Development of an immunoenzymatic assay using a monoclonal antibody against a 50-kDa catabolite from the P126 Plasmodium falciparum protein to the diagnosis of malaria infection

The WHO criterion of defering any donation of blood by a confirmed case of malaria for three years after cessation of therapy can not be applied in areas where malaria in endemic. For this reason we developed an immunoenzymatic assay for the detection of plasmodial antigens for blood screening in malararial endemic areas. So, we tested sera from 191 individuals. Among patients with active disease 100% of the cases of Plasmodium falciparum or mixed infections and 91.7% of those with P. vivax were positive for the presence of plasmodial antigens. The lower parasitaemia detected was 0.0003% for P. vivax malária. When the frequency of positive circulating malarial antigens was evaluated among asymptomatic and symptomatic individuals with negative TBS, positive results were found in respectively 38.7% and 17.7% of the individuals studied in the 30 days after confirmed malaria attack. Data provide by these assays have shown that ELISA seemed to be more sensitive than parasitological examination for malaria diagnosis. This test by virtue of its high sensivity and the facilities in processing a large number of specimens, can prove to be useful in endemic areas for the recognition of asymptomatic malaria and screening of blood donors.

Non-immunologic methods of diagnosis of babesiosis

The diagnosis of tick-borne diseases such as babesiosis still depends on observing the parasite in the infected erythrocyte. Microscopic observation is tedious and often problematic in both early and carrier infections. Better diagnostic methods are needed to prevent clinical disease, especially when susceptible cattle are being moved into disease enzootic areas. This study evaluates two techniques for early diagnosis of Babesia bovis infections in cattle, DNA probes specific for the organism and fluorescent probes specific nucleic acid. The radioisotopically labeled DNA probes are used in slot blot hybridizations whith lysed blood samples, not purified DNA. Thusfar, the probe is specific for B. bovis and can detect as few as 1000 B. bovis parasites in 10µl of blood. The specificity of the fluorescent probe depends on the characteristic morphology of the babesia in whole blood samples, as determined microscopically. The fluorescent probe detects as afew as 10,000 B. bovis parasites in 10*l as blood. The application of each method for alboratory and field use is discussed.

New methods for the diagnosis of Babesia bigemina infection

Accurate diagnosis of Babesia bigemina infection, an economically important tick-transmitted protozoan parasite of cattle, is essential in the management of disease control and in epidemiological studies. The currentlyused methods of diagnosis are blood smear examination and serological tests which include agglutination and immunofluorescence tests. These testes have been used the fild but because they lack sensitivity and specificity, never and improved methods of diagnosis are being developed. The quantitative buffy coat (OBC) method, using microhaematocrit tubes and acridine orange staining allows rapid and quicker diagnosis of B. bigemina and other blood parasites compared to light microscopic examination of stained smears. Parasite specific monoclonal antibodies have been used in antigen/antibody capture enzymelinked immunosorbent assays with grater sensitivity and specificity than previously described serological tests. Similary, DNA probes, derived from a repetitive sequence of the B. bigemina genome, offer a method of detecting very small numbers of parasites which are undetectable by conventional microscopy. An extrachromosomal DNA element, present in all the tick-borne protozoan parasites so far tested, provides an accurate means of diferentiating mixed parasite populations in infected animals. These improved methods will greatly facilitate epidemiological studies.

Comparative cost-effectiveness analyses of cardiovascular magnetic resonance and coronary angiography combined with fractional flow reserve for the diagnosis of coronary artery disease.

