Screening of auramine-stained smears of all fecal samples is a rapid and inexpensive way to increase the detection of coccidial infections.

INTRODUCTION: Coccidia are important causes of diarrhea that is often indistinguishable from other forms of community-acquired diarrhea. However, the detection of oocysts is often only performed when explicitly requested, as part of the ova and parasite (O&P) examination. Reappraisal and understanding of the accurate staining characteristics of auramine O (AuO), which stains nucleic acids, may permit the inexpensive and reliable identification of coccidian oocysts at routine workup of all fecal samples. METHODS: AuO-stained smears were prepared from all stool samples received for stool culture in transport medium (SC) and from concentrated stools received for the ova and parasite (O&P) examination. RESULTS: A total of 3732 samples for stool cultures and 3132 samples for O&P examinations were included. Ninety-one samples (1.3%) from 52 patients yielded Coccidia (45 Cryptosporidium spp and 7 Isospora belli). In seven cases oocysts were only detected in samples sent for stool culture in transport medium. The oocysts showed a typical staining pattern and were easy to recognize. The observation of one smear took only around 30seconds, and the reagents and glass slide for one smear did not exceed US$ 0.03. CONCLUSIONS: The screening of all fecal samples with AuO-stained smears is a rapid and inexpensive way to increase the detection of coccidial infections, which in most laboratories can be incorporated into the microscopic workup for mycobacteria.

Growth of Mycobacterium tuberculosis in conventional BacT/ALERT FA blood culture bottles allows reliable diagnosis of Mycobacteremia.

The conventional BacT/ALERT FA blood cultures supported the ample growth of Mycobacterium tuberculosis in seeding experiments and appeared to perform as reliably as the BACTEC Myco/F-Lytic vials in the recovery of M. tuberculosis from blood in HIV-infected patients. Overall, blood cultures were positive in 39% of patients with tuberculosis.

Smear microscopy to diagnose tuberculosis early and prevent further transmission in a population with a high prevalence of HIV infection.

In western Europe, Portugal has the highest incidence of tuberculosis (TB). The quickest way to diagnose TB is with a positive smear. To evaluate the usefulness of smear microscopy in a population with a high HIV seroprevalence, TB patients admitted during the year 2000 were retrospectively analyzed. Of the 74 TB patients admitted, 45 (61%) had pulmonary TB, 11 having multiresistant (MR)-TB. Considering that only half of the patients with pulmonary TB had a positive smear, a high degree of clinical suspicion is of great importance. Appropriate isolation facilities for TB are much needed, in order to prevent nosocomial TB spread.

Current strategies to avoid misdiagnosis of malaria.

Malaria remains the most important parasitic disease, and tens of thousands of cases are imported into non-endemic countries annually. However, any single institution may see only a very few cases-this is probably the reason why laboratory and clinical misdiagnosis may not be uncommon. In the laboratory, unfamiliarity with microscopic diagnosis may be the main reason, considering the large number of laboratory staff who provide on-call services, often without expert help at hand, as well as the difficulty in detecting cases with low-level parasitemia. Staff should therefore be provided with continuing microscopic training to maintain proficiency. The complementary use of immunochromatographic rapid detection tests (RDTs) may be useful, especially during on-call hours, although, in order to ensure correct interpretation, their inherent limitations have to be well known. Diagnosis based on the polymerase chain reaction is still unsuitable for routine use, due to its long turnaround time, its cost, and its unavailability outside regular hours, although it may be helpful in selected cases. Once the alert clinician has considered the possibility of malaria, and suspicion continues to be high, malaria can be excluded by repeat smears or RDTs. However, the absence of clinical suspicion may not be infrequent, and may have more serious consequences. Depending on the local number of malaria cases seen, laboratory staff should have a low threshold for the decision to perform unsolicited malaria diagnostic tests on suspicious samples, especially if other laboratory tests are abnormal (e.g. thrombocytopenia, presence of atypical lymphocytes, or raised lactate dehydrogenase). The detection of intraleukocytic hemozoin during automated full blood counts is a promising new way to avoid misdiagnosis of clinically unsuspected malaria.

Macrolide resistance in Streptococcus pneumoniae isolated from patients with community-acquired lower respiratory tract infections in Portugal: results of a 3-year (1999-2001) multicenter surveillance study.

