Molecular Signaling and Metabolic Responses during the Interaction between Human Keratinocytes (HaCaT) and the Dermatophyte Trichophyton rubrum.

Trichophyton rubrum is the leading causative agent of dermatophytosis worldwide. Keratinocytes are the first line of defense that drives an immune response against fungal invasion. Host-specific pattern recognition receptors (PRRs) recognize pathogen-associated molecular patterns (PAMPs) to trigger immunological pathways. Fungal cell wall components are the primary sources of fungal PAMPs, and some pathogens increase cell wall rearrangement to evade the immune system. Glycolysis and enhanced lactate levels are critical for improving host immune responses to fungal infections. Using reverse transcription-quantitative polymerase chain reaction (RT-qPCR), we evaluated the transcriptional responses of human genes involved in fungal recognition and glycolytic metabolism and fungal cell-wall-related genes in a co-culture model of human keratinocytes with T. rubrum. We observed the upregulation of several Toll-like receptors (TLRs), NOD-like receptors (NLRs), and glycolytic genes. Complementarily, we measured intra- and extracellular glucose levels and the increase in lactate production in the co-culture supernatant. We noted a distinct transcriptional regulation pattern of fungal cell-wall-related genes from fungal growth on keratin as the primary carbon source compared to co-culture with human keratinocytes. Our results showed new insights into the transcriptional adaptation of keratinocytes, particularly in regulating genes involved in sensing and metabolic processes, during the interaction with T. rubrum.

Addressing Microbial Resistance Worldwide: Challenges over Controlling Life-Threatening Fungal Infections.

Fungal infections are a serious global concern because of their ability to spread and colonize host tissues in immunocompromised individuals. Such infections have been frequently reported worldwide and are currently gaining clinical research relevance owing to their resistant character, representing a bottleneck in treating affected people. Resistant fungi are an emergent public health threat. The upsurge of such pathogens has led to new research toward unraveling the destructive potential evoked by these species. Some fungi-grouped into Candida, Aspergillus, and Cryptococcus-are causative agents of severe and systemic infections. They are associated with high mortality rates and have recently been described as sources of coinfection in COVID-hospitalized patients. Despite the efforts to elucidate the challenges of colonization, dissemination, and infection severity, the immunopathogenesis of fungal diseases remains a pivotal characteristic in fungal burden elimination. The struggle between the host immune system and the physiological strategies of the fungi to maintain cellular viability is complex. In this brief review, we highlight the relevance of drug resistance phenotypes in fungi of clinical significance, taking into consideration their physiopathology and how the scientific community could orchestrate their efforts to avoid fungal infection dissemination and deaths.

The Transcription Factor StuA Regulates the Glyoxylate Cycle in the Dermatophyte Trichophyton rubrum under Carbon Starvation.

Trichophyton rubrum is the primary causative agent of dermatophytosis worldwide. This fungus colonizes keratinized tissues and uses keratin as a nutritional source during infection. In T. rubrum-host interactions, sensing a hostile environment triggers the adaptation of its metabolic machinery to ensure its survival. The glyoxylate cycle has emerged as an alternative metabolic pathway when glucose availability is limited; this enables the conversion of simple carbon compounds into glucose via gluconeogenesis. In this study, we investigated the impact of stuA deletion on the response of glyoxylate cycle enzymes during fungal growth under varying culture conditions in conjunction with post-transcriptional regulation through alternative splicing of the genes encoding these enzymes. We revealed that the ΔstuA mutant downregulated the malate synthase and isocitrate lyase genes in a keratin-containing medium or when co-cultured with human keratinocytes. Alternative splicing of an isocitrate lyase gene yielded a new isoform. Enzymatic activity assays showed specific instances where isocitrate lyase and malate synthase activities were affected in the mutant strain compared to the wild type strain. Taken together, our results indicate a relevant balance in transcriptional regulation that has distinct effects on the enzymatic activities of malate synthase and isocitrate lyase.

Peptidase Regulation in Trichophyton rubrum Is Mediated by the Synergism Between Alternative Splicing and StuA-Dependent Transcriptional Mechanisms.

