Comparación de tres técnicas para subtipificación molecular de Listeria monocytogenes / Comparison of three molecular subtyping techniques for Listeria monocytogenes

Abstract Listeria monocytogenes is a foodborne pathogen. The recent alert for L. monocytogenes in vegetables from Argentina warns about the importance of reinforcing its isolation, characterization and subtyping in food, clinical and environmental samples. The aim of the present study was to compare the discriminatory power of enterobacterial repetitive interpower; genic consensus polymerase chain reaction (ERIC-PCR), automated ribotyping and pulsed-field gel electrophoresis (PFGE) to subtype strains of L. monocytogenes isolated from Argentine meat and environmental samples. Simpson's Diversity Index (DI) was calculated on the basis of based on the dendrograms obtained in the by cluster analysis, showing the following discriminatory power: ApaI-PFGE (0.980), AscI-PFGE (0.966), ribotyping (0.912), ERIC-PCR (0.886). The ID values between ApaI- and AscI-PFGE and between ribotyping and ERIC-PCR were not significantly different. Of the three techniques evaluated, PFGE showed the highest discriminatory power. However, the subtyping techniques should be accompanied by effective food monitoring strategies and reliable clinical and epidemiological studies.

Resumen Listeria monocytogenes es un patógeno alimentario. La reciente alerta por la presencia de L. monocytogenes en vegetales en Argentina advierte sobre la importancia de reforzar el aislamiento, la caracterización y la subtipificación de esta bacteria en muestras clínicas de alimentos y ambientales. El objetivo del presente estudio fue comparar el poder discriminatorio de enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR), la ribotipificación automatizada y la pulsed-field gel electrophoresis (PFGE) para subtipificar cepas de L. monocytogenes aisladas de carne y de muestras ambientales en Argentina. El índice de diversidad (ID) de Simpson, calculado a partir de los dendrogramas obtenidos en el análisis de agrupamiento, mostró los siguientes resultados: Apal-PFGE (0,980), AscI-PFGE (0,966), riboti-pado (0,912), ERIC-PCR (0,886). Los valores obtenidos no fueron significativamente diferentes al comparar entre Apal- y AscI-PFGE, ni entre ribotipadoy ERIC-PCR. De las técnicas evaluadas, la PFGE presentó el mayor poder discriminatorio. Sin embargo, las técnicas de subtipificación deberían acompañarse de estrategias de control de los alimentos efectivas y de estudios clínicos y epidemiológicos confiables.

La microbiota según la topografía gástrica en pacientes con bajo y con alto riesgo de cáncer gástrico en Nariño, Colombia / Microbiota according to gastric topography in patients with low or high risk of gastric cancer in Nariño, Colombia

Resumen Introducción. La inflamación del antro gástrico por Helicobacter pylori aumenta el riesgo de úlcera duodenal, y la del cuerpo gástrico puede producir gastritis atrófica e incrementar la probabilidad de cáncer gástrico. Estas reacciones inflamatorias diferenciadas según su localización, podrían explicarse por la composición de la microbiota gástrica asociada con H. pylori. Objetivo. Identificar y comparar la microbiota del antro y del cuerpo del estómago en individuos de dos poblaciones: una con alto riesgo y otra con bajo riesgo de cáncer gástrico en Nariño, Colombia. Materiales y métodos. Se incluyeron biopsias del cuerpo y el antro gástrico de pacientes con gastritis no atrófica o con gastritis atrófica y metaplasia. La microbiota se definió por secuenciación de la región V3-V4 del gen 16S del ARNr de H. pylori (illumina-MiSeq™). Las unidades taxonómicas operativas se clasificaron utilizando las bases de datos BLASTn y RDPII. Las diferencias entre las poblaciones microbianas del antro y del cuerpo gástrico se evaluaron mediante el análisis de varianza multivariado con base en permutaciones (Permutational Multivariate Analysis of Variance, PERMANOVA) y análisis multivariados. Resultados. La clase Epsilonproteobacteria representada por H. pylori fue más abundante en las biopsias del antro y del cuerpo de los individuos con gastritis no atrófica (>50 %), en tanto que, en los individuos con gastritis no atrófica, esta clase correspondió al 20 % con una mayor diversidad metagenómica. La infección por H. pylori disminuyó significativamente la diversidad metagenómica del antro (p=0,005), en comparación con la del cuerpo gástrico. Conclusiones. Los grupos bacterianos involucrados en la disbacteriosis pueden colonizar ambas regiones topográficas del estómago, independientemente de las reacciones sectorizadas de inflamación. La infección por H. pylori asociada con la microbiota gástrica está relacionada con su localización en el estómago, el tipo de lesión y el mayor o menor riesgo de cáncer gástrico, lo que sugiere su importancia en la disbacteriosis y la de esta en la enfermedad gástrica.

