Modified SureSelectQXT Target Enrichment Protocol for Illumina Multiplexed Sequencing of FFPE Samples.

BACKGROUND: Personalised medicine is nowadays a major objective in oncology. Molecular characterization of tumours through NGS offers the possibility to find possible therapeutic targets in a time- and cost-effective way. However, the low quality and complexity of FFPE DNA samples bring a series of disadvantages for massive parallel sequencing techniques compared to high-quality DNA samples (from blood cells, cell cultures, etc.). RESULTS: We performed several experiments to understand the behaviour of FFPE DNA samples during the construction of SureSelectQXT libraries. First, we designed a quality checkpoint for FFPE DNA samples based on the quantification of their amplification capability (qcPCR). We observed that FFPE DNA samples can be classified according to DIN value and qcPCR concentration into unusable, or low-quality (LQ) and good-quality (GQ) DNA. For GQ samples, we increased the amount of input DNA to 150 ng and the digestion time to 30 min, whereas for LQ samples, we used 50 ng of DNA as input but we decreased the digestion time to 1 min. In all cases, we increased the cycles of the pre-hyb PCR to 10 but decreased the cycles of the post-hyb PCR to 8. In addition, we confirmed that using half of the volume of reagents can be beneficial. Finally, in order to obtain better results, we designed a decision flow-chart to achieve a seeding concentration of 12-14 pM for MiSeq Reagent Kit v2. CONCLUSIONS: Our experiments allowed us to unveil the behaviour of low-quality FFPE DNA samples during the construction of SureSelectQXT libraries. Sequencing results showed that, using our modified SureSelectQXT protocol, the final percentage of usable reads for low-quality samples was increased more than three times allowing to reach median depth/million reads values of 76.35. This value is equivalent to ~ 0.9 and ~ 0.7 of the values obtained for good-quality FFPE and high-quality DNA respectively.

Economic analysis of the robotic approach to inguinal hernia versus laparoscopic: is it sustainable for the healthcare system?

INTRODUCTION: There has been a rapid proliferation of the robotic approach to inguinal hernia, mainly in the United States, as it has shown similar outcomes to the laparoscopic approach but with a significant increase in associated costs. Our objective is to conduct a cost analysis in our setting (Spanish National Health System). MATERIALS AND METHODS: A retrospective single-center comparative study on inguinal hernia repair using a robotic approach versus laparoscopic approach. RESULTS: A total of 98 patients who underwent either robotic or laparoscopic TAPP inguinal hernia repair between October 2021 and July 2023 were analyzed. Out of these 98 patients, 20 (20.4%) were treated with the robotic approach, while 78 (79.6%) underwent the laparoscopic approach. When comparing both approaches, no significant differences were found in terms of complications, recurrences, or readmissions. However, the robotic group exhibited a longer surgical time (86 ± 33.07 min vs. 40 ± 14.46 min, p < 0.001), an extended hospital stays (1.6 ± 0.503 days vs. 1.13 ± 0.727 days, p < 0.007), as well as higher procedural costs (2318.63 ± 205.15 € vs. 356.81 ± 110.14 €, p < 0.001) and total hospitalization costs (3272.48 ± 408.49 € vs. 1048.61 ± 460.06 €, p < 0.001). These results were consistent when performing subgroup analysis for unilateral and bilateral hernias. CONCLUSIONS: The benefits observed in terms of recurrence rates and post-surgical complications do not justify the additional costs incurred by the robotic approach to inguinal hernia within the national public healthcare system. Nevertheless, it represents a simpler way to initiate the robotic learning curve, justifying its use in a training context.

Allergenic profile of nasal polyposis.

