Whole-genome sequencing of Klebsiella pneumoniae MDR circulating in a pediatric hospital setting: a comprehensive genome analysis of isolates from Guayaquil, Ecuador.

BACKGROUND: Klebsiella pneumoniae is the major cause of nosocomial infections worldwide and is related to a worsening increase in Multidrug-Resistant Bacteria (MDR) and virulence genes that seriously affect immunosuppressed patients, long-stay intensive care patients, elderly individuals, and children. Whole-Genome Sequencing (WGS) has resulted in a useful strategy for characterizing the genomic components of clinically important bacteria, such as K. pneumoniae, enabling them to monitor genetic changes and understand transmission, highlighting the risk of dissemination of resistance and virulence associated genes in hospitals. In this study, we report on WGS 14 clinical isolates of K. pneumoniae from a pediatric hospital biobank of Guayaquil, Ecuador. RESULTS: The main findings revealed pronounced genetic heterogeneity among the isolates. Multilocus sequencing type ST45 was the predominant lineage among non-KPC isolates, whereas ST629 was found more frequently among KPC isolates. Phylogenetic analysis suggested local transmission dynamics. Comparative genomic analysis revealed a core set of 3511 conserved genes and an open pangenome in neonatal isolates. The diversity of MLSTs and capsular types, and the high genetic diversity among these isolates indicate high intraspecific variability. In terms of virulence factors, we identified genes associated with adherence, biofilm formation, immune evasion, secretion systems, multidrug efflux pump transporters, and a notably high number of genes related to iron uptake. A large number of these genes were detected in the ST45 isolate, whereas iron uptake yersiniabactin genes were found exclusively in the non-KPC isolates. We observed high resistance to commonly used antibiotics and determined that these isolates exhibited multidrug resistance including ß-lactams, aminoglycosides, fluoroquinolones, quinolones, trimetropins, fosfomycin and macrolides; additionally, resistance-associated point mutations and cross-resistance genes were identified in all the isolates. We also report the first K. pneumoniae KPC-3 gene producers in Ecuador. CONCLUSIONS: Our WGS results for clinical isolates highlight the importance of MDR in neonatal K. pneumoniae infections and their genetic diversity. WGS will be an imperative strategy for the surveillance of K. pneumoniae in Ecuador, and will contribute to identifying effective treatment strategies for K. pneumoniae infections in critical units in patients at stratified risk.

[Construction and validation of a test for verbal auditory screening of cognitive alterations (CAVAC)]. / Construcción y validación de un test para el cribado auditivo verbal de alteraciones cognitivas (CAVAC).

INTRODUCTION: The global burden of dementia will continue to increase and Latin America will be one of the most affected regions. It is urgent to have new and better cognitive screening tools. There are few screenings composed entirely of auditory-verbal items and even more so for Argentine culture. The objective was to build and validate a test for verbal auditory screening of cognitive alterations (CAVAC) in Argentina. SUBJECTS AND METHODS: Evidence based in test content was analyzed through 16 experts collaborated. In addition, evidence based on relationships to other variables, internal consistency and the ability to discriminate between cognitively healthy subjects, Mild Cognitive Impairment (MCI) and dementia were investigated. The CAVAC was applied to 316 Argentines (M age = 68,5). Volunteers from the general population and patients from neurology services from two Argentine hospitals participated. RESULTS: The CAVAC test explores orientation/attention, memory/learning, language and executive functions, and is administered in 10 minutes with a score of 0 to 30. Aiken's V coefficient was satisfactory for all items. The CAVAC achieved a high correlation with the Mini Mental (0.762) and an acceptable Cronbach's alpha (0.746). The CAVAC score differs between controls, mild cognitive impairment (MCI) and dementia, and the ROC curves reveal adequate area values ??to discriminate MCI and dementia (>0.7), and cut-off points with good sensitivity-specificity. CONCLUSIONS: A new auditory-verbal test is made available for the screening of cognitive alterations with acceptable evidence of validity, good reliability, with the capacity to identify MCI and dementia, and free of charge for use.

