Liquid-nitrogen-free CTAB DNA extraction method from silica-dried specimens for next-generation sequencing and assembly.

Next-generation sequencing requires intact and high-quality DNA. However, typical liquid-nitrogen DNA extraction methods are expensive and not practical for field sample collections. Hence, we present a cost-effective method for DNA extraction from silica-dried leaf samples, eliminating the need for liquid nitrogen. Two protocols were evaluated to determine the effectiveness of grinding dried plant samples without liquid nitrogen in comparison to the standard protocol for tissue homogenization and cell lysis. Protocol 1 involved grinding fresh leaf samples with liquid nitrogen, while Protocol 2 entailed incubating dried plant samples at-20 °C for 1 h before grinding in the absence of liquid nitrogen. Both protocols produced comparable DNA yields with an average A260/A280 ratio of 1.78±0.02, suitable for short- and long-read sequencing. Using Protocol 2, we successfully assembled ten plastomes. It also demonstrated versatility as comparable DNA quality was obtained from dried mollusks and actinomycetes, resulting in the successful assembly of two complete mitochondrial genomes. The protocol is advantageous for research workflows involving the collection of samples in the field as a long-term source of genetic material.•Drying: Fresh samples were silica-dried at silica-to-sample ratio of 2:1.•Pre-lysis: Dried samples were frozen at -20 °C for 1 hour before grinding.•Frozen samples were subjected to tissue homogenization followed by the standard CTAB DNA extraction.

Changes in clinical characteristics and outcomes of patients hospitalized with COVID-19 during two years of the pandemic: experience in a venezuelan hospital. / Cambios en las características clínicas y desenlaces de pacientes hospitalizados por COVID-19 durante dos años de pandemia: experiencia en un hospital venezolano.

OBJECTIVES.: To determine changes in the clinical characteristics and in-hospital outcomes of patients hospitalized for COVID-19 in a private hospital in Caracas during two years of the pandemic. MATERIALS AND METHODS.: Retrospective, observational study of patients hospitalized for COVID-19. We evaluated the correspondence between waves of hospital admissions and circulating variants of SARS-CoV-2 in the general population of the Capital District and Miranda state. RESULTS.: A total of 1025 patients (569 men and 456 women) were included, with a mean age of 62.9 SD: 16.2 years. Four waves of hospital admissions were identified: first (March-November 2020) 150/1025 (14.6%) cases; second (December 2020 to May 2021) 415/1025 (40.5%) cases; third (June-December 2021) 344/1025 (33.6%) cases; fourth (January-February 2022) 116/1025 (11.3%) cases. The mean age was higher in the fourth wave (first: 64.0±15.7, second: 61.4±15.8, third: 62.1±16.5, and fourth wave: 68.5±16.4), while the proportion of male patients (first: 66.7%, second: 58.8%, third: 50.3%, and fourth wave: 44.8%), patients with severe-critical illness (first: 65.3%, second: 57%, third: 51.7%, and fourth wave: 44.8%), in-hospital stay (first: 9.1±6.0, second: 9.0±7.3, third: 8.8±7.7, and fourth wave: 6.9±5.0 days), ICU admissions (first: 23.3%, second: 15.7%, third: 14.0%, and fourth wave: 11.2%; p=0.027) and mortality (first: 21. 8%, second: 10.7%, third: 9.1%, and fourth wave: 7.1%; p<0.001) progressively decreased over time. CONCLUSIONS.: The results show lower frequency of severe cases and improvement of in-hospital outcomes in two years of the pandemic. Changes in circulating variants, improvements in disease management and vaccination are likely to have influenced these results.