BACKGROUND: According to recent guidelines, patients with coronary artery disease (CAD) should undergo revascularization if significant myocardial ischemia is present. Both, cardiovascular magnetic resonance (CMR) and fractional flow reserve (FFR) allow for a reliable ischemia assessment and in combination with anatomical information provided by invasive coronary angiography (CXA), such a work-up sets the basis for a decision to revascularize or not. The cost-effectiveness ratio of these two strategies is compared. METHODS: Strategy 1) CMR to assess ischemia followed by CXA in ischemia-positive patients (CMR + CXA), Strategy 2) CXA followed by FFR in angiographically positive stenoses (CXA + FFR). The costs, evaluated from the third party payer perspective in Switzerland, Germany, the United Kingdom (UK), and the United States (US), included public prices of the different outpatient procedures and costs induced by procedural complications and by diagnostic errors. The effectiveness criterion was the correct identification of hemodynamically significant coronary lesion(s) (= significant CAD) complemented by full anatomical information. Test performances were derived from the published literature. Cost-effectiveness ratios for both strategies were compared for hypothetical cohorts with different pretest likelihood of significant CAD. RESULTS: CMR + CXA and CXA + FFR were equally cost-effective at a pretest likelihood of CAD of 62% in Switzerland, 65% in Germany, 83% in the UK, and 82% in the US with costs of CHF 5'794, euro 1'517, £ 2'680, and $ 2'179 per patient correctly diagnosed. Below these thresholds, CMR + CXA showed lower costs per patient correctly diagnosed than CXA + FFR. CONCLUSIONS: The CMR + CXA strategy is more cost-effective than CXA + FFR below a CAD prevalence of 62%, 65%, 83%, and 82% for the Swiss, the German, the UK, and the US health care systems, respectively. These findings may help to optimize resource utilization in the diagnosis of CAD.

Parasitological diagnosis of schistosomiasis mansoni: fecal examination and rectal biopsy

Even with all progress in the search of sensitive and methods for the immunological diagnosis of schistosomiasis, the microscopic detection of eggs of the parasite in the stool still remains the most widely used tool for the actual diagnosis of active infection. Among the coproscopic methods, Kato's technic modified by Katz et al (Kato/Katz) has the advantages of higher sensitivity, the possibility of egg quantification, its low operational cost and its feasibility in areas with minimal infra-structure. The oorgram of the rectal mucosa is valuable in initial clinical trials of schistosomicides, when it is needed to observe egg morphology in tissue. It could be an alternative method for individual diagnosis, being more sensitive than a single stool exam in low intensity infection. However, the increased sensitivity of a higher number of fecal exams makes that invasiveprocedure unnecessary. In the assessment of cure of schistosomiasis, Kato/Katz method (three fecal samples in one, three and six months after treatment) and the rectal biopsy four months after treatment, are equally reliable.

A web-based pilot study of inter-pathologist reproducibility using the ISHLT 2004 working formulation for biopsy diagnosis of cardiac allograft rejection: The European experience.

BACKGROUND: The aim of this study was to assess, at the European level and using digital technology, the inter-pathologist reproducibility of the ISHLT 2004 system and to compare it with the 1990 system We also assessed the reproducibility of the morphologic criteria for diagnosis of antibody-mediated rejection detailed in the 2004 grading system. METHODS: The hematoxylin-eosin-stained sections of 20 sets of endomyocardial biopsies were pre-selected and graded by two pathologists (A.A. and M.B.) and digitized using a telepathology digital pathology system (Aperio ImageScope System; for details refer to http://aperio.com/). Their diagnoses were considered the index diagnoses, which covered all grades of acute cellular rejection (ACR), early ischemic lesions, Quilty lesions, late ischemic lesions and (in the 2005 system) antibody-mediated rejection (AMR). Eighteen pathologists from 16 heart transplant centers in 7 European countries participated in the study. Inter-observer reproducibility was assessed using Fleiss's kappa and Krippendorff's alpha statistics. RESULTS: The combined kappa value of all grades diagnosed by all 18 pathologists was 0.31 for the 1990 grading system and 0.39 for the 2005 grading system, with alpha statistics at 0.57 and 0.55, respectively. Kappa values by grade for 1990/2005, respectively, were: 0 = 0.52/0.51; 1A/1R = 0.24/0.36; 1B = 0.15; 2 = 0.13; 3A/2R = 0.29/0.29; 3B/3R = 0.13/0.23; and 4 = 0.18. For the 2 cases of AMR, 6 of 18 pathologists correctly suspected AMR on the hematoxylin-eosin slides, whereas, in each of 17 of the 18 AMR-negative cases a small percentage of pathologists (range 5% to 33%) overinterpreted the findings as suggestive for AMR. CONCLUSIONS: Reproducibility studies of cardiac biopsies by pathologists in different centers at the international level were feasible using digitized slides rather than conventional histology glass slides. There was a small improvement in interobserver agreement between pathologists of different European centers when moving from the 1990 ISHLT classification to the "new" 2005 ISHLT classification. Morphologic suspicion of AMR in the 2004 system on hematoxylin-eosin-stained slides only was poor, highlighting the need for better standardization of morphologic criteria for AMR. Ongoing educational programs are needed to ensure standardization of diagnosis of both acute cellular and antibody-mediated rejection.