Trichophyton rubrum is the most common causative agent of dermatophytosis worldwide and uses keratinized substrates such as skin and nails as its main source of nutrition during infection. Its pathogenic character relies on colonization and viability maintenance at the target host sites. Since fungal physiology must adapt and respond to host conditions for the successful establishment of infection, biological mechanisms are constantly being triggered by T. rubrum to guarantee its survival in the host environment. The ability of this fungus to sense and modulate the secretion of specific proteases according to environmental pH signaling is considered as a pivotal virulence factor for effective invasion and persistence of infection in the host. Transcriptional regulation of genes encoding specific proteases, such as peptidases, is a key biological process that drives physiological modulation to meet fungal requirements. It accomplishes a robust balance among transcript isoforms that can be directed to perform distinct cellular functions. Thus, alternative splicing mechanisms are suitable for fungal cells to establish a balance toward reprogramming protein translation to impair or boost physiological conditions. In this study, we investigated the role of alternative splicing, especially intron retention events, in generating isoforms of virulence factors in T. rubrum mediated by transcriptional coordination of the protein StuA, a recently described transcription factor in this fungus. By analyzing the previous gene expression data provided by RNA-sequencing and after validation by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR), we observed that two peptidase-coding genes (TERG_00734 and TERG_04614) could be direct targets of alternative splicing in the presence of keratin. Furthermore, protease isoforms generated by alternative splicing in T. rubrum were also detected in a co-culture with human keratinocytes, highlighting the role of these proteins in keratin deconstruction. Our results strongly suggest the influence of StuA on the regulation of virulence factors in T. rubrum and dermatophyte infections by triggering the transcription of the peptidase genes mentioned above in an alternative splicing-independent balance. The results elucidate how fungal cells drive alternate splicing to promote physiological adaptations and show that transcriptional regulation and virulence traits are robust elements required for dermatophyte infection.

Pacientes com lesões crônicas em membros inferiores, atendidos em hospital particular: estudo de prevalência / Patients with chronic lower-limb lesions, seen in a private hospital: a prevalence study / Pacientes con lesiones crónicas en extremidades inferiores, atendidos en un hospital privado: un estudio de prevalencia

Objetivo: Avaliar a população acometida de lesões, atendida em serviço de atenção a feridas crônicas em ambulatório de hospital particular. Método: Estudo descritivo, transversal e analítico. A amostra foi de 320 pacientes com lesões, atendidas neste serviço. Os dados foram coletados via prontuários eletrônicos. Foi calculado taxa de prevalência de lesões de membros inferiores, frequências absoluta e relativa para a descrição das variáveis coletadas, média e desvio padrão. Resultados: A maioria era do sexo feminino, de cor branca, casada, aposentada, com idade média de 68,2 anos, com sobrepeso e portando lesão única. A maior parte tinha úlcera venosa, seguida de lesão em pé diabético e lesão arterial. O restante (lesões em outros locais) somou 33,4%. A prevalência de lesões em membros inferiores foi de 66%. Conclusão: A prevalência de lesão em membros inferiores foi alta, porém na população estudada não havia pacientes hígidos, uma vez que todos apresentavam alguma lesão. (AU).

Objective: To evaluate the population affected by injuries, seen in a service for the care of chronic wounds in an outpatient clinic of a private hospital. Method: Descriptive, cross-sectional, and analytical study. The sample consisted of 320 patients with injuries. Data were collected through the electronic medical records. Calculation of the prevalence rate of lower limb injuries and of absolute and relative frequencies were used to describe the variables collected and mean and standard deviation. Results: The majority was female, white, married, retired, with a mean age of 68.2 years, overweight and with a single lesion. Most had venous ulcers, followed by diabetic foot injury and arterial injury. The remainder totaled 33.4%. The prevalence of lower limb injuries was 66%. Conclusion: The prevalence of injuries in the lower limbs was high, and in the studied population there were no healthy patients since all had some type of chronic injury. (AU).

