophilus influenzae/classificaçãoRESUMO
Objectives: To explore the association of recent hospitalization and asymptomatic carriage of multidrug-resistant Enterobacterales (MDRE) and determine the prevailing strains and antibiotic resistance genes in Siem Reap, Cambodia using WGS. Methods: In this cross-sectional study, faecal samples were collected from two arms: a hospital-associated arm consisted of recently hospitalized children (2-14â years), with their family members; and a community-associated arm comprising children in the matching age group and their family members with no recent hospitalization. Forty-two families in each study arm were recruited, with 376 enrolled participants (169 adults and 207 children) and 290 stool specimens collected from participants. The DNA of ESBL- and carbapenemase-producing Enterobacterales cultured from the faecal samples was subject to WGS on the Illumina NovaSeq platform. Results: Of the 290 stool specimens, 277 Escherichia coli isolates and 130 Klebsiella spp. were identified on CHROMagar ESBL and KPC plates. The DNA of 276 E. coli (one isolate failed quality control test), 89 Klebsiella pneumoniae, 40 Klebsiella quasipneumoniae and 1 Klebsiella variicola was sequenced. CTX-M-15 was the most common ESBL gene found in E. coli (nâ=â104, 38%), K. pneumoniae (nâ=â50, 56%) and K. quasipneumoniae (nâ=â16, 40%). The prevalence of bacterial lineages and ESBL genes was not associated with any specific arm. Conclusions: Our results demonstrate that MDRE is likely to be endemic within the Siem Reap community. ESBL genes, specifically blaCTX-M, can be found in almost all E. coli commensals, indicating that these genes are continuously propagated in the community through various unknown channels at present.
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INTRODUCTION: Bloodstream infections (BSIs) are a leading cause of sepsis, which is a life-threatening condition that significantly contributes to the mortality of bacterial infections. Aminoglycoside antibiotics such as gentamicin or amikacin are essential medicines in the treatment of BSIs, but their clinical efficacy is increasingly being compromised by antimicrobial resistance. The aminoglycoside apramycin has demonstrated preclinical efficacy against aminoglycoside-resistant and multidrug-resistant (MDR) Gram-negative bacilli (GNB) and is currently in clinical development for the treatment of critical systemic infections. METHODS: This study collected a panel of 470 MDR GNB isolates from healthcare facilities in Cambodia, Laos, Singapore, Thailand and Vietnam for a multicentre assessment of their antimicrobial susceptibility to apramycin in comparison with other aminoglycosides and colistin by broth microdilution assays. RESULTS: Apramycin and amikacin MICs ≤ 16 µg/mL were found for 462 (98.3%) and 408 (86.8%) GNB isolates, respectively. Susceptibility to gentamicin and tobramycin (MIC ≤ 4 µg/mL) was significantly lower at 122 (26.0%) and 101 (21.5%) susceptible isolates, respectively. Of note, all carbapenem and third-generation cephalosporin-resistant Enterobacterales, all Acinetobacter baumannii and all Pseudomonas aeruginosa isolates tested in this study appeared to be susceptible to apramycin. Of the 65 colistin-resistant isolates tested, four (6.2%) had an apramycin MIC > 16 µg/mL. CONCLUSION: Apramycin demonstrated best-in-class activity against a panel of GNB isolates with resistances to other aminoglycosides, carbapenems, third-generation cephalosporins and colistin, warranting continued consideration of apramycin as a drug candidate for the treatment of MDR BSIs.
