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Biomedical subjects

J Farrar

Publications and source records attributed to J Farrar.

At least 19 recordsLinked to original sources
PubMed2006

A simple and rapid liquid chromatographic assay for evaluation of potentially counterfeit Tamiflu.

A simple and rapid liquid chromatographic assay for the evaluation of potentially counterfeit oseltamivir (Tamiflu has been developed and assessed. The assay uses approximately 1mg Tamiflu powder when used for authentication and content estimate. The procedure was validated using 50 replicates analysed during five independent series with a total R.S.D. of 11.2%. The assay can also be used to monitor the exact content of oseltamivir in Tamiflu capsules. One Tamiflu capsule was transferred to a 250mL volumetric flask and 150mL water was added. The flask was placed in an ultrasonic bath at 40 degrees C for 20min to dissolve the capsule. The solution was allowed to cool to room temperature before the flask was filled up to the mark (250mL). A small aliquot was centrifuged and then directly injected into the LC-system for quantification. Oseltamivir was analysed by liquid chromatography with UV detection on a Hypersil Gold column (150mmx4.6mm) using a mobile phase containing methanol-phosphate buffer (pH 2.5; 0.1M) (50:50, v/v) at a flow rate of 1.0mL/min. The assay was implemented for the analysis of Tamiflu purchased over the Internet and at local pharmacies in Thailand and Vietnam.

Acetamides↗
PubMed2006

Evolution and adaptation of H5N1 influenza virus in avian and human hosts in Indonesia and Vietnam.

Highly pathogenic avian influenza virus H5N1 is endemic in poultry in East and Southeast Asia with disease outbreaks recently spreading to parts of central Asia, Europe and Africa. Continued interspecies transmission to humans has been reported in Vietnam, Thailand, Cambodia, Indonesia and China, causing pandemic concern. Here, we genetically characterize 82 H5N1 viruses isolated from poultry throughout Indonesia and Vietnam and 11 human isolates from southern Vietnam together with sequence data available in public databases to address questions relevant to virus introduction, endemicity and evolution. Phylogenetic analysis shows that all viruses from Indonesia form a distinct sublineage of H5N1 genotype Z viruses suggesting this outbreak likely originated from a single introduction that spread throughout the country during the past two years. Continued virus activities in Indonesia were attributed to transmission via poultry movement within the country rather than through repeated introductions by bird migration. Within Indonesia and Vietnam, H5N1 viruses have evolved over time into geographically distinct groups within each country. Molecular analysis of the H5N1 genotype Z genome shows that only the M2 and PB1-F2 genes were under positive selection, suggesting that these genes might be involved in adaptation of this virus to new hosts following interspecies transmission. At the amino acid level 12 residues were under positive selection in those genotype Z viruses, in the HA and PB1-F2 proteins. Some of these residues were more frequently observed in human isolates than in avian isolates and are related to viral antigenicity and receptor binding. Our study provides insight into the ongoing evolution of H5N1 influenza viruses that are transmitting in diverse avian species and at the interface between avian and human hosts.

Animals↗
PubMed2004

Comparative pharmacokinetics of intramuscular artesunate and artemether in patients with severe falciparum malaria.

The first-dose pharmacokinetic properties of intramuscular (i.m.) artesunate (ARTS; 2.4 mg/kg immediately [stat], followed by 1.2 mg/kg i.m. daily) and artemether (ARM; 3.2 mg/kg i.m. stat, followed by 1.6 mg/kg i.m. daily) were compared in Vietnamese adults with severe falciparum malaria. A total of 19 patients were studied; 9 received ARTS, and 10 received ARM. ARTS was absorbed very rapidly; concentrations in plasma peaked between 1,362 and 8,388 nmol/liter (median, 5,710 nmol/liter) within 20 min of injection and then declined with a median (range) half-life (t(1/2)) of 30 (3 to 67) min. ARTS was hydrolyzed rapidly and completely to the biologically active metabolite dihydroartemisinin (DHA). Peak DHA concentrations in plasma ranged between 1,718 and 7,080 nmol/liter (median, 3,060 nmol/liter) and declined with a t(1/2) of 52 (26 to 69) min. In contrast, ARM was slowly and erratically absorbed. The absorption profile appeared biphasic. Maximum ARM concentrations in plasma ranged between 67 nmol/liter (a value close to the 50% inhibitory concentration for some Plasmodium falciparum isolates) and 1,631 nmol/liter (median, 574 nmol/liter) and occurred at a median (range) of 10 (1.5 to 24) h. There was relatively little conversion to DHA. After i.m. injection in cases of severe malaria, absorption of the water-soluble ARTS is rapid and extensive, whereas the oil-based ARM is slowly and erratically absorbed, with relatively little conversion to the more active DHA. On the basis of this pharmacological study, parenteral ARTS is preferable to ARM as an initial antimalarial therapy, particularly in the most seriously ill patients. These findings should be formally assessed by a randomized clinical trial.