Objetivo: Evaluar la población afectada por lesiones, tratada en servicio de atención crónica de heridas en un hospital privado. Método: Estudio descriptivo, transversal y analítico. La muestra consistió en 320 pacientes con lesiones. Los datos se obtuvieron de registros médicos electrónicos Fue utilizado cálculo de tasa de prevalencia de lesiones de extremidades inferiores y frecuencias absolutas y relativas para la descripción de las variables recogidas y la media y la desviación estándar. Resultados: La mayoría era mujer, blanca, casada, jubilada, con edad media de 68,2 años, con sobrepeso y portando una sola lesión. La mayoría tenía úlcera venosa, seguida de lesiones de pie diabético y lesiones arteriales. El resto totalizo 33,4%. La prevalencia de lesiones en las extremidades inferiores fue del 66%. Conclusión: La prevalencia de lesiones en las extremidades inferiores fue alta, y en la población estudiada no había pacientes sanos, ya que todos tenían algún tipo de lesión. (AU).

Estimates of Antibacterial Consumption in Timor-Leste Using Distribution Data and Variation in Municipality Usage Patterns.

The association between antimicrobial resistance and antimicrobial usage has become a growing global concern. Many lower-middle income countries including Timor-Leste (TL) have limited information on antimicrobial usage, although recent research suggests increasing resistance rates among human pathogens there. The aim of this study was to use distribution data to estimate antibiotic consumption at both the national and sub-national level in Timor-Leste, stratifying into resistance class and adherence to the national essential medicines list (EML) and WHO AWaRe guidelines. A retrospective review of distribution data from Timor-Leste central medical store (SAMES) was undertaken to give a defined daily dose (DDD)/1000 inhabitants/day using WHO methodology. National antibiotic distribution in the TL EML in 2019 was estimated at 11.1 DDD/1000 inhabitants/day, comparable to consumption rates observed in other lower-middle-income countries using similar methodology. Differences in distribution quantities were noted between municipalities, with 4 of the 13 municipalities notably above the national average and around 32% of listed restricted antimicrobials distributed incongruent with the EML. This study provides insights into estimated antimicrobial consumption in Timor-Leste that has previously been poorly defined. Estimates of consumption can be used to understand emerging resistance in this small island nation, add to the body of knowledge on antimicrobial use to advise policy and guideline development, and help with stewardship activities.

Enhanced IL-6 and IL-12B Gene Expression After SARS-CoV-2 Infection in Leprosy Patients May Increase the Risk of Neural Damage.

Experts have called attention to the possible negative impact of the coronavirus disease 2019 (COVID-19)-related cytokine storm syndrome on the progression of leprosy-related disabilities. We assessed the frequency of reactional states in patients co-infected with Mycobacterium leprae and severe acute respiratory syndrome (SARS) coronavirus (CoV) 2 (SARS-CoV-2). We consecutively included patients during the first peak of the COVID-19 epidemic in Brazil and analyzed the expressions of genes encoding interleukin (IL)-1ß, IL-6, IL-8, IL-10, IL-12A, IL-12B, and tumor necrosis factor-α in peripheral blood mononuclear cells. We included 64 leprosy patients and 50 controls. Twelve of the leprosy patients and 14 of the controls had been diagnosed with COVID-19. Co-infection was associated with increased IL-6 (P = 0.043) and IL-12B (P = 0.017) expression. The median disability grades were higher for leprosy/COVID-19 patients; however, the difference was not significant (P = 0.194). Patients co-infected with M. leprae and SARS-CoV-2 may experience a higher-grade proinflammatory state.

CRZ1 regulator and calcium cooperatively modulate holocellulases gene expression in Trichoderma reesei QM6a.