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Amicacina , Colistina , Aminoglicosídeos , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Sudeste Asiático , Hemocultura , Carbapenêmicos , Cefalosporinas , Colistina/farmacologia , Farmacorresistência Bacteriana Múltipla , Gentamicinas , Bactérias Gram-Negativas , Testes de Sensibilidade Microbiana , Nebramicina/análogos & derivados , Pseudomonas aeruginosa , TobramicinaRESUMO
BACKGROUND: Blood cultures remain the gold standard investigation for the diagnosis of bloodstream infections. In many locations, quality-assured processing of positive blood cultures is not possible. One solution is to incubate blood cultures locally, and then transport bottles that flag positive to a central reference laboratory for organism identification and antimicrobial susceptibility testing. However, the impact of delay between the bottle flagging positive and subsequent sub-culture on the viability of the isolate has received little attention. METHODS: This study evaluated the impact of delays to sub-culture (22 h to seven days) in three different temperature conditions (2-8 °C, 22-27 °C and 35 ± 2 °C) for bottles that had flagged positive in automated detection systems using a mixture of spiked and routine clinical specimens. Ninety spiked samples for five common bacterial causes of sepsis (Escherichia coli, Haemophilus influenzae, Staphylococcus aureus, Streptococcus agalactiae and Streptococcus pneumoniae) and 125 consecutive positive clinical blood cultures were evaluated at four laboratories located in Cambodia, Lao PDR and Thailand. In addition, the utility of transport swabs for preserving organism viability was investigated. RESULTS: All organisms were recoverable from all sub-cultures in all temperature conditions with the exception of S. pneumoniae, which was less likely to be recoverable after longer delays (> 46-50 h), when stored in hotter temperatures (35 °C), and from BacT/ALERT when compared with BACTEC blood culture bottles. Storage of positive blood culture bottles in cooler temperatures (22-27 °C or below) and the use of Amies bacterial transport swabs helped preserve viability of S. pneumoniae. CONCLUSIONS: These results have practical implications for the optimal workflow for blood culture bottles that have flagged positive in automated detection systems located remotely from a central processing laboratory, particularly in tropical resource-constrained contexts.
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Bacteriemia , Hemocultura , Bacteriemia/diagnóstico , Bacteriemia/microbiologia , Bactérias , Técnicas Bacteriológicas/métodos , Meios de Cultura , Escherichia coli , Humanos , Estudos ProspectivosRESUMO
Genomic data contribute invaluable information to the epidemiological investigation of pathogens of public health importance. However, whole-genome sequencing (WGS) of bacteria typically relies on culture, which represents a major hurdle for generating such data for a wide range of species for which culture is challenging. In this study, we assessed the use of culture-free target-enrichment sequencing as a method for generating genomic data for two bacterial species: (1) Bacillus anthracis, which causes anthrax in both people and animals and whose culture requires high-level containment facilities; and (2) Mycoplasma amphoriforme, a fastidious emerging human respiratory pathogen. We obtained high-quality genomic data for both species directly from clinical samples, with sufficient coverage (>15×) for confident variant calling over at least 80% of the baited genomes for over two thirds of the samples tested. Higher qPCR cycle threshold (Ct) values (indicative of lower pathogen concentrations in the samples), pooling libraries prior to capture, and lower captured library concentration were all statistically associated with lower capture efficiency. The Ct value had the highest predictive value, explaining 52â% of the variation in capture efficiency. Samples with Ct values ≤30 were over six times more likely to achieve the threshold coverage than those with a Ct > 30. We conclude that target-enrichment sequencing provides a valuable alternative to standard WGS following bacterial culture and creates opportunities for an improved understanding of the epidemiology and evolution of many clinically important pathogens for which culture is challenging.
