Clonal variation within a mucosal isolate derived from a patient with Leishmania (Viannia) braziliensis infection

Three isolates over 5 years from a patient with persistent relapsing mucosal leishmaniasis due to Leishmania (Viannia) braziliensis and 7 clones from one of these isolates were studied by zymodemes and scrodemes analysis. Results showed evidences of clonal phenotypic variation. Eight isoenzymes markers demonstrated clear differences on Cellulose Acetate (CA) and thin starch gel electrophoresis. Also a panel of specific monoclonal antibodies showed such differences. Our observations provide additional evidence that Leishmania (Viannia) braziliensis is composed by subpopulations of parasites with peculiar biochemical and antigenic characteristics.

Genetic structure of Neisseria meningitidis serogroup C epidemic strains in South Brazil

In the present study we report the results of an analysis, based on serotyping, multilocus enzyme electrophoresis (MEE), and ribotyping of N. meningitidis serogroup C strains isolated from patients with meningococcal disease (MD) in Rio Grande do Sul (RS) and Santa Catarina (SC) States, Brazil, as the Center of Epidemiology Control of Ministry of Health detected an increasing of MD cases due to this serogroup in the last two years (1992-1993). We have demonstrated that the MD due to N.meningitidis serogroup C strains in RS and SC States occurring in the last 4 years were caused mainly by one clone of strains (ET 40), with isolates indistinguishable by serogroup, serotype, subtype and even by ribotyping. One small number of cases that were not due to an ET 40 strains, represent closely related clones that probably are new lineages generated from the ET 40 clone referred as ET 11A complex. We have also analyzed N.meningitidis serogroup C strains isolated in the greater São Paulo in 1976 as representative of the first post epidemic year in that region. The ribotyping method, as well as MEE, could provide useful information about the clonal characteristics of those isolates and also of strains isolated in south Brazil. The strains from 1976 have more similarity with the actual endemic than epidemic strains, by the ribotyping, sulfonamide sensitivity, and MEE results. In conclusion, serotyping with monoclonal antibodies (C:2b:P1.3), MEE (ET 11 and ET 11A complex), and ribotyping by using ClaI restriction enzyme (Rb2), were useful to characterize these epidemic strains of N.meningitidis related to the increased incidence of MD in different States of south Brazil. It is mostly probable that these N.meningitidis serogroup C strains have poor or no genetic corelation with 1971-1975 epidemic serogroup C strains. The genetic similarity of members of the ET 11 and ET 11A complex were confirmed by the ribotyping method by using three restriction endonucleases.

Production of monoclonal antibodies anti-Taenia crassiceps cysticerci with cross-reactivity with Taenia solium antigens

We describe the production of the potential monoclonal antibodies (MoAbs) using BALB/c mice immunized with vesicular fluid (VF)-Tcra (T. crassiceps) antigen. Immune sera presented anti-VF-Tcra (<20kD) IgG and IgM antibodies with cross-reactivity with T. solium (Tso) antigen (8-12, 14, and 18 kD). After cell fusion, we selected 33 anti-Tcra and anti-Tso reactive IgM-clones and 53 anti-Tcra specific IgG-clones, 5 of them also recognizing Tso antigens. Two clones identified the 8-14 and 18kD peptides of VF-Tcra.

Biotyping, serotyping and ribotyping as epidemiological tools in the evaluation of Acinetobacter baumannii dissemination in hospital units, Sorocaba, São Paulo, Brazil

Dissemination of Acinetobacter baumannii strains in different units of a hospital in Sorocaba, São Paulo, Brazil was evaluated over a period of two years. By using biotyping, serotyping and ribotyping, 27 distinct clones were differentiated among 76 strains isolated between 1993-94, from clinical specimens of hospitalized patients. Two clones, 2:O4:A (biotype:serotype:ribotype) and 2:O29:A accounted for the majority of strains widely disseminated in the units during 1993. The introduction in the hospital setting, of a new clone, 6:O13:B, at the end of 1993 and its predominance through 1994 is discussed. Among 15 strains isolated from neonates, 6 (40%) belonged to the same clone, 2:O4:A. Interestingly, this clone was almost all recovered in neonatal intensive care unit, nursery and in pediatric unit. All strains were susceptible to imipenem and polymyxcin B. Multiresistant strains (up to 12 antimicrobial agents) accounted for 66.7% and 84.8% of the strains isolated in 1993 and in 1994, respectively.