TITLE: Construcción y validación de un test para el cribado auditivo verbal de alteraciones cognitivas (CAVAC).Introducción. La carga mundial de demencia continuará en aumento y Latinoamérica será una de las regiones más afectadas. Urge contar con nuevas y mejores herramientas de cribado cognitivo. Son pocos los cribados compuestos íntegramente por ítems auditivos-verbales, y más aún para cultura argentina. El objetivo fue construir y validar un test para el cribado auditivo verbal de alteraciones cognitivas (CAVAC) en Argentina. Sujetos y métodos. Se analizó la evidencia basada en el contenido de la prueba mediante colaboración de 16 jueces. Se investigó, además, la evidencia basada en relaciones con otras variables, la consistencia interna y la capacidad para discriminar entre sujetos cognitivamente sanos, con deterioro cognitivo leve (DCL) y con demencia. Se aplicó el CAVAC a 316 argentinos (media de edad = 68,5). Participaron voluntarios de población general y pacientes de servicios de neurología de dos hospitales argentinos. Resultados. El CAVAC explora la orientación/atención, la memoria/aprendizaje, el lenguaje y las funciones ejecutivas, y se administra en 10 minutos, con una puntuación de 0 a 30. El coeficiente V de Aiken fue satisfactorio para todos los ítems. El CAVAC alcanzó correlación fuerte con el Minimental State Examination (0,762) y un alfa de Cronbach aceptable (0,746). La puntuación del CAVAC difirió entre controles, pacientes con DCL y con demencia, y las curvas ROC revelaron valores de área adecuados para identificar el DCL y la demencia (>0,7), con puntos de corte con buena sensibilidad-especificidad. Conclusiones. Se deja a disposición un nuevo test auditivo-verbal para el cribado de alteraciones cognitivas con aceptables evidencias de validez, buena fiabilidad, capacidad para identificar el DCL y la demencia, y de uso libre y gratuito.

Enhancing biodegradation of aged hydrocarbon-contaminated soils through toluene addition: assessing effects on solid and slurry phase treatments.

The main challenge in treating aged soils highly contaminated with total petroleum hydrocarbons (TPH) is to enhance their bioavailability for microbial degradation. Hydrocarbons in soils undergo chemical changes that make them more resistant to biodegradation. This study investigates toluene's efficacy in enhancing the biodegradation of aged hydrocarbon-contaminated soil containing 292,000 mg TPH kg-1 dry soil. Toluene's effect was compared between solid phase (SOP) and slurry phase (SLP) treatments using a microbial consortium isolated from Cyperus laxus rhizosphere. TPH biodegradation and microbial respiration were measured, the latter to estimate the respiratory quotient (RQ, the ratio between moles of carbon dioxide released and moles of oxygen absorbed during respiration). Toluene significantly accelerated TPH biodegradation in both treatments, achieving ~ 30% higher removal than in a non-solvent control, possibly through improved bioavailability of aromatic compounds and other low molecular weight compounds. According to the RQ analysis, toluene enhanced microbial respiratory processes and hydrocarbon catabolism with higher hydrocarbon mineralization (RQ = ~ 0.5) in both SOP and SLP assays. Our results reveal toluene's potential to increase hydrocarbon availability and microbial degradation efficiency in aged contaminated soils; its use in various bioremediation techniques could be of broad applicability across diverse soil types and pollutants.

[Prevalence, knowledge and control of arterial hypertension in vulnerable neighborhoods of Argentina: A Cross-sectional Study]. / Estudio de prevalencia, conocimiento y control de la hipertensión arterial en barrios vulnerables de Argentina.