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Genômica , Saúde Pública , Animais , Bactérias/genética , Humanos , Sequenciamento Completo do Genoma/métodosRESUMO
Antimicrobial resistance (AMR) surveillance in bloodstream infections (BSIs) is challenging in low/middle-income countries (LMICs) given limited laboratory capacity. Other specimens are easier to collect and process and are more likely to be culture-positive. In 8102 E. coli BSIs, 322,087 E. coli urinary tract infections, 6952 S. aureus BSIs and 112,074 S. aureus non-sterile site cultures from Oxfordshire (1998-2018), and other (55,296 isolates) rarer commensal opportunistic pathogens, antibiotic resistance trends over time in blood were strongly associated with those in other specimens (maximum cross-correlation per drug 0.51-0.99). Resistance prevalence was congruent across drug-years for each species (276/312 (88%) species-drug-years with prevalence within ± 10% between blood/other isolates). Results were similar across multiple countries in high/middle/low income-settings in the independent ATLAS dataset (103,559 isolates, 2004-2017) and three further LMIC hospitals/programmes (6154 isolates, 2008-2019). AMR in commensal opportunistic pathogens cultured from BSIs is strongly associated with AMR in commensal opportunistic pathogens cultured from non-sterile sites over calendar time, suggesting the latter could be used as an effective proxy for AMR surveillance in BSIs.
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Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Escherichia coli/crescimento & desenvolvimento , Sepse/microbiologia , Staphylococcus aureus/crescimento & desenvolvimento , Infecções Urinárias/microbiologia , Gestão de Antimicrobianos , Infecção Hospitalar , Países em Desenvolvimento , Escherichia coli/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Prevalência , Sepse/tratamento farmacológico , Staphylococcus aureus/efeitos dos fármacos , Simbiose , Infecções Urinárias/tratamento farmacológicoRESUMO
Antimicrobial resistance (AMR) is a major threat to global health. Improving laboratory capacity for AMR detection is critically important for patient health outcomes and population level surveillance. We aimed to estimate the financial cost of setting up and running a microbiology laboratory for organism identification and antimicrobial susceptibility testing as part of an AMR surveillance programme. Financial costs for setting up and running a microbiology laboratory were estimated using a top-down approach based on resource and cost data obtained from three clinical laboratories in the Mahidol Oxford Tropical Medicine Research Unit network. Costs were calculated for twelve scenarios, considering three levels of automation, with equipment sourced from either of the two leading manufacturers, and at low and high specimen throughput. To inform the costs of detection of AMR in existing labs, the unit cost per specimen and per isolate were also calculated using a micro-costing approach. Establishing a laboratory with the capacity to process 10,000 specimens per year ranged from $254,000 to $660,000 while the cost for a laboratory processing 100,000 specimens ranged from $394,000 to $887,000. Excluding capital costs to set up the laboratory, the cost per specimen ranged from $22-31 (10,000 specimens) and $11-12 (100,000 specimens). The cost per isolate ranged from $215-304 (10,000 specimens) and $105-122 (100,000 specimens). This study provides a conservative estimate of the costs for setting up and running a microbiology laboratory for AMR surveillance from a healthcare provider perspective. In the absence of donor support, these costs may be prohibitive in many low- and middle- income country (LMIC) settings. With the increased focus on AMR detection and surveillance, the high laboratory costs highlight the need for more focus on developing cheaper and cost-effective equipment and reagents so that laboratories in LMICs have the potential to improve laboratory capacity and participate in AMR surveillance.
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BACKGROUND: Burkholderia pseudomallei is the bacterial causative agent of melioidosis, a difficult disease to diagnose clinically with high mortality if not appropriately treated. Definitive diagnosis requires isolation and identification of the organism. With the increased adoption of MALDI-TOF MS for the identification of bacteria, we established a method for rapid identification of B. pseudomallei using the Vitek MS, a system that does not currently have B. pseudomallei in its in-vitro diagnostic database. RESULTS: A routine direct spotting method was employed to create spectra and SuperSpectra. An initial B. pseudomallei SuperSpectrum was created at Shoklo Malaria Research Unit (SMRU) from 17 reference isolates (46 spectra). When tested, this initial SMRU SuperSpectrum was able to identify 98.2 % (54/55) of Asian isolates, but just 46.7 % (35/75) of Australian isolates. Using spectra (430) from different reference and clinical isolates, two additional SMRU SuperSpectra were created. Using the combination of all SMRU SuperSpectra with seven existing SuperSpectra from Townsville, Australia 119 (100 %) Asian isolates and 31 (100 %) Australian isolates were correctly identified. In addition, no misidentifications were obtained when using these 11 SuperSpectra when tested with 34 isolates of other bacteria including the closely related species Burkholderia thailandensis and Burkholderia cepacia. CONCLUSIONS: This study has established a method for identification of B. pseudomallei using Vitek MS, and highlights the impact of geographical differences between strains for identification using this technique.