Characterization of a clone from an adult worm cDNA library selected with anti-Schistosoma mansoni human antibodies dissociated from immune complexes: a preliminary report

Considering the scarcity of defined antigens, actually useful and reliable for use in the field studies, we propose an alternative method for selection of cDNA clones with potential use in the diagnosis of schistosomiasis. Human antibodies specific to a protein fraction of 31/32 kDa (Sm31/32), dissociated from immune complexes, are used for screening of clones from an adult worm cDNA library. Partial sequencing of five clones, selected through this strategy, showed to be related to Schistosoma mansoni: two were identified as homologous to heat shock protein 70, one to glutathione S-transferase, one to homeodomain protein, and one to a previously described EST (expressed sequence tag) of S. mansoni. This last clone was the most consistently reactive during the screening process with the anti-Sm31/32 antibodies dissociated from the immune complexes. The complete sequence of this clone was obtained and the translation data yielded only one ORF (open reading frame) that code for a protein with 57 amino acids. Based on this amino acid sequence two peptides were chemically synthesized and evaluated separately against a pool of serum samples from schistosomiasis patients and non-schistosomiasis individuals. Both peptides showed strong reactivity only against the positive pool, suggesting that these peptides may be useful as antigens for the diagnosis of schistosomiasis mansoni.

Molecular characterization of vancomycin-resistant Enterococci strains eight years apart from its first isolation in São Paulo, Brazil

E. faecium was the first reported VRE species, carrying the vanA gene in Brazil. In spite of this, vancomycin-resistant E. faecalis has become the predominant species in Brazilian hospitals. The aim of this study was to evaluate the genetic relatedness of VREs isolated in a Brazilian teaching hospital eight years apart from its first isolation. We analyzed 38 VRE strains obtained from 81 surveillance cultures of patients admitted to the four largest intensive care units in Hospital São Paulo in February, 2006. Presence of the vanA gene was assayed by PCR and PFGE analysis was used for molecular characterization. All VRE strains carried the vanA gene. Two distinct clonal groups were observed among vancomycin-resistant E. faecalis. Vancomycin-resistant E. faecium belonged to five distinct clones were demonstrated by molecular typing. All of these clones were different from the first vancomycin-resistant enterococci clone isolated eight years ago in our hospital.

Frequency of Mycoplasma hominis and Ureaplasma urealyticum infections in women with systemic lupus erythematosus

Ureaplasma urealyticum (UU) and Mycoplasma hominis (MH) have been detected in the urine of women with systemic lupus erythematosus (SLE). We evaluated the presence of these mycoplasma in the endocervix of women presenting SLE. A total of 40 SLE patients (mean age 40.2 years), and 51 healthy women (mean age 30.9 years), were studied. Endocervical swabs were cultured in specific liquid media for MH or UU, detected by a quantitative color assay, and considered positive at >10³ dilutions. Statistical analysis was performed using the two-tailed Fisher test. UU was detected in 52.5 % of patients and in 11.8% of controls (p= 0.000059). MH was detected in 20% of patients and 2% controls (p=0.003905). Both mycoplasmas were detected in 7.3% patients and 0% controls (p<0.000001). The results reported here corroborate the association of the mycoplasma infection and SLE. Thus, these agents may stimulate the production of autoreactive clones.

Molecular characterization of clinical multiresistant isolates of Acinetobacter sp. from hospitals in Porto Alegre, State of Rio Grande do Sul, Brazil

INTRODUCTION: Hospitals around the world have presented multiresistant Acinetobacter sp. outbreaks. The spread of these isolates that harbor an increasing variety of resistance genes makes the treatment of these infections and their control within the hospital environment more difficult. This study aimed to evaluate the occurrence and dissemination of Acinetobacter sp. multiresistant isolates and to identify acquired resistance genes. METHODS: We analyzed 274 clinical isolates of Acinetobacter sp. from five hospitals in Porto Alegre, RS, Brazil. We evaluated the susceptibility to antimicrobial, acquired resistance genes from Ambler's classes B and D, and performed molecular typing of the isolates using enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) technique. RESULTS: A high (68%) percentage of multiresistant isolates of Acinetobacter sp. was observed, and 69% were resistant to carbapenems. We identified 84% of isolates belonging to species A. baumannii because they presented the gene blaOXA-51. The gene blaOXA-23 was detected in 62% of the isolates, and among these, 98% were resistant to carbapenems. Using the ERIC-PCR technique, we identified clones of Acinetobacter sp. spread among the four hospitals analyzed during the sampling period. CONCLUSIONS: The data indicate the dissemination of Acinetobacter sp. isolates among hospitals and their permanence in the hospital after one year.

