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This study aims at genetic characterization and phylogenetic relationships of Nocardia brasiliensis focusing by using housekeeping rrs, hsp65, and sodA genes. N. brasiliensis is the species responsible for 80% of cases of actinomycetoma, one form of cutaneous nocardiosis which occurs mainly in tropical regions reaching immunocompetent patients in which the disease can lead to amputation. We analyze 36 indigenous cases of N. brasiliensis that happened in France. Phylogenetic analysis targeting rrs gene showed no robustness at phylogenetic nodes level. However, the use of a concatenation of hsp65 and sodA genes showed that the tested strains surprisingly ranked in 3 well-defined genotypes. Genotypes 2 and 3 were phylogenetically closer to each other and both diverged from genotype 1 sustained by a high bootstrap of 81%. This last genotype hosts all the cases of pulmonary forms (3), the sole cerebral form, and almost all the cases of immunocompromised patients (3 out of 4). Moreover, excepting one of them, all the strains belonging to this group present a susceptibility to imipenem which is not the case in the other genotypes that rarely count among them strains being susceptible to this drug. The haplotype diversity (Hd) of hsp65 (0.927) and sodA (0.885) genes was higher than that of rrs (0.824). For this gene, we obtained 16 polymorphic sites whereas, for hsp65 and sodA genes, up to 27 and 29 were identified, respectively. This study reveals that these two genes have an important genetic discriminatory power for the evaluation of the intraspecies genetic variability of N. brasiliensis and they may be useful for identification purposes at species level. This study also reveals the possible existence of a new species harbored by genotype 1.

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Hepatitis B core antigens (HBcAg) and hepatitis B surface antigens (HBsAg) are the main structural antigens of hepatitis B virus (HBV). Both antigens are potent immunogens for experimental animals as well as in acutely infected patients. A novel formulation based on the combination of HBsAg and HBcAg has been developed as a therapeutic vaccine candidate, aimed at inducing an immune response capable of controlling the infection. An immunization schedule was conducted to evaluate the immunogenicity of this formulation after simultaneous immunization by the intranasal and parenteral routes using different schedules and doses. Humoral and cellular immune responses generated in blood and spleen were evaluated by engyme-linked immunosorbent assay (ELISA) and enzyme-liked immunospot (ELISPOT) assays respectively. A first experiment evaluated two groups of mice simultaneously immunized by intranasal (IN) and subcutaneous (SC) routes, one including alum by SC route and, in the other, the formulation was injected without adjuvant. As a result, alum adjuvant did not increase the immunogenicity under the studied conditions. In fact, the group without alum induced the most potent immune response. The immune response was enhanced by combining IN and SC immunization compared to the SC route alone. In a second experiment, mice were immunized by different mucosal routes at the same time, and compared to the simultaneously (IN/SC) immunized groups. It was demonstrated that there is no improvement on the resulting immune response by using multiple routes of immunizations simultaneously; however, the increase of the antigen dose induced a superior immune response. Interestingly, the increase of antigen dose only by SC route did not favor the resulting immunogenicity. In conclusion, the use of HBsAg transgenic mice has proven useful to optimize the formulation, avoiding the unnecessary use of alum as adjuvant as well as provided information of the role of different mucosal immunization routes and antigen dose on the resulting immune response. How to cite this article: Trujillo H, Blanco A, García D, Freyre F, Aguiar J, Lobaina Y, Aguilar JC. Optimization of a Therapeutic Vaccine Candidate by Studying Routes, Immunization Schedules and Antigen Doses in HBsAg-positive Transgenic Mice. Euroasian J Hepato-Gastroenterol 2014;4(2):70-78.

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Paciente de 14 años, proveniente de Apartadó (Antioquia), remitido al Hospital Pablo Tobón Uribe de Medellín por un cuadro febril agudo asociado a deterioro rápido del estado de conciencia, decantándose posteriormente como causa de su deterioro una ameba de vida libre (Naegleria fowleri) . Las amebas de vida libre son una causa etiológica poco común de meningitis, tienen un curso fulminante y requieren de un alto índice de sospecha para hacer un tratamiento oportuno.

A 14-year-old patient from Apartado (Antioquia, Colombia) was referred to Hospital Pablo Tobon Uribe in Medellin for acute febrile illness associated with headache and rapid loss of consciousness. The etiologic agent causing this deterioration was found to be a free-living amoeba ( Naegleria fowleri ). Free-living amoebae are a rare etiology for meningoencephalitis, have a fulminant course and require a high index of suspicion for early diagnosis.