Adolescent↗
PubMed2002

Assessment of the factors associated with flavivirus seroprevalence in a population in Southern Vietnam.

Dengue and Japanese encephalitis flaviviruses cause severe disease and are hyperendemic in southern Vietnam. This study assesses associations between sociodemographic factors and flavivirus seroprevalence in this region. Sera were collected from 308 community and hospital-based subjects between April 1996 and August 1997 and tested with an indirect ELISA. The factors associated with seroprevalence were assessed using multivariate logistic regression. In this first report of adjusted prevalence odds ratios (POR) for flavivirus infection in Vietnam, seropositivity was associated with increasing age in children (multiple regression coefficients for a child compared to an adult = -4.975 and for age in children = 0.354) and residence in the city compared to surrounding rural districts. The association with age indicates that subjects were most likely to have acquired infection in early childhood. This is key to the design of Vietnamese health education and immunization programmes.

Adolescent↗
PubMed2001

Strong HLA class I--restricted T cell responses in dengue hemorrhagic fever: a double-edged sword?

Dengue is an increasingly important cause of morbidity and mortality in the tropics, but vaccine development has been impeded by a poor understanding of disease pathogenesis and, in particular, of immunologic enhancement. In a large case-control study of Vietnamese patients with dengue hemorrhagic fever (DHF), variation at the HLA-A locus was significantly associated with susceptibility to DHF (P=.02), and specific HLA-A susceptibility and resistance alleles were identified. HLA-A-specific epitopes were predicted from binding motifs, and ELISPOT analyses of patients with DHF revealed high frequencies of circulating CD8 T lymphocytes that recognized both serotype-specific and -cross-reactive dengue virus epitopes. Thus, strong CD8 T cell responses are induced by natural dengue virus infection, and HLA class I genetic variation is a risk factor for DHF. These genetic and immunologic data support both protective and pathogenic roles for dengue virus-specific CD8 T cell responses in severe disease. The potentially pathogenic role of serotype-cross-reactive CD8 T cells poses yet another obstacle to successful dengue vaccine development.

Adolescent↗
PubMed2001

Complete genome sequence of a multiple drug resistant Salmonella enterica serovar Typhi CT18.

Salmonella enterica serovar Typhi (S. typhi) is the aetiological agent of typhoid fever, a serious invasive bacterial disease of humans with an annual global burden of approximately 16 million cases, leading to 600,000 fatalities. Many S. enterica serovars actively invade the mucosal surface of the intestine but are normally contained in healthy individuals by the local immune defence mechanisms. However, S. typhi has evolved the ability to spread to the deeper tissues of humans, including liver, spleen and bone marrow. Here we have sequenced the 4,809,037-base pair (bp) genome of a S. typhi (CT18) that is resistant to multiple drugs, revealing the presence of hundreds of insertions and deletions compared with the Escherichia coli genome, ranging in size from single genes to large islands. Notably, the genome sequence identifies over two hundred pseudogenes, several corresponding to genes that are known to contribute to virulence in Salmonella typhimurium. This genetic degradation may contribute to the human-restricted host range for S. typhi. CT18 harbours a 218,150-bp multiple-drug-resistance incH1 plasmid (pHCM1), and a 106,516-bp cryptic plasmid (pHCM2), which shows recent common ancestry with a virulence plasmid of Yersinia pestis.

Chromosome Mapping↗
PubMed2001

Fake artesunate in southeast Asia.

Artesunate is a key antimalarial drug in the treatment of multidrug-resistant Plasmodium falciparum malaria in southeast Asia. We investigated the distribution of counterfeit artesunate tablets by use of the validated, simple, and inexpensive Fast Red TR dye technique. We also aimed to identify distinguishing characteristics of the fake drugs. Of 104 shop-bought "artesunate" samples from Cambodia, Laos, Myanmar (Burma), Thailand, and Vietnam, 38% did not contain artesunate. Characteristics such as cost and physical appearance of the tablets and packaging reliably predicted authenticity. The illicit trade in counterfeit antimalarials is a great threat to the lives of patients with malaria. The dye test will assist national malaria control authorities in urgently needed campaigns to stop this murderous trade.