Abstract Introduction: Inflammation in the gastric antrum caused by Helicobacter pylori increases the risk of duodenal ulcer while inflammation in the body generates atrophic gastritis and increased risk of gastric cancer. These inflammatory responses according to gastric topography could be explained by the composition of the gastric microbiota associated with H. pylori. Objective: To identify and compare the microbiota of the gastric antrum and body of individuals from two populations, one with high risk and one with low risk of gastric cancer from Nariño, Colombia. Materials and methods: Biopsies of the gastric antrum and body of patients with non-atrophic gastritis or metaplastic atrophic gastritis were included. The microbiota was defined by sequencing the 16S rRNA gene, V3-V4 region, (illumina-MiSeq™). The operational taxonomic units were classified using the BLASTn and RDPII databases. The differences among microbial populations were evaluated with the PERMANOVA and multivariate analyses. Results: The Epsilonproteobacteria class represented by H. pylori was more abundant in the antrum and body biopsies of individuals with metaplastic atrophic gastritis (>50%) while in individuals with non-atrophic gastritis it was 20 % and had greater metagenomic diversity. Helicobacter pylori infection significantly decreases the metagenomic diversity of the gastric antrum (p=0.005) compared to that of the body. Conclusions: The bacterial groups involved in the dysbiosis can colonize both topographic regions of the stomach, regardless of the sectorized inflammation responses. Helicobacter pylori infection associated with the gastric microbiota is related to its localization in the stomach, the type of lesion, and the population at risk of gastric cancer, which suggests its importance in microbial dysbiosis and gastric disease.

Melioidosis en Colombia, descripción de un caso clínico y consideraciones epidemiológicas / Melioidosis in Colombia, description of a clinical case and epidemiological considerations

Resumen La melioidosis es una enfermedad infecciosa causada por Burkholderia pseudomallei cuyo diagnóstico clínico puede ser difícil debido a su variada presentación clínica y a las dificultades del diagnóstico microbiológico, por lo cual pueden requerirse técnicas moleculares para su adecuada identificación una vez se sospecha su presencia. Son pocos los antibióticos disponibles para el tratamiento de esta enfermedad y, además, deben usarse durante un tiempo prolongado. Aunque se conoce por ser endémica en Tailandia, Malasia, Singapur, Vietnam y Australia, en Colombia se han reportado algunos pocos casos. Se presenta un caso de melioidosis en la región norte de Colombia, se hace una revisión de las características clínicas y el tratamiento, y se describe la epidemiología local de esta enfermedad.

Abstract Melioidosis is an infectious disease caused by Burkholderia pseudomallei whose clinical diagnosis can be difficult due not only to its varied clinical presentation but also to the difficulties in the microbiological diagnosis.Thus, it may be necessary to use molecular techniques for its proper identification once it is suspected. There are few antibiotics available for the treatment of this disease, which must be used over a long period of time. Although it is known to be endemic in Thailand, Malaysia, Singapore, Vietnam, and Australia, in Colombia there are few reported cases. We describe a case of melioidosis in the northern region of Colombia. Additionally, we review its clinical characteristics and treatment and we describe the local epidemiology of this disease.