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Estudou-se a ossificação endocontral de 18 embriões e 12 fetos de até três meses de gestação, os quais foram coletados de úteros gestantes em frigoríficos e abatedouros. Os úteros foram dissecados e, em seguida, realizou-se uma incisão dorsal até o cérvix para avaliações macroscópicas dos embriões e fetos. Para o estudo microscópico foram realizadas técnicas de inclusão, seguidas de marcação dos depósitos de cálcio e fósforo, responsável pela ossificação dos moldes de cartilagem. Foram identificados hipertrofia da cartilagem e morte dos condrócitos e aumento da área de depósito de cálcio e fósforo, por volta da 10ª semana gestacional (74 dias). Durante a 11ª semana de gestação (81 dias), os grupamentos de carbonato de cálcio e fósforo espalharam-se por todo o osso, sendo mais intenso na diáfise.

The endocondral ossification process was analyzed in 18 embryos and 12 fetus until three months of pregnancy, which had been collected from pregnant uterus at slaughterhouse. Later, the uterus was separated followed by a dorsal incision from the cervix for better gross evaluations of the embryos and fetus. For the microscopical study, histological techniques had been performed followed by staining of the deposits of calcium and phosphorus match, responsible for the ossification of the cartilage casts, used as Von Kossa stain. Cartilage hypertrophy and death of chondrocytes, enlargement of the area of calcium and phosphorus deposit were foundon the 10th week of pregnancy (74 d). During the 11th week of pregnancy (81 d), the calcium carbonate and phosphorus had spread to all bones, being more intense in diaphysis.

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Estudou-se a ossificação endocontral de 18 embriões e 12 fetos de até três meses de gestação, os quais foram coletados de úteros gestantes em frigoríficos e abatedouros. Os úteros foram dissecados e, em seguida, realizou-se uma incisão dorsal até o cérvix para avaliações macroscópicas dos embriões e fetos. Para o estudo microscópico foram realizadas técnicas de inclusão, seguidas de marcação dos depósitos de cálcio e fósforo, responsável pela ossificação dos moldes de cartilagem. Foram identificados hipertrofia da cartilagem e morte dos condrócitos e aumento da área de depósito de cálcio e fósforo, por volta da 10ª semana gestacional (74 dias). Durante a 11ª semana de gestação (81 dias), os grupamentos de carbonato de cálcio e fósforo espalharam-se por todo o osso, sendo mais intenso na diáfise.(AU)

The endocondral ossification process was analyzed in 18 embryos and 12 fetus until three months of pregnancy, which had been collected from pregnant uterus at slaughterhouse. Later, the uterus was separated followed by a dorsal incision from the cervix for better gross evaluations of the embryos and fetus. For the microscopical study, histological techniques had been performed followed by staining of the deposits of calcium and phosphorus match, responsible for the ossification of the cartilage casts, used as Von Kossa stain. Cartilage hypertrophy and death of chondrocytes, enlargement of the area of calcium and phosphorus deposit were foundon the 10th week of pregnancy (74 d). During the 11th week of pregnancy (81 d), the calcium carbonate and phosphorus had spread to all bones, being more intense in diaphysis.(AU)

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Congenital malformations correspond to one of the main causes of embryonic loss during the gestational process. They result from interaction of several factors such as multifactor heredity, chromosomal and genetic alterations and environmental agents; however, unknown aetiology also can be present. In this article, we have used 10 embryos, from a frigorific area of Dracena, SP, Brazil, which were fixed in Bouin solution for a macro- and micro-scopic description. We could verify the presence of an encephalic tissue mass on the embryo's dorsal cranial area, resulting from the non-formation of part of the cranial cap and from the non-closing of cephalic neuropore and consequent neuroepithelial cells disorganization. In the abdominal area, the embryos did not show the complete fusion of the body lateral pleats during the abdominal wall formation, and the liver extruded into the amniotic cavity without involvement of the intestine.