INTRODUCTION: Hypertension (HTN) represents the primary individual risk factor, contributing significantly to the global burden of cardiovascular diseases (CVD). In our country, epidemiological research has highlighted substantial variations in the prevalence of these risk factors across different populations. However, there is a lack of epidemiological studies assessing exclusive cardiovascular risk factors within vulnerable neighborhoods characterized by extremely limited economic resources, sociocultural challenges, and inadequate healthcare access. METHODS: A multicenter cross-sectional observational study was conducted among individuals residing in economically deprived and marginalized communities, including informal settlements and underprivileged neighborhoods. Simple random sampling of households was employed. Blood pressure measurements, anthropometric assessments, and epidemiological, economic, and sociocultural questionnaires were administered. Results encompass prevalence rates, awareness levels, and blood pressure control across diverse regions. Logistic regression was utilized to identify independent variables influencing primary outcomes. RESULTS: A total of 989 participants were analyzed. The overall prevalence of hypertension was 48.2%. About 82% had a body mass index (BMI) >25. Approximately 45.3% had less than 6 years of formal education. Independent association was established between education levels below 6 years and higher hypertension prevalence. Among hypertensive individuals, 44% were unaware of their condition, with only 17.2% achieving control, correlated with having health insurance and a higher educational background. Merely 24% were receiving combined therapy. CONCLUSION: The prevalence of hypertension within vulnerable neighborhoods is alarmingly high, surpassing rates in other social strata. Knowledge, treatment, and control levels of hypertension are suboptimal, comparable to other populations. Inadequate use of combination therapy was observed. This study underscores the urgent need for targeted interventions addressing cardiovascular risk factors in poor areas to mitigate the burden of CVD.

Evaluating the performance of WHO and CDC case definitions for influenza-like illness in diagnosing influenza during the 2022-2023 flu season in Mexico.

OBJECTIVE: This study aimed to evaluate the performance of the World Health Organization (WHO) and Centers for Disease Control and Prevention (CDC) case definitions for influenza-like illness (ILI) in diagnosing influenza during the 2022-2023 flu season in Mexico. STUDY DESIGN: We conducted a cross-sectional analysis of national epidemiological surveillance data in Mexico, focusing on respiratory viral pathogens. METHODS: We analyzed data from 6027 non-hospitalized patients between 5 and 65 years old who underwent molecular testing for respiratory viral pathogens. The performance of both case definitions was evaluated in terms of sensitivity, specificity, and the area under the receiver operating characteristic (AUROC) curve. RESULTS: Overall, the diagnostic accuracy of the evaluated ILI definitions in identifying influenza patients was low, particularly among older patients. When compared to the CDC, the WHO definition had a lower sensitivity but a higher specificity, resulting in a higher AUROC (P = 0.031) for the WHO criteria. CONCLUSIONS: Our findings suggest that the WHO and CDC ILI case definitions have limited accuracy for diagnosing influenza in non-hospitalized patients and highlight the need for more specific diagnostic tools to improve the detection of influenza cases during the flu season.

Restoration of a Vital Tooth With Extensive Crown Destruction Using Glass Fiber and Polyethylene Fiber-reinforced Composite Resin: A Clinical Case.

Conservative restorative dentistry has been evolving in the last 25 years, focusing mainly on the development of direct restorative materials. Resin-based composites remain an excellent conservative alternative for restoration of teeth with extensive caries lesions. Over time, several strategies have been proposed to improve the mechanical properties of these composites so that they can adequately withstand masticatory forces. Glass fiber-reinforced resin-based composites and their use in situations where there is great loss of tooth structure have gained popularity due to their favorable mechanical properties. Combined techniques with polyethylene fibers can further enhance their clinical performance. This study presents a brief review of their most important qualities and potential use in direct restorative procedures. In addition, a clinical case is described where a vital tooth with extensive coronal destruction was restored using polyethylene fibers embedded in glass fiber-reinforced flowable resin under resin-based composite. The reinforcement of both the remaining tooth structure and the restoration with fibers is a valid treatment option since the network structure formed by the fiber reinforcement can increase the longevity of the direct composite restorations. There are few reports in the literature describing the use of a combined technique using polyethylene fibers embedded in glass fiber-reinforced flowable resin under resin-based composite. Thus, clinical follow-up of this case is required.

Neotropical sisterhood: new species of Gyrodactylus von Nordmann, 1832 (Platyhelminthes: Monogenea) infecting Rhamdia guatemalensis and Rhamdia laticauda (Siluriformes: Heptapteridae) in Mexico.