Rapid and accurate diagnosis of Clostridium difficile infection by real-time polymerase chain reaction

BACKGROUND/AIMS: The incidence and severity of Clostridium difficile infection (CDI) have increased worldwide, resulting in a need for rapid and accurate diagnostic methods. METHODS: A retrospective study was conducted to compare CDI diagnosis methods between January 2014 and December 2014. The stool samples, which were obtained in presumptive CDI patients, were compared for their diagnostic accuracy and rapidity, including real-time polymerase chain reaction (PCR) of toxin genes, C. difficile toxin assay, and culture for C. difficile. RESULTS: A total of 207 cases from 116 patients were enrolled in this study and 117 cases (56.5%) were diagnosed as having CDI. Among the 117 cases, the sensitivities of real-time PCR, C. difficile toxin assay, and culture for C. difficile were 87.2% (102 cases; 95% CI, 80.7%–92.8%), 48.7% (57 cases; 95% CI, 41.0%–59.8%), and 65.0% (76 cases; 95% CI, 60.2%–78.5%), respectively (P < 0.005). Notably, 34 cases (29.0%) were diagnosed with CDI by real-time PCR only. The time required to obtain results was 2.27 hours (136.62±82.51 minutes) for real-time PCR, 83.67 hours (5,020.66±3,816.38 minutes) for toxin assay, and 105.79 hours (6,347.68±3,331.46 minutes) for culture (P < 0.005), respectively. CONCLUSIONS: We confirmed that real-time PCR of toxin genes is the most effective diagnostic method for accurate and early diagnosis of CDI. It also helps to diagnose hypervirulent CDI, such as ribotype 027 infection.

Simultaneous detection and characterization of toxigenic Clostridium difficile directly from clinical stool specimens / 医学前沿

We employed a multiplex polymerase chain reaction (PCR) coupled with capillary electrophoresis (mPCR-CE) targeting six Clostridium difficile genes, including tpi, tcdA, tcdB, cdtA, cdtB, and a deletion in tcdC for simultaneous detection and characterization of toxigenic C. difficile directly from fecal specimens. The mPCR-CE had a limit of detection of 10 colony-forming units per reaction with no cross-reactions with other related bacterial genes. Clinical validation was performed on 354 consecutively collected stool specimens from patients with suspected C. difficile infection and 45 isolates. The results were compared with a reference standard combined with BD MAX Cdiff, real-time cell analysis assay (RTCA), and mPCR-CE. The toxigenic C. difficile species were detected in 36 isolates and 45 stool specimens by the mPCR-CE, which provided a positive rate of 20.3% (81/399). The mPCR-CE had a specificity of 97.2% and a sensitivity of 96.0%, which was higher than RTCA (x = 5.67, P = 0.017) but lower than BD MAX Cdiff (P = 0.245). Among the 45 strains, 44 (97.8%) were determined as nonribotype 027 by the mPCR-CE, which was fully agreed with PCR ribotyping. Even though ribotypes 017 (n = 8, 17.8%), 001 (n = 6, 13.3%), and 012 (n = 7, 15.6%) were predominant in this region, ribotype 027 was an important genotype monitored routinely. The mPCR-CE provided an alternative diagnosis tool for the simultaneous detection of toxigenic C. difficile in stool and potentially differentiated between RT027 and non-RT027.

Detección de unidades discretas de tipificación de Trypanosoma cruzi en triatominos recolectados en diferentes regiones naturales de Perú / Identifying Trypanosoma cruzi discreet typing units in triatomines collected in different natural regions of Perú

Resumen Introducción. Trypanosoma cruzi se ha dividido en seis unidades taxonómicas discretas (Discreet Typing Units, DTU) denominadas TcI, TcII, TcIII, TcIV, TcV y TcVI. Aún se desconocen los factores determinantes de la dinámica de la transmisión vectorial de los genotipos de T. cruzi en las diferentes regiones geográficas de distribución de la enfermedad de Chagas en Perú. Objetivo. Detectar y tipificar las unidades taxonómicas discretas de T. cruzi en las heces de siete especies de triatominos (Panstrongylus chinai, P. geniculatus, P. herreri, Rhodnius robustus, R. pictipes, Triatoma carrioni y T. infestans), capturados en ocho departamentos de diferentes regiones naturales de Perú. Materiales y métodos. Se examinaron 197 insectos para la detección de tripanosomas. Se extrajo el ADN del contenido intestinal de cada insecto y se amplificó mediante reacción en cadena de la polimerasa (PCR) de los genes kDNA, SL-IR, 24Sa rRNA y 18Sa RNA para detectar las DTU de T. cruzi. Resultados. Se detectaron cinco infecciones con T. rangeli y 113 con T. cruzi. De estas últimas, fue posible identificar 95 de TcI (dos en P. chinai, una en P. geniculatus, 68 en P. herreri, cuatro en R. pictipes, siete en R. robustus, una en T. carrioni, y 12 en T. infestans); cinco de TcII (cuatro en P. herreri, una en T. infestans); cuatro de TcIII (tres en P. herreri, una en R. robustus) y cuatro infecciones de TcIV en P. herreri. Conclusión. Este es el primer trabajo de caracterización a gran escala de T. cruzi en el intestino de vectores de importancia epidemiológica en Perú, orientado a generar información básica que permita entender la dinámica de la transmisión vectorial de T. cruzi en esta región del continente.