Trichoderma reesei is the main filamentous fungus used in industry to produce cellulases. Here we investigated the role of CRZ1 and Ca2+signaling in the fungus T. reesei QM6a concerning holocellulases production. For this, we first searched for potential CRZ1 binding sites in promoter regions of key genes coding holocellulases, as well as transcriptional regulators and sugar and calcium transporters. Using a nearly constructed T. reeseiAcrz1 strain, we demonstrated that most of the genes expected to be regulated by CRZ1 were affected in the mutant strain induced with sugarcane bagasse (SCB) and cellulose. In particular, our data demonstrate that Ca2+ acts synergistically with CRZ1 to modulate gene expression, but also exerts CRZ1-independent regulatory role in gene expression in T. reesei, highlighting the role of the major regulator Ca2+ on the signaling for holocellulases transcriptional control in the most part of cellulases genes here investigated. This work presents new evidence on the regulatory role of CRZ1 and Ca2+ sensing in the regulation of cellulolytic enzymes in T. reesei, evidencing significant and previously unknown function of this Ca2+sensing system in the control key transcriptional regulators (XYR1 and CRE1) and on the expression of genes related to sugar and Ca2+ transport.

Reverse Engineering of an Aspirin-Responsive Transcriptional Regulator in Escherichia coli.

Bacterial transcription factors (TFs) are key devices for the engineering of complex circuits in many biotechnological applications, yet there are few well-characterized inducer-responsive TFs that could be used in the context of an animal or human host. We have deciphered the inducer recognition mechanism of two AraC/XylS regulators from Pseudomonas putida (BenR and XylS) for creating a novel expression system responsive to acetyl salicylate (i.e., aspirin). Using protein homology modeling and molecular docking with the cognate inducer benzoate and a suite of chemical analogues, we identified the conserved binding pocket of BenR and XylS. By means of site-directed mutagenesis, we identified a single amino acid position required for efficient inducer recognition and transcriptional activation. Whereas this modification in BenR abolishes protein activity, in XylS, it increases the response to several inducers, including acetyl salicylic acid, to levels close to those achieved by the canonical inducer. Moreover, by constructing chimeric proteins with swapped N-terminal domains, we created novel regulators with mixed promoter and inducer recognition profiles. As a result, a collection of engineered TFs was generated with an enhanced response to benzoate, 3-methylbenzoate, 2-methylbenzoate, 4-methylbenzoate, salicylic acid, aspirin, and acetylsalicylic acid molecules for eliciting gene expression in E. coli.

Synthetic and minimalist vectors for Agrobacterium tumefaciens-mediated transformation of fungi.

We present a collection of minimalist binary vectors for transformation through ATMT applicable to several fungi species. pLUO plasmid binary vectors consist of a reporter module containing fluorescent proteins, mCherry or eGFP, flanked by a multiple cloning site and a transcription terminator site. They also present a synthetic gene allowing resistance to Hygromicin B flanked by alternate promoters, one for yeast and another for filamentous fungi. Left and right borders were added for Agrobacterium tumefaciens recognition, and a minimal broad-host range RK2 replication origin. Transformation was validated in the pathogenic fungus Paracoccidioides lutzii. Hence, we developed an efficient and reliable molecular tool for fungal transformation: minimalist, synthetic, modular, and available in four different versions, and these can still be readily modified using a few primers and few cloning steps.

Lower limb wounds in diabetic and non-diabetic patients: survival analysis. / Feridas em membros inferiores em diabéticos e não diabéticos: estudo de sobrevida.

OBJECTIVE: To evaluate the survival of wounds in lower limbs of diabetic and non-diabetic patients. METHOD: Retrospective cohort study of patients with lower limb ulcers treated at a specialized center between 2011 and 2013. Outcome: healing of lower limb injuries in days. The survival function of lower limb wounds and the differences between diabetic and non-diabetic were analyzed. The Log-rank test was used to compare the survival curves between the study groups. RESULTS: In up to 600 days, 23% of the diabetic patients presented wound healing, while 63% of the non-diabetic patients had their wounds healed, with a statistically significant difference in survival curves in comparison between the groups. The Hazard Ratios (RH) of healing were lower for diabetic patients (HR = 0.13, 95% CI = 0.02-0.97). CONCLUSION: The results show that there is a delay in wound healing in diabetic patients.

The art of vector engineering: towards the construction of next-generation genetic tools.