BACKGROUND: Nasal polyposis is highly prevalent in the general population. Its exact origin is unknown, although several factors are involved in the etiology and development of this condition. Clinical patterns, a history of atopy, environmental exposure, eosinophil-mediated inflammation, the presence of inflammatory mediators, and sensitization to some allergens indicate that nasal polyposis is associated with allergic phenomena. The aim of this study was to identify the association between nasal polyposis and allergic factors by examining hypersensitivity reactions to common allergens and environmental exposure that could lead to the development of atopy. METHODS: We conducted a comparative study of 190 patients with nasal polyposis and 190 healthy individuals. The study included clinical and epidemiological variables, environmental exposure factors, and an allergology workup using skin prick tests with 18 inhaled allergens. RESULTS: A total of 121 patients (63.7%) of the 190 were male; 62.1% had a family history of allergy. The incidence of asthma was 48.9% among the patients and only 2.3% among the controls (P < .001). The factor most frequently involved in the patients' symptoms was weather changes (67.4%). Skin prick tests were positive in 63.2% of the patients and 31.1% of the controls. The allergens that most frequently elicited a reaction from the patients in the prick tests were Dermatophagoides pteronyssinus (27.7%), Dermatophagoides farinae (21.3%), and Olea europaea (21.1%). The difference between these results and those of the controls was statistically significant. CONCLUSIONS: Patients with nasal polyposis are sensitive to the most common allergens in our environment and exhibit a clear-cut correlation with other allergic factors, as confirmed by personal and family histories, the presence of chronic rhinitis, and the results of in vivo tests.

Fungal sensitization in nasal polyposis.

BACKGROUND: There are indications that polyposis is somehow related to allergic phenomena. Fungal sensitization in substantial proportions of patients has been cited as a trigger of inflammatory mechanisms involving either an immunoglobulin (Ig) E-mediated reaction to fungal colonization or fungal invasion of tissues. OBJECTIVE: To confirm whether fungi were involved in the development of polyposis by examining sensitivity to fungal allergens and potential local contamination by fungal species. METHODS: We performed a study of 190 patients with polyposis and 190 controls in which we compared the results of skin prick tests to 12 fungi, total IgE, and specific IgE to 15 fungal extracts and nasal fungal cultures. RESULTS: The specific fungi eliciting a reaction from the largest proportion of patients in the skin prick tests were Fusarium solani (13.7%), Penicillium frequentans (12.6%), Trichophyton mentagrophytes (11.1%), and Candida albicans (8.4%) (P < .001). The proportion of individuals that tested positive for fungal-specific IgE was 22.4% (38/170) for patients and 10.1% (19/189) for controls (P = .04). The respective proportions of positive responses to fungal cultures were 58.7% and 60%. Furthermore, no significant differences between patients and controls were found for the results of in vitro tests with cultured fungal allergens. CONCLUSIONS: Although the patients with polyposis exhibited sensitization to fungal allergens, we found that nasal colonization by fungi was similar in patients and the general population. We were also unable to find a correlation between a positive response to the cultures and the presence of fungal allergen-specific IgE. It therefore seems that nasal colonization by fungi does not induce fungal sensitization.

Cerebrospinal fluid leucocyte counts in healthy neonates.

OBJECTIVES: To determine the cerebrospinal fluid (CSF) white blood cell (WBC) count of normal term neonates, and compare the CSF WBC profile of normal and symptomatic infants without infection of the central nervous system (CNS). METHOD: Neonates were included if (a) they were at risk of congenital Toxoplasma infection and had undergone a lumbar puncture to assess CNS involvement, and (b) serial specific serum IgG and IgM determinations had ruled out congenital infection. According to neonatal chart reviews, 30 consecutive patients without CNS infection were classified as normal (absolutely asymptomatic) or symptomatic (any kind of symptoms). RESULTS: CSF WBC count was higher in 11 symptomatic (7/mm(3), 0-30/mm(3)) than in 19 normal (1/mm(3), 0-5/mm(3)) neonates (p<0.01). CONCLUSION: Normal neonatal CSF contains up to 5 WBCs/mm(3). Mild pleocytosis can be found in symptomatic infants without CNS infection.

[Acute palsy of twelfth cranial nerve]. / Parálisis aguda del XII par craneal.