OBJETIVOS.: Determinar los cambios en las características clínicas y desenlaces intrahospitalarios de los pacientes hospitalizados por COVID-19 en un hospital privado de Caracas durante dos años de pandemia. MATERIALES Y MÉTODOS.: Estudio retrospectivo, observacional, de pacientes hospitalizados por COVID-19. Se investigó la correspondencia entre las olas de ingresos hospitalarios con las variantes circulantes del SARS-CoV-2 en la población general del Distrito Capital y estado Miranda. RESULTADOS.: Se incluyeron 1025 pacientes (569 hombres y 456 mujeres), con edad promedio de 62,9 DE: 16,2 años. Cuatro olas de ingresos hospitalarios fueron identificadas: primera (marzo-noviembre 2020) 150/1025 (14,6%) casos; segunda (diciembre-2020 a mayo-2021) 415/1025 (40,5%) casos; tercera (junio-diciembre 2021) 344/1025 (33,6%) casos; cuarta (enero-febrero 2022) 116/1025 (11,3%) casos. La edad promedio fue mayor en la cuarta ola (primera 64,0±15,7, segunda 61,4±15,8, tercera 62,1±16,5, y cuarta ola 68,5±16,4), mientras que la proporción de pacientes masculinos (primera 66,7%, segunda 58,8%, tercera 50,3%, y cuarta 44,8%), los pacientes con enfermedad grave-crítica (primera 65,3%, segunda 57%, tercera 51,7% y cuarta 44,8%), la estadía intrahospitalaria (primera 9,1±6,0, segunda 9,0±7,3, tercera 8,8±7,7, y cuarta 6,9±5,0 días), los ingresos a la UCI (primera 23,3%, segunda 15,7%, tercera 14,0%, y cuarta 11,2%; p=0,027) y la mortalidad (primera 21.8%, segunda 10,7%, tercera 9,1%, y cuarta 7,1%; p<0,001) disminuyeron progresivamente con el tiempo. CONCLUSIONES.: Los resultados muestran menor frecuencia de casos severos y mejoría de los desenlaces intrahospitalarios en dos años de pandemia. Es probable que los cambios en las variantes circulantes, las mejoras del manejo de la enfermedad y la vacunación hayan influido sobre estos resultados.

Cambios en las características clínicas y desenlaces de pacientes hospitalizados por COVID-19 durante dos años de pandemia: experiencia en un hospital venezolano / Changes in clinical characteristics and outcomes of patients hospitalized with COVID-19 during two years of the pandemic: experience in a venezuelan hospital

RESUMEN Objetivos. Determinar los cambios en las características clínicas y desenlaces intrahospitalarios de los pacientes hospitalizados por COVID-19 en un hospital privado de Caracas durante dos años de pandemia. Materiales y métodos. Estudio retrospectivo, observacional, de pacientes hospitalizados por COVID-19. Se investigó la correspondencia entre las olas de ingresos hospitalarios con las variantes circulantes del SARS-CoV-2 en la población general del Distrito Capital y estado Miranda. Resultados. Se incluyeron 1025 pacientes (569 hombres y 456 mujeres), con edad promedio de 62,9 DE: 16,2 años. Cuatro olas de ingresos hospitalarios fueron identificadas: primera (marzo-noviembre 2020) 150/1025 (14,6%) casos; segunda (diciembre-2020 a mayo-2021) 415/1025 (40,5%) casos; tercera (junio-diciembre 2021) 344/1025 (33,6%) casos; cuarta (enero-febrero 2022) 116/1025 (11,3%) casos. La edad promedio fue mayor en la cuarta ola (primera 64,0±15,7, segunda 61,4±15,8, tercera 62,1±16,5, y cuarta ola 68,5±16,4), mientras que la proporción de pacientes masculinos (primera 66,7%, segunda 58,8%, tercera 50,3%, y cuarta 44,8%), los pacientes con enfermedad grave-crítica (primera 65,3%, segunda 57%, tercera 51,7% y cuarta 44,8%), la estadía intrahospitalaria (primera 9,1±6,0, segunda 9,0±7,3, tercera 8,8±7,7, y cuarta 6,9±5,0 días), los ingresos a la UCI (primera 23,3%, segunda 15,7%, tercera 14,0%, y cuarta 11,2%; p=0,027) y la mortalidad (primera 21.8%, segunda 10,7%, tercera 9,1%, y cuarta 7,1%; p<0,001) disminuyeron progresivamente con el tiempo. Conclusiones. Los resultados muestran menor frecuencia de casos severos y mejoría de los desenlaces intrahospitalarios en dos años de pandemia. Es probable que los cambios en las variantes circulantes, las mejoras del manejo de la enfermedad y la vacunación hayan influido sobre estos resultados.