When recombinant DNA technology was developed more than 40 years ago, no one could have imagined the impact it would have on both society and the scientific community. In the field of genetic engineering, the most important tool developed was the plasmid vector. This technology has been continuously expanding and undergoing adaptations. Here, we provide a detailed view following the evolution of vectors built throughout the years destined to study microorganisms and their peculiarities, including those whose genomes can only be revealed through metagenomics. We remark how synthetic biology became a turning point in designing these genetic tools to create meaningful innovations. We have placed special focus on the tools for engineering bacteria and fungi (both yeast and filamentous fungi) and those available to construct metagenomic libraries. Based on this overview, future goals would include the development of modular vectors bearing standardized parts and orthogonally designed circuits, a task not fully addressed thus far. Finally, we present some challenges that should be overcome to enable the next generation of vector design and ways to address it.

Feridas em membros inferiores em diabéticos e não diabéticos: estudo de sobrevida / Heridas en miembros inferiores en diabéticos y no diabéticos: estudio de supervivencia / Lower limb wounds in diabetic and non-diabetic patients: survival analysis

Resumo OBJETIVO Avaliar a sobrevida de feridas em membros inferiores de pacientes diabéticos e não diabéticos. MÉTODO Estudo de coorte retrospectivo de pacientes com úlceras de membros inferiores tratados em centro especializado entre 2011 e 2013. Desfecho: cicatrização de lesões de membros inferiores em dias. Realizou-se análise da função de sobrevida das feridas de membros inferiores e das diferenças entre diabéticos e não diabéticos. Aplicou-se o teste Log-rank para comparação das curvas de sobrevida entre os grupos de estudo. RESULTADOS Em até 600 dias, 23% dos diabéticos apresentaram cicatrização das feridas, enquanto 63% dos não diabéticos tiveram suas feridas cicatrizadas, com diferença estatística das curvas de sobrevida na comparação entre os grupos. Os Hazard Ratios (HR) de cicatrização foram menores para pacientes diabéticos (HR = 0,13, IC95% = 0,02-0,97). CONCLUSÃO Os resultados mostram que há retardo na cicatrização de feridas em pacientes diabéticos.

Resumen OBJETIVO Evaluar la supervivencia de las heridas en miembros inferiores de pacientes diabéticos y no diabéticos. MÉTODO Estudio de cohorte retrospectivo de pacientes con úlceras de miembros inferiores tratados en centro especializado entre 2011 y 2013. CONCLUSIÓN cicatrización de lesiones de miembros inferiores, en días. Se realizó un análisis de la función de supervivencia de las heridas de miembros inferiores y de las diferencias entre diabéticos y no diabéticos. Se aplicó el test Log-rank para la comparación de las curvas de supervivencia entre los grupos de estudio. RESULTADOS En un total de 600 días, el 23% de los diabéticos presentaron cicatrización de las heridas, mientras que el 63% de los no diabéticos tuvieron sus heridas cicatrizadas, con diferencia estadística de las curvas de sobrevida en la comparación entre los grupos. Los Hazard Ratios (HR) de cicatrización fueron menores para pacientes diabéticos (HR = 0,13, IC95% = 0,02-0,97). CONCLUSIÓN Los resultados muestran que hay retraso en la cicatrización de heridas en pacientes diabéticos.

Abstract OBJECTIVE To evaluate the survival of wounds in lower limbs of diabetic and non-diabetic patients. METHOD Retrospective cohort study of patients with lower limb ulcers treated at a specialized center between 2011 and 2013. Outcome: healing of lower limb injuries in days. The survival function of lower limb wounds and the differences between diabetic and non-diabetic were analyzed. The Log-rank test was used to compare the survival curves between the study groups. RESULTS In up to 600 days, 23% of the diabetic patients presented wound healing, while 63% of the non-diabetic patients had their wounds healed, with a statistically significant difference in survival curves in comparison between the groups. The Hazard Ratios (RH) of healing were lower for diabetic patients (HR = 0.13, 95% CI = 0.02-0.97). CONCLUSION The results show that there is a delay in wound healing in diabetic patients.

Acrania-exencephaly-anencephaly sequence phenotypic characterization using two- and three-dimensional ultrasound between 11 and 13 weeks and 6 days of gestation.