We describe two new species of monogenean parasites of the genus Gyrodactylus von Nordmann, 1832 infecting Neotropical catfishes (Siluriformes) in southern Mexico: Gyrodactylus chulini n. sp. from 'chulín', Rhamdia laticauda collected in Oaxaca; and Gyrodactylus juili n. sp. from 'juil', Rhamdia guatemalensis from Veracruz. Morphologically, both new taxa are similar to Gyrodactylus spp. infecting catfishes (Siluriformes) in South America. Sequences of the internal transcribed spacers (ITS1-5.8S-ITS2 rDNA), the D2+D3 domains of the large ribosomal subunit (28S rDNA) and the cytochrome oxidase II (COII) gene were obtained from multiple parasite specimens and analysed using Bayesian inference. Phylogenetic hypotheses using ITS rDNA and COII genes, recovered two new Gyrodactylus species from Rhamdia spp.: G. chulini n. sp.; and Gyrodactylus juili n. sp., which are sister species to Gyrodactylus lilianae, a parasite of Rhamdia quelen in Brazil, and show strong affinity to other gyrodactytlids infecting Neotropical catfishes. This suggests that these new taxa, the first gyrodactylids described from Rhamdia spp. in Mexico, co-migrated to Tropical Middle America with their Neotropical catfish hosts, after the emergence of the Isthmus of Panama.

Tasas de egreso hospitalario por torsión ovárica en el período 2018-2021 en Chile según grupo etario: un estudio descriptivo / Ovarian torsion hospital discharge rates in the period 2018-2021 in Chile according to age group: a descriptive study

Introducción: La torsión ovárica (TO) es la rotación completa o parcial del ovario por sobre su propio pedículo vascular. El objetivo es comparar descriptivamente las tasas de egreso hospitalario (TEH) por TO en el período 2018-2021 en Chile. Materiales y métodos: Estudio descriptivo, transversal. Los datos de egresos hospitalarios por TO en Chile entre 20182021 (n=1.599) según grupo etario y días de estadía hospitalaria se obtuvieron del Departamento de Estadísticas e Información de Salud. Se calculó la TEH. No se requirió comité de ética. Resultados: Se determinó una TEH por TO de 4,33/100.000 habitantes entre los años 2018 y 2021, siendo el año 2021 la mayor con 5,92. Los grupos etarios de 10-14 y 15-19 años registraron las mayores TEH de 8,5 y 7,55, respectivamente. El promedio de días de estadía hospitalaria por TO entre los años 2018-2021 fue de 2,15 días, siendo el año 2018 el mayor con 2,3 días. Pacientes de 80 años y más presentaron la mayor duración de estancia hospitalaria con 3,37 días. Discusión: La TEH por TO en Chile aumentó durante los últimos años. En 2020-2021 las mayores TEH por TO se encontraron entre los 10-19 años, probablemente por aumentos en la prevalencia de síndrome de ovario poliquístico en Chile. Los grupos de mayor edad registraron la mayor duración de estancia hospitalaria, pudiendo deberse a una mayor susceptibilidad a complicaciones postquirúrgicas. Considerando la escasa fuente de información al respecto, nuestro estudio permite dar a conocer un perfil epidemiológico nacional actualizado.

Introduction: Ovarian Torsion (OT) is complete or partial rotation of the ovary above its vascular pedicle. The objective is to descriptively compare the Hospital Discharge Rates (HDR) for OT in the period 2018-2021 in Chile. Material and methods: Descriptive, cross-sectional study. The data of hospital discharge for OT in Chile between 20182021 (n=1,599) according to age group and days of hospital stay were obtained from the Department of Health Information Statistics. HDR was calculated. No ethics committee was required. Results: An HDR for OT of 4.33/100.000 population was determined between the years 2018 and 2021, with 2021 being the highest with 5.92. The age groups of 10-14 and 15-19 years old registered the highest HDR of 8.5 and 7.55, respectively. The average number of days of hospital stay for OT between the years 2018-2021 was 2.15 days, with 2018 being the longest year with 2.3 days. Patients 80 years and older had the longest hospital length of stay with 3.37 days. Discussion: The HDR for OT in Chile has increased in recent years. In 2020-2021 the highest HDR for OT were found between 10-19 years old, probably due to increases in the prevalence of polycystic ovary syndrome in Chile. The oldest age groups recorded the longest average hospital length of stay, which may be due to increased susceptibility to postoperative complications. Considering the limited source of information in this regard, our study allows us to present an updated national epidemiological profile.