Abstract Introduction: Trypanosoma cruzi has been divided by international consensus into six discrete typing units (DTU): TcI, TcII, TcIII, TcIV, TcV y TcVI. The factors determining the dynamics of T. cruzi genotypes vector transmission of Chagas' disease in the different geographical regions of Perú are still unknown. Objective: To detect and type T. cruzi DTUs from the faeces of seven species of triatomines (Panstrongylus chinai, P. geniculatus, P. herreri, Rhodnius robustus, R. pictipes, Triatoma carrioni and T. infestans) captured in eight departments from different natural regions of Perú. Materials and methods: We examined 197 insects for detecting trypanosomes. DNA was extracted from each insect intestinal contents and PCR amplification of kDNA, SL-IR, 24Sa rRNA and 18Sa RNAwas performed for detecting T. cruzi DTUs. Results: Five T. rangeli and 113 T. cruzi infections were detected; 95 of the latter were identified as TcI (two in P. chinai, one in P. geniculatus, 68 in P. herreri, four in R. pictipes, seven in R. robustus, one in T. carrioni, 12 in T. infestans), five as TcII (four in P. herreri, one in T. infestans), four as TcIII (three in P. herreri, one in R. robustus) and four TcIV infections in P. herreri. Conclusions: This is the first study which has attempted a large-scale characterization of T. cruzi found in the intestine of epidemiologically important vectors in Perú, thus providing basic information that will facilitate a better understanding of the dynamics of T. cruzi vector transmission in Perú.

Tipificación molecular de aislados de vibrio cholerae O1 causantes de dos brotes de cólera en Venezuela / Molecular typing of vibrio cholerae O1 isolates causing two outbreaks of cholera in Venezuela

En Venezuela, en junio de 1996, se reportó que los casos de cólera eran causados por V. cholerae O1 serotipo Ogawa. A finales de 1998 se detectó un segundo brote de cólera causado por V. cholerae O1 serotipo Inaba resistente a la ampicilina y el trimetoprim-sulfametoxazol. Para estudiar las relaciones entre las cepas se examinaron veinticinco aislados de Vibrio cholerae O1 obtenidos desde 1996 a 2000 en Venezuela, para determinar la presencia de genes de virulencia y perfiles genómicos. Mediante la reacción en cadena de la polimerasa se determinó la presencia de genes de virulencia. Para determinar el perfil genómico de los aislamientos se utilizó ribotipificación y electroforesis en gel de campo pulsado (PFGE). Todos los aislados resultaron positivos para los genes ctxA, ctxB, zot y ace. El análisis RFLP de los genes RNAr mostró un único patrón de ribotipo V. El análisis de PFGE mostró una similitud de 91,5% independientemente del año o lugar de aislamiento, lo que indica la relación genómica entre los aislados. En conjunto, los datos sugieren que la cepa de V. cholerae O1 resistente a los antibióticos que apareció en 1998 surgió de la cepa epidémica anterior o de otro estrechamente relacionado con el clon anterior, con cambio de serotipo y ganancia de determinantes de resistencia a antibióticos. Es muy importante monitorear continuamente la aparición de la variantes porque mejorará la comprensión de la evolución de nuevos clones de V. cholerae