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This randomized, observer-blind, multicenter, parallel-group study compared the clinical and bacteriologic efficacy and safety of amoxycillin, 45 mg/kg/day b.d. and amoxycillin, 40 mg/kg/day t.d.s. after 7 days of treatment in 517 children with acute bacterial tonsillopharyngitis. At the end of treatment, a successful clinical response was recorded in more than 96% of patients in each of the treatment groups. A similar result was obtained at follow-up. Among those patients who were bacteriologically evaluable at the end of treatment, a successful bacteriologic response was achieved in more than 94% in each treatment group. Both treatments were well tolerated. Drug-related adverse events were recorded in just 12 patients (4.6%) in the b.d. group and six (2.4%) in the t.d.s. group. The study demonstrated that a twice-daily regimen of amoxycillin, 45 mg/kg/day, was as effective and as well tolerated as the standard three-times-daily regimen of amoxycillin, 40 mg/kg/day, in the treatment of acute bacterial tonsillopharyngitis in children.

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OBJECTIVE: To identify the frequency of drug resistant tuberculosis and to determine risk factors for its development in five health districts in the State of Chiapas, Mexico. DESIGN: A cross-sectional study. EMPLACEMENT: Primary care. PARTICIPANTS: Patients with pulmonary tuberculosis diagnosed between September 1 and December 31, 1995. Each patient responded to a questionnaire regarding epidemiological, diagnostic and treatment variables. Sputa were cultured, drug sensitivities and typing of micobacteria were obtained. MEASUREMENTS AND MAIN RESULTS: There were 142 cases of pulmonary tuberculosis confirmed by acid fast smear. The frequency of resistant M. tuberculosis in 61 positive cultures was 24.2%. Of these, 33.3% was resistant to one drug and 66.7% at two or more drugs. Resistance to isoniazide and rifampicin together was present in 66.7% of the cases. The most frequent resistance, 86.7%, was to isoniazide. A history of multitreatments, either completed or not, resulted in a risk factor 6.48 times greater for drug resistance and patients with a history of abandoning treatment presented an increased risk factor of 5.71. CONCLUSIONS: Drug resistant tuberculosis y a serious problem in Chiapas that requires urgent attention if future control of the disease is to be maintained. The frequency of resistant tuberculosis is high, 24.2%, and the risk factors identified indicate a worsening of the situation with the accumulation of patients inadequeately treated without follow up until cure.

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Neste estudo observador-cego, cento e setenta e um pacientes com evidência clínica de faringotonsilite foram randomizados para serem submetidos ao tratamento com amoxicilina administrada duas vezes ao dia(bd)ou três vezes ao dia (tid). Este é um relatório descritivo preliminar, até o momento, a característica "observador-cego" do estudo. Assim, os grupos de pacientes estäo sendo caracterizados como "Tratamento A" e "Tratamento B". Ao final do tratamento, foi obtido um índice de cura clínica de noventa e seis e cinco por cento para os pacientes do grupo "Tratamento A" e noventa e seis e cinco por cento para pacientes do grupo "Tratamento B", sugerindo equivalência terapêutica entre os grupos quanto à eficácia clínica. Os dois esquemas posológicos foram bem tolerados. Em conclusäo, os dados apresentados sugerem que o regime terapêutico de amoxilina administrada duas vezes ao dia (bd) seja täo eficaz quanto o regime de amoxicilina administrada três vezes ao dia (tid) para o tratamento destes pacientes, com a vantagem de poder determinar maior aderência dos doentes ao tratamento

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SETTING: Radiometric technology and molecular biology are used in rapid diagnosis of tuberculosis in laboratories around the world. However, these technologies increase costs and are not available in laboratories where economic resources are limited. OBJECTIVE: To compare sensitivity and time for detection of positive cultures in a microcolony method, Middlebrook 7H11 thin layer agar plate (TL7H11), and a conventional culture, Lowenstein-Jensen (L-J). DESIGN: A total of 761 clinical samples were processed using acid-fast smear and culture on TL7H11 plates and L-J tubes. TL7H11 plates were checked microscopically for microcolony growth twice weekly for 4 weeks, and L-J tubes were checked once a week for 8 weeks. RESULTS: Overall positivity was 11.0%. More than 60% of the positive samples were detected within the first 10 days on TL7H11, and none on L-J. After 2 weeks, more than 80% were positive on TL7H11 compared to 10% on L-J. In paucibacillary samples, TL7H11 detected 2.18% and L-J 4.57% (P < 0.001). Microcolony morphology was 100% distinctive for Mycobacterium tuberculosis on TL7H11. The calculated cost of TL7H11 prepared in the laboratory was US$2.90 per unit. CONCLUSION: The TL7H11 method is an inexpensive, rapid and reliable alternative for diagnosing M. tuberculosis infection. It is therefore a valuable option for laboratories in low income countries.

