Mycobacterium tuberculosis strains of the Beijing genotype are rarely observed in tuberculosis patients in South America

The frequency of the Beijing genotype of Mycobacterium tuberculosis as a cause of tuberculosis (TB) in South America was determined by analyzing genotypes of strains isolated from patients that had been diagnosed with the disease between 1997 and 2003 in seven countries of the subcontinent. In total, 19 of the 1,202 (1.6%) TB cases carried Beijing isolates, including 11 of the 185 patients from Peru (5.9%), five of the 512 patients from Argentina (1.0%), two of the 252 Brazilian cases (0.8%), one of the 166 patients from Paraguay (0.6%) and none of the samples obtained from Chile (35), Colombia (36) and Ecuador (16). Except for two patients that were East Asian immigrants, all cases with Beijing strains were native South Americans. No association was found between carrying a strain with the Beijing genotype and having drug or multi-drug resistant disease. Our data show that presently transmission of M. tuberculosis strains of the Beijing genotype is not frequent in Latin America. In addition, the lack of association of drug resistant TB and infection with M. tuberculosis of the Beijing genotype observed presently demands efforts to define better the contribution of the virulence and lack of response to treatment to the growing spread of Beijing strains observed in other parts of the world.

The development of the Quality Indicator for Rehabilitative Care (QuIRC): a measure of best practice for facilities for people with longer term mental health problems

BACKGROUND: Despite the progress over recent decades in developing community mental health services internationally, many people still receive treatment and care in institutional settings. Those most likely to reside longest in these facilities have the most complex mental health problems and are at most risk of potential abuses of care and exploitation. This study aimed to develop an international, standardised toolkit to assess the quality of care in longer term hospital and community based mental health units, including the degree to which human rights, social inclusion and autonomy are promoted. METHOD: The domains of care included in the toolkit were identified from a systematic literature review, international expert Delphi exercise, and review of care standards in ten European countries. The draft toolkit comprised 154 questions for unit managers. Inter-rater reliability was tested in 202 units across ten countries at different stages of deinstitutionalisation and development of community mental health services. Exploratory factor analysis was used to corroborate the allocation of items to domains. Feedback from those using the toolkit was collected about its usefulness and ease of completion. RESULTS: The toolkit had excellent inter-rater reliability and few items with narrow spread of response. Unit managers found the content highly relevant and were able to complete it in around 90 minutes. Minimal refinement was required and the final version comprised 145 questions assessing seven domains of care. CONCLUSIONS: Triangulation of qualitative and quantitative evidence directed the development of a robust and comprehensive international quality assessment toolkit for units in highly variable socioeconomic and political contexts

Records of Chrysomya albiceps in Northern Italy: an ecological and forensic perspective

Knowledge of the carrion-breeding insects present at a local level is important and necessary for defining the post-mortem interval. Climate changes and globalisation are affecting species ranges and population dynamics. In this note, we report the incidence of Chrysomya albiceps (Diptera: Calliphoridae) on dead human bodies and carrion in Northern Italy. These data confirm the spread of this species in the Northern regions. The partial sequencing of a 583-bp region of the cytochrome oxidase subunit 1 gene of an Adriatic population did not reveal any difference compared to the same genomic region in the African and South American populations of this species.

Microsatellite markers for population genetic studies of the blowfly Chrysomya putoria (Diptera: Calliphoridae)

The investigation of the genetic variation and population structure of Chrysomya species is of great interest for both basic and applied research. However, very limited genetic information is available for this genus across its geographical distribution. Here, we describe 12 polymorphic microsatellite loci isolated from Chrysomya putoria with expected heterozygosities ranging from 0.1402-0.8312. These markers are of potential applied interest for forensic entomologists and for the characterisation of the genetic structure of C. putoria from recently colonised regions, with great promise for understanding the colonisation dynamics and spread of the genus Chrysomya in the New World.