The study presents a pictorial essay of acrania-exencephaly-anencephaly sequence using two-(2D) and three-dimensional (3D) ultrasonography, documenting the different phenotypic characterization of this rare disease. Normal and abnormal fetuses were evaluated during the first trimester scan. The International Society of Ultrasound in Obstetrics and Gynecology practice guidelines were adopted to standardize first trimester anatomical ultrasound screening. The guidelines outline the importance of systematic fetal head and brain examination including the formation of cranial bones, choroid-plexus and ventricles. Acrania-exencephaly-anencephaly sequence and/or other neural tube defects, such as meningoencephalocele, may be identified during a routine 11-14 week scan. Early first trimester detection of acrania-exencephaly-anencephaly sequence with the characterization of different related phenotypes, 2D and 3D ultrasound imaging as well as differential diagnosis are also presented in this pictorial essay. The main diagnostic ultrasound features of the disease may be characterized by findings of acrania with increased amniotic fluid echogenicity; "Mickey-Mouse" bi-lobular face, cystic, elongated, irregular and overhanging head morphology. Lightening techniques have also been added to 3D ultrasound to enhance anatomical details. Moreover, discordant amniotic fluid echotexture in the setting of twin pregnancies may be the first sign of acrania-exencephaly-anencephaly sequence. Extracranial malformations, aneuploidy and genetic syndromes associated with acrania-exencephaly-anencephaly sequence are also reported and described. First trimester neuroscan by an expert sonographer with appropriate training together with the application of standardized protocol are essential for a high detection rate of this rare type of neural tube defect malformation during a scan performed at 11 and 13 weeks and 6 days.

Systems and Synthetic Biology Approaches to Engineer Fungi for Fine Chemical Production.

Since the advent of systems and synthetic biology, many studies have sought to harness microbes as cell factories through genetic and metabolic engineering approaches. Yeast and filamentous fungi have been successfully harnessed to produce fine and high value-added chemical products. In this review, we present some of the most promising advances from recent years in the use of fungi for this purpose, focusing on the manipulation of fungal strains using systems and synthetic biology tools to improve metabolic flow and the flow of secondary metabolites by pathway redesign. We also review the roles of bioinformatics analysis and predictions in synthetic circuits, highlighting in silico systemic approaches to improve the efficiency of synthetic modules.

Fetoscopic laser photocoagulation for twin-to-twin transfusion syndrome treatment: initial experience in tertiary reference center in Brazil.

OBJECTIVE: To evaluate the initial maternal and perinatal outcomes of fetoscopic laser photocoagulation for the treatment of twin-to-twin transfusion syndrome (TTTS) in a referral center in Brazil. METHODS: This prospective observational study analyzed 24 fetoscopic laser photocoagulation procedures at 18-26 weeks of gestation. TTTS severity was determined using the Quintero classification. Blood vessels that crossed the interamniotic membrane were nonselectively photocoagulated. The χ2 test and Mann-Whitney U test were used for the statistical analysis. RESULTS: The mean (±standard deviation) age of pregnant women, gestational age at surgery, surgical time, gestational age at birth, and newborn weight were 32.2±4.1 years, 20.7±2.9 weeks, 51.8±16.7 minutes, 30.5±4.1 weeks, and 1,531.0±773.1 g, respectively. Using the Quintero classification, there was a higher percentage of cases in stage III (54.2%), followed by stages IV (20.8%), II (16.7%), and I (8.3%). Ten (41.7%) donor fetuses died and 14 (58.3%) donor fetuses survived until the end of gestation. Placental insertion location (anterior vs. posterior) did not affect the incidence of iatrogenic septostomy, surface bleeding, and premature rupture of membranes until the end of gestation. The death rate of donor and recipient fetuses before 24th gestational week increased with severity of TTTS. CONCLUSION: The maternal and perinatal outcomes resulting from the implementation of a new minimally invasive surgical technique are in line with those obtained in major centers worldwide, considering the learning curves and infrastructures.