Phenotypic and molecular characterization of antimicrobial resistance and virulence factors in Pseudomonas aeruginosa clinical isolates from Recife, State of Pernambuco, Brazil

INTRODUCTION: The emergence of carbapenem resistance mechanisms in Pseudomonas aeruginosa has been outstanding due to the wide spectrum of antimicrobial degradation of these bacteria, reducing of therapeutic options. METHODS: Sixty-one clinical strains of P. aeruginosa isolated from five public hospitals in Recife, Pernambuco, Brazil, were examined between 2006 and 2010, aiming of evaluating the profiles of virulence, resistance to antimicrobials, presence of metallo-β-lactamase (MBL) genes, and clonal relationship among isolates. RESULTS: A high percentage of virulence factors (34.4% mucoid colonies; 70.5% pyocyanin; 93.4% gelatinase positives; and 72.1% hemolysin positive) and a high percentage of antimicrobial resistance rates (4.9% pan-resistant and 54.1% multi-drug resistant isolates) were observed. Among the 29 isolates resistant to imipenem and/or ceftazidime, 44.8% (13/29) were MBL producers by phenotypic evaluation, and of these, 46.2% (6/13) were positive for the blaSPM-1 gene. The blaIMP and blaVIM genes were not detected. The molecular typing revealed 21 molecular profiles of which seven were detected in distinct hospitals and periods. Among the six positive blaSPM-1 isolates, three presented the same clonal profile and were from the same hospital, whereas the other three presented different clonal profiles. CONCLUSIONS: These results revealed that P. aeruginosa is able to accumulate different resistance and virulence factors, making the treatment of infections difficult. The identification of blaSPM-1 genes and the dissemination of clones in different hospitals, indicate the need for stricter application of infection control measures in hospitals in Recife, Brazil, aiming at reducing costs and damages caused by P. aeruginosa infections.

Community-acquired methicillin-resistant Staphylococcus aureus carrying SCCmec type IV in southern Brazil

INTRODUCTION: Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen commonly associated with nosocomial infections. However, it has also been associated with community-acquired skin and soft tissue infections (CA-MRSA). There are few data on the identification and prevalence of CA-MRSA infections in Brazil. METHODS: This is a cross-sectional study of 104 patients with community-acquired skin infections attending two health care centers in Porto Alegre, southern Brazil. MRSA isolates were characterized by molecular methods, including detection of the mecA gene by PCR, gene SCCmec typing, Panton-Valentine leukocidin (PVL) detection, pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST). RESULTS: From the 104 samples, 58 Staphylococcus aureus isolates were obtained, of which five (8.6%) had a CA-MRSA-resistant profile. All five isolates had the mecA gene and amplified to SCCmec type IV. Analysis of chromosomal DNA by PFGE revealed the presence of two clusters related to international clones (OSPC and USA 300), with a Dice similarity coefficient >80%. The study was complemented by MLST, which detected three different strains: ST30, ST8, and ST45, the latter not presenting any relation with the clones compared in PFGE. CONCLUSIONS: The presence of CA-MRSA reveals an important change in the epidemiology of this pathogen and adds new elements to the knowledge of the molecular biology of infections by MRSA with SCCmec type IV in southern Brazil.

The occurrence and dissemination of methicillin and vancomycin-resistant Staphylococcus in samples from patients and health professionals of a university hospital in Recife, State of Pernambuco, Brazil

Introduction Methicillin-resistant Staphylococcus aureus (MRSA) strains have been responsible for many nosocomial outbreaks. Within hospitals, colonized employees often act as reservoirs for the spread of this organism. This study collected clinical samples of 91 patients admitted to the intensive care unit (ICU), hemodialysis/nephrology service and surgical clinic, and biological samples from the nasal cavities of 120 professionals working in those environments, of a University Hospital in Recife, in the State of Pernambuco, Brazil. The main objective of this study was to determine the occurrence and dissemination of methicillin- and vancomycin-resistant Staphylococcus spp. Methods The isolates obtained were tested for susceptibility to oxacillin and vancomycin and detection of the mecA gene. In addition, the isolates were evaluated for the presence of clones by ribotyping-polymerase chain reaction (PCR). Results MRSA occurrence, as detected by the presence of the mecA gene, was more prevalent among nursing technicians; 48.1% (13/27) and 40.7% (11/27) of the isolates were from health professionals of the surgical clinic. In patients, the most frequent occurrence of mecA-positive isolates was among the samples from catheter tips (33.3%; 3/9), obtained mostly from the hemodialysis/nephrology service. Eight vancomycin-resistant strains were found among the MRSA isolates through vancomycin screening. Based on the amplification patterns, 17 ribotypes were identified, with some distributed between patients and professionals. Conclusions Despite the great diversity of clones, which makes it difficult to trace the source of the infection, knowledge of the molecular and phenotypic profiles of Staphylococcus samples can contribute towards guiding therapeutic approaches in the treatment and control of nosocomial infections.