Sex pheromone and period gene characterization of Lutzomyia longipalpis sensu lato (Lutz & Neiva) (Diptera: Psychodidae) from Posadas, Argentina

Lutzomyia longipalpis s.l. is the primary vector of Leishmania (L.) infantum in the New World. In this study, male Lutzomyia longipalpis specimens from Posadas, Argentina were characterized for two polymorphic markers: the male sex pheromone and the period (per) gene. The male sex pheromone was identified as (S)-9-methylgermacrene-B, the same compound produced by Lu. longipalpis from Paraguay and many populations from Brazil. The analysis of per gene sequences revealed that the population from Argentina is significantly differentiated from previously studied Brazilian populations. Marker studies could contribute to the understanding of the distribution and spread of urban American visceral leishmaniasis, thus aiding in the design of regional surveillance and control strategies.

Sequence analysis of coding DNA fragments of pfcrt and pfmdr-1 genes in Plasmodium falciparum isolates from Odisha, India

The global emergence and spread of malaria parasites resistant to antimalarial drugs is the major problem in malaria control. The genetic basis of the parasite's resistance to the antimalarial drug chloroquine (CQ) is well-documented, allowing for the analysis of field isolates of malaria parasites to address evolutionary questions concerning the origin and spread of CQ-resistance. Here, we present DNA sequence analyses of both the second exon of the Plasmodium falciparum CQ-resistance transporter (pfcrt) gene and the 5' end of the P. falciparum multidrug-resistance 1 (pfmdr-1) gene in 40 P. falciparum field isolates collected from eight different localities of Odisha, India. First, we genotyped the samples for the pfcrt K76T and pfmdr-1 N86Y mutations in these two genes, which are the mutations primarily implicated in CQ-resistance. We further analyzed amino acid changes in codons 72-76 of the pfcrt haplotypes. Interestingly, both the K76T and N86Y mutations were found to co-exist in 32 out of the total 40 isolates, which were of either the CVIET or SVMNT haplotype, while the remaining eight isolates were of the CVMNK haplotype. In total, eight nonsynonymous single nucleotide polymorphisms (SNPs) were observed, six in the pfcrt gene and two in the pfmdr-1 gene. One poorly studied SNP in the pfcrt gene (A97T) was found at a high frequency in many P. falciparum samples. Using population genetics to analyze these two gene fragments, we revealed comparatively higher nucleotide diversity in the pfcrt gene than in the pfmdr-1 gene. Furthermore, linkage disequilibrium was found to be tight between closely spaced SNPs of the pfcrt gene. Finally, both the pfcrt and the pfmdr-1 genes were found to evolve under the standard neutral model of molecular evolution.

Sequence analysis of the 2009 pandemic influenza A H1N1 virus haemagglutinin gene from 2009-2010 Brazilian clinical samples

In this paper, we analysed the haemagglutinin (HA) gene identified by polymerase chain reaction from 90 influenza A H1N1 virus strains that circulated in Brazil from April 2009-June 2010. A World Health Organization sequencing protocol allowed us to identify amino acid mutations in the HA protein at positions S220T (71%), D239G/N/S (20%), Y247H (4.5%), E252K (3.3%), M274V (2.2%), Q310H (26.7%) and E391K (12%). A fatal outcome was associated with the D239G mutation (p < 0.0001). Brazilian HA genetic diversity, in comparison to a reference strain from California, highlights the role of influenza virus surveillance for study of viral evolution, in addition to monitoring the spread of the virus worldwide.

Molecular markers and genetic diversity of Plasmodium vivax

Enhanced understanding of the transmission dynamics and population genetics for Plasmodium vivax is crucial in predicting the emergence and spread of novel parasite phenotypes with major public health implications, such as new relapsing patterns, drug resistance and increased virulence. Suitable molecular markers are required for these population genetic studies. Here, we focus on two groups of molecular markers that are commonly used to analyse natural populations of P. vivax. We use markers under selective pressure, for instance, antigen-coding polymorphic genes, and markers that are not under strong natural selection, such as most minisatellite and microsatellite loci. First, we review data obtained using genes encoding for P. vivax antigens: circumsporozoite protein, merozoite surface proteins 1 and 3α, apical membrane antigen 1 and Duffy binding antigen. We next address neutral or nearly neutral molecular markers, especially microsatellite loci, providing a complete list of markers that have already been used in P. vivax populations studies. We also analyse the microsatellite loci identified in the P. vivax genome project. Finally, we discuss some practical uses for P. vivax genotyping, for example, detecting multiple-clone infections and tracking the geographic origin of isolates.