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Streptococcus pneumoniae is the leading bacterial cause of childhood pneumonia in the developing world. This study describes the type distribution and antimicrobial susceptibility of invasive pneumococcal isolates from Colombian children and is part of the Sistema Regional de Vacunas (SIREVA), a PAHO regional initiative designed to determine the ideal serotype composition of a protein polysaccharide pneumococcal conjugate vaccine for use in children less than 5 years old in Latin America. In Colombia, during the study period, centres in Bogota, Medellin, and Cali collected 324 S. pneumoniae isolates from invasive diseases, 238 (73.5%) from children under the age of 2. Pneumonia was the clinical diagnosis in 41.3% cases, meningitis in 41%, and sepsis in 11.2%. The seven most frequent types included 14(21.9%), 5(10.5%), 23F(9.6%), 1(9%), 6B(9%), 19F(7.1%), and 6A(6.2%). The frequency of diminished susceptibility to penicillin (DSP) was 12%, with 8.9% of isolates showing intermediate level resistance and 3.1% showing high level resistance. Among DSP isolates, 23% were also resistant to cefotaxime, 33.3% to erythromycin, 48.7% to chloramphenicol, and 74.3% to trimethoprim/sulfamethoxazole. Multiple resistance was detected in 59% of the isolates that have DSP. Penicillin resistance was associated with types 23F (53.8%) and 14 (25.6%). These data provides information on capsular types prevalent in Colombia that will not only allow the formulation of an ideal vaccine for the region but also reinforce the need for ongoing regional surveillance.

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PIP: In Colombia, health workers obtained a nasopharyngeal wash from 103 infants aged less than 12 months hospitalized for acute lower respiratory infection (ALRI) at the General Hospital of Medellin during April 1994 to April 1995 so researchers could determine the frequency of ALRI caused by respiratory syncytial virus (RSV) in hospitalized children. Immunofluorescence detected RSV infection in 43 (41.7%) patients. The presence of the following signs and symptoms allowed a clinical diagnosis of a viral infection: rhinorrhea, prolonged expiration, expiratory wheezing, interstitial infiltrates, and hyperinflation on chest radiographs as well as negative tests for 3 or 4 acute phase reactants. The physicians initiated antibiotic therapy (for 1-3 days) in 12 cases (27.9%) based on acute phase reactant findings who actually had an RSV infection. When the physicians learned that the laboratory confirmed RSV infection, they stopped antibiotic therapy. Antibiotics were continued in 16 (37.2%) other RSV infected infants, all of whom were less than 2 months old, due to mixed pneumonia (viral and bacterial). 39.4% of RSV-infected children whose clinical findings strongly suggested RSV received no antibiotics. None of these children or other ALRI patients with a viral disease suffered complications. They required less hospitalization time--since no further diagnostic tests were needed--than ALRI patients with a bacterial infection. Admissions for both ALRI and RSV infection peaked during November to January. RSV incidence peaked in January (23.3%). The leading reasons for hospitalization were pneumonia and bronchiolitis. These findings show that RSV diagnosis is useful and it lessens the indiscriminate use of antibiotics.^ieng

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OBJECTIVES: To describe the clinical characteristics and laboratory diagnosis of seven children with disseminated histoplasmosis and evaluate the effectiveness of itraconazole therapy in this severe form of the mycosis as well as to determine the long term results of such treatment. METHODS: The diagnosis of histoplasmosis was based on the direct observation of Histoplasma capsulatum var. capsulatum and/or on the isolation of the fungus from pathologic materials; the results of the serologic tests were taken into consideration. Chest roentgenograms also contributed to the diagnosis. PATIENTS: The patients were seven rural children, five girls and two boys, ages 1 to 14 years (mean, 4.6), with a confirmed diagnosis of disseminated histoplasmosis and who had no underlying disease other than malnourishment. RESULTS: The seven children experienced a subacute febrile syndrome for 4 months accompanied by anorexia, weight loss and signs of reticuloendothelial involvement such as lymph node hypertrophy, hepatomegaly and/or splenomegaly. The lung revealed roentgenographic alterations consisting mainly of nodular infiltrates. All patients received itraconazole orally in a mean dosage of 7.2 mg/kg/day, for variable periods (3 to 12 months), depending on the individual response and the toxic effects of the medication. One of the patients who was improving after 1 month of treatment was taken from the hospital by his guardian against medical advice and died shortly afterward. The remaining six patients responded to the treatment with marked clinical improvement and showed negative cultures and decreases in anti-H. capsulatum antibody titers after 3 months of treatment. Only one patient, the youngest and most severely affected child, exhibited hepatotoxicity, which subsided when itraconazole was discontinued. Extended follow-up studies revealed no relapses. CONCLUSION: The results of this study indicate that itraconazole is effective for treatment of disseminated childhood histoplasmosis. More studies should be performed to determine the most appropriate dosage and the optimal duration of itraconazole treatment in children.