In Venezuela, cholera reported in June 1996 was caused by V. cholerae O1 serotype Ogawa. Second outbreak of cholera caused by V. cholerae O1 serotype Inaba, resistant to ampicillin and trimethoprim- Sulfamethoxazole, was notify at the end of 1998. Twenty-five isolates of Vibrio cholerae O1 obtained from 1996 to 2000 in Venezuela were examined to study the relationships between strains, presence of virulence genes and genomic profiles. Presence of virulence genes was detected by Polymerase Chain Reaction. Ribotyping and pulsed-field gel electrophoresis (PFGE) were used to determine the genomic profile of isolates. All isolates shown PCR product for ctxA, ctxB, zot and ace genes. RFLP analysis of rRNA gene showed one unique pattern from ribotype V. PFGE analysis revealed a similarity of 91.5%, regardless year or place of isolation, suggesting genomic relatedness among them. Overall, these data suggest that antibiotic resistant V. cholerae O1 El Tor strain that appeared in 1998 emerged from the previous epidemic strain or from another closely related to the previous clone. It is important the continuous monitor the emergence of variants because it will improve our understanding of the evolution of new clones V. cholerae

Effect of medicinal plants, Heavy metals and antibiotics against P pathogenic bacteria isolated from raw, Boiled and pasteurized milk

Present study has been undertaken to isolate and identify the bacterial flora in raw, boiled and pasteurized milk. Agar disc diffusion method was used to determine their sensitivity using medicinal plants, antibiotics and heavy metals. Methylene blue reduction test was used to test the quality of milk samples. Total 10 pathogenic strains were isolated, five strains were isolated from raw milk, three from boiled milk and 2 two from pasteurized milk. To determine optimum conditions for growth, these pathogenic microorganisms were incubated at various temperatures and pH. Gram's staining and biochemical tests revealed that these pathogenic bacteria include Lactobacillus sp., E. coll, Salmonella sp., Pseudomonas sp., Streptococcus sp. and Staphylococcus. Ribotyping revealed S2 as Pseudomonas fluorescens, S5 as Lactococcus lactis and S9 as Lactobacillus acidophilus. Prevalence of pathogenic organisms provided the evidence that contamination of milk arises during milking, transportation and storage of milk. Raw milk is more contaminated than other two types of milk because it contains highest percentage of pathogenic organisms and pasteurized milk was found to be of best quality among three types. So it is recommended to drink milk after proper boiling or pasteurization. Proper pasteurization and hygienic packing of milk is essential to minimize contamination in milk which can save human beings from many milk borne diseases. Our study suggests that antimicrobial use in animal husbandry should be minimized to reduce the hazard of antibiotic resistance. Plant extracts are better alternative against pathogenic bacteria in milk

Isolation of recombinant antibodies directed against surface proteins of Clostridium difficile

Abstract Clostridium difficile has emerged as an increasingly important nosocomial pathogen and the prime causative agent of antibiotic-associated diarrhoea and pseudomembranous colitis in humans. In addition to toxins A and B, immunological studies using antisera from patients infected with C. difficile have shown that a number of other bacterial factors contribute to the pathogenesis, including surface proteins, which are responsible for adhesion, motility and other interactions with the human host. In this study, various clostridial targets, including FliC, FliD and cell wall protein 66, were expressed and purified. Phage antibody display yielded a large panel of specific recombinant antibodies, which were expressed, purified and characterised. Reactions of the recombinant antibodies with their targets were detected by enzyme-linked immunosorbent assay; and Western blotting suggested that linear rather than conformational epitopes were recognised. Binding of the recombinant antibodies to surface-layer proteins and their components showed strain specificity, with good recognition of proteins from C. difficile 630. However, no reaction was observed for strain R20291—a representative of the 027 ribotype. Binding of the recombinant antibodies to C. difficile M120 extracts indicated that a component of a surface-layer protein of this strain might possess immunoglobulin-binding activities. The recombinant antibodies against FliC and FliD proteins were able to inhibit bacterial motility.