Sentinel network for monitoring in vitro susceptibility of Plasmodium falciparum to antimalarial drugs in Colombia: a proof of concept

Drug resistance is one of the principal obstacles blocking worldwide malaria control. In Colombia, malaria remains a major public health concern and drug-resistant parasites have been reported. In vitro drug susceptibility assays are a useful tool for monitoring the emergence and spread of drug-resistant Plasmodium falciparum. The present study was conducted as a proof of concept for an antimalarial drug resistance surveillance network based on in vitro susceptibility testing in Colombia. Sentinel laboratories were set up in three malaria endemic areas. The enzyme linked immunosorbent assay-histidine rich protein 2 and schizont maturation methods were used to assess the susceptibility of fresh P. falciparum isolates to six antimalarial drugs. This study demonstrates that an antimalarial drug resistance surveillance network based on in vitro methods is feasible in the field with the participation of a research institute, local health institutions and universities. It could also serve as a model for a regional surveillance network. Preliminary susceptibility results showed widespread chloroquine resistance, which was consistent with previous reports for the Pacific region. However, high susceptibility to dihydroartemisinin and lumefantrine compounds, currently used for treatment in the country, was also reported. The implementation process identified critical points and opportunities for the improvement of network sustainability strategies.

Changing the epidemiology of carbapenem-resistant Pseudomonas aeruginosa in a Brazilian teaching hospital: the replacement of São Paulo metallo-β-lactamase-producing isolates

In Brazil, carbapenem-resistant Pseudomonas aeruginosa isolates are closely related to the São Paulo metallo-β-lactamase (SPM) Brazilian clone. In this study, imipenem-resistant isolates were divided in two sets, 2002/2003 and 2008/2009, analysed by pulsed field gel electrophoresis and tested for the Ambler class B metallo-β-lactamase (MBL) genes blaSPM-1, blaIMP and blaVIM. The results show a prevalence of one clone related to the SPM Brazilian clone in 2002/2003. In 2008/2009, P. aeruginosa isolates were mostly MBL negative, genetically diverse and unrelated to those that had been detected earlier. These findings suggest that the resistance to carbapenems by these recent P. aeruginosa isolates was not due to the spread of MBL-positive SPM-related clones, as often observed in Brazilian hospitals.

Oral live attenuated human rotavirus vaccine (RotarixTM) offers sustained high protection against severe G9P[8] rotavirus gastroenteritis during the first two years of life in Brazilian children

In a large Phase III trial conducted in 10 Latin American countries, the safety and efficacy of the live attenuated monovalent rotavirus vaccine RIX4414 was evaluated in 15,183 healthy infants followed up during the first two years of life. Belém was the only site in Brazil included in this multicentre trial. The study in Belém included a subset of 653 infants who were followed up until 24 months of age for protection against severe rotavirus gastroenteritis. These subjects were randomly assigned in a 1:1 ratio to receive two doses of vaccine (n = 328) or two doses of placebo (n = 325) at approximately two and four months of age. Of the 653 enrolled infants, 23 dropped out during the study period. For the combined two-year period, the efficacy of RIX4414 was 72.3% [95% confidence interval (CI) 37.5-89.1%] against severe rotavirus-related gastroenteritis, reaching a protection rate of 81.8% (95% CI 36.4-96.6%) against circulating wild-type G9 rotavirus strains. It is concluded that two doses of RIX4414 are highly efficacious against severe rotavirus gastroenteritis in Belém during the first two years of life and provide high protection against the worldwide emergence and spread of G9P[8] strains.

High rates of undiagnosed leprosy and subclinical infection amongst school children in the Amazon Region

Leprosy in children is correlated with community-level factors, including the recent presence of disease and active foci of transmission in the community. We performed clinical and serological examinations of 1,592 randomly selected school children (SC) in a cross-sectional study of eight hyperendemic municipalities in the Brazilian Amazon Region. Sixty-three (4%) SC, with a mean age of 13.3 years (standard deviation = 2.6), were diagnosed with leprosy and 777 (48.8%) were seropositive for anti-phenolic glycolipid-I (PGL-I). Additionally, we evaluated 256 house-hold contacts (HHCs) of the students diagnosed with leprosy; 24 (9.4%) HHC were also diagnosed with leprosy and 107 (41.8%) were seropositive. The seroprevalence of anti-PGL-I was significantly higher amongst girls, students from urban areas and students from public schools (p < 0.0001). Forty-five (71.4%) new cases detected amongst SC were classified as paucibacillary and 59 (93.6%) patients did not demonstrate any degree of physical disability at diagnosis. The results of this study suggest that there is a high rate of undiagnosed leprosy and subclinical infection amongst children in the Amazon Region. The advantages of school surveys in hyperendemic areas include identifying leprosy patients at an early stage when they show no physical disabilities, preventing the spread of the infection in the community and breaking the chain of transmission.