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Burkholderia pseudomallei/química , Burkholderia pseudomallei/isolamento & purificação , Melioidose/diagnóstico , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Técnicas Bacteriológicas/instrumentação , Técnicas Bacteriológicas/normas , Melioidose/microbiologia , Reprodutibilidade dos Testes , Especificidade da EspécieRESUMO
BACKGROUND: Blood cultures are one of the most important tests performed by microbiology laboratories. Many hospitals, particularly in low and middle-income countries, lack either microbiology services or staff to provide 24 h services resulting in delays to blood culture incubation. There is insufficient guidance on how to transport/store blood cultures if delays before incubation are unavoidable, particularly if ambient temperatures are high. This study set out to address this knowledge gap. METHODS: In three South East Asian countries, four different blood culture systems (two manual and two automated) were used to test blood cultures spiked with five common bacterial pathogens. Prior to incubation the spiked blood culture bottles were stored at different temperatures (25 °C, in a cool-box at ambient temperature, or at 40 °C) for different lengths of time (0 h, 6 h, 12 h or 24 h). The impacts of these different storage conditions on positive blood culture yield and on time to positivity were examined. RESULTS: There was no significant loss in yield when blood cultures were stored < 24 h at 25 °C, however, storage for 24 h at 40 °C decreased yields and longer storage times increased times to detection. CONCLUSION: Blood cultures should be incubated with minimal delay to maximize pathogen recovery and timely result reporting, however, this study provides some reassurance that unavoidable delays can be managed to minimize negative impacts. If delays to incubation ≥ 12 h are unavoidable, transportation at a temperature not exceeding 25 °C, and blind sub-cultures prior to incubation should be considered.
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Hemocultura/normas , Manejo de Espécimes/normas , Sudeste Asiático , Bactérias/classificação , Bactérias/isolamento & purificação , Hemocultura/estatística & dados numéricos , Serviços de Laboratório Clínico/normas , Serviços de Laboratório Clínico/estatística & dados numéricos , Humanos , Manejo de Espécimes/estatística & dados numéricos , Temperatura , Fatores de TempoRESUMO
Background: Antimicrobial resistance (AMR) / drug resistant infections (DRIs) are a major global health priority. Surveillance data is critical to inform infection treatment guidelines, monitor trends, and to assess interventions. However, most existing AMR / DRI surveillance systems are passive and pathogen-based with many potential biases. Addition of clinical and patient outcome data would provide considerable added value to pathogen-based surveillance. Methods: The aim of the ACORN project is to develop an efficient clinically-oriented AMR surveillance system, implemented alongside routine clinical care in hospitals in low- and middle-income country settings. In an initial pilot phase, clinical and microbiology data will be collected from patients presenting with clinically suspected meningitis, pneumonia, or sepsis. Community-acquired infections will be identified by daily review of new admissions, and hospital-acquired infections will be enrolled during weekly point prevalence surveys, on surveillance wards. Clinical variables will be collected at enrolment, hospital discharge, and at day 28 post-enrolment using an electronic questionnaire on a mobile device. These data will be merged with laboratory data onsite using a flexible automated computer script. Specific target pathogens will be Streptococcus pneumoniae, Staphylococcus aureus, Salmonella spp ., Klebsiella pneumoniae, Escherichia coli, and Acinetobacter baumannii. A bespoke browser-based app will provide sites with fully interactive data visualisation, analysis, and reporting tools. Discussion: ACORN will generate data on the burden of DRI which can be used to inform local treatment guidelines / national policy and serve as indicators to measure the impact of interventions. Following development, testing and iteration of the surveillance tools during an initial six-month pilot phase, a wider rollout is planned.