Molecular analysis of methicillin-resistant Staphylococcus aureus dissemination among healthcare professionals and/or HIV patients from a tertiary hospital

Abstract INTRODUCTION: Methicillin-resistant Staphylococcus aureus (MRSA) is a nosocomial pathogen in community settings. MRSA colonized individuals may contribute to its dissemination; the risk of MRSA infection is increased in human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS) patients, although the prevalence of colonization in this group is not well established. The present study addressed this issue by characterizing MRSA isolates from HIV/AIDS patients and their healthcare providers (HCPs) to determine whether transmission occurred between these two populations. METHODS: A total of 24 MRSA isolates from HIV-infected patients and five from HCPs were collected between August 2011 and May 2013. Susceptibility to currently available antimicrobials was determined. Epidemiological typing was carried out by pulsed-field gel electrophoresis, multilocus sequence typing, and Staphylococcus cassette chromosome (SCCmec) typing. The presence of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) and heterogeneous daptomycin-resistant Staphylococcus aureus (hDRSA) was confirmed by population analysis profile. Isolates characterized in this study were also compared to isolates from 2009 obtained from patients at the same hospital. RESULTS: A variety of lineages were found among patients, including ST5-SCCmecII and ST30-SCCmecIV. Two isolates were Panton-Valentine leukocidin-positive, and hVISA and hDRSA were detected. MRSA isolates from two HCPs were not related to those from HIV/AIDS patients, but clustered with archived MRSA from 2009 with no known relationship to the current study population. CONCLUSIONS: ST105-SCCmecII clones that colonized professionals in 2011 and 2012 were already circulating among patients in 2009, but there is no evidence that these clones spread to or between HIV/AIDS patients up to the 7th day of their hospitalization.

Estudo comparativo do balanço hídrico em um clone diplóide (IAN-873) e outro poliplóide (IAC-222) de Hevea brasiliensis Muell. Arg

Foi montado um experimento em condições de campo, envolvendo clones diplóides e poliplóides de Hevea brasiliensis Muell. Arg., objetivando o estudo comparativo de balanço hídrico com base na análise dos seguintes parâmetros: potencial hídrico foliar, resistência estomática, transpiração, conteúdo percentual de água, déficit de saturação foliar, análise de microestrutura epidérmica e do índice de eficiência hídrica. De um modo geral, os resultados sugerem maior capacidade hídrica ao clone poliplóide IAC-222, conseqüente do seu maior índice de eficiência hídrica, podendo ser definido como vantajoso sobre o diplóide por se re-hidratar mais rapidamente após períodos críticos.

Associação entre características da parte aérea e do sistema radicular em mudas de guaranazeiro por correlações canônicas

Este trabalho teve como objetivo quantificar o grau de asociação entre variáveis de parte aérea e de raízes em mudas clonadas de guaranazeiro, utilizando-se correlações canônicas, a fim de aperfeiçoar o procedimento de seleção de mudas para garantir o aumento da porcentagem de sobrevivência das mudas após o plantio. Foram avaliados dois grupos de variáveis em mudas aptas ao plantio definitivo de 36 clones de guaranazeiro. O delineamento usado foi o aleatorizado em blocos com cinco repetições e 10 plantas por parcela, sob condições de viveiro. Os caracteres avaliados foram submetidos à análise de correlações canônicas. Utilizou-se a análise de correlações canônicas. O grupo de variáveis da parte aérea não se mostrou independente do grupo de variáveis do sistema radicular. Através de seleção baseada em variáveis da parte aérea pode-se melhorar o sistema radicular, principalmente através do maior comprimento do ramo (CRA). A seleção de clones de guaraná para maior peso de raiz pode ser efetuada de forma indireta, realizando-se mensurações do comprimento dos ramos, o que evita a necessidade de se destruir as mudas.

Resistência do morangueiro (Fragaria híbridos) à Mycosphaerella fragariae (TUL.) LIND. (Ramularia tulasnei SACC.)

Foram testados 49 clones de morangueiro quanto a reação à incidência natural de Ramularia tulasnei sob condições de campo, em Piracicaba, SP. A avaliação foi realizada de acordo com uma escala de 1 (ausência de sintomas) a 6 (severa incidência) na fase de maior severidade da doença e os clones classificados em resistentes (1,00 a 2,30), moderadamente resistentes (2,31 a 3,70) e suscetíveis (3,71 a 5,00). Identificaram-se 11 clones resistentes, 31 moderadamente resistentes e 7 suscetíveis. Os resistentes foram "I-2008" (grau 1,00 ± 0,00), "IAC-2713", "Camanducaia (IAC-3530)", "A. Brucknner (I-2492)","IAC-3113 x (IAC-2712 x 1-2008-1)10", "IAC-4326", "IAC-3530 x IAC-2747-2", "Kon-woy (1-3846)", "Atibaia (IAC-4325)", "1-4896 -4" e "IAC-4749" e o mais suscetível foi "Jundiaí (IAC-4204)" (grau 5,00 ± 0,41).