A nationwide multicenter study (including 31 laboratories) of the antimicrobial susceptibility of 1210 Streptococcus pneumoniae isolates from patients with community-acquired lower respiratory tract infections (LRTI) was carried out over 3 years (1999-2001) in Portugal. Testing of all isolates was undertaken in a central laboratory. Overall macrolide resistance was 13.1%. Decreased susceptibility to penicillin was 24.5% (15.5% low-level and 9.0% high-level resistance). Taken into consideration, the resistance rates reported in a previous surveillance study of 1989-1993, a six-fold increase of erythromycin resistance in the last decade was documented. Resistance to erythromycin, clarithromycin, and azithromycin was higher in pediatric patients than in adults. The overwhelming majority (82.3%) of macrolide-resistant isolates were multidrug resistant, although 44.9% were fully susceptible to penicillin. Most macrolide-resistant isolates (80.4%) showed the MLSB phenotype (76.6% MLSB-constitutive resistance, and 3.8% MLSB-inducible resistance) and were also resistant to clindamycin, tetracycline, and co-trimoxazole. The M phenotype was seen in 19.6% isolates and these had MIC90 values of 8 mg/L for erythromycin and clarithromycin, and of 12 mg/L for azithromycin. The clinical significance of macrolide resistance in the management of LRTI is discussed. Because of the specific situation concerning macrolide resistance described in S. pneumoniae, careful use of macrolide antibiotics in therapy and cautious monitoring of macrolide resistance should be continued in Portugal.

Rapid immunochromatographic malarial antigen detection unreliable for detecting Plasmodium malariae and Plasmodium ovale.

In order to determine the reliability of two commercial tests for the rapid detection of plasmodial antigen in cases of infection with Plasmodium ovale and Plasmodium malariae, the products were evaluated in four centers and a search of the relevant literature was performed. The results of the present and previous studies were compared. With overall sensitivities ranging between 18.8% and 47.6% for Plasmodium malariae and between 20% and 31.3% for Plasmodium ovale, it is evident that neither test is reliable for the detection of Plasmodium ovale and Plasmodium malariae infections.

Feasibility of introducing rejection criteria for stool cultures in a teaching hospital in Portugal.

The possible introduction of rejection criteria for stool cultures (

Automated detection of malaria pigment in white blood cells for the diagnosis of malaria in Portugal.

A novel automated method (Cell-Dyn 3500) allows malaria diagnosis by detecting malaria pigment in white blood cells during routine full blood counts. In Portugal, 174 samples from 148 patients who presented to the emergency department were analyzed. Compared with microscopy the sensitivity was 95% and the specificity was 88%. In 5 cases, false-positive Cell-Dyn 3500 results were from patients who had a recent history of treated malaria, indicating that the method may remain positive during convalescence. Six patients were diagnosed due to the changes observed with the automated method only, because clinicians had not requested malaria smears. This instrument appears to provide a promising method for the diagnosis of malaria, especially where automated full blood counts are routine in the work-up of febrile patients.

Automated malaria diagnosis using pigment detection.

Several new methods of malaria diagnosis have recently been developed, but these all rely on clinical suspicion and, consequently, an explicit clinical request. Although some methods lend themselves to automation (eg. PCR), no technique can yet be used for routine clinical automated screening. Detection of birefringent haemozoin has been used to diagnose malaria since the turn of the 20th century. A new generation of full blood count analysers, used widely in clinical laboratories, have the potential to detect haemozoin in white blood cells and probably erythrocytes. Thomas Hänscheid, Emilia Valadas and Martin Grobusch here describe this novel technique for malaria diagnosis and discuss its potential applications.

Malaria diagnosis with the haematology analyser Cell-Dyn 3500: What does the instrument detect?

The Cell-Dyn 3500 instrument could become a sensitive and specific tool in the diagnosis of malaria. The instrument appears to detect malaria-pigment within monocytes and granulocytes. A case of P. vivax malaria in a patient with increased osmotically resistant erythrocytes illustrates the potential of the instrument to detect intraerythrocytic parasites with pigment. However, in most malaria-patients with normal red cell osmotic resistance the observed phenomena seem rather to represent intraleukocytic pigment. This can remain in the circulation of clinically and parasitologically cured individuals and thus may not indicate acute infection. While the instrument can indicate those patients who have been infected a diagnosis of acute malaria must be established independently.

Diagnosis of malaria: a review of alternatives to conventional microscopy.

Malaria causes significant morbidity and mortality worldwide, including countries with mainly imported malaria. In developing nations, scarce resources lead to inadequate diagnostic procedures. In affluent countries, poor familiarity with malaria may cause clinical and laboratory misdiagnosis. Microscopy of Giemsa-stained thick and thin films remains the current standard for diagnosis. Although it has good sensitivity and allows species identification and parasite counts, it is time consuming, requires microscopical expertise and maintenance of equipment. Microscopy with fluorescent stains (QBC), dipstick antigen detection of HRP2 and pLDH (Parasight-F, ICT Malaria Pf, OptiMAL), polymerase chain reaction assays and some automated blood cell analysers offer new approaches and are reviewed here, with emphasis on clinical relevance and their potential to complement conventional microscopy, especially in countries with imported malaria.