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BACKGROUND: Klebsiella pneumoniae is a leading cause of bloodstream infection (BSI). Strains producing extended-spectrum beta-lactamases (ESBLs) or carbapenemases are considered global priority pathogens for which new treatment and prevention strategies are urgently required, due to severely limited therapeutic options. South and Southeast Asia are major hubs for antimicrobial-resistant (AMR) K. pneumoniae and also for the characteristically antimicrobial-sensitive, community-acquired "hypervirulent" strains. The emergence of hypervirulent AMR strains and lack of data on exopolysaccharide diversity pose a challenge for K. pneumoniae BSI control strategies worldwide. METHODS: We conducted a retrospective genomic epidemiology study of 365 BSI K. pneumoniae from seven major healthcare facilities across South and Southeast Asia, extracting clinically relevant information (AMR, virulence, K and O antigen loci) using Kleborate, a K. pneumoniae-specific genomic typing tool. RESULTS: K. pneumoniae BSI isolates were highly diverse, comprising 120 multi-locus sequence types (STs) and 63 K-loci. ESBL and carbapenemase gene frequencies were 47% and 17%, respectively. The aerobactin synthesis locus (iuc), associated with hypervirulence, was detected in 28% of isolates. Importantly, 7% of isolates harboured iuc plus ESBL and/or carbapenemase genes. The latter represent genotypic AMR-virulence convergence, which is generally considered a rare phenomenon but was particularly common among South Asian BSI (17%). Of greatest concern, we identified seven novel plasmids carrying both iuc and AMR genes, raising the prospect of co-transfer of these phenotypes among K. pneumoniae. CONCLUSIONS: K. pneumoniae BSI in South and Southeast Asia are caused by different STs from those predominating in other regions, and with higher frequency of acquired virulence determinants. K. pneumoniae carrying both iuc and AMR genes were also detected at higher rates than have been reported elsewhere. The study demonstrates how genomics-based surveillance-reporting full molecular profiles including STs, AMR, virulence and serotype locus information-can help standardise comparisons between sites and identify regional differences in pathogen populations.
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Farmacorresistência Bacteriana Múltipla/genética , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/genética , Resistência beta-Lactâmica/genética , Sudeste Asiático , Proteínas de Bactérias/genética , Genoma Bacteriano , Humanos , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/patogenicidade , Mutação , Virulência/genética , beta-Lactamases/genéticaRESUMO
We describe 6 clinical isolates of Elizabethkingia anophelis from a pediatric referral hospital in Cambodia, along with 1 isolate reported from Thailand. Improving diagnostic microbiological methods in resource-limited settings will increase the frequency of reporting for this pathogen. Consensus on therapeutic options is needed, especially for resource-limited settings.
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Bacteriemia/diagnóstico , Infecções por Flavobacteriaceae/diagnóstico , Flavobacteriaceae/isolamento & purificação , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Bacteriemia/tratamento farmacológico , Ciprofloxacina/farmacologia , Ciprofloxacina/uso terapêutico , Feminino , Flavobacteriaceae/efeitos dos fármacos , Flavobacteriaceae/genética , Infecções por Flavobacteriaceae/tratamento farmacológico , Humanos , Recém-Nascido , Testes de Sensibilidade Microbiana , Vancomicina/farmacologia , Vancomicina/uso terapêuticoRESUMO
BACKGROUND: There is a pressing need to understand better the extent and distribution of antimicrobial resistance on a global scale, to inform development of effective interventions. Collation of datasets for meta-analysis, mathematical modelling and temporo-spatial analysis is hampered by the considerable variability in clinical sampling, variable quality in laboratory practice and inconsistencies in antimicrobial susceptibility testing and reporting. METHODS: The Microbiology Investigation Criteria for Reporting Objectively (MICRO) checklist was developed by an international working group of clinical and laboratory microbiologists, infectious disease physicians, epidemiologists and mathematical modellers. RESULTS: In keeping with the STROBE checklist, but applicable to all study designs, MICRO defines items to be included in reports of studies involving human clinical microbiology data. It provides a concise and comprehensive reference for clinicians, researchers, reviewers and journals working on, critically appraising, and publishing clinical microbiology datasets. CONCLUSIONS: Implementation of the MICRO checklist will enhance the quality and scientific reporting of clinical microbiology data, increasing data utility and comparability to improve surveillance, grade data quality, facilitate meta-analyses and inform policy and interventions from local to global levels.