[Microbiological characterization and antimicrobial susceptibility pattern of infections associated with febrile neutropenia in pediatric hemato-oncological patients.] / Caracterización microbiológica y de susceptibilidad antimicrobiana de las infecciones asociadas a neutropenia febril en pacientes hemato-oncológicos pediátricos.

INTRODUCTION: Febrile neutropenia (FN) is the most frequent hemato-oncological emergency, with high morbidi ty and mortality in pediatrics. The objective of the study was the microbiological characterization and antimicrobial susceptibility of infections associated with FN in pediatric hemato-oncological patients. PATIENTS AND METHOD: Retrospective cohort study with patients aged between 1 month and 18 years, with onco-hematological pathology according to ICD-10 codes, hospitalized in a tertiary healthcare center in Bucaramanga, Colombia. Based on the medical records of the period 2013-2017, the episodes of FN were identified, and the isolated microorganisms and their susceptibility pattern were described. Biochemical identification and antimicrobial susceptibility testing were performed with the Dade Behring Microscan« automated system. The resistant microorganism classification was performed based on the Minimum Inhibitory Concentration (MIC) and the interpretation of the laboratory according to the cut-off points of the Clinical and Laboratory Standards Institute recommendations. RESULTS: Of 130 patients, 14.7% of the cultures obtained were positive. Bloods tream infection was observed in 17.5% of the episodes. The isolated microorganisms were mainly Gram-negative bacteria (75.8%). Enterobacteriaceae (EB) were the most frequent, led by Klebsiella pneumoniae, Escherichia coli, followed by Pseudomonas aeruginosa and coagulase-negative Staphylo cocci. Of the EBs, 40.5% showed resistance to Piperacillin/Tazobactam, 33.3% to Cefepime, and 8.2% to Meropenem. According to the antimicrobial resistance pattern, it was observed that 16.4% of the positive EB cultures had an extended-spectrum beta-lactamase pattern and 5% a pattern suggestive of carbapenemases. All Gram-positive cocci were sensitive to Vancomycin. CONCLUSION: In the studied patients, the predominant pathogenic microorganisms were Gram-negative ones with resistance in dices similar to those of developing countries.

Aquatic Macrophytes Hosting Immature Mansonia (Mansonia) Blanchard, 1901 (Diptera, Culicidae) in Porto Velho, Rondonia State, Brazil.

High abundance of hematophagous mosquitoes of the genus Mansonia Blanchard, 1901 (Diptera: Culicidae) threatens human and domestic animal health and well-being. Knowledge of the biology of nuisance mosquito species is necessary to understand specific ecological and biological factors to enable rapid and effective monitoring measures for sustainable control programs. The establishment and dispersion of Mansonia species are associated with the occurrence of aquatic macrophytes species, which are indispensable for the development of larvae and pupae. To increase knowledge of the host plants for Mansonia immature stages in Porto Velho, Rondonia State, Brazil, specimens of four plant species, which occur across the tributaries of the Madeira River were sampled and inspected for the presence of egg batches, larvae, and pupae. A total of 1,386 larvae and pupae of Mansonia spp. were collected attached to the roots of Eichhornia crassipes (Mart.) Solms (Commelinales: Pontederiaceae), Pistia stratiotes L. (Alismatales: Araceae), and Limnobium laevigatum (Humb. and Bonpl. Ex Willd.) Heine (Alismatales: Hydrocharitaceae). The novel association of Mansonia species with L. laevigatum is presented. Egg batches of Mansonia spp. were found only on Salvinia molesta D.S. Mitch. (Salviniales: Salviniaceae). Possible differences in the roles played by E. crassipes and S. molesta in the reproductive cycle of Mansonia spp. in the surveyed area are discussed. All species of host plants including E. crassipes, P. stratiotes, S. molesta, and L. laevigatum should be considered when planning macrophyte management for the control of Mansonia species.

