High incidence of Pneumocystis jirovecii pneumonia in oncological patients: a 19-year study.

AIM: In the past, Pneumocystis jirovecii belonged to the Protozoa group, but is currently taxonomically included in the kingdom Fungi. P. jirovecii is an opportunistic pathogen, responsible for pneumocystis pneumonia with frequent complications of immunocompromised patients. Delayed initiation of appropriate therapy increases the risk of death in immunocompromised patient. The aim of this work was to determine and evaluate the reliability of methods of laboratory diagnosis of pneumocystosis used in routine laboratories as well as the occurrence of this disease in patients from Slovakia during 19 years. MATERIAL AND METHODS: The diagnosis is based on microscopic examination (Giemsa- and Gram-Weigert-staining) and detection of parasite DNA by classical or real-time PCR in bronchoalveolar lavage and sputum. RESULTS: Pneumocysts were detected in 190 persons (5.7%) from the whole group of patients. Cancer patients represented the riskiest group in terms of pneumocystosis, which was confirmed by the highest percentage (57.9%) of individuals infected with P. jirovecii. Compared with the PCR, 33.7% sensitivity and 100% specificity of microscopy was calculated by using a binary classification test. Molecular methods are more sensitive in the detection of P. jirovecii compared to microscopic evidence and currently represent a reliable detection system in the diagnosis of pneumocystosis. CONCLUSION: In view of the increasing number of immunocompromised persons, diagnostics of P. jirovecii in patients with pulmonary complications is essential. This was also confirmed in our study, where the number of examinations and detection of this opportunistic pathogen increased over the years.

Unusual Toxoplasma infection of the eye and central nervous system in an HIV-positive patient.

OBJECTIVE: To report on a unique combination of cerebral toxoplasmosis and ocular toxoplasmosis in an HIV-positive patient in Slovakia. METHODS: A 35-year-old heterosexual patient who presented with headache and major seizures underwent computed tomography (CT) and magnetic resonance imaging (MRI). Based on clinical findings, serological tests for toxoplasmosis were performed on serum and ocular fluid specimens. PCR was also used to detect Toxoplasma gondii and cytomegalovirus DNA. Goldmann and Witmer coefficient calculation was applied to demonstrate the synthesis of intraocular IgG antibodies. RESULTS: CT and MRI revealed cystic lesions suspected of metastasis in the occipital and temporal regions, and we searched for the primary tumor. After vision loss in the left eye, which rapidly progressed to complete blindness, an eye examination detected macular edema. Anti-edema treatment was initiated. HIV positivity with a very low CD4 T-cell count (20/μL) was found, and the viral load was 100 400 HIV-RNA copies/ml. The serum was positive for anti-Toxoplasma IgG antibodies (> 200 IU/mL), IgM negative, and IgA borderline. As toxoplasmic encephalitis and retinitis were suspected, antitoxoplasmic therapy with pyrimethamine, spiramycin, and folinic acid was started. The ophthalmologist considered cytomegalovirus retinitis, which was not confirmed by serology or PCR. In contrast, the presence of IgG antibodies in ocular fluid and serum with the calculation of the Goldmann-Witmer coefficient (GW = 32) as well as PCR DNA positivity pointed to Toxoplasma gondii as the etiological agent. Follow-up MRI scan confirmed regression of the pathological lesions, neurological deficit also improved, CD4 T-lymphocytes increased above 200/μL, but blindness of the left eye persisted. CONCLUSION: CT and MRI scans offered no clue as to Toxoplasma etiology of the brain and eye involvement in an HIV-positive patient, which was only confirmed by laboratory tests. Due to the delay in the diagnosis of toxoplasmosis, time from the epileptic seizure to treatment initiation was 16 days, which assumedly caused irreversible blindness in the patient.

Imported new world cutaneous leishmaniasis in a traveller from Slovakia.

We present a case of imported leishmaniasis in a 31-year-old woman from Slovakia, who visited the countries of South America for three months in 2011. On 29 and 31 August 2011, she was probably infected with Leishmania parasites in the jungles of Ecuador. Approximately one week after returning to Slovakia, a small papules appeared on patient's left leg. Another wound was found after two weeks. Both ulcers were enlarging. We proved amastigote forms of Leishmania spp. only in repeated dermal scrapings from the edge of the ulcer by Giemsa staining after negative results from examination of a wound scrape and biopsy specimen. We identified the species Leishmania (Viannia) panamensis as a causative agent by using the polymerase chain reaction (PCR) method and subsequent sequencing of the ITS region. Closure of wounds and scab formation were observed after 20 days of treatment with sodium stibogluconate. In the control microscopic examination after the end of the treatment, parasites were not present, and the PCR confirmed the negative result (Fig. 2, Ref. 31).

Histological and molecular confirmation of the fourth human case caused by Dirofilaria repens in a new endemic region of Slovakia.