A new, automated, four-colour interphase FISH approach for the simultaneous detection of specific aneuploidies of diagnosis and prognosis significance in high hyperdiploid ALL

In hyperdiploid acute lymphoblastic leukaemia (ALL), the simultaneous occurrence of specific aneuploidies confers a more favourable outcome than hyperdiploidy alone. Interphase (I) FISH complements conventional cytogenetics (CC) through its sensitivity and ability to detect chromosome aberrations in non-dividing cells. To overcome the limits of manual I-FISH, we developed an automated four-colour I-FISH approach and assessed its ability to detect concurrent aneuploidies in ALL. I-FISH was performed using centromeric probes for chromosomes 4, 6, 10 and 17. Parameters established for automatic nucleus selection and signal detection were evaluated (3 controls). Cut-off values were determined (10 controls, 1000 nuclei/case). Combinations of aneuploidies were considered relevant when each aneuploidy was individually significant. Results obtained in 10 ALL patients (1500 nuclei/patient) were compared with those by CC. Various combinations of aneuploidies were identified. All clones detected by CC were observed by I-FISH. I-FISH revealed numerous additional abnormal clones, ranging between 0.1 % and 31.6%, based on the large number of nuclei evaluated. Four-colour automated I-FISH permits the identification of concurrent aneuploidies of prognostic significance in hyperdiploid ALL. Large numbers of cells can be analysed rapidly by this method. Owing to its high sensitivity, the method provides a powerful tool for the detection of small abnormal clones at diagnosis and during follow up. Compared to CC, it generates a more detailed cytogenetic picture, the biological and clinical significance of which merits further evaluation. Once optimised for a given set of probes, the system can be easily adapted for other probe combinations.

Prenatal diagnosis of facial clefts : evaluation of a specialised counseling

Résumé: Questions étudiées / principes: Le diagnostic prénatal des fentes faciales et palatines a soulevé maintes questions concernant la meilleure manière d'informer les parents et d'offrir une aide appropriée durant les semaines de grossesse restant, afin de faire de la naissance une expérience positive. Le conseil prénatal diffère selon la personne qui en est responsable et ses connaissances concernant les aspects pratiques des soins ainsi que l'impact de cette malformation sur la qualité de vie de l'enfant. Le but de cette étude est d'évaluer nos techniques actuelles et de les modifier de façon appropriée. Méthodes: 29 couples ayant bénéficié d'un diagnostic prénatal ont été priés de répondre rétrospectivement à un questionnaire. L'histoire obstétrique personnelle ainsi que l'histoire familiale, le déroulement des évènements, la qualité de l'information fournie et leurs impressions à la naissance ont été considérées. Résultats: Les parents utilisent de multiples sources d'information: l'obstétricien, le conseil du chirurgien pédiatrique au sein de I « équipe fente », les groupes de soutien de parents et Internet. 93% des parents se sont sentis bien préparés à la naissance de leur enfant, autant psychologiquement que concernant les aspects pratiques des soins. 54% ont été soulagés de voir leur enfant moins affecté qu'ils ne l'imaginaient. Finalement, 96% considèrent que le diagnostic prénatal est un bénéfice. Conclusion: Une information de base devrait être apportée rapidement après l'ultrason confirmant le diagnostic, suivie de discussions techniques progressivement plus détaillées durant le temps de grossesse restant. Summary: Questions under study / principles: Prenatal diagnosis of cleft lip and palate has raised many questions concerning the best way to inform the parents and offer appropriate support during the remaining pregnancy to help prepare a positive birth experience. Prenatal counseling differs according to whose is responsible and that person's knowledge of practical aspects of care and the impact of the malformation on the child's quality of life. The aim of the study was to evaluate our current techniques and modify them when appropriate. Methods: 29 couples having experienced prenatal diagnosis were asked to respond retrospectively to a questionnaire. Personal obstetric and family history, timing and quality of information provided and their impressions at birth were considered. Results: The parents used several sources of information: the obstetrician, the counsel by the cleft team surgeon, the parents support groups and Internet. 93% of the parents felt well prepared for the birth of their child psychologically and concerning practical aspects of care. 54% felt relieved that their child was less affected than imagined. 96% considered prenatal diagnosis a benefit. Conclusion: Basic information should be provided soon after ultrasound confirmation, followed by progressively more detailed technical discussions over the remaining pregnancy.