Isolation and characterization of arsenic resistant bacteria from wastewater

The present study proposed the isolation of arsenic resistant bacteria from wastewater. Only three bacterial isolates (MNZ1, MNZ4 and MNZ6) were able to grow in high concentrations of arsenic. The minimum inhibitory concentrations of arsenic against MNZ1, MNZ4 and MNZ6 were 300 mg/L, 300 mg/L and 370 mg/L respectively. The isolated strains showed maximum growth at 37 ºC and at 7.0 pH in control but in arsenite stress Luria Bertani broth the bacterial growth is lower than control. All strains were arsenite oxidizing. All strains were biochemically characterized and ribotyping (16S rRNA) was done for the purpose of identification which confirmed that MNZ1 was homologous to Enterobacter sp. while MNZ4 and MNZ6 showed their maximum homology with Klebsiella pneumoniae. The protein profiling of these strains showed in arsenic stressed and non stressed conditions, so no bands of induced proteins appeared in stressed conditions. The bacterial isolates can be exploited for bioremediation of arsenic containing wastes, since they seem to have the potential to oxidize the arsenite (more toxic) into arsenate (less toxic) form.

Rich bacterial assemblages from Maritime Antarctica (Potter Cove, South Shetlands) reveal several kinds of endemic and undescribed phylotypes / Una rica colección de bacterias marinas antárticas (Caleta Potter, islas Shetlands del Sur) revela diversos filotipos endémicos y previamente no descritos

.

Bacterial richness in maritime Antarctica has been poorly described to date. Phylogenetic affiliation of seawater free-living microbial assemblages was studied from three locations near the Argentinean Jubany Station during two Antarctic summers. Sixty 16S RNA cloned sequences were phylogenetically affiliated to Alphaproteobacteria (30/60 clones), Gammaproteobacteria(19/60 clones), Betaproteobacteria and Cytophaga-Flavobacteriia- Bacteroides (CFB), which were (2/60) and (3/60) respectively. Furthermore, six out of 60 clones could not be classified. Both, Alphaproteobacteria and Gammaproteobacteria, showed several endemic and previously undescribed sequences. Moreover, the absence of Cyanobacteria sequences in our samples is remarkable. In conclusion, we are reporting a rich sequence assemblage composed of widely divergent isolates among themselves and distant from the most closely related sequences currently deposited in data banks.

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. .

Clostridium difficile Infection: What's New?

Since 2000, Clostridium difficile infection has increased substantially in both hospital-acquired and community-acquired diarrhea, not only in North America but also in Europe. There was a steady increase in the incidence and severity of C. difficile infection over the past decade, associated with significantly higher morbidity and mortality. The major risk factors for C. difficile infection appear to be the use of new antimicrobial therapy, long-term hospitalization in old age and emerging hypervirulent strains, such as various ribotypes. Rapid and accurate diagnosis of C. difficile infection is necessary for appropriate treatment as well as reliable epidemiological data. Currently available treatment options are withdrawal of the suspected offending antibiotics and then treating patients with highly effective antibiotics for C. difficile. Multiple recurrence or acute fulminant C. difficile infection could be treated with fecal microbiota transplantation. Promising therapies for treating C. difficile infection should always be equipped with high efficacy and safety in the future.

Clinical and Microbiologic Characteristics of Clostridium difficile Infection Caused by Binary Toxin Producing Strain in Korea