Antimalarials↗
PubMed2001

Identification of a matrix-degrading phenotype in human tuberculosis in vitro and in vivo.

Tuberculous meningitis is characterized by cerebral tissue destruction. Monocytes, pivotal in immune responses to Mycobacterium tuberculosis, secrete matrix metalloproteinase-9 (MMP-9), which facilitates leukocyte migration across the blood-brain barrier, but may cause cerebral injury. In vitro, human monocytic (THP-1) cells infected by live, virulent M. tuberculosis secreted MMP-9 in a dose-dependent manner. At 24 h, MMP-9 concentrations increased 10-fold to 239 +/- 75 ng/ml (p = 0.001 vs controls). MMP-9 mRNA became detectable at 24--48 h. In contrast, tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) gene expression and secretion were similar to constitutive levels from controls at 24 h and increased just 5-fold by 48 h. In vivo investigation revealed MMP-9 concentration per leukocyte in cerebrospinal fluid (CSF) from tuberculous meningitis patients (n = 23; median (range), 3.19 (0.19--31.00) ng/ml/cell) to be higher than that in bacterial (n = 12; 0.23 (0.01--18.37) ng/ml/cell) or viral meningitis (n = 20; 0.20 (0.04--31.00) ng/ml/cell; p < 0.01). TIMP-1, which was constitutively secreted into CSF, was not elevated in tuberculous compared with bacterial meningitis or controls. Thus, a phenotype in which MMP-9 activity is relatively unrestricted by TIMP-1 developed both in vitro and in vivo. This is functionally significant, since MMP-9 concentrations per CSF leukocyte (but not TIMP-1 concentrations) were elevated in fatal tuberculous meningitis and in patients with signs of cerebral tissue damage (unconsciousness, confusion, or neurological deficit; p < 0.05). However, MMP-9 activity was unrelated to the severity of systemic illness. In summary, M. tuberculosis-infected monocytic cells develop a matrix-degrading phenotype, which was observed in vivo and relates to clinical signs reflecting cerebral injury in tuberculous meningitis.

Adult↗
PubMed2001

Typhoid fever and genetic polymorphisms at the natural resistance-associated macrophage protein 1.

Control of Salmonella enterica serovar Typhimurium (S. typhimurium) infection in the mouse model of typhoid fever is critically dependent on the natural resistance-associated macrophage protein 1 (Nramp1). In this study, we examined the role of genetic polymorphisms in the human homologue, NRAMP1, in resistance to typhoid fever in southern Vietnam. Patients with blood-culture-confirmed typhoid fever and healthy control subjects were genotyped for 6 polymorphic markers within and near NRAMP1 on chromosome 2q35. Four single base-pair polymorphisms (274 C/T, 469+14 G/C, 1465-85 G/A, and D543N), a (GT)(n) repeat in the promoter region of NRAMP1 and D2S1471, and a microsatellite marker approximately 130-kb downstream of NRAMP1 were examined. The allelic and genotypic frequencies for each polymorphism were compared in case patients and control subjects. No allelic association was identified between the NRAMP1 alleles and typhoid fever susceptibility. In addition, neither homozygotes nor heterozygotes for any NRAMP1 variants were at increased risk of typhoid fever.

Alleles↗
PubMed2001

An outbreak of Salmonella serotype Thompson associated with fresh cilantro.

An outbreak of Salmonella serotype Thompson in California was identified through laboratory-based surveillance and investigated with case-control, traceback, and laboratory studies. There were 35 "sporadic" cases and a restaurant-associated outbreak of 41 cases with onset between 6 March and 31 March 1999. Three case patients were hospitalized. A case-control study found a significant association between illness and eating cilantro at a restaurant (63% of case patients vs. 34% of control subjects; odds ratio, 3.5; 95% confidence interval, 1.1-11.4). Although common distributors of cilantro were identified, inadequate records prohibited the identification of a single farm supplying cilantro. At room temperature, Salmonella Thompson grew more rapidly and to a higher concentration on chopped cilantro, compared with whole-leaf cilantro. Freshly made salsa (pH 3.4) supported growth of Salmonella Thompson. Cilantro should be served promptly after chopping. Accurate records of the distribution of produce should be available, and bacterial contamination of produce should be prevented in retail and wholesale establishments, in packing sheds, and on farms.

Adolescent↗
PubMed2001

Acute management of dengue shock syndrome: a randomized double-blind comparison of 4 intravenous fluid regimens in the first hour.