Caregivers' knowledge and perceptions are associated with children's TB preventive treatment completion.

SETTING: Forty-six health centers in south Lima, Peru. OBJECTIVE: To assess the association between caregivers' knowledge and perceptions around isoniazid preventive therapy (IPT) and whether their children complete IPT. DESIGN: We conducted a retrospective medical record review of children who initiated IPT during 2017-2018. We administered structured surveys to caregivers of the children about their knowledge about and perceptions of IPT. We used a modified Poisson regression to determine factors associated with IPT completion. RESULTS: We included 550 children, of whom 31% did not complete IPT. Independent factors associated with not completing IPT were low caregiver knowledge about TB and IPT (adjusted risk ratio [aRR] 1.41, 95% CI 1.06-1.78), low caregiver perception of the importance of IPT (aRR 1.76, 95% CI 1.30-2.39), low caregiver satisfaction with the health services (aRR 1.57, 95% CI 1.14-2.16), experience of adverse events (aRR 2.08, 95% CI 1.51-2.87), and living in a household with moderate or severe family dysfunction (aRR 1.53, 95% CI 1.07-2.19). CONCLUSION: IPT completion among children was associated with the knowledge and perceptions of their caregivers, as well as the experience of adverse events. To improve IPT completion among children, health care providers should prioritize education and counseling for caregivers, promote positive interpersonal relationships with them, and monitor adverse events.

CONTEXTE: Quarante-six centres de santé au Sud de Lima, Pérou. OBJECTIF: Evaluer l'association entre les connaissances et les perceptions relatives au traitement préventif par l'isoniazide (TPI) des responsables des enfants et le fait que leurs enfants aient achevé le TPI. SCHÉMA: Nous avons réalisé une revue rétrospective des dossiers médicaux des enfants qui ont démarré le TPI entre 2017 et 2018. Nous avons administré aux responsables des enfants des questionnaires structurés relatifs à leurs connaissances et perceptions du TPI. Nous avons utilisé une régression de Poisson modifiée afin de déterminer les facteurs associés à l'achèvement du TPI. RÉSULTATS: Nous avons inclus 500 enfants dont 31% n'ont pas achevé le TPI. Les facteurs indépendants associés au non achèvement étaient faibles connaissances relatives à la TB et au TPI (risque relatif ajusté [RRa] 1,41 ; IC 95% 1,06­1,78), faibles perception par les responsables de l'importance du TPI (RRa 1,76 ; IC 95% 1,30­2,39), faible taux de satisfaction des responsables par rapport aux services de santé (RRa 1,57 ; IC 95% 1,14­2,16), survenue d'effets secondaires (RR 2,08 ; IC 95% 1,51­2,87) et dysfonction familiale modérée ou grave (RRa 1,53 ; IC 95% 1,07­2,19). CONCLUSION: L'achèvement du TPI chez les enfants a été associé aux connaissances et perceptions de leurs responsables, ainsi qu'à la survenue d'effets secondaires. Pour améliorer l'achèvement du TPI chez les enfants, les prestataires de soins de santé devraient accorder la priorité à l'éducation des responsables, promouvoir des relations inter personnelles positives avec eux et surveiller les effets secondaires.

Survival in adult pneumonia inpatients fulfilling suspected COVID-19 criteria and baseline negative RT-qPCR.