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Serviços de Laboratório Clínico , Confiabilidade dos Dados , Interpretação Estatística de Dados , Técnicas Microbiológicas , Projetos de Pesquisa , Lista de Checagem/normas , Serviços de Laboratório Clínico/normas , Serviços de Laboratório Clínico/estatística & dados numéricos , Conjuntos de Dados como Assunto , Humanos , Técnicas Microbiológicas/métodos , Técnicas Microbiológicas/normas , Técnicas Microbiológicas/estatística & dados numéricos , Guias de Prática Clínica como Assunto , Editoração/normas , Projetos de Pesquisa/normas , Relatório de Pesquisa/normasRESUMO
'Candidatus Ornithobacterium hominis' represents a new member of the Flavobacteriaceae detected in 16S rRNA gene surveys of people from South-East Asia, Africa and Australia. It frequently colonizes the infant nasopharynx at high proportional abundance, and we demonstrate its presence in 42â% of nasopharyngeal swabs from 12-month-old children in the Maela refugee camp in Thailand. The species, a Gram-negative bacillus, has not yet been cultured, but the cells can be identified in mixed samples by fluorescent hybridization. Here, we report seven genomes assembled from metagenomic data, two to improved draft standard. The genomes are approximately 1.9 Mb, sharing 62â% average amino acid identity with the only other member of the genus, the bird pathogen Ornithobacterium rhinotracheale. The draft genomes encode multiple antibiotic-resistance genes, competition factors, Flavobacterium johnsoniae-like gliding motility genes and a homologue of the Pasteurella multocida mitogenic toxin. Intra- and inter-host genome comparison suggests that colonization with this bacterium is both persistent and strain exclusive.
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Farmacorresistência Bacteriana/genética , Infecções por Flavobacteriaceae/microbiologia , Interações Hospedeiro-Patógeno , Nasofaringe/microbiologia , Ornithobacterium/classificação , Ornithobacterium/genética , Adulto , Proteínas de Bactérias/genética , Toxinas Bacterianas/genética , Estudos de Coortes , Feminino , Humanos , Lactente , Recém-Nascido , Metagenoma , RNA Ribossômico 16S/genética , Estudos Retrospectivos , TailândiaRESUMO
Malaria rapid diagnostic tests (RDTs) primarily detect Plasmodium falciparum antigen histidine-rich protein 2 (HRP2) and the malaria-conserved antigen lactate dehydrogenase (LDH) for P. vivax and other malaria species. The performance of RDTs and their utility is dependent on circulating antigen concentration distributions in infected individuals in a population in which malaria is endemic and on the limit of detection of the RDT for the antigens. A multiplexed immunoassay for the quantification of HRP2, P. vivax LDH, and all-malaria LDH (pan LDH) was developed to accurately measure circulating antigen concentration and antigen distribution in a population with endemic malaria. The assay also measures C-reactive protein (CRP) levels as an indicator of inflammation. Validation was conducted with clinical specimens from 397 asymptomatic donors from Myanmar and Uganda, confirmed by PCR for infection, and from participants in induced blood-stage malaria challenge studies. The assay lower limits of detection for HRP2, pan LDH, P. vivax LDH, and CRP were 0.2 pg/ml, 9.3 pg/ml, 1.5 pg/ml, and 26.6 ng/ml, respectively. At thresholds for HRP2, pan LDH, and P. vivax LDH of 2.3 pg/ml, 47.8 pg/ml, and 75.1 pg/ml, respectively, and a specificity ≥98.5%, the sensitivities for ultrasensitive PCR-confirmed infections were 93.4%, 84.9%, and 48.9%, respectively. Plasmodium LDH (pLDH) concentration, in contrast to that of HRP2, correlated closely with parasite density. CRP levels were moderately higher in P. falciparum infections with confirmed antigenemia versus those in clinical specimens with no antigen. The 4-plex array is a sensitive tool for quantifying diagnostic antigens in malaria infections and supporting the evaluation of new ultrasensitive RDTs.