Dengue hemorrhagic fever is an important cause of morbidity among Asian children, and the more severe dengue shock syndrome (DSS) causes a significant number of childhood deaths. DSS is characterized by a massive increase in systemic capillary permeability with consequent hypovolemia. Fluid resuscitation is critical, but as yet there have been no large trials to determine the optimal fluid regimen. We undertook a randomized blinded comparison of 4 fluids (dextran, gelatin, lactated Ringer's, and "normal" saline) for initial resuscitation of 230 Vietnamese children with DSS. All the children survived, and there was no clear advantage to using any of the 4 fluids, but the longest recovery times occurred in the lactated Ringer's group. The most significant factor determining clinical response was the pulse pressure at presentation. A comparison of the colloid and crystalloid groups suggested benefits in children presenting with lower pulse pressures who received one of the colloids. Further large-scale studies, stratified for admission pulse pressure, are indicated.

Adolescent↗
PubMed2001

Cellular stress and injury responses in the brains of adult Vietnamese patients with fatal Plasmodium falciparum malaria.

Immunohistochemical techniques have been used to investigate specific patterns of potentially reversible cellular injury, DNA damage, and apoptosis in the brainstems of Vietnamese patients who died of severe Plasmodium falciparum malaria. The degree and pattern of neuronal and glial stress responses were compared between patients with cerebral and non-cerebral malaria (CM), and appropriate non-malaria infected controls. The following markers were examined: (i) heat shock protein 70 (HSP70), for reversible injury; (ii) heme oxygenase-1, for oxidative stress; (iii & iv) two DNA-repair proteins, poly(ADP) ribose polymerase (PARP) and DNA-dependent protein kinase catalytic subunit; (v) poly(ADP) ribose, an end-product of PARP activity; and (vi) caspase-3-active, for apoptosis. Stress responses were found in a range of cell types as reflected by the widespread expression of HSP70. Oxidative stress predominated in the vicinity of vessels and haemorrhages. Some degree of DNA damage was found in the majority of malaria patients, but the distribution and frequency of the damage was much less than that observed in controls with irreversible neuronal injury. Similarly, caspase-3-active expression, as a measure of apoptosis, was no higher in the majority of malaria patients than the negative control cases, although 40% of CM cases expressed caspase-3-active in a small number of neurones of the pontine nuclei or within swollen axons of the pontocerebellar and corticospinal tracts. In conclusion, cells within the brainstem of all patients who died from severe malaria showed staining patterns indicative of considerable stress response and reversible neuronal injury. There was no evidence for a specific pattern of widespread irreversible cell damage in those patients with cerebral malaria.

Adult↗
PubMed2001

Quantitation of bacteria in bone marrow from patients with typhoid fever: relationship between counts and clinical features.

Enteric fever is the only bacterial infection of humans for which bone marrow examination is routinely recommended. A prospective study of the concentrations of bacteria in the bone marrow and their relationship to clinical features was conducted with 120 Vietnamese patients with suspected enteric fever, of whom 89 had confirmed typhoid fever. Ninety-three percent of the Salmonella enterica serovar Typhi samples isolated were resistant to ampicillin, chloramphenicol, and co-trimoxazole. For 81 patients with uncomplicated typhoid and satisfactory bone marrow aspirates, the number of serovar Typhi CFU in bone marrow aspirates was a median value of 9 (interquartile range [IQR], 1 to 85; range, 0.1 to 1,580) compared to 0.3 (IQR, 0.1 to 10; range, 0.1 to 399) CFU/ml in simultaneously sampled blood. The ratio of individual blood counts to bone marrow counts was 10 (IQR, 2.3 to 97.5). The number of bacteria in blood but not bone marrow was correlated inversely with the duration of preceding fever. Thus, with increasing duration of illness the ratio of bone marrow-to-blood bacterial concentrations increased; the median ratio was 4.8 (IQR, 1 to 27.5) during the first week compared with 158 (IQR, 60 to 397) during the third week. After lysing the host cells, the median ratio of viable bone marrow to blood increased, reflecting the higher concentration of intracellular serovar Typhi in the bone marrow. Effective antibiotic pretreatment had a significantly greater effect in reducing blood counts compared to bone marrow counts (P < 0.001). Thus, bacteria in the bone marrow of typhoid patients are less affected by antibiotic treatment than bacteria in the blood. The numbers of bacteria in bone marrow correlated negatively with the white blood cell (R = -0.3, P = 0.006) and platelet counts (R = -0.32, P = 0.01) and positively with fever clearance time after treatment (R = 0.4, P < 0.001). The bacterial load in bone marrow therefore may reflect the clinical course of the infection, and high levels may suppress neutrophil proliferation.