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We are reporting the most frequent urologic injuries of the gynecologic and urogynecologic surgeries at the National Institute of Perinatology in Mexico City. From the period of March 1993 to February 1995. There were performed 3,452 surgeries. Of this, 2,971 were gynecologic and 481 urogynecologic cases. We found 20 patients with injury to the inferior urinary tract. The most frequent type of injury found was by puncture and blunt, in eight cases respectively and cutting in four. The transoperatory diagnostic of injury was performed in 17 patients and in three few days after. The late complications of the injury was urethral vaginal fistula in two, one vesicovaginal communication and one ureterovaginal, one stone formation and one transurethral catheter retention. The more affected organ was the bladder in 18 cases, urether and urethra. The injury to the inferior urinary tract represents 4.15% of the urogynecologic surgical complications, and 0.67% of the gynecological ones. The main aspect in the prognosis of the injury to the inferior urinary tract, is the early diagnostic during the surgical procedures.

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Two hundred and twenty-nine patients operated for genuine stress urinary incontinence by two surgical techniques, were studied. Risk factors were analyzed: age, gesta, para, and added gynecological pathology. They were placed in two groups. Group I formed by operated patients with Pereyra's technique (136), and Group II operated by Burch's technique (93). There were no differences as far as general characteristics of patients in both groups. More frequent added pathologies for Group I were pelvic statics changes, and for Group II, uterine myomatosis. Technically there were no advantages for one technique as compared with the other one, independently of indications for each one.

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We are reporting the institutional experience of March 1992 to May 1993. Eleven patients were studied with a diagnostic of urogenital fistula, one of them was vesico-uterine, two uretero-vaginal and eight vesico-vaginal. Only to eight of the patients surgery correction was realized, one of them was aboarded by vaginal tract and seven by abdominal tract. Of the patients with abdominal aboarded, two cases were ureteroneocystostomies. The failure of the surgery was presented in two cases, both with abdominal aboarded. These data clears up that the urogenital fistula is a frequent complication in patients that are submitted to surgical gynecological procedures. The surgical success probably depends more of the surgical technique rather than aboarded tract.

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Patient's histories of urinary incontinence and results of several standard clinical tests, were correlated with final diagnosis obtained by multichannel urodynamic testing. One hundred and thirty one histories of women with urinary incontinence who were in the Urogynecology Clinics of the Instituto Nacional de Perinatología in the period from july to december 1992, were evaluated including dates of standardized questionnaire and structural clinical examination with the urodynamic testing to obtain a final diagnosis. Although the objective urinary loss and the cystocele II-III were significantly associated with genuine stress incontinence and mixed incontinence, in patients with detrusor instability did not. Women with complaints of urinary incontinence, especially those for whom surgery is contemplated, should undergo complete urodynamic evaluation.

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The uterine leiomyomas (UL) are tumors compound by smooth muscle connective tissue. Growth depends mainly of two basic mechanisms: hypertrophy of myometrial cells and connective tissue deposit. The quantification of the basic elements of the tumor permits to try its application as biochemical markers of disease, auxiliary diagnostic criteria, or in medical therapeutical monitorization, alternative that lately has become very important. In the present study 33 patients are included, in two groups. Group I women with uterine leiomyomatosis (n = 17) and Group II (n = 16) without UL. A prospective, double blind, transversal of cases and control study. Connective tissue concentration was evaluated based on collagen determination. The evaluation of muscular tissue was done by desoxyribonucleic acid (DNA) measurement. The results showed a significant increase in connective tissue concentration (until 500%) and a significant diminution of cellularity (DNA) in women with UL, as compared with control group. The main biochemical and clinical implications of these findings, are commented upon.

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