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Antígenos de Protozoários/sangue , Infecções Assintomáticas , Proteína C-Reativa/análise , Imunoensaio/métodos , Malária/sangue , Malária/diagnóstico , Adulto , Infecções Assintomáticas/epidemiologia , Criança , Pré-Escolar , Testes Diagnósticos de Rotina , Doenças Endêmicas , Humanos , Lactente , L-Lactato Desidrogenase/sangue , Malária/epidemiologia , Mianmar/epidemiologia , Plasmodium/imunologia , Proteínas de Protozoários/sangue , Sensibilidade e Especificidade , Uganda/epidemiologiaRESUMO
Sporadic cases of melioidosis have been diagnosed in Myanmar since the disease was first described in Yangon in 1911. Published and unpublished cases are summarized here, along with results from environmental and serosurveys. A total of 298 cases have been reported from seven states or regions between 1911 and 2018, with the majority of these occurring before 1949. Findings from soil surveys confirm the presence of Burkholderia pseudomallei in the environment in all three regions examined. The true epidemiology of the disease in Myanmar is unknown. Important factors contributing to the current gaps in knowledge are lack of awareness among clinicians and insufficient laboratory diagnostic capacity in many parts of the country. This is likely to have led to substantial under-reporting.
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Sensitive field-deployable diagnostic tests can assist malaria programs in achieving elimination. The performance of a new Alere™ Malaria Ag P.f Ultra Sensitive rapid diagnostic test (uRDT) was compared with the currently available SD Bioline Malaria Ag P.f RDT in blood specimens from asymptomatic individuals in Nagongera, Uganda, and in a Karen Village, Myanmar, representative of high- and low-transmission areas, respectively, as well as in pretreatment specimens from study participants from four Plasmodium falciparum-induced blood-stage malaria (IBSM) studies. A quantitative reverse transcription PCR (qRT-PCR) and a highly sensitive enzyme-linked immunosorbent assay (ELISA) test for histidine-rich protein II (HRP2) were used as reference assays. The uRDT showed a greater than 10-fold lower limit of detection for HRP2 compared with the RDT. The sensitivity of the uRDT was 84% and 44% against qRT-PCR in Uganda and Myanmar, respectively, and that of the RDT was 62% and 0% for the same two sites. The specificities of the uRDT were 92% and 99.8% against qRT-PCR for Uganda and Myanmar, respectively, and 99% and 99.8% against the HRP2 reference ELISA. The RDT had specificities of 95% and 100% against qRT-PCR for Uganda and Myanmar, respectively, and 96% and 100% against the HRP2 reference ELISA. The uRDT detected new infections in IBSM study participants 1.5 days sooner than the RDT. The uRDT has the same workflow as currently available RDTs, but improved performance characteristics to identify asymptomatic malaria infections. The uRDT may be a useful tool for malaria elimination strategies.