ABSTRACT Objectives. To determine changes in the clinical characteristics and in-hospital outcomes of patients hospitalized for COVID-19 in a private hospital in Caracas during two years of the pandemic. Materials and Methods. Retrospective, observational study of patients hospitalized for COVID-19. We evaluated the correspondence between waves of hospital admissions and circulating variants of SARS-CoV-2 in the general population of the Capital District and Miranda state. Results. A total of 1025 patients (569 men and 456 women) were included, with a mean age of 62.9 SD: 16.2 years. Four waves of hospital admissions were identified: first (March-November 2020) 150/1025 (14.6%) cases; second (December 2020 to May 2021) 415/1025 (40.5%) cases; third (June-December 2021) 344/1025 (33.6%) cases; fourth (January-February 2022) 116/1025 (11.3%) cases. The mean age was higher in the fourth wave (first: 64.0±15.7, second: 61.4±15.8, third: 62.1±16.5, and fourth wave: 68.5±16.4), while the proportion of male patients (first: 66.7%, second: 58.8%, third: 50.3%, and fourth wave: 44.8%), patients with severe-critical illness (first: 65.3%, second: 57%, third: 51.7%, and fourth wave: 44.8%), in-hospital stay (first: 9.1±6.0, second: 9.0±7.3, third: 8.8±7.7, and fourth wave: 6.9±5.0 days), ICU admissions (first: 23.3%, second: 15.7%, third: 14.0%, and fourth wave: 11.2%; p=0.027) and mortality (first: 21. 8%, second: 10.7%, third: 9.1%, and fourth wave: 7.1%; p<0.001) progressively decreased over time. Conclusions. The results show lower frequency of severe cases and improvement of in-hospital outcomes in two years of the pandemic. Changes in circulating variants, improvements in disease management and vaccination are likely to have influenced these results.

Pentamycin Biosynthesis in Philippine Streptomyces sp. S816: Cytochrome P450-Catalyzed Installation of the C-14 Hydroxyl Group.

Pentamycin is a polyene antibiotic, registered in Switzerland for the treatment of vaginal candidiasis, trichomoniasis, and mixed infections. Chemical instability has hindered its widespread application and development as a drug. Here, we report the identification of Streptomyces sp. S816, isolated from Philippine mangrove soil, as a pentamycin producer. Genome sequence analysis identified the putative pentamycin biosynthetic gene cluster, which shows a high degree of similarity to the gene cluster responsible for filipin III biosynthesis. The ptnJ gene, which is absent from the filipin III biosynthetic gene cluster, was shown to encode a cytochrome P450 capable of converting filipin III to pentamycin. This confirms that the cluster directs pentamycin biosynthesis, paving the way for biosynthetic engineering approaches to the production of pentamycin analogues. Several other Streptomyces genomes were found to contain ptnJ orthologues clustered with genes encoding polyketide synthases that appear to have similar architectures to those responsible for the assembly of filipin III and pentamycin, suggesting pentamycin production may be common in Streptomyces species.

Group B Streptococcus early-onset disease and observation of well-appearing newborns.