BACKGROUND: Binary toxin-producing Clostridium difficile infections (CDI) are known to be more severe and to cause higher case fatality rates than those by binary toxin-negative isolates. There has been few data of binary toxin-producing CDI in Korea. Objective of the study is to characterize clinical and microbiological trait of CDI cause by binary-toxin producing isolates in Korea. MATERIALS AND METHODS: From September 2008 through January 2010, clinical characteristics, medication history and treatment outcome of all the CDI patients were collected prospectively. Toxin characterization, PCR ribotyping and antibiotic susceptibility were performed with the stool isolates of C. difficile. RESULTS: During the period, CDI caused by 11binary toxin-producing isolates and 105 toxin A & toxin B-positive binary toxin-negative isolates were identified. Comparing the disease severity and clinical findings between two groups, leukocytosis and mucoid stool were more frequently observed in patients with binary toxin-positive isolates (OR: 5.2, 95% CI: 1.1 to 25.4, P = 0.043; OR: 7.6, 95% CI: 1.6 to 35.6, P = 0.010, respectively), but clinical outcome of 2 groups did not show any difference. For the risk factors for acquisition of binary toxin-positive isolates, previous use of glycopeptides was the significant risk factor (OR: 6.2, 95% CI: 1.4 to 28.6, P = 0.019), but use of probiotics worked as an inhibitory factor (OR: 0.1, 95% CI: 0.0 to 0.8; P = 0.026). PCR ribotypes of binary toxinproducing C. difficile showed variable patterns: ribotype 130, 4 isolates; 027, 3 isolates; 267 and 122, 1 each isolate and unidentified C1, 2 isolates. All 11 binary toxin-positive isolates were highly susceptible to clindamycin, moxifloxacin, metronidazole, vancomycin and piperacillin-tazobactam, however, 1 of 11 of the isolates was resistant to rifaximin. CONCLUSIONS: Binary toxin-producing C. difficile infection was not common in Korea and those isolates showed diverse PCR ribotypes with high susceptibility to antimicrobial agents. Glycopeptide use was a risk factor for CDI by those isolates.

Seroepidemiología del paludismo en un grupo de migrantes en tránsito (Chiapas, 2008)

OBJETIVO: Conocer la prevalencia del paludismo y los factores asociados con la infección de migrantes en la frontera sur de México, durante 2008. MATERIAL Y MÉTODOS: En 706 migrantes, se investigó la infección activa mediante prueba rápida y PCR o pasada, mediante serología y se aplicó un cuestionario para investigar las condiciones asociadas con la infección. RESULTADOS: 85.6% provenía de Centroamérica. Ninguno presentó infección activa; 4.2% fue seropositivo y la mayoría provenía de los países con mayor incidencia de paludismo en la región. La seropositividad se asoció con el número de episodios previos de paludismo (RM=1.44; IC95% 1.04-2.00), años de permanencia en su comunidad de origen (RM=1.03; IC95% 1.00 -1.07) y conocimiento y automedicación con antipalúdicos (RM=3.38; IC95% 1.48-7.67). CONCLUSIONES: La exposición previa de migrantes al paludismo y las dificultades para su detección indican la necesidad de nuevas estrategias para la vigilancia epidemiológica para estas poblaciones.

OBJECTIVE: To know the prevalence of malaria and the factors associated with the infection in migrants in the southern border of Mexico, during 2008. MATERIALS AND METHODS: In 706 migrants, active malaria infection was investigated using a rapid diagnostic test and PCR and past infection using serology. A questionnaire was applied to investigate the conditions associated to infection. RESULTS: 85.6% originated from Central America, none presented an active infection, although 4.2% were seropositive, most of these came from the countries with the highest malaria incidence in the region. Seropositivity was associated with the number of previous malaria episodes (OR=1.44; IC95% 1.04-2.00), years living in their community of origin (OR=1.03; IC95% 1.00-1.07), and knowledge and self-medication with anti-malaria drugs (OR=3.38; IC95% 1.48-7.67). CONCLUSIONS:. The previous exposure of migrants and the difficulties for their detection indicate the need of new strategies for the epidemiological surveillance for these populations.

Establishment and comparison of pulsed-field gel electrophoresis, multiple-locus variable number tandem repeat analysis and automated ribotyping methods for subtyping of Citrobacter strains / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To establish and compare the pulsed-field gel electrophoresis (PFGE), multiple-locus variable number tandem repeat analysis (MLVA) and automated ribotyping for subtyping of Citrobacter strains.</p><p><b>METHODS</b>PFGE protocol was optimized in terms of plug preparation procedure, restriction enzymes and configuration of electrophoretic parameters. MLVA method was evaluated by finding variable number tandem repeats in two genomes of Citrobacter strains. The ribotyping was performed by using the automated RiboPrinter system.</p><p><b>RESULTS</b>We optimized the plug preparation procedure, focused on the cell suspension concentration (turbidity of 2.5 to 3.5), SDS addition (no SDS needed) and lysis time (1 h), and selected the appropriate restriction enzyme (XbaI) and the electrophoretic parameters (1.0 s-20.0 s for 19 h) of PFGE. There was nearly no discriminatory power of MLVA between Citrobacter strains. For 51 Citrobacter strains, automated ribotyping gave a D-value of 0.9945, while PFGE gave a D-value of 0.9969. Both PFGE and automated ribotyping clustered strains from the same sources (with the same species from the same place at the same time identified as the same source) and divided strains from different sources (from different years, places and hosts) into different subtypes.</p><p><b>CONCLUSION</b>PFGE protocol established in this paper and automated ribotyping are suitable for application in Citrobacter subtyping.</p>