The hypoglossal nerve or Twelfth-nerve palsy is a rare damage with different causes: tumors or metastases in skull base, cervicals tumors, schwannoma, dissection or aneurysm carotid arteries, stroke, trauma, idiopathic cause, radiation, infections (mononucleosis) or multiple cranial neuropathy. Tumors were responsible for nearly half of the cases in different studies. We studied a female with hypoglossal nerve acute palsy. We made a differential diagnostic with others causes and a review of the literature.

Helicobacter pylori second-line rescue therapy with levofloxacin- and bismuth-containing quadruple therapy, after failure of standard triple or non-bismuth quadruple treatments.

BACKGROUND: The most commonly used second-line Helicobacter pylori eradication regimens are bismuth-containing quadruple therapy and levofloxacin-containing triple therapy, both offering suboptimal results. Combining bismuth and levofloxacin may enhance the efficacy of rescue eradication regimens. AIMS: To evaluate the efficacy and tolerability of a second-line quadruple regimen containing levofloxacin and bismuth in patients whose previous H. pylori eradication treatment failed. METHODS: This was a prospective multicenter study including patients in whom a standard triple therapy (PPI-clarithromycin-amoxicillin) or a non-bismuth quadruple therapy (PPI-clarithromycin-amoxicillin-metronidazole, either sequential or concomitant) had failed. Esomeprazole (40 mg b.d.), amoxicillin (1 g b.d.), levofloxacin (500 mg o.d.) and bismuth (240 mg b.d.) was prescribed for 14 days. Eradication was confirmed by (13) C-urea breath test. Compliance was determined through questioning and recovery of empty medication envelopes. Incidence of adverse effects was evaluated by questionnaires. RESULTS: 200 patients were included consecutively (mean age 47 years, 67% women, 13% ulcer). Previous failed therapy included: standard clarithromycin triple therapy (131 patients), sequential (32) and concomitant (37). A total of 96% took all medications correctly. Per-protocol and intention-to-treat eradication rates were 91.1% (95%CI = 87-95%) and 90% (95%CI = 86-94%). Cure rates were similar regardless of previous (failed) treatment or country of origin. Adverse effects were reported in 46% of patients, most commonly nausea (17%) and diarrhoea (16%); 3% were intense but none was serious. CONCLUSIONS: Fourteen-day bismuth- and levofloxacin-containing quadruple therapy is an effective (≥90% cure rate), simple and safe second-line strategy in patients whose previous standard triple or non-bismuth quadruple (sequential or concomitant) therapies have failed.

Risk factors for HIV-2 infection in Guinea-Bissau.

To define the epidemiology of HIV-2 infection, we conducted a case-control study among hospitalized patients at an acute care hospital in Bissau, Guinea-Bissau, a country with endemic HIV-2 infection. Among 128 patients with various diagnoses, 23 (18%) were positive for HIV-2 by ELISA and Western blot. One of these patients was serologically reactive for HIV-1 also, but PCR and viral culture revealed the presence of HIV-2 only. To study risk factors, behaviors, and AIDS knowledge related to the acquisition of HIV infection, 22 HIV-2-seropositive and 21 seronegative hospitalized patients were given a physical examination and administered a questionnaire. Among women, transfusion was associated with HIV-2 infection (OR = 14.4, p = 0.02); among men, sex with a prostitute was the principal risk factor (OR = undefined, p = 0.02). Although 79% of HIV-infected patients and controls had heard of AIDS, only 17% of all study participants and 50% of males reporting sex with prostitutes had used condoms in the previous year. These data suggest that the risk factors for HIV-2 infection are similar to those for HIV-1 and support previous studies showing that HIV-2 is the predominant HIV in Guinea-Bissau. Efforts to decrease transmission of HIV-2 should include screening for HIV-2 in blood for transfusion in endemic areas (now done in Bissau) and education about the risk of sexual transmission.

Mononucleosis caused by Epstein-Barr virus and cytomegalovirus in children: a comparative study of 124 cases.