BACKGROUND: International guidelines lack a substantial consensus regarding management of asymptomatic full-term and late preterm neonates at risk for early-onset disease (EOS). Large cohorts of newborns are suitable to increase the understanding of the safety and efficacy of a given strategy. METHODS: This is a prospective, area-based, cohort study involving regional birth facilities of Emilia-Romagna (Italy). We compared cases of EOS (at or above 35 weeks' gestation) registered in 2003-2009 (baseline period: 266,646 LBs) and in 2010-2016, after introduction of a new strategy (serial physical examinations, SPEs) for managing asymptomatic neonates at risk for EOS (intervention period: 265,508 LBs). RESULTS: There were 108 cases of EOS (baseline period, n = 60; intervention period, n = 48). Twenty-two (20.4%) remained asymptomatic through the first 72 hours of life, whereas 86 (79.6%) developed symptoms, in most cases (52/86, 60.5%) at birth or within 6 hours. The median age at presentation was significantly earlier in the intrapartum antibiotic prophylaxis (IAP)-exposed than in the IAP-unexposed neonates (0 hours, IQR 0.0000-0.0000 vs 6 hours, IQR 0.0000-15.0000, p<0.001). High number of neonates (n = 531) asymptomatic at birth, exposed to intrapartum fever, should be treated empirically for each newborn who subsequently develops sepsis. IAP exposed neonates increased (12% vs 33%, p = 0.01), age at presentation decreased (median 6 vs 1 hours, p = 0.01), whereas meningitis, mechanical ventilation and mortality did not change in baseline vs intervention period. After implementing the SPEs, no cases had adverse outcomes due to the strategy, and no cases developed severe disease after 6 hours of life. CONCLUSIONS: Infants with EOS exposed to IAP developed symptoms at birth in almost all cases, and those who appeared well at birth had a very low chance of having EOS. The risk of EOS in neonates (asymptomatic at birth) exposed to intrapartum fever was low. Although definite conclusions on causation are lacking, our data support SPEs of asymptomatic newborns at risk for EOS. SPEs seems a safe and effective alternative to laboratory screening and empirical antibiotic therapy.

Strategies for preventing early-onset sepsis and for managing neonates at-risk: wide variability across six Western countries.

Objective: Group B streptococcus (GBS) early-onset sepsis (EOS) has declined after widespread intrapartum antibiotic prophylaxis. However, strategies for preventing EOS may differ across countries. The analysis of their strategies allows to compare the effectiveness of prevention in different countries and suggests opportunities for improvement. Methods: We compared six western countries. Prevention strategies, incidence rates of EOS and approaches for managing neonates at-risk were analysed. Countries were selected because of availability of recommendations for prevention and sufficient epidemiological data for comparison. Results: Five of six countries recommend antenatal vagino-rectal screening. The decline of GBS cases is relevant in most countries, particularly in those with a screening-based strategy, which have reached incidence rates from 0.1 to 0.3/1000 live births and zero or close to zero mortality in full-term newborns. The recommendation for managing asymptomatic neonates at risk for EOS varies according to gestational age and ranges from observation only to laboratory testing plus empirical antibiotics. Chorioamnionitis (suspected or confirmed) is the main indication for carry out laboratory testing and for administering empirical antibiotics. Conclusions: Wide variations exists in preventing EOS. They depend on national epidemiology of GBS infections, compliance, cost, and feasibility of the strategy. The extreme variability of approaches for managing neonates at risk for EOS reflects the even greater uncertainty regarding this issue, and may explain the persisting, great use of resources to prevent a disease that has become very rare nowadays.

Near-Complete Genome Sequence of Ralstonia solanacearum T523, a Phylotype I Tomato Phytopathogen Isolated from the Philippines.

Ralstonia solanacearum strain T523 is the major phytopathogen causing tomato bacterial wilt in the Philippines. Here, we report the complete chromosome and draft megaplasmid genomes with predicted gene inventories supporting rhizosphere processes, extensive plant virulence effectors, and the production of bioactive signaling metabolites, such as ralstonin, micacocidin, and homoserine lactone.

Complete Genome Sequence of Rhizobium sp. Strain 11515TR, Isolated from Tomato Rhizosphere in the Philippines.