OBJECTIVES: The objective of this study was to evaluate the survival experience of suspicion COVID-19 hospitalized patients with pneumonia and negative baseline reverse-transcription quantitative polymerase chain reaction (RT-qPCR) test results. STUDY DESIGN: We conducted a nationwide retrospective cohort study in Mexico. METHODS: Adult pneumonia inpatients fulfilling suspected COVID-19 criteria, and hospital entry from March to August 2020, were enrolled. The Kaplan-Meier method was to use to compare survival estimates among patients with negative RT-qPCR nasopharyngeal or oropharyngeal swabs and those with a baseline positive test. RESULTS: Data from 64,624 individuals fulfilling suspected COVID-19 criteria were analyzed and 1.6% of them had negative RT-qPCR tests. The overall mortality rate was higher among laboratory-positive patients (48.5% vs. 34.2%, P < 0.001) and, at any given threshold, the survival estimates were higher among RT-qPCR-negative pneumonia inpatients. CONCLUSIONS: The pathogenic mechanism of COVID-19 remains poorly understood and suspected cases with pneumonia and negative laboratory results represent a major challenge for healthcare systems. Our findings suggest that RT-qPCR-negative inpatients may have an improved disease prognosis, but the in-hospital mortality was still high among them. However, further research is needed to clarify the clinical and epidemiological implications of our results.

Optimization and application of a high-resolution melting protocol in the characterization of avian infectious laryngotracheitis virus

Resumen En un estudio epidemiológico realizado previamente en Argentina, se analizó la secuencia de un fragmento del gen US5 del virus de la laringotraqueítis infecciosa (ILTV), lo que permitió diferenciar las cepas de campo de las vacunales. También esto permitió definir cinco haplotipos del ILTV, con variaciones específicas en las posiciones 461, 484, 832, 878 y 894 del gen US5. La caracterización de las cepas virales también puede lograrse mediante el análisis de la disociación de alta resolución o high-resolution melting analysis (HRMA), descripto como un método efectivo, rápido y sensible para detectar mutaciones en productos de PCR. En el presente estudio se desarrolló un protocolo de disociación de alta resolución con el objetivo de caracterizar cepas del ILTV circulantes en Argentina. Para ello,se confirmó la especificidad de esta herramienta en diferentes diluyentes del ADN de las muestras, sin observarse interferencias en presencia de ADN heterólogo u otros metabolitos celulares. Asimismo, la concentración de sales en el buffer de elución utilizado durante la extracción de ADN no alteró los perfiles de las curvas. Se obtuvieron perfiles bien definidos con concentraciones de ADN más elevadas (Ct = 26.0), mientras que concentraciones más bajas presentaron curvas heterogéneas (Ct = 32.5). El HRMA mostró una concordancia del 97.49% con la técnica de referencia, la secuenciación. El protocolo de disociación de alta resolución amplifica el ADN antes de su caracterización, por lo que esta técnica podría ser eventualmente utilizada para confirmar la presencia del ILTV y, al mismo tiempo, distinguir haplotipos, optimizando su valor como herramienta de diagnóstico. Esta característica implica una reducción significativa en el tiempo dedicado al procesamiento de muestras.

Atypical phenotype and response of B cells in patients with seropositive rheumatoid arthritis.

Patients with rheumatoid arthritis (RA) may be classified as seropositive or seronegative according to the presence of autoantibodies. An abnormal B cell phenotype and function could be one of the main components of the immunopathology of seropositive patients; however, there is little information regarding B cell defects in these patients. This study shows a broad characterization of the B cell phenotype and function in patients with seropositive RA. We focused mainly on the evaluation of subsets, the expression of modulatory molecules of cell activation (CD22, FcÉ£RIIb and FcµR), calcium mobilization, global tyrosine phosphorylation, expression of activation markers, cytokine and immunoglobulin (Ig) production, proliferation and the in-vitro generation of plasma cells. Increased frequency of CD27- IgM- IgD- and CD21- B cells was observed in patients with seropositive RA compared with healthy donors (HD). Decreased expression of CD22 was primarily found in memory B cells of patients with RA regardless of seropositivity. B cells from seropositive patients exhibited normal proliferation, calcium mobilization kinetics and global tyrosine phosphorylation, but showed an increased frequency of CD86+ B cells compared with HD. B cells of seropositive patients secrete less interleukin-10 after in-vitro activation and showed a decreased frequency of plasma cell differentiation and IgM production compared with HD. Our data indicate that patients with seropositive RA have an increased frequency of atypical B cell populations previously associated with chronic activation and antigen exposure. This may result in the observed low responsiveness of these cells in vitro.