We present 124 children who had mononucleosis. The patients were selected according to strict clinical features. Twenty (16.1%) of the 124 children were proved to have cytomegalovirus mononucleosis and 104 (83.8%) children had Epstein-Barr virus mononucleosis. The symptoms were similar in both groups. Significant differences were found only for the presence of cervical lymphadenopathy, which was more frequent in the Epstein-Barr group (83.2%) compared with the cytomegalovirus group (75%). Fever was the most frequent symptom in both groups. Cytomegalovirus mononucleosis was significantly more frequent in children younger than 4 years.

Influence of recent antibiotic therapy on antimicrobial resistance of Streptococcus pneumoniae in children with acute otitis media in Spain.

BACKGROUND: Despite the high prevalence of penicillin resistance among Streptococcus pneumoniae strains in Spain (40 to 60% with MIC > or = 0.1 microg/ml), the data on acute otitis media (AOM) isolates are scarce. We conducted a prospective, longitudinal study to determine the rates of antimicrobial resistance of S. pneumoniae isolates from children with AOM in our country and to analyze the effect of previous antibiotic therapy on these rates. METHODS: Tympanocentesis was performed on 169 children diagnosed with AOM (age range, 1 month to 14 years). Two groups were considered: Group A, 113 patients with non-antibiotic-treated AOM, subdivided into Group A1 (collected from 1989 to 1992) and Group A2 (1992 to 1996); Group B, 56 patients from the period 1992 to 1996, with AOM clinical failure, defined as worsening or persistent symptoms after at least 2 days of appropriate antibiotic therapy. Amoxicillin-clavulanate was the most frequent antibiotic used (68%), followed by azithromycin (21%), cefaclor and cefixime (11%). RESULTS: A total of 63 S. pneumoniae isolates were recovered, 42 in Group A and 21 in Group B. Resistance to penicillin (MIC > or = 0.1 microg/ml) was found in 38% of strains in Group A (32% in A1 and 50% in A2), but in Group B the rate of resistance reached 90% (P = 0.0002). Erythromycin resistance was also increased from 35% (Group A2) to 62% (Group B), and trimethoprim-sulfamethoxazole resistance rose from 64% to 81%. CONCLUSIONS: Resistance to penicillin among S. pneumoniae AOM isolates is frequent and is increasing in Spain. After failure of standard antibiotic therapy, the rates of penicillin resistance reached 90% of the isolates.

Tolerance of pyrazinamide in short course chemotherapy for pulmonary tuberculosis in children.

BACKGROUND: This prospective study was performed to evaluate the tolerance of pyrazinamide in short course chemotherapy in children. METHODS: A total of 114 children ages 6 months to 15 years (4.5 +/- 3.4 years) with diagnosed pulmonary tuberculosis from 1985 to 1995 entered the trial. A 2-month regimen of isoniazid, rifampin and pyrazinamide, followed by rifampin and isoniazid for the remaining 4 months, was administered orally to all children. Clinical adverse effects specifically investigated were gastrointestinal disturbances, rash, signs of hepatotoxicity and arthralgias. Laboratory toxicity data (number of leukocytes, erythrocyte sedimentation rate, aspartate aminotransferase, alanine aminotransferase and serum uric acid) were collected before treatment and 1, 3 and 5 months after the beginning of chemotherapy. RESULTS: Clinical adverse effects were mild in all cases. Three children (2.6%) had fever and 5 (4.4%) had gastrointestinal disturbances. Aspartate aminotransferase and alanine aminotransferase mean values showed no differences along time and no patients had clinical signs of hepatotoxicity. Only 11 children (19.6%) showed a slight increase in alanine aminotransferase (< 194 units/l). Serum uric acid increased in 92.2% of the children compared with pretreatment values. This increase remained within the normal range in all but 9.8% of patients. There was a significant increase in uric acid mean concentrations after 1 month of therapy (from 3.7 +/- 0.7 mg/dl to 5.7 +/- 1.6 mg/dl, P < 0.05), which fell again (4.0 +/- 1.1) 1 month after pyrazinamide was stopped. There were no signs of gout or arthralgias. In no case was the treatment interrupted. CONCLUSION: The addition of pyrazinamide in chemotherapy for pulmonary tuberculosis in children was found to be safe. The slight increase in uric acid concentration during its administration had no recognized adverse consequences.