Infección diseminada por Penicillium marneffei en un paciente HIV-positivo: Primera observación en la República Argentina / Disseminated infection due to Penicillium marneffei related to HIV infection: First observation in Argentina

Se presenta el primer caso humano de peniciliosis por Penicillium marneffei observado en la República Argentina. El paciente era un joven de 16 años, HIV-positivo, procedente de un área rural del sur de China. El paciente fue internado en el Hospital "F. J. Muñiz" por padecer una neumonía grave con insufciencia respiratoria aguda. El agente causal fue aislado de un lavado broncoalveolar y se lo observó en un citodiagnóstico de piel. La identifcación de P. marneffei fue confrmada por las características fenotípicas del aislamiento y la amplifcación del ADNr. El enfermo padecía una infección muy avanzada por HIV que condujo a la aparición simultánea de infecciones por citomegalovirus, Pneumocystis jirovecii y procesos bacterianos nosocomiales. Este complejo cuadro derivó en una evolución fatal.

The frst case observed in Argentina of AIDS-related human penicillosis is herein presented. The patient was a six- teen year-old young man coming from a rural area of southern China. He was admitted at the F. J. Muñiz Hospital of Buenos Aires city with severe pneumonia and adult respiratory distress. Penicillium marneffei was isolated from bronchoalveolar lavage fuid and was microscopically observed in a skin cytodiagnosis. P. marneffei identifcation was confrmed by rRNA amplifcation and its phenotypic characteristics. The patient suffered an advanced HIV infection and also presented several AIDS-related diseases due to CMV, nosocomial bacterial infections and Pneumocystis jirovecii which led to a fatal outcome.

Chemotactic responses to gas oil of Halomonas spp. strains isolated from saline environments in Argentina / Respuesta quimiotáctica hacia gas oil de cepas de Halomonas spp. aisladas de ambientes salinos de Argentina

In this study, two halophilic bacterial strains isolated from saline habitats in Argentina grew in the presence of gas oil. They were identified as Halomonas spp. and Nesterenkonia sp. by 16S ribosomal RNA sequencing. Chemotaxis towards gas oil was observed in Halomonas spp. by using swimming assays.

En el presente trabajo se aislaron dos cepas bacterianas halofílicas a partir de muestras obtenidas en ambientes salinos de Argentina, que crecieron en presencia de gasoil como única fuente de carbono. Las cepas aisladas se identificaron como Halomonas spp. y Nesterenkonia sp. mediante secuenciación del gen del ARN ribosomal 16S. En ensayos de swimming, las cepas del genero Halomonas spp. mostraron una respuesta quimiotáctica hacia el gas oil.

Clostridium difficile Infections in Children / 이화의대지

During the past decade, rates of Clostridium difficile infection (CDI) increased worldwide. Hypervirulent strains of C. difficile such as NAP1/BI/027 and PCR ribotype 078 have emerged that have changed the epidemiology of CDI. Especially, CDI rates also have increased in the community, in children previously thought to be at low risk. Recently, the use of gastric acid suppressant that facilitates intestinal transit of the bacteria and presence of inflammatory bowel disease has been reported as risk factors. Treatment for CDI usually relies on metronidazole or vancomycin, but recurrence rates remains high. New treatment options for multiple recurrence are challenging. In this article, we reviewed recent epidemiological changes, current knowledge of virulence factors, reasonable approach to the diagnosis, and optimal treatment of CDI. But, clinical guidelines for pediatric C. difficile disease have not been defined. It seems that the consensus and recommendations for managing pediatric CDI are urgently needed.