Rhizobium sp. strain 11515TR was isolated from the rhizosphere of tomato in Laguna, Philippines. The 7.07-Mb complete genome comprises three replicons, one chromosome, and two plasmids, with a G+C content of 59.4% and 6,720 protein-coding genes. The genome encodes gene clusters supporting rhizosphere processes, plant symbiosis, and secondary bioactive metabolites.

An area-based study on intrapartum antibiotic prophylaxis for preventing group B streptococcus early-onset disease: advances and limitations.

INTRODUCTION: The prevalence of maternal group-B-streptococcus (GBS) colonization and risk factors (RFs) for neonatal early-onset disease (EOD) in Europe are poorly defined. Large-scale information concerning adherence to recommendations for preventing GBS-EOD are lacking. MATERIALS AND METHODS: This was a 3-month retrospective area-based study including all regional deliveries ≥35 weeks' gestation (in 2012). The sensitivity, specificity, positive and negative predictive values, odds ratio and receiver operating characteristic (ROC) curve for intrapartum antibiotic prophylaxis (IAP) among full-term and preterm deliveries and prolonged membrane rupture (PROM) were calculated. RESULTS: Among 7133 women, 259 (3.6%) were preterm (35-36 weeks' gestation). Full-term women were 6874, and 876 (12.7%) had at least 1 RF. Most women (6495) had prenatal screening and 21.4% (1390) were GBS positive. IAP was given to 2369 (33.2%) women (preterm, n = 166; full term, n = 2203). Compared to full-term, preterm women were less likely to receive IAP when indicated (73.2% versus 90.3%, p < 0.01). Full-term women represented the largest area under the curve (AUC, 0.87). PROM showed the highest sensitivity (98.6%), but the lowest specificity (6.9%) and AUC (0.53). CONCLUSIONS: Large-scale prenatal screening and IAP are feasible. Women delivering preterm are less likely to receive IAP when indicated. Most unnecessary antibiotics are given in cases of PROM.

Docencia en medicina centrada en el estudiante: ¿como realizar un foro con estudiantes? / Teaching student-centered medicine: how to make a forum with students?

El artículo médico: La Clase Dialogada publicado por la profesora Luisa Teresa Silva en 1982 es pionero en la Facultad de la Docencia en Medicina Centrada en el estudiante (DMCE). Este artículo tiene como propósitos: 1) transcribir el desarrollo de un foro donde se evaluó con las Escuelas de la Facultad de Medicina, la percepción de esta estrategia docente, que cada día tiene más adeptos entre estudiantes y profesores, 2) honrar y recordar a la profesora Silva por su memorable contribución pedagógica. 3) describir, evaluar y analizar la metodología empleada en la ejecución de un foro con estudiantes de Medicina y 4) organizar sus resultados para la publicación como artículo médico en una revista de difusión científica. El evento tuvo lugar el 30 de enero de 2008, en la Escuela de Medicina “JM Vargas”. Se elaboró una encuesta con cuatro preguntas cerradas, sobre: cómo se enteraron de la actividad, la manera cómo fue organizada, participación en la docencia centrada en el estudiante y cuál estrategia docente prefiere, la cual se distribuyó a los asistentes. El foro se inició con una tormenta de ideas, prosiguió una mesa redonda sobre el devenir histórico y las estrategias docentes. Seguidamente se oyó la participación de los profesores asistentes de las diferentes escuelas de la Facultad, una sesión de preguntas y respuestas y el cierre se realizó con la clase magistral “La autonomía universitaria en las Escuelas de Ciencias de la Salud” a cargo de la Profesora Cecilia García Arocha, para la fecha Secretaria de la UCV. De los 262 inscritos en el foro, 64 (24,4 %) entregaron la encuesta completada. Del análisis de las respuestas se pudo concluir que la DMCE incorpora activamente al alumno y al docente en la búsqueda activa del conocimiento, lo cual supera el tradicional esquema de docencia centrada en el profesor, por el rol pasivo del estudiante en esta última modalidad...