Adolescent↗
PubMed2000

Genes of the class II and class III major histocompatibility complex are associated with typhoid fever in Vietnam.

The influence of genes of the major histocompatibility complex (MHC) class II and class III loci on typhoid fever susceptibility was investigated. Individuals with blood culture-confirmed typhoid fever and control subjects from 2 distinct geographic locations in southern Vietnam were genotyped for HLA-DRB1 and HLA-DQB1 alleles, the gene that encodes tumor necrosis factor (TNF)-alpha (TNFA [-238] and TNFA [-308]), the gene that encodes lymphotoxin-alpha, and alleles of the TNF-alpha microsatellite. HLA-DRB1*0301/6/8, HLA-DQB1*0201-3, and TNFA*2 (-308) were associated with susceptibility to typhoid fever, whereas HLA-DRB1*04, HLA-DQB1*0401/2, and TNFA*1 (-308) were associated with disease resistance. The frequency of all possible haplotypes of the 3 individually associated loci were estimated and were found to be significantly different in typhoid case patients and control subjects (chi2=55.56, 32 df; P=.006). Haplotypes that were either protective (TNFA*1 [-308].DRB1*04) or predisposed individuals to typhoid fever (TNFA*2 [-308].DRB1*0301) were determined. This report identifies a genetic association in humans between typhoid fever and MHC class II and III genes.

Alleles↗
PubMed2000

Blood-brain barrier function in cerebral malaria and CNS infections in Vietnam.

BACKGROUND: The intraerythrocytic parasite Plasmodium falciparum induces the life-threatening neurologic syndrome of cerebral malaria (CM) from within cerebral blood vessels, without entering the brain parenchyma. OBJECTIVES: 1) To assess the use of CSF as an indicator of specific pathologic processes occurring in the brain during CM; 2) to compare this with other neurologic and infectious diseases to understand the distinct pathogenic features of CM; 3) to test the hypothesis that CM involves a specific functional breakdown of the blood-brain barrier (BBB). METHODS: 1) Radial immunodiffusion assays to detect albumin and IgG in matched plasma and CSF samples as indicators of BBB integrity and intrathecal IgG production; and 2) ELISA for soluble intracellular adhesion molecule-1 and sE-selectin, the cytokines tumor necrosis factor-alpha and transforming growth factor-beta1, and the matrix metalloproteinase MMP-9, to detect cellular activation and inflammatory responses within the brain. RESULTS: Albumin and IgG indices implied only minimal degree of BBB breakdown in a few cases of CM, with most remaining within the normal range. In contrast, cryptococcal, tubercular, and acute bacterial meningitis produced detectable changes in the composition of the CSF and evidence of BBB breakdown. CONCLUSIONS: CM appears to involve only subtle functional changes in BBB integrity with minimal intraparenchymal inflammatory responses compared with other neurologic infections. This focuses attention on local events within and around the cerebral microvasculature in CM, rather than indicating widespread parenchymal disease.

Adolescent↗
PubMed2000

High-performance liquid chromatographic determination of morphine and its 3- and 6-glucuronide metabolites by two-step solid-phase extraction.

To provide more accurate measurement of morphine and its metabolites for a study of the genetic differences on morphine response, a method for the analysis of morphine and its metabolites is described which has the advantages of increased sensitivity and specificity by using a cleaner extraction. The new extraction method involves both the hydrophobic isolation on a carbon cartridge and ion-exchange isolation on ion-exchange resin which has not preliminary been described for morphine analysis. The combination of these two steps successfully purified drugs from human plasma with maximum removal of interfering substance comparing with a conventional C18 cartridge alone. The analytes are quantified by high-performance liquid chromatography on a reversed-phase C18 column employing a mobile phase consisting of 25% acetonitrile in 0.05 M phosphate buffer (pH 2.1), and 2.5 mM sodium dodecyl sulfate as the pairing ion with a combination of electrochemical and fluorometric detections. The recoveries for morphine (M), morphine-3-glucuronide (M3G), morphine-6-glucuronide (M6G) and hydromorphone after the SPE procedure were 86+/-7.1%, 82+/-6.9%, 79+/-6.0% and 85+/-6.0%, respectively. Limits of detection for this method are 0.1 ng/ml for M, and 0.18 ng/ml for M3G and M6G. Limits of quantitation were approximately 0.25 ng/ml for M, and 0.45 ng/ml for M3G and M6G. The present assay was applied to measure M, M3G and M6G content in human plasma to test the applicability and suitability of this method for clinical and research use.

Chromatography, High Pressure Liquid↗