The roles of PKCδ and Src kinases in the glucocorticoid induction and the TGF-β superinduction of the mouse mammary tumor virus transcription in B lymphocytes

Résumé : Le virus tumoral de la glande mammaire de la souris (MMTV) est un rétrovirus provoquant le développement de tumeurs dans les glandes mammaires des souris susceptibles femelles. Au cours de son évolution, le virus s'est adapté et s'exprime dans des cellules spécialisées. Les lymphocytes B sont les premières cellules infectées et elles sont essentielles pour la propagation de l'infection aux glandes mammaires. Dans notre étude, le virus MMTV a été utilisé afin d'examiner les voies de signalisation induites par les glucocorticoïdes (dexaméthasone (dex), une hormone stéroïdienne) et le transforming growth factor-f3 (TGF-P, une cytokine), deux molécules impliquées dans l'activation de la transcription à partir du promoteur du MMTV dans les cellules B. Le TGF-P seul n'influence pas l'activité du promoteur du MMTV. Par contre, en synergie avec dex, le TGF-P provoque une super-induction de l'expression du promoteur par rapport à une stimulation par le glucocorticoïde seul. Cette super-induction est régulée par une famille de protéines, les Smads. Ainsi, dans les lymphocytes B, l'utilisation du MMTV a permis de mettre en évidence une nouvelle synergie entre les glueocortieoïdes et le TGF-p. pans ce travail, l'utilisation d'inhibiteurs pharmacologiques et de mutants « dominant-négatifs » nous a pet mis de démontrer qu'une Protéine Kinase C delta (PKC5) active est impliquée dans la transduction du signal lors de la réponse au dex ainsi que celle au TGF-P. Néanmoins, la PKC5 est régulée différemment dans chaque voie spécifique : la voie du TGF-p nécessitait l'activation du PKC5 par diacylglycerol (DAG) et la phosphorylation de tyrosines spécifiques, alors que la voie impliquant les glucocorticoïdes ne le nécessitait pas. Nous avons aussi démontré qu'une tyrosine kinase de la famille Src est responsable de la phosphorylation des tyrosines sur la PKC5. Les essais de kinase in vitro nous ont permis de découvrir que plusieurs Src kinases peuvent phosphoryler la PKC6 dans les cellules B et qu'elles étaient constitutivement actives. Enfin, nous avons montré qu'il existe une interaction protéine - protéine induite par dex, entre le récepteur aux glucocorticoïdes (GR) et la PKC5 dans les cellules B, une association qui n'a pas été démontrée auparavant. Par ailleurs, nous avons analysé les domaines d'interactions entre PKC5 et GR en utilisant les essais de «GST pull-down». Nos résultats montrent que le domaine régulateur de la PKC5 et celui qui interagit avec l'ADN du GR sont impliqués. En résumé, nous avons trouvé que dans une lignée lymphocytaire B, le virus MMTV utilise des mécanismes pour réguler à la fois la transcription et la voie de signalisation qui sont différents de ceux utilisés dans les cellules mammaires épithéliales et les fibroblastes. Nos découvertes pourraient être utilisées comme modèles pour l'étude de gènes cellulaires impliqués dans des processus tels qu'inflammation, immunité ou cancérogénèse. Summary: Mouse Mammary Tumor Virus (MMTV) is a retrovirus that causes tumors in the mammary glands of susceptible female mice and has adapted evolutionarily to be expressed in specialized cells. The B lymphocytes are the first cells to be infected by the MMTV and are essential for the spread of infection to the mammary glands. Here, we used the MMTV as a model system to investigate the signalling cascade induced by giucocorticoids (dexamethasone, "dex", a steroid hormone), and by Transforming Growth Factor-beta (TGF-P, a cytokine) leading to its transcriptional activation in B lymphocytes. By itself, TGF-I3 does not affect the basal activity of the MMTV promoter. However, TGF-13 significantly increases glucocorticoid-induced expression, through its effectors, the Smad factors. Thus, MMTV in B cells demonstrates a novel synergism between glucocorticoids and TGF-16. In this thesis project, we present evidence, based on the use of pharmacological inhibitors and of dominant-negative mutants, that an active Protein Kinase C delta (PKC6) is required as a signal transducer for the dex response and for the TGF-P superinduction as well. The PKC6 is differentially regulated in each specific pathway: whereas the TGF-13 superinduction required PKC6 to be activated by diacylglycerol (DAG) and to be phosphorylated at specific tyrosine residues, the glueocorticoid-induced pathway did not. We also showed that a protein tyrosine kinase of the Src family is responsible for the phosphorylation of tyrosines on PKC6. By performing in vitro kinase assays, we found that several Src kinases of B cells were able to phosphorylate PKC6 and that they were constitutively active. Finally, we demonstrate a dex-dependent functional protein-protein interaction between the glucocorticoid receptor (GR) and PKC6 in B cells, an association that has not been previously described. We further analysed the interacting domains of PKG6 and GR using in vitro GST pull-down assays, whereby the regulatory domain of PKC6 and the extended DNA-binding domain of the GR were involved. In summary, we found that in B-lymphoid cell lines, MMTV uses novel mechanisms of transcriptional control and signal transduction that are different from those at work in mammary epithelial or fibroblastic cells. These findings will be used as model for cellular genes involved in cellular processes such as immune functions, inflammation, or oncogenic transformation that may have a similar pattern of regulation.