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Infecções Assintomáticas/epidemiologia , Malária Falciparum/diagnóstico , Malária Falciparum/epidemiologia , Antígenos de Protozoários/sangue , Criança , Pré-Escolar , Testes Diagnósticos de Rotina , Ensaio de Imunoadsorção Enzimática , Humanos , Lactente , Mianmar/epidemiologia , Plasmodium falciparum , Proteínas de Protozoários/sangue , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , Manejo de Espécimes , Uganda/epidemiologiaRESUMO
Toxoplasma gondii primary infection in pregnancy is associated with poor obstetric outcomes. This study aimed to determine the seroprevalence of Toxoplasma infection in pregnant migrant and refugee women from Myanmar attending antenatal care in Thailand. A random selection of 199 residual blood samples from first antenatal screen in 2014-2015 was tested for Toxoplasma IgG and IgM antibodies. Seroprevalence of Toxoplasma infection was 31.7% (95% confidence interval = 25.6-38.4). Avidity testing in the three positive IgM cases indicated all were past infections. Multiparity (≥ 3 children) was significantly associated with higher Toxoplasma seropositivity rates. Seroprevalence of T. gondii infection in this pregnant population is similar to the only other report from Myanmar, where multiparity was also identified as a significant association. Toxoplasma infection is important in pregnant women. Nevertheless, in this marginalized population, this infection may be given less priority, due to resource constraints in providing the most basic components of safe motherhood programs.
Assuntos
Anticorpos Antiprotozoários/sangue , Refugiados , Estudos Soroepidemiológicos , Toxoplasma/imunologia , Toxoplasmose/sangue , Migrantes , Adolescente , Adulto , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Pessoa de Meia-Idade , Mianmar/epidemiologia , Gravidez , Tailândia/epidemiologia , Toxoplasmose/epidemiologia , Adulto JovemRESUMO
BACKGROUND: Primaquine is the only drug consistently effective against mature gametocytes of Plasmodium falciparum. The transmission blocking dose of primaquine previously recommended was 0.75 mg/kg (adult dose 45 mg) but its deployment was limited because of concerns over haemolytic effects in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency. G6PD deficiency is an inherited X-linked enzymatic defect that affects an estimated 400 million people around the world with high frequencies (15-20%) in populations living in malarious areas. To reduce transmission in low transmission settings and facilitate elimination of P. falciparum, the World Health Organization now recommends adding a single dose of 0.25 mg/kg (adult dose 15 mg) to Artemisinin-based Combination Therapies (ACTs) without G6PD testing. Direct evidence of the safety of this low dose is lacking. Adverse events and haemoglobin variations after this treatment were assessed in both G6PD normal and deficient subjects in the context of targeted malaria elimination in a malaria endemic area on the North-Western Myanmar-Thailand border where prevalence of G6PD deficiency (Mahidol variant) approximates 15%. METHODS AND FINDINGS: The tolerability and safety of primaquine (single dose 0.25 mg base/kg) combined with dihydroartemisinin-piperaquine (DHA-PPQ) given three times at monthly intervals was assessed in 819 subjects. Haemoglobin concentrations were estimated over the six months preceding the ACT + primaquine rounds of mass drug administration. G6PD deficiency was assessed with a phenotypic test and genotyping was performed in male subjects with deficient phenotypes and in all females. Fractional haemoglobin changes in relation to G6PD phenotype and genotype and primaquine round were assessed using linear mixed-effects models. No adverse events related to primaquine were reported during the trial. Mean fractional haemoglobin changes after each primaquine treatment in G6PD deficient subjects (-5.0%, -4.2% and -4.7%) were greater than in G6PD normal subjects (0.3%, -0.8 and -1.7%) but were clinically insignificant. Fractional drops in haemoglobin concentration larger than 25% following single dose primaquine were observed in 1.8% of the population but were asymptomatic. CONCLUSIONS: The single low dose (0.25mg/kg) of primaquine is clinically well tolerated and can be used safely without prior G6PD testing in populations with high prevalence of G6PD deficiency. The present evidence supports a broader use of low dose primaquine without G6PD testing for the treatment and elimination of falciparum malaria. TRIAL REGISTRATION: ClinicalTrials.gov NCT01872702.