The paper “The Dialogued Class” published by Professor Luisa Teresa Silva in 1982 is considered in the Faculty of Medicine as a pioneer of Medical Teaching Centered in the Student (MTSC). The present publication has the following objectives: 1) Transcribe the forum carried out in order to evaluate the perception held by different members of the Faculty regarding about these teaching strategies, which have gained more acceptance in students and professors every day, 2) Honor and remember Professor Silva for her memorable pedagogic contribution, 3) Describe, evaluate and analyze the methodology used in the execution of a forum made with medical students, and 4) Organize the results for publication as a medical article in a scientific magazine. The event took place on the 30th of January 2008, in the Medical School “Jose María Vargas”. A questionnaire was made with four closed questions: 1) How the assistants learned about the activity, 2) Considerations about the quality of the event organization, 3)Participation and preferences in the teaching strategy centered in the student. The questionnaire was delivered to the assistants. The forum was initiated with a brainstorm session and continued with a round table discussion about the historical outcome of the teaching strategies, followed by comments from professors of the different Health Science Schools in the Faculty, a session with questions and answers, and a closure with the lecture: “The university autonomy in all the Science Health Schools”, by Professor Cecilia Garcia Arocha, by the time, Secretary of the UCV. There were 262 persons registered in the forum and only 64 (24.4 %) returned the filled questionnaire. In conclusion, the MTSC actively incorporates students and professors in an active search for knowledge that overcomes the traditional teaching system, centered on the professor, with a passive role from the student...

Universal antenatal screening for group B streptococcus in Emilia-Romagna.

BACKGROUND: Group B streptococcus (GBS) is a leading cause of severe infections in newborns. Intrapartum antibiotic chemoprophylaxis (IAP) reduces the rate of early-onset disease. The aim of this study is to determine the degree of clinicians' compliance with the suggested protocol for GBS prevention in Emilia-Romagna (Italy). METHODS: Characteristics of each delivery were prospectively recorded in the period between October 2005 to December 2005. Standardized pro-forma were used to collect data. RESULTS: Among 5118 babies, 7.2% (369) were preterm and 92.3% were born at term (4749). Antenatal screening was performed in 86.6% of women who delivered at term, of which 18.1% were GBS culture-positive. Information regarding culture site was available in 93.2% of women screened and recto-vaginal cultures were documented in 42.7%. IAP was administered to 28.7% of 3937 women at term who had either spontaneous delivery or emergency caesarean section. In this cohort, 15.9% were diagnosed GBS culture-positive, of which 92.6% received IAP. Prophylaxis was also administered to 8.4% (331) of women for no apparent reason. Compared with tertiary level hospitals, women delivering in primary/secondary hospitals were more likely to be both GBS screened (P < 0.0001; OR 3.04; CI 2.33-3.97) and to receive prophylaxis ≥4 hours before delivery (P = 0.0025; OR 1.57; CI 1.17-2.12). CONCLUSIONS: GBS screening was performed in >85% of women and >90% of culture-positive women received prophylaxis. However, there is a need to educate clinicians about protocol adherence, as most cultures were suboptimal and cases of unnecessary IAP were administered. The screening was more effective in hospitals with fewer deliveries.

Group B Streptococcus early-onset disease in Emilia-romagna: review after introduction of a screening-based approach.

BACKGROUND: Group B Streptococcus (GBS) is a leading cause of neonatal bacterial infections. Early-onset infections have decreased in recent years but, despite considerable efforts poured into prevention, cases continue to occur. OBJECTIVES: To analyze trends and identify determining factors for the persistence of the GBS infections. To evaluate the impact of antenatal screening and intrapartum chemoprophylaxis on the clinical presentation of the infection. METHODS: A prospective cohort, population-based study has been ongoing in Emilia-Romagna (Italy) since 2003. Invasive GBS infections, observed between 2003 and 2008 in infants aged < 7 days were analyzed. RESULTS: Among 214,120 live births, 61 early-infections were observed. Fourteen infants (23.0%) were born preterm. Among 47 infants who were delivered at term, 28 were born to mothers who had no risk factors and 7 were born to mothers who had none other than GBS colonization. Forty-one women at term had been screened prenatally; among them, only 10 were documented as GBS culture-positive.Disease severity was highest in infants at lower gestational ages, but most meningitis cases were observed in term infants born to mothers who were GBS culture-negative at screening.Nine newborns had culture-proven infection despite having received intrapartum antibiotics. They were born to mothers with > or =1 obstetrical risk factors and 5 mothers had been treated during labor with macrolides. CONCLUSION: Most infections presented in infants whose mothers had been screened as GBS culture-negative. Missed opportunities for prevention contributed more than prophylaxis failures to the early-onset disease burden.