Optimization and application of a high-resolution melting protocol in the characterization of avian infectious laryngotracheitis virus.

A previous sequence analysis of a US5 gene fragment of infectious laryngotracheitis virus (ILTV) performed in an Argentinian epidemiological study allowed to differentiate between wild and vaccine strains. This analysis also defined five ILTV haplotypes with specific variations at positions 461, 484, 832, 878 and 894 of the US5 gene. This characterization of viral strains may also be accomplished using the High-Resolution Melting Analysis (HRMA), which has been described as an effective, fast and sensitive method to detect mutations in PCR products. In the present study, an HRM protocol was developed with the aim of characterizing the circulating ILTV strains in Argentina. The specificity of this tool was confirmed in different DNA diluents, without interference from heterologous DNA or other cellular metabolites. Additionally, the salt concentration in the elution buffer used for DNA extraction did not alter the curve profiles. Higher concentrations of DNA (Ct≅26.0) displayed well-defined curve profiles, whereas lower concentrations (Ct≅32.5) exhibited more heterogeneous curves. The HRMA showed 97.49% concordance with the reference technique, i.e., sequencing. The HRM protocol has the capability to perform DNA amplification prior to its characterization. Thus, eventually this technique may be used simultaneously as a diagnostic tool. This advantage implies a significant reduction in the time and effort involved in sample processing.

Chemical signals act as the main reproductive barrier between sister and mimetic Heliconius butterflies.

Colour pattern is the main trait that drives mate recognition between Heliconius species that are phylogenetically close. However, when this cue is compromised such as in cases of mimetic, sympatric and closely related species, alternative mating signals must evolve to ensure reproductive isolation and species integrity. The closely related species Heliconius melpomene malleti and H. timareta florencia occur in the same geographical region, and despite being co-mimics, they display strong reproductive isolation. In order to test which cues differ between species, and potentially contribute to reproductive isolation, we quantified differences in the wing phenotype and the male chemical profile. As expected, the wing colour pattern was indistinguishable between the two species, while the chemical profile of the androconial and genital males' extracts showed marked differences. We then conducted behavioural experiments to study the importance of these signals in mate recognition by females. In agreement with our previous results, we found that chemical blends and not wing colour pattern drive the preference of females for conspecific males. Also, experiments with hybrid males and females suggested an important genetic component for both chemical production and preference. Altogether, these results suggest that chemicals are the major reproductive barrier opposing gene flow between these two sister and co-mimic species.

Effectiveness of the use of an algorithm in the diagnostic approach of joint pain patients by primary care physicians.

There is a high percentage of error in the approach of patients with joint pain by primary care physicians. An algorithm can help improve this misdiagnosis problem. Our study seeks to determine the effectiveness of an algorithm when used by primary care physicians for the diagnosis of cases of joint pain patients. A randomized clinical experiment was carried out. Primary care physicians from five cities in Colombia developed a series of clinical cases, which were presented to them through a website on their personal cell phones. Half of the doctors developed the cases using the diagnostic algorithm, and the other half developed the cases without the use of the algorithm. Main measures were proportion of correct diagnosis, number, type of laboratory and diagnostic images requested for the diagnostic approach of clinical cases. Two hundred and twenty-four primary care physicians participated. The overall proportion of cases correctly diagnosed was 37.3% higher in the intervention group; we found a greater difference in cases of spondyloarthritis (60.8%), followed by systemic lupus erythematosus with joint involvement (32.2%), rheumatoid arthritis (30.3%) and osteoarthritis (25.9%). The average number of tests requested to develop clinical cases was lower in the intervention group than in the control group, both globally and for each of the four diseases, with statistically significant differences for each of the comparisons. The diagnostic algorithm proved to be an effective tool when used by primary care physicians; the proportion of correct diagnoses increased, and the number of tests requested in the development of the cases decreased.