Dirofilariosis is considered to be the arthropod vector-borne disease with the fastest spread in Europe. Slovakia belongs to new endemic regions for canine and human infections. This paper reports the fourth human case, where diagnosis was confirmed using computed tomography (CT), histological and molecular examinations - for the first time in this endemic region. The epidemiological history of this case indicated the autochthonous origin, and infection manifested by intense swelling in the periocular region, where a subcutaneous nodule was localized. Microscopic analysis of cross-sections of the surgically removed nodule confirmed the presence of a single male worm of Dirofilaria repens, indicated by the external cuticular ridges, the robust muscle cells and a single male sex organ seen as one tube beside the intestine. Considering that the worm morphology was partially damaged, molecular study was performed using DNA isolated from formalin-fixed and paraffin-embedded tissue sections of the worm. In polymerase chain reactions (PCR) a set of primers specific for D. repens DNA for the CO1-encoding gene amplified the expected 246 bp product using a high concentration of DNA template. Our diagnostic approach, involving molecular techniques, showed that identification of D. repens from excised tissues is possible even when the morphology of the worm and DNA are damaged during tissue processing. It is expected that the spread of this disease will continue due to climatic changes in central Europe. This warrants higher awareness among clinicians, who will initially be approached by patients, and better co-operation with parasitologists in newly endemic countries.

Weak cytotoxic activity of miltefosine against clinical isolates of Acanthamoeba spp.

Hexadecylphosphocholine (miltefosine) is an anticancer drug active in vitro against various protozoan parasites, and recently used for the treatment of disseminated Acanthamoeba infection. In the present study, we present results of weak cytotoxic activity of this potential amoebicidal agent for 2 of 3 clinical isolates of Acanthamoeba spp. Although the inhibition effect for all tested concentrations was apparent, and showed 100% eradication of trophozoites of Acanthamoeba castellanii strain at a concentration of 62.5 µM after 24 hr, the strains Acanthamoeba sp. and Acanthamoeba lugdunensis exhibited low sensitivity to hexadecylphosphocholine, even in high concentrations. The determined minimal trophocidal concentrations were 250 µM for Acanthamoeba sp. and 500 µM for A. lugdunensis after 24 hr of exposure. Although hexadecylphosphocholine is a potential agent for treatment of Acanthamoeba keratitis and systemic infections, in clinical practice the possible insusceptibility of the amoebic strain should be considered for optimizing therapy.

[Seroprevalence of Toxoplasma gondii among pregnant women in Slovakia]. / Séroprevalencia Toxoplasma gondii u gravidných zien na Slovensku.

Toxoplasmosis is caused by a protozoan parasite, Toxoplasma gondii. Primary infection with Toxoplasma gondii in pregnant women can lead to adverse outcomes. Since initially asymptomatic, toxoplasmosis is difficult to diagnose. The aims of this study were to determine the seroprevalence of toxoplasmosis in pregnant women in Slovakia and to identify possible risk factors. As many as 145 (22.1%) of 656 pregnant women screened for T. gondii were seropositive. The prevalence of toxoplasmosis significantly increases with age and the highest seropositivity rate (35.4 %) was found in the age group of 35-44 years. A significantly higher seropositivity rate was observed in women living in the surroundings of Bratislava (63.5%) as compared to Bratislava (36.6%, p < 0.001; OR = 0.53, 95% CI = 0.36-0.79) Among the epidemiological factors, contact with soil appeared statistically significant (35.3%, p = 0.014; OR = 1.67, 95% CI = 1.09-2.56). Fatigue (32.1%, p = 0.026; OR = 1.84, 95% CI = 1.03-3.27), miscarriages (44.1%, p = 0.002; OR = 2.22, 95% CI=1.3-3.8) and toxoplasmosis in the family (10.7%, p = 0.002) were most commonly reported in history of the study subjects.

The significance of complement fixation test in clinical diagnosis of toxoplasmosis.

BACKGROUND: The complement fixation test (CFT) is the basic method in the diagnosis of toxoplasmosis. Despite its standard and reproducible results, it is rarely used in routine diagnosis of toxoplasmosis where the detection of IgG by means of EIA tests is widely used. OBJECTIVE: The study is focused on the verification of CFT value within the spectrum of specific classes of immunoglobulins IgG, IgM, IgA antibodies and avidity of IgG antibodies, as well as on the clinical diagnosis of toxoplasmosis. METHOD: The study analysed 1705 samples of serum from patients suspected to be infected by Toxoplasma gondii. Out of these patients 451 suffered from acute lymphadenopathic toxoplasmosis (6 with seroconversion of both CFT and IgG antibodies) and 1254 serum samples were collected from patients with latent infection, out of whom 176 were with long-term persistence of IgM, or IgA antibodies after the elimination of disease. CONCLUSION: CFT is a reliable indicator of Toxoplasma infection and as opposed to IgG antibodies its levels enable a more objective determination of its course. However from the aspect of the determination of the stage of toxoplasmosis from a single sample of serum, the combination of CFT with IgM or IgA antibodies with quantitative order or test of avidity of IgG antibodies must be performed. (Tab. 4, Fig. 4, Ref. 30.).