Fetoscopic laser photocoagulation for twin-to-twin transfusion syndrome treatment: initial experience in tertiary reference center in Brazil

OBJECTIVE: To evaluate the initial maternal and perinatal outcomes of fetoscopic laser photocoagulation for the treatment of twin-to-twin transfusion syndrome (TTTS) in a referral center in Brazil. METHODS: This prospective observational study analyzed 24 fetoscopic laser photocoagulation procedures at 18–26 weeks of gestation. TTTS severity was determined using the Quintero classification. Blood vessels that crossed the interamniotic membrane were nonselectively photocoagulated. The χ2 test and Mann-Whitney U test were used for the statistical analysis. RESULTS: The mean (±standard deviation) age of pregnant women, gestational age at surgery, surgical time, gestational age at birth, and newborn weight were 32.2±4.1 years, 20.7±2.9 weeks, 51.8±16.7 minutes, 30.5±4.1 weeks, and 1,531.0±773.1 g, respectively. Using the Quintero classification, there was a higher percentage of cases in stage III (54.2%), followed by stages IV (20.8%), II (16.7%), and I (8.3%). Ten (41.7%) donor fetuses died and 14 (58.3%) donor fetuses survived until the end of gestation. Placental insertion location (anterior vs. posterior) did not affect the incidence of iatrogenic septostomy, surface bleeding, and premature rupture of membranes until the end of gestation. The death rate of donor and recipient fetuses before 24th gestational week increased with severity of TTTS. CONCLUSION: The maternal and perinatal outcomes resulting from the implementation of a new minimally invasive surgical technique are in line with those obtained in major centers worldwide, considering the learning curves and infrastructures.

Prenatally diagnosed fetal tumors of the head and neck: a systematic review with antenatal and postnatal outcomes over the past 20 years.

AIM: The aim of this study was to review prenatally diagnosed tumors of the head and neck in the fetus and to report antenatal and postnatal outcomes. METHODS: PubMed/Medline, EMBASE/SCOPUS, Cochrane database and Google Scholar were reviewed over the last 20 years. No language or article type restriction was used. RESULTS: A total of 1940 record were retrieved. Of the 713 records screened, 566 full-text articles were assessed for eligibility. After 445 articles were excluded for specified reasons, 111 studies met the research criteria and were included for qualitative analysis. Overall, 306 cases of fetal tumors of the head and neck were reviewed. Maternal age was an independent factor. The mean maternal age was 28.2 years and gestational age at prenatal diagnosis was 27.1 weeks. Conventional 2D ultrasound was the standard diagnostic procedure in 27.9% of cases and was implemented in 27.3% of cases by 3D ultrasound and fetal magnetic resonance imaging (MRI). Diagnostic evaluation of intracranial spreading and high-airway obstructions was greatly enhanced by fetal MRI. The more common type of fetal tumor was hemangioma/lymphangioms (42.1%), followed by teratomas (29.7%), tumors of the gingiva (10.1%) and lymphatic venous malformations (9.1%), respectively. Fetal karyotyping was performed only in 9.8% of cases; within fetuses undergoing karyotype, chromosomal abnormalities accounted for 20% of cases. The most common pregnancy complication was polyhydramnios (26.3%). Ex utero intrapartum treatment (EXIT) procedure was performed in 30.1% of cases while surgical excision was used in 22.9% during postnatal life. The survival rate was 35.35%. CONCLUSION: Fetal tumors of the head and neck are rare congenital malformations. Two-dimensional ultrasound is diagnostic in almost all cases; however, MRI may be an important diagnostic adjunct in targeted cases and help patient selection for immediate intubation at the time of delivery. EXIT procedure and surgical removal of the tumor was associated with good prognosis.

Assessment of ultrasound and Doppler parameters in the third trimester of pregnancy as predictors of adverse perinatal outcome in unselected pregnancies.