Group B streptococcal infections in a northern region of Italy.

BACKGROUND: Group B streptococcus is a leading cause of neonatal bacterial infections. Despite adoption of preventive strategies, cases of infection continue to occur and there is concern that widespread antimicrobial prophylaxis might delay rather than prevent disease onset, increasing the rates of late-onset diseases. OBJECTIVES: The purpose of this study was to determine the incidence and clinical features of early- and late-onset group B streptococcus disease in a northern region of Italy where a screening-based approach had been proposed. METHODS: A population-based study was prospectively conducted in Emilia-Romagna, Italy. Infections that occurred during 2003-2005 in infants aged <3 months were analyzed. RESULTS: Among 112,933 live births, 56 cases of invasive disease (30 early- and 26 late-onset disease) were observed, giving an annual group B streptococcus disease incidence of 0.50 per 1000 live births. Eleven infants with early-onset disease showed no signs of illness or were mildly ill, whereas 19 had moderate-to-severe symptoms, and culture-proven meningitis was found in 2. Risk factors were detected in 12 women. Twenty-two mothers had antenatal screening; 5 were group B streptococcus colonized, but 17 were culture-negative. Prophylaxis was administered in 3 women. Three infants with late-onset diseases were mildly ill, whereas 23 had moderate-to-severe symptoms. Risk factors were found in 7 mothers. Late-onset diseases were clinically more severe than early-onset diseases; meningitis was diagnosed in 12 infants, and 4 of 26 died. CONCLUSIONS: The incidence of early-onset disease was low. Some early infections were still observed because of negative screening results or missed opportunity for prevention. Late-onset diseases accounted for most meningitis cases and deaths. Strict adherence to protocols and adoption of optimal culture methods would further improve prevention of early-onset disease, but the aim of future strategies should be the prevention of all invasive diseases.

Neonatal cytomegalovirus blood load and risk of sequelae in symptomatic and asymptomatic congenitally infected newborns.

OBJECTIVE: Human cytomegalovirus (CMV) is a ubiquitous human-specific DNA virus and is the main cause of congenital virus infection in developed countries leading to psychomotor impairment and deafness. Diagnostic techniques for CMV detection have greatly improved during recent years with the advent of sophisticated serological and virological methods. The aim of the present study was to assess the diagnostic and prognostic value of detection and quantification of virus in neonatal blood samples of symptomatic and asymptomatic newborns with CMV congenital infection. METHODS: Between January 1997 and December 2003, we studied 99 newborns who were born to women with primary, recurrent, and undefined CMV infection during pregnancy. CMV congenital infection was identified by isolation of the virus in urine within the second week of life. Fifty-eight of 99 infants were infected and were assessed clinically for disease in the newborn period and classified as having symptomatic or asymptomatic infection on the basis of physical, instrumental, and laboratory findings. The infants were followed up from birth according to a protocol of the tertiary NICU at the University of Bologna in a prospective study of long-term sequelae of congenital infection. Forty-seven blood samples were obtained from 47 infants in the neonatal period: 34 were examined for pp65 antigenemia test and 44 for qualitative and quantitative polymerase chain reaction (PCR and qPCR). Sequelae at 12 months were evaluated in a group of 50 infants. RESULTS: Antigenemia was positive in only 10 of 34 samples of infected newborns (29.4% sensitivity). PCR was performed in 44 samples of infected newborns and was positive in all (100% sensitivity). qPCR showed a finding of > or =100 copies per 10(5) of polymorphonuclear leukocytes (PMNLs) in 39 of 44 samples; in the other 5 cases, the number of copies per 10(5) PMNLs was <100. Between symptomatic and asymptomatic newborns, the mean values of viral blood load determined by qPCR turned out to be significantly higher in symptomatic newborns. Mean values of neonatal blood viral load were statistically higher in newborns who developed sequelae than in those who did not. Of 20 children with a neonatal viral blood load of <1000 copies per 10(5) PMNLs, 19 did not develop sequelae (negative predictive value: 95%), whereas 2 of 3 with a viral blood load of >10,000 copies did develop sequelae. CONCLUSIONS: Different viremia value ranges are correlated to a different risk of sequelae: approximately 70% sequelae were found in newborns with a qPCR higher than 10,000 copies per 10(5) PMNLs. Low neonatal viral blood load detected by pp65 antigenemia test and qPCR was highly predictive of absence of sequelae: DNAemia <1000 copies per 10(5) PMNLs has a negative predictive value of 95%. As an independent predictive factor of outcome, neonatal viremia is another useful element for neonatal counseling and therapeutic choices in symptomatic and asymptomatic newborns.