Novel methicillin-resistant coagulase-negative Staphylococcus clone isolated from patients with haematological diseases at the Blood Bank Centre of Amazon, Brazil

Methicillin-resistant Staphylococcus remains a severe public health problem worldwide. This research was intended to identify the presence of methicillin-resistant coagulase-negative staphylococci clones and their staphylococcal cassette chromosome mec (SCCmec)-type isolate from patients with haematologic diseases presenting bacterial infections who were treated at the Blood Bank of the state of Amazonas in Brazil. Phenotypic and genotypic tests, such as SCCmec types and multilocus sequence typing (MLST), were developed to detect and characterise methicillin-resistant isolates. A total of 26 Gram-positive bacteria were isolated, such as: Staphylococcus epidermidis (8/27), Staphylococcus intermedius (4/27) and Staphylococcus aureus (4/27). Ten methicillin-resistant staphylococcal isolates were identified. MLST revealed three different sequence types: S. aureus ST243, S. epidermidis ST2 and a new clone of S. epidermidis, ST365. These findings reinforce the potential of dissemination presented by multi-resistant Staphylococcus and they suggest the introduction of monitoring actions to reduce the spread of pathogenic clonal lineages of S. aureus and S. epidermidis to avoid hospital infections and mortality risks.

Genetics of chloroquine-resistant malaria: a haplotypic view

The development and rapid spread of chloroquine resistance (CQR) in Plasmodium falciparum have triggered the identification of several genetic target(s) in the P. falciparum genome. In particular, mutations in the Pfcrt gene, specifically, K76T and mutations in three other amino acids in the region adjoining K76 (residues 72, 74, 75 and 76), are considered to be highly related to CQR. These various mutations form several different haplotypes and Pfcrt gene polymorphisms and the global distribution of the different CQR- Pfcrt haplotypes in endemic and non-endemic regions of P. falciparum malaria have been the subject of extensive study. Despite the fact that the Pfcrt gene is considered to be the primary CQR gene in P. falciparum , several studies have suggested that this may not be the case. Furthermore, there is a poor correlation between the evolutionary implications of the Pfcrt haplotypes and the inferred migration of CQR P. falciparum based on CQR epidemiological surveillance data. The present paper aims to clarify the existing knowledge on the genetic basis of the different CQR- Pfcrt haplotypes that are prevalent in worldwide populations based on the published literature and to analyse the data to generate hypotheses on the genetics and evolution of CQR malaria.