[Diagnosis of toxoplasmosis and toxocariasis using fluid from the anterior ocular chamber]. / Diagnostika toxoplazmózy a toxokarózy z moku prednej ocnej komory.

The authors discuss methods of improvement of the etiologic diagnosis of the toxoplasma and toxocara associated posterior uveititis, by examining specific antibodies in the aqueous humor. Paired samples of aqueous humor and serum were analyzed in 23 patients. Chorioretinitis of infectious etiology was suspected in 15 patients, 3 patients were treated for panuveitis, 4 patients for intermediate uveitis, and in 1 patient the sample was obtained to rule out malignant melanoma. The local ocular antibody production was calculated according to Desmonts. Toxoplasma etiology was confirmed by local antibody production analysis in 1 patient, toxocara etiology in 5 patients. In 20 patients in the control group local specific antibody production was not demonstrated.

[Acanthamoeba keratitis after use of soft contact lenses--case report]. / Acanthamoebová keratitis po aplikácii mäkkých kontaktných sosoviek--kazuistika.

The case history of a 39-year patient suffering from a deep inflammation of cornea and not responding to conventional antibiotic treatment is presented. The patient was using soft contact lenses during the period of initial symptoms; moreover, he was bathing in thermal bathing pool. A cultivation examination of smears from the area of corneal defect revealed the presence of Acanthamoeba lugdunensis in combination with bacterial infection by Pseudomonas aeruginosa. The available data indicate that it is the first case of acanthamoeba karatitis (AK) after the application of contact lenses in Slovakia. A long-term local treatment with propamidin isethionate (Brolene gtt, ung.) resulted in healing up. The subsequent vision after 16 months since the initial symptoms proved to be 6/12 (0.5). The healing of the centrally localized defect changed the curvature of cornea with consequent hypermetropic shift. The defect completely corrected the patient's myopia (-8.5). The paper describes present possibilities of AK therapy.

Trichinellosis in the Slovak Republic.

The occurrence of trichinellosis in farm or wildlife animals in some areas pose the permanent risk of infection in humans. In Slovakia, where only the sylvatic trichinellosis exists, the humans trichinellosis is sporadic. The most common cause of infection is wild boar meat. Unusual local eating customs may also favour an outbreak of disease in humans. The outbreak of trichinellosis in Slovakia in 1998, when 336 people were affected, was of the same kind. Trichinella britovi was the agent of the infection.

[Laboratory diagnosis of toxoplasmosis]. / Postupy v laboratórnej diagnostike toxoplazmózy.

Toxoplasmosis is a world-wide spread parasitosis. The disease potentially highly affects two groups of patients: foetus and immunosuppressed patients. The determination of diagnosis and therapy on the basis of a single serum examination is very important; possible on the basis of a single serum sample. In most cases, it is possible to differentiate between recent and latent infections using a combination of suitable methods, which permit us to confirm particular antibody classes. In the presented paper the authors suggest diagnostic procedures for 4 groups of patients: pregnant, neonates with suspected congenital toxoplasmosis, immunodeficient patients and immunocompetent patients. The diagnostic methods consist of a combination of basic and supplemented diagnostic methods. Each patient's serum should be tested by basic tests which include the detection of total antibodies with CFT or IFT and specific classes of IgM and IgG antibodies by ELISA. The potential activity of toxoplasma infection can be determined by supplementary methods of e.g. IgG avidity antibodies, establishment of IgA antibodies, western blotting method and monitoring of antibodies production. For each situation the authors present interpretations of suspected cases including proposals for clinicians. These procedures are suggested for practical use in laboratories of various diagnostic levels in order to help to the diagnostic procedures in a particular situation as well as for clinical evaluation of established results. (Fig. 4, Ref. 65).

[A rare case of human autochthonous dicrocoeliasis in Czechoslovakia]. / Ojedinelý prípad humánnej autochtónnej dikroceliózy v CSSR.

The first case of not imported dicroceliasis in Czechoslovakia is reported in an eleven-year-old boy. The patient suffered from gastrointestinal complaints for about half a year. Diarrhea frequently alternated with constipation, the abdominal pain was mainly in the region of the pancreas. With the exception of eosinophilia (16%), all other laboratory findings (blood count, ELFO, urinalysis) were within normal values. Eggs of Dicrocoelium dendriticum were repeatedly found in stool specimens. The patient was successfully treated with Bithin (4,6-dichlorpentol). Examination of the other members of the family proved negative. At present the patient is without complaints, stool examinations for parasites were repeatedly negative. Contact with sheep or consumption of contaminated liver was not demonstrated. The parasitic infestation may have occurred via an intermediate host, as the patient was keen on collecting different animals, particularly snails.