Prenatal diagnosis of congenital lung malformations.

Prenatal diagnosis of congenital lung anomalies has increased in recent years as imaging methods have benefitted from technical improvements. The purpose of this pictorial essay is to illustrate typical imaging findings of a wide spectrum of congenital lung anomalies on prenatal US and MRI. Moreover, we propose an algorithm based on imaging findings to facilitate the differential diagnosis, and suggest a follow-up algorithm during pregnancy and in the immediate postnatal period.

The use and limitations of ultrasonography in the diagnosis of liver morbidity attributable to Schistosoma mansoni infection in community-based surveys

The objective of this population-based study was to estimate the liver morbidity attributable to Schistosoma mansoni infection by ultrasonography adopting the proposed standard protocols of the Cairo Meeting on Ultrasonography, 1991. We examined 2384 individuals representing 20 of the households of the rural population of the Ismailia Governorate, East of Delta, Egypt. Prevalence of S. mansoni and S. haematobium infections were 40.3 and 1.7 respectively. Portal tract thickening (PTT) grade 1, 2 and 3 considered diagnostic of schistosomal liver morbidity was detected in 35.1, 1.3 and 0.2 individuals respectively. Generally, ultrasonographically-detected pathological changes increased with age, but correlated with intensity of infection only in age group 20-59 years. Comparing individuals with and without S. mansoni infections in an endemic and a non-endemic community indicated no significant difference between the former and the latter in either case. In conclusion: ultrasonography had a limited value in estimating schistosomal liver morbidity in our population-based study where early grades of liver morbidly were prevalent. The criteria of diagnosing grade I portal fibrosis need to be revised as well as the staging system proposed by the Cairo Meeting on ultrasonography in schistosomiasis.

Diagnosis of malaria by acridine orange fluorescent microscopy in an endemic area of Venezuela

Fluorescent (acridine orange) microscopical examination of capillary centrifuged blood (quantitative buffy coat [QBC®] analysis) and Giemsa stained thick blood smears (GTS) were compared for diagnosis of malaria in blood specimens from adults living in malaria transmission areas of the States of Bolivar and Amazonas in southeastern and south Venezuela, respectively. Of a total of 198 GTS examined, 95 subjects (48%) showed parasitaemia. Among the 95 blood films with a positive GTS, 94 were judged positive by the QBC. However, positive QBC tubes were found in 29 out of 103 blood specimens with a negative GTS. Thus, relative to a GTS standard, the sensitivity and specificity of the QBC-test was 99.2% and 72%, respectively. Young trophozoites of Plasmodium vivax and P. falciparum could not be distinguished with certainty. It is confirmed that the QBC offers many advantages compared with the standard diagnosis of malaria parasites, specifically in the speed of staining and ease of interpretation. However, in places where P. falciparum and P. vivax occur, species and stage differentiation should be confirmed with the GTS.