Congenital infection with human herpesvirus 6 variant B associated with neonatal seizures and poor neurological outcome.

Human herpesvirus 6 (HHV 6) has neurotropic and neuroinvasive properties. The virus has been found in the cerebrospinal fluid of many children with aseptic meningoencephalitis. Intrauterine transmission has been documented by HHV 6 DNA detection in cord blood specimens of apparently healthy newborns and in fetuses following spontaneous abortions. A patient is described with early neonatal afebrile seizures resulting from a congenital HHV 6 variant B infection disclosed by repeated detection of viral genome by polymerase chain reaction (PCR) in cerebrospinal fluid in the first days of life. At follow-up, magnetic resonance imaging (MRI) studies disclosed hyperintensities in the periventricular white matter and basal ganglia, associated with cerebral atrophy. Further follow-up at 18 months revealed poor neurological outcome with mild neurodevelopmental retardation, strabismus and hypertonia of legs. This report provides evidence of neurological involvement after HHV 6 vertical transmission, and the association with neurological sequelae.

HLA DR13 and HCV vertical infection.

Risk factors affecting vertical hepatitis C virus (HCV) transmission are not completely known, if we exclude maternal HIV coinfection. We hypothesized that immunogenetic factors related to maternal or neonatal HLA profiles may affect HCV vertical transmission. HLA typing (microcytotoxicity assay on blood samples) was performed in 18 infants affected by vertically transmitted HCV infection and in 17 serum-reverted infants. (Serum-reversion is defined as antibody negative by 1 year of age and persistently HCV-RNA negative.) Moreover, HLA typing was performed in 20 mothers. Logistic regression analysis showed a significant negative association between children's HLA-DR13 antigens and risk of HCV vertical transmission (p < 0.01). This association persisted in a model including the maternal HIV status: HLA DR13 and maternal HIV coinfection showed a separate, opposite effect on vertical HCV infection (p < 0.01 and p < 0.001, respectively). The relative risk estimate for the ratio of not-infected to infected children in the presence of DR13 was 8.4 (95% confidence bounds, 1.1-60.8). Breast-feeding did not affect the risk of vertical HCV transmission. Maternal HLA profile did not relate to vertical infection. The present study reveals a significant association between HLA-DR13 and the likelihood of seroreversion in infants born to HCV-infected mothers. The findings of the present study could help in better understanding the pathogenesis of vertical HCV infection and in better identifying the cases at higher risk, which would be useful for the development of prevention strategies. It is possible that DR13 modulates the immune response to viruses, enhancing their clearance and, thus, in the case of HCV, exerting a protective role against the development of vertical infection.