OBJECTIVES: The aim of the study was to investigate ultrasound and Doppler parameters in the third trimester of pregnancy as possible predictors of adverse perinatal outcome in unselected pregnancies. MATERIAL AND METHODS: We performed a retrospective cross-sectional study including unselected pregnant women be-tween 27 and 36 + 6 weeks of gestation. The following ultrasound and Doppler parameters were assessed: estimated fetal weight (EFW) [g], EFW percentile, placental maturity grade (Grannum classification), single vertical deepest pocket (SVDP) of amniotic fluid [cm], amniotic fluid index (AFI) [cm], mean uterine artery (UtA) pulsatility index (PI), umbilical artery (UA) PI, middle cerebral artery (MCA) PI, MCA peak systolic velocity (PSV) [cm/s], and cerebroplacental ratio (CPR). Adverse perinatal outcome was defined as Apgar score of < 7 at 1 min, birth weight of < 2500 g at delivery, and gestational age of < 37 weeks at delivery. The unpaired t test was used to compare the groups. RESULTS: AFI (p = 0.01), mean UtA PI (p = 0.04) and mean UA PI (p = 0.03) were significantly different with regard to the Apgar score at 1 min. EFW, EFW percentile, SVDP of amniotic fluid, AFI, mean UtA PI, UA PI, and MCA PI were significantly different (p < 0.001) in terms of birth weight. Placental maturity grade (p = 0.02), SVDP of the amniotic fluid (p < 0.001), AFI (p < 0.001), mean UtA PI (p < 0.001), UA PI (p = 0.001), and MCA PI (p < 0.001) were significantly different as far as gestational age at delivery is concerned. CONCLUSION: Ultrasound and Doppler parameters may predict adverse perinatal outcomes in unselected pregnancies in the third trimester of pregnancy.

Mitotic activity of the brain ganglia in different subphases of the last larval instar of Melipona quadrifasciata lepeletier (Hymenoptera, apidae, meliponini) / Atividade mitótica do gânglio cerebral em diferentes fases do último instar larval de melipona quadrifasciata (hymenoptera, apidae, meliponini) / MITOTIC ACTIVITY OF THE BRAIN GANGLIA IN DIFFERENT SUBPHASES OF THE LAST LARVAL INSTAR OF MELIPONA QUADRIFASCIATA LEPELETIER (HYMENOPTERA, APIDAE, MELIPONINI)

In recent years, the number of cytogenetic studies on Melipona species has increased considerably. However, most cytogenetic techniques used for these studies require preparations with a great number of metaphase cells for reliable analysis of the karyotypes. The present study seeks to evaluate which subphase of the last larval instar of Melipona quadrifasciata Lepeletier provides the greatest number of metaphases, which is here considered a direct measure of mitotic activity. A total of 25 defecating larvae were selected based on the quantity of feces in their intestines, so as to maintain five larvae in each of the five different developmental subphases. The brain ganglia of each larva were extracted and used for cytogenetic preparation. The number of metaphase mitotic cells per preparation was counted. An analysis of variance (ANOVA) model, with Tukey's post hoc tests, was conducted. It was observed that larvae in the second subphase, defined here as the subphase in which feces were visible below the segment VII, provided the greatest number of metaphases. Therefore, it is the most appropriate developmental subphase for cytogenetic preparations of brain glanglia in M. quadrifasciata and possibly in other Melipona species.

Estudos citogenéticos envolvendo o gênero Melipona vêm aumentando nos últimos anos. Entretanto, a utilização de várias técnicas para o estudo do cariótipo exigem preparações com um grande número de células em metáfase para uma análise confiável das características citogenéticas das espécies. O presente estudo teve como principal objetivo avaliar, para Melipona quadrifasciata, o instar do desenvolvimento larval mais adequado para estudos citogenéticos, no que se refere à atividade mitótica. Foram selecionadas 25 larvas defecantes divididas em cinco subfases de acordo com a quantidade restante de fezes no intestino. Os gânglios cerebrais das larvas foram extraídos e utilizados para a obtenção dos cromossomos mitóticos metafásicos. O número de metáfases por lâmina foi contabilizado para cada indivíduo e os dados submetidos à análise de variância (ANOVA) e ao teste de TUKEY. Foi observado que larvas da segunda subfase, definidas aqui como a subfase na qual as fezes se encontram na altura do VII segmento apresentaram o maior número de metáfases. Logo, esta é a subfase mais indicada para obtenção de grande número de metáfases em células do gânglio cerebral de Melipona quadrifasciata e, possivelmente, para outras espécies do gênero Melipona.