Secretion of the Escherichia coli K-12 SheA hemolysin is independent of its cytolytic activity.

The Escherichia coli K-12 sheA gene encodes a pore-forming hemolysin that is secreted to the medium by a hitherto unidentified mechanism. To study SheA secretion, we constructed fusions between SheA and the mature form of the periplasmic enzyme beta-lactamase, and performed site-directed mutagenesis on these constructs. The SheA-Bla and Bla-SheA hybrid proteins displayed hemolytic activity and were efficiently exported to the extracellular medium. Our results with mutant hybrid proteins show that secretion of SheA is independent of its cytolytic activity, that secretion is paralleled by a transient leakage of periplasmic contents to the extracellular medium, and that deletion of the 11 C-terminal residues of SheA has no effect on its secretion and cytolytic activity.

[Fiberoptic bronchoscopy in childhood endobronchial tuberculosis]. / Fibrobroncoscopia en la tuberculosis endobronquial.

BACKGROUND: Endobronchial tuberculosis is a classical manifestation of primary tuberculosis in childhood. Fiberoptic bronchoscopy (FB) is an ancillary diagnostic tool, but its utility and indications are not well established. OBJECTIVES: To analyze the FB performed over 11 years (1992-2003) in children with a diagnosis of tuberculosis and to review the literature. A further aim was to establish the current usefulness of FB in children with tuberculosis and propose criteria to determine the indications for FB in this population. METHODS: We report a retrospective series (n 5 16). FB was indicated in children who showed any of the following findings in chest roentgenogram: a) persistent parenchymal consolidation; b) lymphadenopathy and consolidation; c) hyperinsufflation (emphysema); d) atelectasias, and e) airway narrowing caused by lymphadenopathies. Published series on the topic were reviewed and six studies were suitable for comparison with our own. RESULTS: Endobronchial tuberculosis was found in seven children (43 %) and extrinsic compression was found in three (18 %). The microbiological results obtained from FB samples were not superior to those obtained from classical diagnostic methods. The sensitivity of the proposed criteria for suspicion of endobronchial tuberculosis was 71 %. Endoscopic findings justified a change in therapy in 50 % of the children (addition of corticoids or surgery) and this percentage was similar to that reported in other series. CONCLUSIONS: FB does not usually improve microbiological diagnosis of tuberculosis but can be useful when choosing the most appropriate therapy in children with suspected endobronchial tuberculosis. In some cases, computed tomography may make FB unnecessary, but in others this procedure can be therapeutic (obstruction due to caseum, atelectasias). Establishing the indications for FB in childhood tuberculosis is difficult, but the proposed criteria may be an acceptable guide to identifying which patients could benefit most from this procedure. Not all children with endobronchial tuberculosis require corticoids.

[Endoscopic nasal surgery in sellar tumors]. / Cirugía endoscópica nasal de procesos selares.

The advances in endoscopy have improved the approach to the sellar region throught nasal cavity. We studied 20 patients with pituitary adenomas operated on using an endoscope through nasal cavity with transseptal- trassphenoidal approach during the last two years. Average age was 45.6 years, and 75% were females. 30% of the patients had non functioning adenomas, 30% acromegaly, 25% Cushing's disease and 10% prolactinomas. No complications occurred during surgery. The more common complications were diabetes insipidus (two cases) and CSF leak (one case). Two patients needed hormonal treatment because panhypopituitarism. No patients developed septal perforation, nasal deformity, epistaxis, meningitis, lip numbness or oronasal fistula.

[Metastatic neck adenopathies without known primary tumor. Incidence study 1996-2001]. / Adenopatías metastásicas cervicales sin tumor previo conocido. Estudio de incidencia 1996-2001.

We made a retrospective descriptive study reviewing 92 patients with cervical lymph node metastases from unknown primary tumour. The study was accomplished during 6 years. (January 1996 to December 2001) in the Department of Otorhinolaryngology of the Hospital Reina Sofia, Córdoba. The median age was 61.7 years old, male 96.7% and heavy smoker (89.1%) and drinkers most of them. According to TNM categories, 72.2% patients were in phase N2a and half of them were localized Level II. Squamous cell carcinoma was the predominant histologic type. The patients were treated with surgery in 70.6% of the cases with or without radiation therapy and/or chemotherapy.

[Forestier-Rotes Querol syndrome]. / Síndrome de Forestier-Rotes Querol.

The diffuse idiopathic skeletal hyperostosis (DISH) or Forestier's disease is characterized by an hyperostosis of the anterolateral margin of the vertebral bodies. It is an uncommon cause of dysphagia in old patients. In the differential diagnosis of mechanical dysphagia we must have present this disease. Cervico-dorsal spine radiographies reveal large anterior osteophytes creating pharyngeal encroachment.

[Rhinocerebral mucormycosis: differential diagnosis of invasive fungal sinusitis]. / Mucormicosis rinocerebral: diagnóstico diferencial de las sinusitis fúngicas invasivas.

Mucormycosis is an opportunistic infection caused by molds Mucoraceae of the family Phycomycetes. This invasive FS is found in debilitated hosts, most frequently in patients with hematologic malignancies, burn patients and diabetes mellitus. There are several differents forms: rhino-cerebral, pulmonary or disseminated. We describe a case of invasive fungal sinusitis (FS): Rhino-orbital-cerebral mucormycosis, treated at our hospital with radical surgery and medical treatmen We reviewed the clinical presentation of the different types.

[Non invasive fungal sinusitis: mycetoma and alergical fungal sinusitis]. / Sinusitis fúngicas no invasivas: micetoma y sinusitis fúngica alérgica.

We have made a study about different cases of non-invasive fungal sinusitis (FS) treated at our hospital with surgery (endoscopic sinus approach) and medical treatment. We review two cases: Mycetoma and allergic fungal sinusitis, following the recent classification based on physiopathology, treatment and prognosis. We review the clinical presentation of the different types.

[Trans-septal endoscopic approach of pituitary tumors]. / Abordaje endoscópico transseptal de tumores hipofisarios.

The advances in endoscopic instruments have eased the approach to the sellar region through the nasal cavity. We carry out an analysis of the surgical results on 20 patients that underwent surgery for sellar tumours through a transeptal-transphenoidal approach in the last 2 years in our hospital. The average was 45.6 years old, and 75% were females. 30% of cases were pituitary adenomas and another 30% acromegaly, 25% Cushing's disease and 10% prolactinomas. No complications were encountered during surgery being the most common postoperative complications, diabetes insipida in two cases (10%) and CSF leak in one case. At present 2 patients are having hormonal treatment for panhypopituitarism. No patients developed a septal perforation, nasal deformity, epistaxis, meningitis, lip numbness or oronasal fistula. The rest did have good results noith no recurrence and hormonal values back to normal.

[Endoscopic surgery of nasopharyngeal angiofibroma]. / Cirugía endoscópica del angiofibroma nasofaríngeo.

Juvinile nasopharyngeal angiofibromas are highly vascular benign tumors but with an agressive behaviour with invasion of skull base and vital structures. Our study includes 11 male patients that underwent surgery for angiofibromas between 1992-2002 with an endoscopic surgical approach and embolization before surgery. The average age was 14.7 years old. Using Fisch staging, 8 patients were stage II. All cases achieved symptomatic remission, with no complications but the recurrence was 36.3%. The endoscopic excision is a safe and effective minimally invasive surgical treatment in patients in stage I and II. It should be considered as a first-choice option for these cases.